Kainate Receptors in Health and Disease

Neuron

Review Kainate Receptors in Health and Disease Juan Lerma1,* and Joana M. Marques1 1Instituto de Neurociencias, CSIC-UMH, San Juan de Alicante, 03550 Spain *Correspondence: [email protected] http://dx.doi.org/10.1016/j.neuron.2013.09.045

Our understanding of the molecular properties of kainate receptors and their involvement in synaptic physiology has progressed significantly over the last 30 years. A plethora of studies indicate that kainate receptors are important mediators of the pre- and postsynaptic actions of glutamate, although the mechanisms underlying such effects are still often a topic for discussion. Three clear fields related to their behavior have emerged: there are a number of interacting proteins that pace the properties of kainate receptors; their activity is unconventional since they can also signal through G proteins, behaving like metabotropic receptors; they seem to be linked to some devastating brain diseases. Despite the significant progress in their importance in brain function, kainate receptors remain somewhat puzzling. Here we examine discoveries linking these receptors to physiology and their probable implications in disease, in particular mood disorders, and propose some ideas to obtain a deeper understanding of these intriguing proteins. A Historical Overview Most excitatory synapses in the brain use the amino acid glutamate as a neurotransmitter. Since the excitatory properties of glutamate were postulated nearly 40 years ago, an extraordinary wealth of data has accumulated on the types of synaptic responses triggered by this neurotransmitter. Glutamate acts on a variety of receptor proteins, initially classified by the mechanisms that they use to transmit signals (i.e., metabotropic versus ionotropic). A more precise specification of ionotropic receptors into three types was subsequently proposed, based on the agonist that activates or binds to them. Thus, AMPA, kainate, and NMDA receptors (AMPARs, KARs, and NMDARs, respectively) are recognized as the main effectors of glutamate at synapses. We now know that this classification is misleading, since there is certain cross-reactivity between agonists and receptors and only recently have some new compounds enriched the pharmacological armamentarium (see Jane et al., 2009 for a review). Unlike other receptors, studies of KARs suffered from the lack of specific compounds to activate or block these proteins. First of all, kainate is derived from the seaweed known as ‘‘kaininso’’ in Japanese, and it is a mixed agonist that can also activate AMPARs. This fact led to certain misinterpretations of the role of KARs in the brain and, even nowadays, some related errors can be detected in the literature. In addition, the prototypical AMPAR agonist, AMPA, can also activate diverse KARs. Like the AMPARs and NMDARs, KARs are tetrameric combinations of a number of subunits: named GluK1, GluK2, GluK3, GluK4, and GluK5 (previously known as GluR5–GluR7 and KA1 and KA2, respectively). Of these, GluK1–GluK3 may form functional homomeric or heteromeric receptors, while GluK4 and GluK5 only participate in functional receptors when partnering any of the GluK1–GluK3 subunits. The structural repertoire of KAR subtypes is further extended by editing of the GluK1 and GluK2 receptor subunit pre-mRNAs at the so-called Q/R site of the second membrane domain. More isoforms also arise from the alternative splicing of GluK1–GluK3 subunits, while GluK4 and GluK5 seem not to be subjected to this type of processing. 292 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

The absence of specific antibodies against different KAR subunits has been a significant limitation in terms of exploring receptor distribution. Thus, most of the information regarding their tissue expression comes from in situ hybridization studies that, although informative, cannot reveal the subcellular distribution of a given subunit. Relatively good and specific antisera against the KAR subunits GluK2/3 and GluK5 are now available, although not all work properly in immunocytochemistry. Nevertheless, some general rules could be extracted from all these studies. GluK2 subunits are mostly expressed by principal cells (hippocampal pyramidal cells; both hippocampal and cerebellar granule cells; cortical pyramidal cells), while GluK1 is mainly present in hippocampal and cortical interneurons (Paternain et al., 2003) as well as in Purkinje cells and sensory neurons. GluK3 is poorly expressed, appearing in layer IV of the neocortex and dentate gyrus in the hippocampus (Wisden and Seeburg, 1993). GluK4 is mainly expressed in CA3 pyramidal neurons, dentate gyrus, neocortex, and Purkinje cells, while GluK5 is expressed abundantly in the brain (Bahn et al., 1994). The functional description of KARs within the CNS (Lerma et al., 1993) and the molecular identification of KAR subunits represented real breakthroughs in the study of these receptors, as did the discovery that GYKI53655, a 2,3, benzodiazepine, was essentially inactive at KARs (Paternain et al., 1995; Wilding and Huettner, 1995) (with the exception of a few particular assemblies on which it may act at high concentrations; see Perrais et al., 2009), and constitute the foundation upon which our understanding of KARs has been constructed. On the basis of the data collected over the last 30 years of research, how do we now envisage the physiological role of KARs? A comprehensive analysis of the profuse yet often controversial literature on KARs leads us to conclude that these receptors play significant roles in the brain at three main levels. In the first place, they mediate postsynaptic depolarization and they are responsible for carrying some of the synaptic current, although this only happens at some synapses. Second, KARs can modulate the synaptic release of neurotransmitters such

Neuron

Review Table 1. The Kainate Receptor Interactome Interactor

KAR Subunit

Direct

Role

Actinfilin

GluK2

Yes

Receptor degradation through ubiquitination

References Salinas et al., 2006

b-catenin

GluK2

No

Plasma membrane dynamics

Coussen et al., 2005

Cadherin

GluK2

ND

Receptor trafficking/subcellular localization

Coussen et al., 2005

Calcineurin

GluK2

Yes

Ca2+-regulation of channel function

Coussen et al., 2005

Calmodulin

GluK2

Yes

ND

Coussen et al., 2005 Coussen et al., 2005

Contactin

GluK2

Yes

ND

COPI

GluK5

Yes

Receptor trafficking

Vivithanaporn et al., 2006

Dynamin-1

GluK2

Yes

ND

Coussen et al., 2005 Coussen et al., 2005

Dynamitin

GluK2

Yes

ND

14.3.3

GluK1, GluK2, GluK5

Yes

Receptor trafficking

Coussen et al., 2005; Vivithanaporn et al., 2006

4.1N

GluK1, GluK2

Yes

Receptor trafficking

Copits and Swanson, 2013b Hirbec et al., 2003

GRIP

GluK2, K5

Yes

Receptor trafficking

KRIP6

GluK2

Yes

Receptor gating

Laezza et al., 2007

NETO1

GluK1-3

Yes

Ion channel function

Zhang et al., 2009; Copits et al., 2011; Straub et al., 2011a; Tang et al., 2011

NETO2

GluK1-3

Yes

Ion channel function

Zhang et al., 2009; Copits et al., 2011

NSF

GluK2

Yes

ND

Coussen et al., 2005

PICK1

GluK2, GluK5

Yes

Receptor trafficking

Hirbec et al., 2003

Profillin II

GluK2

Yes

Receptor trafficking

Coussen et al., 2005; Mondin et al., 2010

PSD95

GluK2, GluK5

Yes

Alters receptor function by reducing desensitization

Garcia et al., 1998

SAP102

GluK2, GluK5

Yes

Receptor clustering;

Garcia et al., 1998

SAP90

GluK2, GluK5

Yes

Receptor clustering; modulation of desensitization

Garcia et al., 1998

SAP97

GluK2

Yes

Receptor clustering

Garcia et al., 1998

SNAP25

GluK5

ND

Receptor trafficking

Selak et al., 2009

Spectrin

GluK2

Yes

ND

Coussen et al., 2005

Syntenin

GluK1, GluK2

Yes

Plasma membrane dynamics

Hirbec et al., 2003

VILIP-1

GluK2

Yes

ND

Coussen et al., 2005

VILIP-3

GluK2

Yes

ND

Coussen et al., 2005

ND, not determined.

as GABA and glutamate at different sites. Third, they play an influential role in the maturation of neural circuits during development. These roles are frequently fulfilled in an unconventional way given that KARs can signal by activating a G protein, behaving more like a metabotropic receptor than an ion channel. This noncanonical signaling is totally unexpected considering that the three iGluRs share a common molecular design, as recently revealed by their crystal structure (Mayer, 2005; Furukawa et al., 2005; Gouaux, 2004). It is difficult to do justice to the literature generated on KARs over the years in the short space available, and indeed, there are several reviews that have described many of the molecular, biophysical, pharmacological, and functional aspects of these receptors (Rodrigues and Lerma, 2012; Contractor et al., 2011; Lerma et al., 2001, Lerma, 2003, 2006; Copits and Swanson, 2012; Vincent and Mulle, 2009; Coussen and Mulle, 2006; Pinheiro and Mulle, 2006; Tomita and Castillo, 2012; Jaskolski et al., 2005; Matute, 2011). Hence, in this Review we will focus primarily on the data that have influenced our notion of KAR func-

tion and the wealth of new data available implicating KARs in brain pathology. The Explosion of KAR Interacting Proteins To date, and like many other receptors and channels, a whole set of proteins have been identified that can interact with KAR subunits (Table 1). Indeed, the identification of these proteins has changed our view on how KARs function and provided insight into the discrepancies between native and recombinant KAR properties. While the exact role of these interactions still remains to be unambiguously established, the role of KARs in physiology will be difficult to understand without taking into account the contribution of these proteins. For instance, KARs and many of these proteins seem to undergo transient interactions that promote receptor trafficking, regulating their surface expression. PDZ motif-containing proteins such as postsynaptic density protein 95 (PSD-95), protein interacting with C kinase-1 (PICK1), and glutamate receptor interacting protein (GRIP) seem to be relevant for the stabilization of KARs at the synaptic membrane Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 293

Neuron

Review (Hirbec et al., 2003). However, PDZ-binding motifs in the C terminus of KAR subunits are also present in other glutamate receptors. Thus, these interacting proteins are not selective for KARs. Although interactions with PDZ domains cannot entirely account for the subcellular distribution of KARs, the interaction with PDZ proteins produce apparently different outcomes for these receptors, as these proteins prevent AMPAR internalization but facilitate KAR internalization (Hirbec et al., 2003). It was recently demonstrated that the SNARE protein SNAP25 is a KAR-interacting protein (Selak et al., 2009). In MF-CA3 synapses, activity-dependent stimulation of PKC intensifies this interaction and triggers the internalization of KARs, leading to a specific long-term depression (LTD) of KAR-mediated synaptic transmission. Interestingly, these results implied that SNAP25, classically regarded as a member of the exocytotic machinery, may be also involved in endocytosis (Selak et al., 2009). This view has been recently supported by a report defining a role for SNAP25 in clathrin-dependent endocytosis at conventional synapses (Zhang et al., 2013). KARs at these synapses may also contain GluK2 subunits and, recently, it was proposed that this mechanism of LTD requires the synergistic SUMOylation of GluK2 subunits, initiated by PKC phosphorylation (Chamberlain et al., 2012). This new mechanism expands the repertoire of events associated with synaptic plasticity. The possibility of modifying information transfer at this level has been further illustrated by the recent observation that CaMKII-mediated phosphorylation of GluK5 subunits also depresses a KAR-mediated synaptic component at CA3 synapses (Carta et al., 2013). A spike timing-dependent plasticity protocol, known to activate CaMKII in a number of synapses and induce AMPAR LTP, induces phosphorylation of GluK5-containing receptors in MF-CA3 synapses, resulting in LTD of the KARmediated synaptic component. Rather than involving endocytosis of KARs, this depression is evoked by the lateral diffusion of these receptors upon uncoupling of the PSD-95 scaffolding protein at the postsynaptic density (Carta et al., 2013; see also Copits and Swanson, 2013a). Additional proteins that interact and directly modulate the properties of KARs have also been identified. These include proteins such as kainate receptor interacting protein for GluR6 (KRIP6; Laezza et al., 2007), a protein that belongs to the BTB/ kelch family and that binds to a C-terminal motif distinct to the PDZ binding motif. Coexpression of KRIP6 with GluK2 reduces both the peak current and steady-state desensitization in recombinant systems, as well as that of native KARs. Interestingly, KRIP6 does not affect the surface expression of GluK2 receptors, indicating that the interaction with this protein only affects channel gating. Another BTB/kelch family member, actinfilin, is also thought to interact with GluK2 subunits (Salinas et al., 2006), this protein promoting the degradation of GluK2 receptors by acting as a scaffold to link this subunit to the E3 ubiquitinligase complex. In this way, actinfilin regulates the synaptic expression of receptors containing GluK2 (Salinas et al., 2006), although more work will be necessary to reveal what is the physiological impact of these BTB/kelch proteins. For instance, it is known that KRIP6 can interact with PICK1, forming clusters that lack GluK2 and preventing the mutual regulation of GluK2 containing KARs (Laezza et al., 2008). 294 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

A number of studies have identified trafficking and targeting motifs in KAR subunits and increased our knowledge of the mechanisms controlling KAR targeting and surface expression (see reviews by Pinheiro and Mulle, 2006 and Contractor et al., 2011). However, many other aspects of KAR biology still remain to be defined. For instance, many of the mechanisms supporting the polarized targeting of KARs to different neuronal populations are unknown. Recently, two integral membrane proteins have been identified that seem to be true auxiliary subunits of KARs (Zhang et al., 2009; Straub et al., 2011a; Tang et al., 2011). Neuropilin Tolloid-like 1 and Neuropilin Tolloid-like 2 (Neto1 and Neto2) are auxiliary proteins of native KARs that exert an important influence on their function. Indeed, these proteins radically alter the gating properties of KARs, accounting for a number of previously unexplained properties of these receptors (see Copits and Swanson, 2012; Lerma, 2011; Tomita and Castillo, 2012 for recent reviews). Neto1 and Neto2 share an identical and unique domain structure, representing a novel subfamily of transmembrane proteins containing CUB and LDLa domains. Neto1 was first identified as a protein that interacts with the NMDA receptor (Ng et al., 2009), although a number of studies then illustrated that it has a more striking influence on the function of KARs. In general, the coexpression of Neto1 and Neto2 with KARs in recombinant systems alters the gating properties of the latter. The most obvious effect is that the onset of the desensitization of kainate-evoked responses decelerates (Copits et al., 2011; Straub et al., 2011b; Fisher and Mott, 2013), while recovery from the desensitized state accelerates. This modulation implies that the kainate-induced steady current persists for longer periods in the presence of an agonist (e.g., Fisher and Mott, 2013). This effect is evident for all subunits and reconciles the properties of recombinant receptors with the reported action of kainate in more physiological preparations, where it behaves as a strong depolarizing agent. Moreover, the rapid deactivation of kainate-induced currents upon agonist removal is also decelerated in the presence of Neto, suggesting an increase in the steady-state affinity of KARs when associated to Neto. Indeed, equilibrium agonist affinity substantially increased in the presence of Neto, again reconciling the properties of recombinant and native KARs. A prominent feature of KAR-mediated excitatory postsynaptic currents (EPSCKARs) is that they are characteristically slower and smaller than AMPAR-mediated EPSCs (Castillo et al., 1997; Vignes et al., 1998; Frerking et al., 1998). This cannot be anticipated from the properties of recombinant receptors, since single KARs and AMPARs have similar affinity and activation-inactivation kinetics (see Lerma, 1997). A prominent perisynaptic localization of KARs was also ruled out (Castillo et al., 1997) and if both receptor subtypes colocalize at the synapse, one would expect similar kinetics for the KAR- and AMPAR-mediated synaptic responses. Although the subunit composition of KARs may have an influence in EPSC kinetics (Contractor et al., 2001; Barberis et al., 2008; Fernandes et al., 2009), no convincing explanation for this discrepancy had been available until the discovery that Neto proteins confer higher agonist affinity to KARs and impose significantly slower deactivation kinetics. Curiously, the kinetics of KARs imposed by Neto protein is remarkably similar to those

Neuron

Review A

B

MF->CA3

-60 mV; No drugs

-70 mV; GYKI53655

KAR NMDAR AMPAR

-30 mV; CNQX

50 ms

C

100 pA

D

Scaled responses

EPSCAMPAR NETO1 KO

EPSCNMDAR

100 ms

EPSCKAR

E

-Ctrl

F

-UBP310

0.5 s

WT

10 mV

0

GluK2-/-

wild-type

5 10 15 20 25 30 0

5 10 15 20 25 30

s

of NMDARs (see Figure 1). The difference between NMDAR activation kinetics and that of the faster AMPARs provides adequate timing for activation, since the Mg2+ blockade implies that NMDAR activation would not be operative until sufficient membrane depolarization is attained. In contrast, the functional significance of the slower kinetics of KARs is starting to be illustrated by examples that provide comprehensive roles for such a prolonged current in synaptic integration (Frerking and OhligerFrerking, 2002; Goldin et al., 2007; Sachidhanandam et al., 2009; Pinheiro et al., 2013; see Figure 1). Another striking action of Neto1 and Neto2 is that association with these proteins greatly reduces inward rectification of KARmediated currents without modifying Ca2+ permeability (Fisher and Mott, 2012). It seems that three positive charges (RKK) in the C-terminal of Neto proteins preclude internal polyamine blockade of KAR channel. This effect is reminiscent of stargazin in AMPARs (Soto et al., 2007). However, the functional implication of this action remains to be defined. Apart from the clear effect of Netos on KAR channel gating and on current amplitudes (see Copits and Swanson, 2012), it remains unclear whether Neto proteins are involved in KAR targeting to the synapse, although there is weak evidence indicating that this may be possible. Cultured hippocampal neurons express native KARs, but these are not targeted to synapses (Lerma et al., 1997). However, a small proportion of ESPCs may be mediated by KARs when such cells are transfected with Neto2 and GluK1, indicating that exogenous Neto2 may target a small proportion of exogenous GluK1 to synapses (Copits et al., 2011). Similar effects were observed in cerebellar granule cells and with GluK2 (Zhang et al., 2009). However, although GluK2 association with PSD95 is reduced in Neto2 null mice (Tang et al., 2012), the lack of Neto2 expression does not prevent the presence of endogenous GluK1 or

s

Figure 1. Kinetic Similarities between NMDAR and KARs (A) NMDARs, AMPARs, and KARs coexist in Mossy Fiber to CA3 neuron synapses. (B) Synaptic responses mediated by AMPAR are much faster than the KAR and NMDAR components, as can be seen after their pharmacological isolation. (C) Responses mediated by AMPARs, NMDARs, and KARs are superimposed once scaled. Note how KARs and NMDARs generate responses with a remarkable similar onset and decay time courses. (D) The slow time course of KARs is accounted for by their interaction with Neto proteins. Note how KAR-mediated EPSCs are accelerated in the Neto1 KO mice, approaching the characteristics of the AMPAR-mediated component (records reproduced with permission from Straub et al., 2011a). (E) Participation of postsynaptic KARs to MF-CA3 synaptic transmission: averaged traces illustrate the effect of KARs antagonist UBP310 on the depolarizing envelope (shaded area) induced by a train of stimuli. KARs blockade reduces summation and prevents neuron firing (spikes are shown truncated) (from Pinheiro et al., 2013). (F) CA3 pyramidal cell firing in response to a physiological granule cell firing pattern of stimulation (extracted from an in vivo recording of a freely moving mouse) in WT and a GluK2-deficient mouse, revealing the contribution of postsynaptic KARs to neuronal output (from Sachidhanandam et al., 2009).

GluK2 in synaptic contacts, despite the fact that synaptic KARs are normally associated with Netos in hippocampal slices. Indeed, KAR-mediated EPSCs in brain slices display distinct kinetics in Neto-deficient animals and EPSCKARs are present in mice even deficient for the two Neto proteins, yet with fast kinetics, consistent with the idea that Netos are not key elements in the targeting of KARs to the synapse (Tang et al., 2011). From these data, it is clear that Netos exert an important influence on KARs, which may vary depending on the subunit combination. However, Neto proteins are not specific to KARs. Indeed, Neto1 was initially identified as a NMDAR interactor. While Neto2 was originally thought to partner KARs only, it was recently reported to also interact with distant proteins, such as the neuron-specific K+-Cl
cotransporter KCC2, which is essential to maintain Cl
homeostasis in neurons (Ivakine et al., 2013). These data raise the possibility that Neto proteins play a more wide-ranging role than initially anticipated. Nevertheless, the interesting question that remains to be answered is whether the association of Neto1/2 with KARs could be regulated by physiological signals, and under what circumstances this occurs. Since KARs are fully operational in the absence of Neto, it is possible that two populations of KARs might exist, those with and without Neto probably fulfilling complementary functional roles. Recently, the group of Maricq has identified in the worm C. elegans SOL-2, a CUB-domain protein that associates with both the related auxiliary subunit SOL-1 and with the GLR-1 AMPAR (Wang et al., 2012). Like Neto1, SOL-2 contributes to the kinetics of receptor desensitization and is an essential component of AMPAR complexes at worm synapses. These data indicate that several different interacting proteins could form the receptor complex at synapses. Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 295

Neuron

Review Figure 2. Kainate Receptors Signal through Two Different and Independent Pathways On the one hand, the gating of the channel (canonical pathway, a) is responsible for membrane depolarization and the synaptic responses (b) and probably for the facilitation of neurotransmitter release at some synapses. These receptors bear ancillary proteins, like Neto. On the other hand, KARs activate G proteins (c), signaling through the stimulation of phospholipase C and PKC in a manner independent of ion flux. It is unknown whether these receptors are coupled to Neto proteins and whether they require an intermediate protein to couple to G protein. The main effects so far documented are: the increase in neuronal excitability through the inhibition of afterhypopolarizing current (IAHP, d), leading to an increase in the firing frequency of these neurons (e); the facilitation and inhibition of transmitter release, probably through the modulation of calcium currents (ICa2+, f); and their role in the maturation of neuronal circuits during development.

The Uniqueness of Ion-Dependent KAR Gating One unique feature of KARs is that their channel gating requires external monovalent cations and anions. This ion-dependent channel gating differentiates KARs from other ligand-gated channels, including the closely related NMDARs and AMPARs (Paternain et al., 2003; Bowie, 2002; see Bowie, 2010 for a review). Indeed, crystallographic studies have revealed the existence of an ion binding pocket in KAR subunits (Plested and Mayer, 2007; Plested et al., 2008). The absolute requirement of ion binding for channel opening indicates that KAR activity would be abolished if this binding site remained unoccupied, prompting the suggestion that this site might be used as a target for specific allosteric modulation of KARs by external agents. The question as to what might be the physiological role of such a strict dependence of the channel gate has not been answered yet but prompted some possibilities. For instance, under intense neuronal activity, a situation under which external Na+ levels drop, activation of KARs would be limited, constituting a brake for tissue damage. Indeed, a large fraction of KARs seems to have unoccupied the cation binding site at physiological salt concentrations, making them insensitive to activation by released glutamate (Plested et al., 2008). Much work remains to be done to figure out whether this fraction of incompetent KARs could be modulated up and down as a way to regulate the weight of these receptors in, for instance, synaptic transmission. A Receptor with Two Modes of Signaling High-resolution structural analysis has revealed many similarities between the three glutamate receptor families. However, unlike AMPA and NMDA receptors, KARs appear to also signal through 296 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

an unconventional metabotropic mechanism involving G proteins and second messengers at inhibitory CA1 hippocampal synapses (Rodrı´guez-Moreno et al., 1997; Clarke et al., 1997). This signaling follows presynaptic inhibition of GABA release and is dependent on G protein activation and PKC activity (Rodrı´guezMoreno and Lerma, 1998; Rodrı´guez-Moreno et al., 2000). Later, this nonconventional mode of signaling was compellingly established in dorsal root ganglion neurons and was shown to be independent of ion flux (Rozas et al., 2003). Since then, an increasing number of metabotropic actions triggered by KARs have been described in many cell types and in different regions of the CNS, particularly in association with the presynaptic control of neurotransmitter release or the postsynaptic regulation of neuronal excitability (see Rodrigues and Lerma, 2012 for a recent review and Figure 2). However, key aspects of the molecular mechanisms underlying this noncanonical signaling still remain unclear, including how KARs activate G proteins to trigger these effects and what determines the mode of action of KARs (i.e., conventional ionotropic versus noncanonical metabotropic signaling). The evidence for a direct interaction between KARs and G proteins is limited. Prior to describing the metabotropic behavior of KARs, the Pertussis toxin (PTx)-sensitive binding of an agonist to goldfish-purified KARs was demonstrated biochemically, providing a link between KARs and PTx-sensitive proteins (Ziegra et al., 1992). While similar PTx-sensitive KAR agonist-binding was also observed in hippocampal membranes (Cunha et al., 1999), this kind of interaction does not seem to be that related to the functional signal transduced by KAR activation through G protein activity. It is expected that undergoing proteomic analysis of KAR subunits identify partners that could account for the coupling between an ion channel receptor and a G protein. Initially, it was unclear which subunits might engage this activity and, still, the search to identify the KAR subunit that mediates this noncanonical signaling is not free of controversy. In dorsal

Neuron

Review root ganglia (DRG) neurons that exclusively express GluK1 and GluK5 subunits, noncanonical signaling was dependent on GluK1 rather than GluK5 (Rozas et al., 2003). Subsequent studies found that KAR-mediated modulation of IAHP, an action provoked by the noncanonical signaling of KARs (Melyan et al., 2002), was absent in GluK2- (Fisahn et al., 2005) or GluK5(Ruiz et al., 2005) deficient mice. However, more recent studies reported that noncanonical signaling persisted in GluK5 and GluK4–GluK5 knockout (KO) animals (Fernandes et al., 2009). Indeed, expression of GluK1 in SHSY5 neuroblastoma cells was sufficient to reconstitute metabotropic activity of KARs, as evaluated by the G protein and PKC activation inducing internalization of KARs from the membrane (Rivera et al., 2007). Recent experiments confirmed the involvement of GluK1 in the metabotropic control of glutamate release (Segerstra˚le et al., 2010; Salmen et al., 2012). However, a biochemical interaction between GluK5 and a Gaq protein was identified in biochemical experiments (Ruiz et al., 2005). These data would in principle indicate that the subunit interacting with G proteins might be GluK5, either directly or indirectly. There are additional issues that appear at odds with this idea. The involvement of Gq protein does not fit with the PTx sensitivity of the metabotropic actions of KARs described to date (see Rodrigues and Lerma, 2012 and references therein) but, rather, the PTx sensitivity suggests that Gi or Go proteins are likely to be involved in the metabotropic actions of KARs. However, the concomitant involvement of PLC and PKC in most of the metabotropic effects described to date rules out the participation of Gi, leaving the Go protein as the only strong candidate to mediate these effects (e.g., Rozas et al., 2003). Nevertheless, some effects induced by KA are contingent on the inhibition of adenylate cyclase and the subsequent reduction in cAMP would involve Gi protein activation, as also described (Gelsomino et al., 2013; Negrete-Dı´az et al., 2006). Available data clearly show that subunit composition alone cannot define the signaling mode triggered by KARs, pointing to interacting partners as candidates likely to determine the mode of action of KARs. However, the existence of proteins that functionally couple KARs and G proteins remains to be demonstrated. It should be also taken into account that some at odds data has been published pointing out that at least part of the noncanonical signaling triggered by KARs may be indirect (Lourenc¸o et al., 2011). Regardless of the specific mechanisms, it is now clear that KARs can no longer be considered simply as ligandgated ion channels. The increasing number of activities known to be mediated by KARs through this noncanonical signaling, as described below, indicates that this dual signaling is one of the main factors underlying the diverse actions of KARs reported over the years. Roles in Synaptic Transmission and Network Excitability Postsynaptic Kainate Receptors Unlike AMPAR-mediated currents, the activation of postsynaptic KARs by synaptically released glutamate yields small amplitude EPSCs, with slow activation and deactivation kinetics (see Figure 1; Castillo et al., 1997). Moreover, while AMPARs and NMDARs are localized to the postsynaptic density of the vast majority of glutamatergic synapses in the brain, EPSCs mediated

by KARs have only been found in a few central synapses, such as in MF to CA3 pyramidal neurons (Castillo et al., 1997; Vignes and Collingridge, 1997), the contacts between Schaffer collaterals and CA1 hippocampal interneurons (Cossart et al., 1998; Frerking et al., 1998), between parallel fibers and Golgi cells in the cerebellum (Bureau et al., 2000), at thalamocortical connections (Kidd and Isaac, 1999), in the basolateral amygdala (Li and Rogawski, 1998), in the synapses between afferent sensory fibers and dorsal horn neurons in the spinal cord (Li et al., 1999), and those of parallel fibers and cerebellar Golgi cells (Bureau et al., 2000). In all these synapses, the characteristic slow kinetics is a predominant feature of the EPSCKAR, probably providing integrative capacities to information transfer clearly unfulfilled by other glutamate receptors (Frerking and OhligerFrerking, 2002; Pinheiro et al., 2013). For instance, O-LM interneurons are a somatostatin interneuronal subtype at the stratum oriens that processes glutamatergic inputs through KARs, which endow these cells with the ability to follow inputs at the theta frequency (Goldin et al., 2007). In addition, recent data indicate that GluK1-containing KARs in a subset of stratum radiatum interneurons mediate feedforward inhibition of pyramidal cells. The output of these interneurons is enhanced during both lowfrequency-evoked stimulation and natural-type firing patterns. During this activity, the threshold for the induction of theta-burst LTP is raised. In this way, such KAR-mediated input promotes a shift in the dynamics of synaptic transmission in favor of interneuronal output onto CA1 pyramidal neurons (Clarke et al., 2012). A striking impact on neuronal excitability of postsynaptic KARs, acting through their noncanonical signaling, is provided by the regulatory action of the slow afterhyperpolarization current (IsAHP: Melyan et al., 2002, 2004). The IsAHP activates upon bursts of action potentials and it is generated by voltage-sensitive Ca2+-dependent K+ channels. It has a slow decay time as it may last for several seconds, it is activated in proportion to the number and frequency of action potentials (Lancaster and Adams, 1986), and it underlies spike frequency adaptation (Figure 2). At Schaffer-CA1 pyramidal cell synapses, at which no EPSCKAR has been documented (Lerma et al., 1997; Castillo et al., 1997; Frerking et al., 1998; Cossart et al., 1998), nanomolar concentrations of KA cause long-lasting inhibition of IAHP through the direct activity of KARs. This effect is mimicked by synaptic glutamate released from excitatory afferents at the CA1 synapses (Melyan et al., 2004; Chamberlain et al., 2013). Pharmacological evidence indicates that this inhibition involves the noncanonical signaling engaging Gi/o protein and PKC activation (Melyan et al., 2002) and probably PKA and downstream activation of MAP kinases (Grabauskas et al., 2007). The inhibition of both the slow and medium IAHP by KAR activation increases the firing frequency of these neurons, largely enhancing circuit excitability (Fisahn et al., 2005; Ruiz et al., 2005). Like KAR-mediated EPSCs, inhibition of IAHP has been observed in MF-CA3 pyramidal cell synapses (Ruiz et al., 2005; Fisahn et al., 2005) and, therefore, both signaling modes can coexist within the same synapses. Thus, a short train of stimuli to the mossy fibers could not only directly depolarize the postsynaptic membrane but also increase neuronal excitability by preventing spike adaptation. Interestingly, both the ionotropic and Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 297

Neuron

Review A

MF-CA3 EPSCs

B Untreated

GABAergic IPSCs +Glu uptake inhibitor

Figure 3. Presynaptic KARs Modulate Synaptic Neurotransmitter Release in a Bidirectional Manner and Both Inotropic and Noncanonical Metabotropic Activity Are Involved

(A) In the mossy fiber to CA3 synapses, activation of KARs is responsible of part of the frequency facilitation, such that the inclusion of a KAR Control Control Control antagonist reduces the synaptic responses to the second and subsequent stimuli but not the first +KAR EPSC. At the bottom of each panel, the variation in Antagonist ΔQ ( Cont-Anta) synaptic current charge (DQ) due to the activation of KARs is indicated. (B) Bidirectional modulation of evoked IPSCs by KARs. Superimposed traces illustrating evoked IPSCs obtained under control conditions (black traces) and in the presence of a KAR inhibitor (UBP302) (blue traces) in slices untreated or treated with D,L-TBOA, an inhibitor of glutamate reuptake (based on Bonfardin et al., 2010). In the first case, GABA release is facilitated by tonic activity of KARs, while in the second (larger concentrations of extracellular glutamate) GABA release is inhibited. +KAR Antagonist

KAR Antagonist

metabotropic actions are segregated in the neurite tree and are independent of each other (Rozas et al., 2003), a result further supported by the use of mice lacking individual KAR subunits (Ruiz et al., 2005; Fernandes et al., 2009) or pharmacologically antagonized ion channel activity (Pinheiro et al., 2013). This reinforces the idea that KARs may engage metabotropic and ionotropic signaling in an independent manner. Together, the evidence provided so far demonstrates that postsynaptic KARs regulate neuronal excitability both by producing long-lasting depolarization and by inhibiting IAHP through a segregated G protein-coupled pathway. The efficiency of KARs in the regulation of neuronal excitability seems to rely on repetitive synaptic activation rather than on single impulses, indicating that postsynaptic KARs are designed to modulate the temporal integration of excitatory circuits. Similarly, there is now compelling evidence that KARs elicit sufficient charge transfer to have a substantial impact on synaptic function wherever they are expressed. For example, the kinetics of the EPSP mediated by KARs is sufficiently slow to allow substantial tonic depolarization during even modest presynaptic activity (Frerking and Ohliger-Frerking, 2002; Sachidhanandam et al., 2009; see Figure 1). But not only has the long ionotropic activity had an impact on synaptic integration. The importance of the metabotropic actions of KARs has also been recently put forward by showing that the plastic changes in the KAR-mediated synaptic component could modify the degree of inhibition of IAHP in CA3 pyramidal neurons. Chamberlain and associates (Chamberlain et al., 2013) showed that induction of LTD of the KAR-mediated EPSC induced by natural pattern of stimulation relieves the KARinduced inhibition of IAHP, resulting in further attenuation of neuronal responses to subsequent inputs. These data indicate that KARs may exert a major role in regulating neuron excitability and that although long-lasting plastic modulation of these receptors does alter their ionotropic function, their concomitant metabotropic activity becomes a dominant factor, at least under certain experimental conditions such as high-frequency (10–20 Hz) activity. Also, KARs have been recently shown to be subject to homeostatic plasticity (Yan et al., 2013) in that the KAR-mediated EPSC at mossy fiber to cerebellar granule cell synapses was enhanced after network activity blockade (either by TTx or genetically removing AMPARs). This phenomenon relies on the enhanced expression of GluK5 subunits that produces receptors with a 298 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

higher affinity for glutamate, efficiently maintaining spike generation at granule cells. Such effects should be explored at different synapses given that this homeostatic regulation has also been observed in climbing fibers to Purkinje cell synapses (Yan et al., 2013), which may indicate it to be a more universal mechanism than originally thought. Presynaptic Kainate Receptors The unconventional hypothesis that KARs could also play a role as presynaptic modulators of neurotransmitter release was prompted by the observation that the pharmacological activation of KARs modulated Ca2+-dependent glutamate release from synaptosomes, which are structures devoid of somatodendritic elements (Chittajallu et al., 1996). Since then, much effort has been devoted to determine the presynaptic role of KARs and it is now widely accepted that functional presynaptic KARs play a crucial role in the control of neurotransmitter release (Lerma, 2003). Indeed, it is now known that presynaptic KARs modulate neurotransmitter release in a bidirectional manner, not only at excitatory but also at inhibitory synapses. KAR activation modulates GABAergic transmission in a complex cellular and subcellular manner, and both depression and facilitation of GABA release have been reported (Figure 3). The question then arises as to which event takes preference over the other and under what circumstances? Early indications of KA-induced depression of inhibition in the hippocampus (Sloviter and Damiano, 1981) were confirmed by the demonstration that KARs can inhibit GABA release (Rodrı´guez-Moreno et al., 1997; Vignes et al., 1998). The depression of inhibition induced was shown to be sensitive to PTx and to inhibitors of both PLC and PKC, leading to the postulate that KARs participated in unconventional events at presynaptic sites that most likely involve a metabotropic signaling pathway rather than ion flux (Rodrı´guez-Moreno and Lerma, 1998). This idea was later supported by measuring GABA release in synaptosomes (Cunha et al., 1997, 2000; Perkinton and Sihra, 1999) and it has been observed in other structures such as the amygdala (e.g., Braga et al., 2004), neocortex (Ali et al., 2001), globus pallidus (Jin and Smith, 2007), and hypothalamic supraoptic nucleus (Bonfardin et al., 2010). However, CA1 interneurons become overactivated by exogenous KA through somatodendritic KARs, leading to the paradox of KA inducing both overflow (Frerking et al., 1998; Cossart et al., 1998) and inhibition of GABA release. Presynaptic and somatodendritic KARs seem to coexist, presenting

Neuron

Review A

B

Figure 4. Kainate Depresses Hippocampal Inhibition and Generates Epileptic Activity In Vivo

(A) Arrangement of recording and stimulating electrodes in vivo. In this experiment, a microdialysis probe implanted in the region allowed kainate to be slowly introduced into the extracellular fluid. The degree of GABAergic inhibition could be tested by the paired-pulse test, recording field potentials from the CA1 pyramidal layer through an extracellular electrode. (B) Stimulation of the Schaffer collateral pathway evokes the synchronous firing of a large number of C CA1 pyramidal cells (the sharp negative wave indicated by an asterisk in the first evoked potential). This initial firing of the neuronal population activates a population of inhibitory interneurons that feedback onto the CA1 pyramidal neurons, such that a second stimulus arriving during the inhibitory phase (40 ms later) is unable to induce CA1 neuron firing, evident by the absence of a population spike in response to the second stimulus under control conditions. After perfusion of kainate through the dialysis probe (3 mM in the perfusate, probably 10% of this concentration reaches the extracellular fluid), the population spike in the second response progressively develops, indicating a failure in the inhibition of pyramidal cells. (C) After prolonged perfusion of kainate, ongoing epileptic activity develops that is characterized by the presence of interictal spikes in the EEG. The insert in yellow represents a 1 s period over an expanded time base (modified from Rodrı´guez-Moreno et al., 1997).

distinct pharmacological profiles and subunit compositions and using different signaling pathways (Rodrı´guez-Moreno et al., 2000; Mulle et al., 2000; Christensen et al., 2004; Maingret et al., 2005). Thus, while somatodendritic KARs mediate part of the synaptic input from Schafer collaterals, presynaptic KARs are activated by synaptically released glutamate and they reduce the inhibitory input to pyramidal cells (Min et al., 1999). Thus, KARs play a fundamental role in the performance of neuronal circuits, as exemplified in the hippocampus. As for other aspects of KAR activity, the mechanism by which KARs modulate inhibitory input to pyramidal neurons is not free of controversy. It has been reported that inhibition of GABA release from CB1 receptor-expressing interneurons induced by KAR activation depended, at least in part, on cannabinoid-1 (CB1) and GABAB receptors strategically situated at the presynaptic GABA boutons (Lourenc¸o et al., 2010, 2011). It was proposed that synaptically released glutamate induced the postsynaptic release of endocannabinoids and the extracellular accumulation of GABA. This conclusion contradicts the observation of the inhibitory effect of KA on GABA release in interneuronpyramidal cell pairs in the presence of antagonists of CB1 (e.g., AM251) or GABAB (e.g., CGP55845) receptors (Daw et al., 2010). Perhaps these disparate conclusions may come from the combined or independent antagonism of both types of receptors, but even when both receptors were blocked, a fraction of inhibition of GABA release seems to rely on KAR activity (Lourenc¸o et al., 2011). Whatever the fraction of GABA release affected by KAR activation, one can consider that presynaptic KARs will be activated by glutamate released during intense periods of activity in vivo. This will result in the depression of GABA release, likely leading to a state of overexcitability that could have important consequences on the circuit performance, making it more seizurogenic by dampening inhibition. Indeed, this has been proven to occur in the hippocampus in vivo when a low concentration of KA is slowly dialyzed into the extracellular fluid. This causes a depression of synaptic inhibition and the appearance

of interictal epileptic spikes in the electroencephalogram (EEG) (see Figure 4; Rodrı´guez-Moreno et al., 1997). Evidence that presynaptic KARs mediate tonic inhibition of GABAergic transmission has also been obtained from the hypothalamic supraoptic nucleus, where it involves a PLC-dependent metabotropic pathway (Bonfardin et al., 2010). During development, tonic activation of presynaptic GluK1-containing KARs also depresses GABA release from hippocampal MF in a G protein- and PLC-dependent manner (Caiati et al., 2010). Thus, regardless of the mechanism, there is a growing body of evidence that activation of presynaptic KARs depresses GABAergic transmission, either after exogenous agonist application or through endogenous released glutamate. However, paired recordings of interneurons and CA1 pyramidal cells also show that endogenous activation of KARs can facilitate GABA release at some synapses (Jiang et al., 2001). Further evidence that presynaptic KARs exert such a facilitatory effect on GABA release has been gathered from hippocampal interneurons (Mulle et al., 2000; Cossart et al., 2001), as well as in several other regions of the CNS like the neocortex (Mathew et al., 2008) and hypothalamus (Liu et al., 1999). In contrast to the inhibitory effects, the facilitation of GABA release by presynaptic KARs is likely to involve the conventional ionotropic activity of these receptors (reviewed in Rodrigues and Lerma, 2012). This seems to at least be the case for endogenous activation of KARs in the hypothalamic supraoptic nucleus. It was shown in an elegant study that the increase in the ambient levels of glutamate, either associated to a physiological reduction in astrocyte cover or due to the blockade of glutamate transporters, switches facilitation to inhibition of GABAergic release (Bonfardin et al., 2010) (Figure 3). Interestingly, while facilitation was prevented by philantotoxin (a blocker of Ca2+-permeable receptors), the inhibitory effect involved PLC. Thus, the net result of activating presynaptic KARs by endogenous glutamate may depend on the glutamate concentration actually reaching the presynaptic KARs, which will ultimately depend on the magnitude of glutamate spillover Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 299

Neuron

Review arising from different patterns of synaptic activity coupled to the astrocyte uptake capacity. This hypothesis is further substantiated by data from the cerebellum, where bidirectional modulation of transmitter release has also been found. Synaptically activated presynaptic KARs facilitate and depress transmission at parallel fiber synapses (Delaney and Jahr, 2002). Activation of presynaptic KARs by synaptically released glutamate at parallel fibers facilitates glutamate release to both interneurons (e.g., stellate or basket cells) and Purkinje cells when these fibers are subjected to a regime of low-frequency stimulation. By contrast, with high-frequency stimulation, the synapses onto inhibitory interneurons are depressed, while synapses at Purkinje cells are still facilitated. Such differential sensitivities to the frequency of these two synapses may regulate the excitation/inhibition balance of Purkinje cells and, therefore, cerebellar output. Thus, at some structures, KARs bestow computational properties to circuits according to the activity regime of afferent inputs. Presynaptic regulation of excitatory transmission by KARs has been studied extensively at MF-CA3 synapses. At these synapses, presynaptic KARs are implicated in the characteristic frequency-dependent facilitation of MF excitatory transmission (Schmitz et al., 2001; Lauri et al., 2001; Contractor et al., 2001; Pinheiro et al., 2007), a phenomenon initially ascribed to the residual intraterminal calcium. Since KAR antagonists attenuate the potentiation of the second EPSC during high-frequency trains (e.g., 25 Hz; Schmitz et al., 2001), the synaptic activation of presynaptic KARs must be quite fast (10–30 ms), indicating that KARs should be found near the active zone. Indirect evidence suggests that the facilitation of glutamate release may occur through the depolarization of presynaptic terminals (Schmitz et al., 2001) that should enhance action potential-driven Ca2+ influx (Kamiya et al., 2002; Lauri et al., 2003). The reduction of synaptic facilitation by a blocker of Ca2+-permeable KARs (Lauri et al., 2003) also points to a contribution of direct Ca2+ entry through these receptor channels, although Ca2+ mobilization from intracellular stores may also add to this use-dependent facilitation of glutamate release (Lauri et al., 2003; Scott et al., 2008). Nevertheless, these conclusions have been challenged by the indication that presynaptic KARs are insufficient to facilitate glutamate release at MF-CA3 synapses, attributing the phenomenon to the activation of recurrent CA3 network activity (Kwon and Castillo, 2008). However, blockade of postsynaptic KARs at MF-CA3 synapses with newly available compounds (e.g., UBP310) had no effect on presynaptic facilitation (Pinheiro et al., 2013), a result similar to that observed in double GluK4/ GluK5 mice, in which there is no deficit in short-term plasticity, whereas postsynaptic KAR-mediated responses are totally lost (Fernandes et al., 2009). Therefore, in the absence of further evidence against it, it should be concluded that part of the synaptic facilitation observed at MF-CA3 synapses is due to the activation of presynaptic facilitatory KARs. Considering that presynaptic KAR function has been assessed indirectly, direct electrophysiological recording from these presynaptic structures may clarify the issue of whether or not ionotropic facilitatory KARs are present at MF boutons. Conclusive evidence indicates that this mechanism imposes associative properties to MF-LTP, since the activity in neigh300 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

boring MF synapses influences the threshold to induce LTP at these synapses (Schmitz et al., 2003). NMDARs implement the associative properties of LTP. However, the contribution of NMDARs to the induction of LTP in the CA3 field is quite modest and one might think that the presence of KARs at these synapses maintains the general properties of LTP unaltered. While the facilitation of glutamate release has clear functional implications, it remains unclear under what circumstances the suppression of glutamate release by KARs may fulfill a significant role. Interestingly, it seems that during development, the inhibitory modulation of glutamate release may shape synaptic properties (Lauri et al., 2006; see below), and it has been observed that long and strong trains of afferent activity depress rather than facilitate synaptic transmission (Schmitz et al., 2001), a mechanism that may be active under physiological conditions. Facilitation of glutamate release at MF-CA3 synapses is mimicked by applying low concentrations of exogenous KA (Schmitz et al., 2000; Kamiya and Ozawa, 2000). Higher concentrations of KA depress synaptic transmission not only at MF-CA3 synapses but also at synapses between Schaffer collaterals and CA1 pyramidal cells (Chittajallu et al., 1996; Kamiya and Ozawa, 1998; Vignes et al., 1998; Frerking et al., 2001) and those of the associational/commissural pathway terminating on CA3 neurons (Salmen et al., 2012). This inhibition is accompanied by a reduction in presynaptic Ca2+ (Kamiya and Ozawa, 1998; Salmen et al., 2012), and since it is sensitive to G protein blockers, this inhibition is unlikely to involve presynaptic depolarization, but it is more likely to be contingent on noncanonical signaling (Frerking et al., 2001; Negrete-Dı´az et al., 2006; Salmen et al., 2012). The inhibition of glutamate release at Schaffer collaterals-CA1 synapses does not involve protein kinases but, rather, a membrane-delimited action that probably involves the direct inhibition of presynaptic Ca2+ channels by G protein gb subunits (De Waard et al., 1997). The inhibitory effects of KARs upon tonic activation by endogenous glutamate at these two hippocampal synaptic populations have been observed during development and involve KAR metabotropic signaling (Lauri et al., 2005, 2006; Sallert et al., 2007). Tonic activation of presynaptic KARs in the adult brain also inhibits glutamate release in the rat globus pallidus, once again mediated by a presynaptic Gi/o-proteincoupled and PKC-dependent mechanism (Jin and Smith, 2007). What now seems clear is that presynaptic KARs modulate transmitter release in a bidirectional manner: facilitation probably occurs through their ionotropic activity, while inhibition seems to involve noncanonical metabotropic signaling. It is possible that the threshold to activate one or other KAR signaling pathways would determine physiological responses. In summary, the presynaptic modulation of both glutamate and GABA release together with the postsynaptic regulation of neuronal excitability clearly demonstrates that KARs are endowed with diverse capacities. These activities enable them to fine-tune synaptic function and regulate neuronal network activity in the adult brain, bestowing them with a much broader role at synapses than the simple transfer of information. Role of Kainate Receptors during Development KARs are expressed strongly in the brain during development in a complex cell-type-specific manner. The properties of synapses during development differ significantly from those at mature

Neuron

Review stages and it is now known that presynaptic KARs contribute to these changes. The expression of KARs, and particularly of GluK1 subunits, increases markedly and peaks during the first week of life in rodents (Bahn et al., 1994). In immature hippocampal CA1 synapses, the tonic activation of KARs by ambient glutamate keeps the probability of release low (Lauri et al., 2006). However, a burst of synaptic activity produces strongly facilitating postsynaptic responses, a facilitation that is not only abolished in the presence of a KAR antagonist but also later in development. Interestingly, this change is recapitulated by inducing LTP and it seems that the tonic stimulation of KARs by ambient glutamate is no longer possible under each of these circumstances, changing the shortterm synaptic dynamics. It is possible that the lower ambient glutamate contributes to the lack of tonic KAR activation after the first postnatal week. However, it does seem that the affinity of the receptor actually changes, making it less sensitive to the agonist. Indeed, it has been claimed that a change in an isoform of the GluK1 subunit in KARs is responsible for this lower affinity (Vesikansa et al., 2012). The inhibition of synaptic release under these circumstances seems to be mediated by noncanonical KAR signaling. Another interesting situation in which a change in the properties of KARs takes place during development is in CA3 interneurons, where the firing rate is controlled by the KAR-mediated tonic inhibition of IAHP during the first postnatal week (Segerstra˚le et al., 2010). One more example of how KARs may control network activity during development is provided by the reduced glutamatergic input to CA3 pyramidal cells following tonic KAR activation and the simultaneous facilitation of glutamate release onto CA3 interneurons (Lauri et al., 2005). This action permits network bursting in the developing hippocampus. All in all, these data imply a role for KARs in driving network activity during maturation, when synchronous neuronal oscillations are important for the development of synaptic circuits (e.g., Zhang and Poo, 2001). KARs also seem to contribute to the development of neuronal connectivity by guiding the morphological development of the neuronal synaptic network (i.e., the tracks and the formation of early synaptic contacts). In GluK2-deficient animals, the functional maturation of MF-CA3 synaptic contacts that normally occurs between postnatal day 6 (P6) and P9 is delayed (Lanore et al., 2012). In the early contact and rearrangement stages, growth cone motility is essential for the axon to explore its environment and find its appropriate synaptic targets (Goda and Davis, 2003). In the developing hippocampus, KARs bidirectionally regulate the motility of filopodia in a developmentally regulated and concentration-dependent manner, increasing filopodia motility upon activation with low concentrations of KA and decreasing it in the presence of high concentrations of KA (Tashiro et al., 2003). These data support a two-step model of synaptogenesis, whereby low concentrations of glutamate early in development enhance motility by activating KARs to promote the localization of synaptic targets. Having established the nascent synapse, the increase in glutamate concentrations as a consequence of the reduction in extracellular volume may then reduce filopodia motility, prompting stabilization of the contact (Tashiro et al., 2003). This model is also consistent with the observation that filopodia motility is related to the free extracel-

lular space in which it is found, displaying lower motility as the free extracellular space diminishes (Tashiro et al., 2003). In this regard, KARs may represent sensors for the axonal filopodia to probe their immediate environment and, hence, it may be essential for guidance and the formation of synaptic contacts. Together, these data demonstrate a critical role for KARs in the development of synaptic connectivity and in the maturation of neuronal networks. In particular, how altering KAR activity during development highlights the key role fulfilled by these receptors when synaptic networks are established. However, we are only just beginning to understand the role of KARs in neuronal development and maturation, and the exact mechanisms underlying such events remain not defined. Putative Roles of KARS in Brain Disease Abnormalities in glutamatergic neurotransmission are considered to be an important factor contributing to neurodegenerative and mental disorders (e.g., Frankle et al., 2003). Kainate receptors have been linked to a number of brain disorders such as epilepsy, schizophrenia, and autism, yet their role in brain pathologies appears at times contradictory. Although the experimental data now available indicate a number of putative roles for KARs in mood disorders, the data available are not free of caveats (see Table 2). Mood Disorders Perhaps the most fascinating results come from the studies that potentially connect KARs with schizophrenia and bipolar disorders. On the one hand, postmortem studies provided evidence of a change in KAR subunits in schizophrenic brains (Benes et al., 2001), although these were not corroborated in other studies. For instance, a careful quantitative study of glutamate receptor mRNA expression failed to detect any change in KAR subunit expression in dissected thalamic nuclei from the brains of subjects diagnosed with schizophrenia (Dracheva et al., 2008). On the other hand, postmortem gene expression profiling indicated that in the hippocampus, parahippocampus, and the prefrontal cortex, at least, there is a decrease in the mRNAencoding GluK1 subunits (Scarr et al., 2005). Obviously it is difficult to evaluate the availability of protein from mRNA quantification, and given the absence of a specific GluK1 antibody, these data await further verification. Recent GWAS studies of thousands of cases indicated a polygenic basis to schizophrenia, identifying SNPs that are shared with bipolar disorder but not with other nonpsychiatric diseases (Ripke et al., 2011; Sklar et al., 2011). The common involvement of several genes in a disease complicates the reproduction of those diseases in experimental models, as it would not be expected that a single mutation could fully reproduce the syndrome. In the case of KARs, this is exemplified by the fact that an SNP for Grik4 (rs1954787) is more abundant in subjects responding to antidepressant treatment with a serotonin uptake inhibitor (citalopram) than in patients that do not (Paddock et al., 2007). This SNP is located in the 30 region of the first intron of Grik4 gene and, while it does not directly affect the protein sequence, it seems to alter gene expression. Similarly, there are data suggesting that Grik3 might be a susceptibility gene for major depressive disorder, whereby the SNP T928G (rs6691840) that causes an S to A alteration in the extracellular Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 301

Neuron

Review Table 2. KARs Most Salient Linkage to Mood Disorders Gene Data

Linked Disease

Behavioral Test in KO

References

Grik1 Upregulated expression

Epilepsy

No

Sander et al., 1997; Izzi et al., 2002; Li et al., 2010; Lucarini et al., 2007

Grik2 Modest linkage

Autism

No

Jamain et al. 2002; Shuang et al., 2004; Szatmari et al., 2007; Freitag 2007; but see Dutta et al., 2007

Grik2 Deletion of exons 7 and 8

Mania, mild mental No retardation

Motazacker et al., 2007; Shaltiel et al., 2008; Lanore et al., 2012

Grik2 Mapping susceptibility locus Schizophrenia

Yes

Beneyto et al., 2007; Shaltiel et al., 2008; but see Shibata et al., 2002, 2006

Grik2 mapping

Huntington

No

MacDonald et al., 1999; Chattopadhyay et al., 2003; but see Lee et al., 2012 and Diguet et al., 2004

Grik3 SNP T928G (rs6691840)

Schizophrenia

No

Begni et al., 2002; Kilic et al., 2010; Ahmad et al., 2009;

Grik3 SNP T928G (rs6691840)

Major depression

No

Schiffer and Heinemann, 2007; Wilson et al., 2006

Grik4 Treatment response

Depression

No

Paddock et al., 2007

Grik4 14 bp deletion/insertion variant

Bipolar disorder

Yes

Pickard et al., 2008; Catches et al., 2012; Lowry et al., 2013

Grik4 SNPs rs2282586 and rs1944522

Protection against Schizophrenia

Yes

Pickard et al., 2006

domain of GluK3, is in linkage disequilibrium with recurrent major depressive disorder patients (Schiffer and Heinemann, 2007) and subjects with schizophrenia (Begni et al., 2002; Kilic et al., 2010; Djurovic et al., 2009; Ge´cz et al., 1999). Indeed, the distribution of the GG homozygous genotype was significantly higher in schizophrenia patients than in controls (Ahmad et al., 2009; Kilic et al., 2010). However, this amino acid change does not have any detectable functional consequence in the receptor (Schiffer et al., 2000), although it could convey aberrant gene dosage and/or unequal allele expression (Schiffer et al., 2000; Wilson et al., 2006). Indeed, mRNAs for GluK3 and other glutamate receptors are reduced in the frontal cortex of schizophrenic subjects (Sokolov, 1998; but see Meador-Woodruff et al., 2001). As for other subunits, GluK3 gene expression is developmentally regulated and aberrant gene dosage during development may impact disease in adulthood (Wilson et al., 2006). Thus, further experiments using transgenic animals are warranted. A clear example of gene dosage is provided by trisomy of chromosome 21, leading to Down syndrome. Grik1, the gene coding for GluK1 subunits, is located on human chromosome 21q22.1, and genetic mapping places Grik1 in the vicinity of genes coding for APP and super oxide dismutase (SOD1; Gregor et al., 1994). However, linkage analysis failed to detect any association with familial amyotrophic lateral sclerosis and there are no data indicating any role for GluK1 gene disequilibrium dosage in Down syndrome. Based on multiple regression analyses, it appears that the effects of anxiety and depression treatment are significantly and independently associated with the Grik4 gene (Paddock et al., 2007). An association was also observed in female patients with markers in Grik1. Together, these data indicate that reduced expression of Grik1, Grik4, and other genes encoding KAR subunits could be implicated in mood disorders (but see Li et al., 2008). However, the sign of this implication is clearly elusive and these linkages may be circumstantial given that causal mutations have not yet been identified through linkage or candidate gene association studies. It is becoming clear that no con302 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

clusions can be reached without more precise information of the role of these subunits in general brain physiology. However, recent studies using experimental models have started to assess how the absence of one of these genes affects behavior. The ablation of Grik4 in mice results in marked hyperactivity (Catches et al., 2012; Lowry et al., 2013), one of the endophenotypes of patients with bipolar disorders, which has been interpreted as if lack of GluK4 activity has an anxiolytic and antidepressivelike effect (Catches et al., 2012). Anxiety and depression are concurrent with bipolar disorders, and these data would in principle support the hypothesis that GluK4 hyperactivity could be a hallmark of bipolar phenotypes. However, genetic data from bipolar patients seem to refute this conclusion. In a case control association study, two SNP haplotypes (rs2282586 and rs1944522) exhibited a protective effect against bipolar disorder in a diverse Scottish population (Pickard et al., 2006). Subsequent studies identified a 14 bp deletion/insertion variant in the 30 UTR of the Grik4 gene in subjects carrying a protective haplotype with a significant association to the risk of bipolar disorders (Pickard et al., 2008). Surprisingly, the deletion allele was negatively associated with bipolar disorder and it resulted in an increase in the abundance of both GluK4 mRNA (Pickard et al., 2008) and protein in the hippocampus and prefrontal cortex (Knight et al., 2012). This resulted from the fact that the mRNA bearing the deletion seems to be more stable and persistent than that bearing the insertion, resulting in an increase in GluK4 protein of up to 90% in the hippocampus and 40% in the cortex of the human brain. Consequently, one would expect that mice either deficient for this gene or with GluK4 hypoactivity would display behavior associated to bipolar disorders rather than expressing an antianxiolytic or antidepressive phenotype. Indeed, in the forced swimming test, immobility is reduced by a number of antidepressant drugs in normal mice, indicating that such immobility may be read out of depressive-like behavior. As in GluK4 KO mice, GluK2-deficient animals show less immobility than wildtype (WT) mice, although chronic lithium treatment reduced

Neuron

Review immobility in these GluK2-deficient mice to the same extent as in WT mice (Shaltiel et al., 2008). If the lack of GluK2 were antidepressive, it should occlude the action of lithium, as lithium has no effect in normal subjects. Therefore, it is possible that less immobility would reflect anxiogenicity rather than less depression. Actually, this kind of test of behavioral despair was designed as a test for the primary screening of antidepressant drugs (Porsolt et al., 1977). Therefore, when using this test, it is difficult to deduce an antidepressive state through less immobility without directly checking the action of the antidepressants. Nevertheless, mice in which Grik4 is deleted also display a schizophrenic phenotype and, indeed, GluK4 KO mice show impaired paired-pulse inhibition, mirroring one of the endophenotypes of patients with schizophrenia (Lowry et al., 2013). These data are in keeping with the presence of three SNPs of the Grik4 gene in a patient with chronic schizophrenia and mild mental retardation (Blackwood et al., 2007, 2008; Pickard et al., 2006). This patient was found to present a complex rearrangement of a segment of chromosome 11, involving chromosomes 2 and 8. The Grik4 gene was disrupted at a breakpoint situated at 11q23.3 and the expected outcome was the truncation of all putative transcripts such that the protein encoded would not be functional (Blackwood et al., 2007; Pickard et al., 2006). This means that knocking out Grik4 would result in symptoms of schizophrenia and/or mental disability. Indeed, the GluK4 KO does present learning deficits (Lowry et al., 2013; but see Catches et al., 2012). Interestingly, treatment with neuroleptic drugs seems to restore levels of GluK4 mRNA expression that are abnormally low (ca. 50%) in the frontal cortex of schizophrenics (Sokolov, 1998). Clearly, SNPs found in the Grik4 gene as markers of schizophrenia and bipolar disorders confirm a role for this gene as a risk factor for mood disorders. The SNPs vary with the disease, with SNPs at the center of the gene at chromosome 6q11 associated with schizophrenia and SNPs at the gene’s 30 end associated with bipolar disorders. Clarification of aspects mentioned above is critical for envisioning therapeutic opportunities. On the one hand, data from patients suggest that a pharmacologically mediated increase in KAR activity might be beneficial to protect against bipolar disorders, while on the other hand, behavioral data from mice (e.g., GluK4-deficient mice) may open the door to therapeutic opportunities for antagonists (e.g., of GluK4). However, this latter approach would be detrimental to other phenotypes, such as schizophrenia. The uncertainty of interpreting behavioral data in mice must also be born in mind and, as mentioned above, reduced immobility of KO mice in the forced swimming test has been interpreted as antidepressant in some cases and as a sign of mania in others. A significant decrease in GluK2 mRNA expression has been reported in schizophrenic subjects (Porter et al., 1997). Interestingly, this gene maps close to a locus of schizophrenia susceptibility on chromosome 6 (6q16.3-q21) (Bah et al., 2004), although no association between this gene and schizophrenia could be demonstrated after studying 15 SNPs evenly distributed over the entire Grik2 region, ruling out a major role of GluK2 in the pathogenesis of schizophrenia (Shibata et al., 2002). However, several genome-wide studies have shown significant linkage between bipolar disorders and chromosome

6q21 (McQueen et al., 2005), where Grik2 maps, and GluK2 mRNA expression is also reduced in the brain of bipolar patients (Beneyto et al., 2007). Interestingly, Grik2 KO mice exhibit a variety of behaviors, including hyperactivity, aggressiveness, and sensitivity to psychostimulants, reproducing in mice the behavioral symptoms of mania in humans (Shaltiel et al., 2008). However, it is not currently possible to infer whether GluK2 is involved in the pathophysiology of mania and/or susceptibility to bipolar disorders, or if it is just related to some features of their symptoms. Mental Retardation In one of the eight genomic loci linked to nonsyndromic autosomal recessive mental retardation in a study of 78 consanguineous Iranian families, gene defects were revealed precisely in an interval on chromosome 6116.1-q21. This locus contains 25 annotated genes, including Grik2, which was screened for DNA mutations in patients with mental retardation. Only one single nonpolymorphic sequence change was detected, involving a deletion that removed exons 7 and 8 of the Grik2 gene (Motazacker et al., 2007). Although a truncated GluK2 subunit was generated, it was unable to contribute to functional receptors, suggesting a severe hypofunction in glutamatergic signaling through KARs that would alter local brain circuits and induce cognitive impairment. An attractive hypothesis proposed in this study was that defects in GluK2 signaling may be important for correct circuit maturation in areas important for higher brain functions, such as the hippocampus. This hypothesis was recently tested in GluK2-deficient animals in which a delay in the functional maturation of MF to CA3 synaptic contacts occurs between P6 and P9 (Lanore et al., 2012). Although this transient defect in synaptic transmission may be critical for correct postnatal development, it is not clear how it may affect hippocampal performance in adults beyond the fact that the absence of KARs at MF to CA3 synapses alters the integrative properties of the hippocampal trisynaptic circuit, which is important for memory acquisition in novel environments and sometimes for memory recall (Nakashiba et al., 2008). Grik2 has also been associated with autism (Jamain et al., 2002; Shuang et al., 2004; Freitag, 2007). However, these findings were inconclusive and could not be substantiated by further genome-wide linkage studies in which only a modest positive linkage could be seen in families containing females (Szatmari et al., 2007). Indeed, other studies also failed to find associated markers in individuals with autism spectrum disorder (Dutta et al., 2007) and GluK2 KO mice have not been studied in terms of this pathology. Huntington Disease A number of studies have shown that glutamine repeats in Huntington disease (HD) only account for 50%–60% of the variance at the age of onset (e.g., Snell et al., 1993). This fact prompted a search for functional consequences in the age of onset of polymorphisms associated with HD chromosomes. Among other genes, and given that excitoxicity is considered a potential mechanism for the cell death seen in HD, the Grik2 gene was examined for its potential influence on the age of onset, particularly since it maps to chromosome 6q. More than 4% of the total variance could be attributed to variation in the GluK2 genotype, representing 13% of the variance in the age of onset of HD not Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 303

Neuron

Review accounted for by CAG repeats (Rubinsztein et al., 1997). Therefore, it was concluded that a younger onset of HD is in linkage disequilibrium with a variant of GriK2 or another gene in the region (MacDonald et al., 1999; Chattopadhyay et al., 2003). To reproduce these data in a larger population, the HD CAG repeats and the Grik2 TTA repeats were genotyped in a large (>2,900) population of HD subjects (Snell et al., 1993; Lee et al., 2012). No evidence of an influence of Grik2 polymorphisms on age at motor onset was found and, therefore, there was no support for the role of Grik2 as a genetic modifier of the age of onset in HD. Similarly, a study of the susceptibility of GluK2-deficient mice to 3-nitropropionic acid, a drug used to induce metabolic failure and secondary excitotoxic HD-like damage, was also inconclusive (Diguet et al., 2004). Therefore, despite efforts to link GluK2 to HD, it seems unlikely that KARs are involved in the direct pathogenesis of this disease. Epilepsy There are several lines of evidence strongly suggesting that KARs might be involved in the excitatory to inhibitory imbalances linked to epilepsy. Actually, KA injection has served as an animal model that reproduces details of human temporal lobe epilepsy (TLE). The inhibition of GABA release and the activation of postsynaptic KARs might account for the acute epileptogenic effect of KA (Rodrı´guez-Moreno et al., 1997), although these events do not explain the chronic epilepsy generated months after KA treatment. Actually, the seizures provoked initiate a number of molecular changes and morphological rearrangements in structures with a low epileptogenic threshold, such as the hippocampus. For instance, it is well known that sprouting of glutamatergic fibers takes place in both the KA model of TLE and in human patients and, accordingly, a large number of aberrant synapses are established de novo. These functional aberrant synapses made on granule cells of the dentate gyrus are sprouted MFs and they incorporate KARs, which provide a substantial component of the excitatory input (Epsztein et al., 2005). Thus, aberrant KAR-operated synapses formed under pathological conditions represent a morphological substrate likely to participate in the pathogenesis of TLE (Artinian et al., 2011). The data available from human epileptic tissue indicates an upregulation of GluK1 in the hippocampus of pharmacoresistant TLE patients (e.g., Li et al., 2010), suggesting that rearrangements in neural circuits involving KARs could also take place in humans suffering epilepsy. Although these data should be considered with care due to the poor specificity of some KAR antibodies (e.g., GluK1), it raises the possibility of designing antiepileptic therapies based on the antagonism of KARs. Consistent with KARs influencing this imbalance, the genetic elimination of GluK2 subunits in mice reduced their sensitivity to develop seizures after KA injections (Mulle et al., 1998), illustrating that these receptors contribute to the establishment of overexcitability by exogenous KA that leads to epilepsy. Similarly, exogenous KA reduced GABA release in slices (Clarke et al., 1997; Rodrı´guez-Moreno et al., 1997), dramatically preventing the recurrent inhibition of hippocampal principal neurons in vivo and provoking epileptic activity (Rodrı´guez-Moreno et al., 1997). According to these data, constituting the strongest evidence of the potential therapeutic utility of KARs, a consortium of academic and industry groups (Smolders et al., 2002) showed 304 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

that antagonists of GluK1 (i.e., LY377770 and LY382884) prevent the development of epileptic activity in the CA3 area of hippocampal slices in a model of pilocarpine-induced epileptiform activity. Interestingly, GluK1 antagonists were equally effective in abolishing epileptic activity in slices and in preventing seizures as well as in abolishing seizures once established in vivo. In contrast, others (Khalilov et al., 2002) have indicated a potential for GluK1 agonists as antieplieptic based on the overinhibition largely mediated by GluK1-containing receptors, which are enriched in hippocampal interneurons. The muscarinic agonist pilocarpine is used as a standard model to generate epileptiform activity in order to evaluate the potential of anticonvulsant drugs (cf. Smolders et al., 2002 and references therein). One of the advantages of this model is that it does not involve direct stimulation of KARs, thereby allowing the evaluation of the contribution of tonic KAR activation by ambient glutamate to the epileptic phenomena. It is likely that multiple mechanisms may account for the involvement of KARs in epilepsy. It is possible that the glutamate released due to circuit hyperactivity may provoke both tonic activation of CA3 neurons and KAR-mediated depression of synaptic inhibition. These two actions would be sufficient to generate a drastic imbalance between excitation and inhibition, leading to hippocampal seizures. A similar mechanism has been invoked in the amygdala to account for the therapeutic effects of topiramate (Braga et al., 2009), an approved antiepileptic medicine. A linkage study of 20 families found a significant excess of the Grik1 tetranucleotide polymorphism (nine ‘‘AGTA’’ repeats) in members of families affected by idiopathic juvenile absence epilepsy (Sander et al., 1997). This allelic variant of Grik1 probably confers susceptibility to juvenile absence epilepsy, when superimposed on a background of strong polygenic effects. The tetranucleotide polymorphism maps to the noncoding region of the gene, close to regulatory sequences, and although it does not seem to affect receptor structure (Izzi et al., 2002), it could alter gene expression. However, as there is no evidence of this to date, this association may also be due to a hypothetical epilepsy gene in this region in linkage disequilibrium with Grik1 tetranucleotide repeats (Lucarini et al., 2007). Despite all the evidence linking KARs to epilepsy, to our knowledge no antiepileptic drugs have been developed to date based on KAR antagonists. Pain KARs are expressed strongly in DRG cells and dorsal horn neurons, pointing to a specific role for these receptors in sensory transmission and pain. Indeed, KARs were targeted as potential elements involved in pain transmission and kainate was demonstrated to depolarize primary afferents (Agrawal and Evans, 1986). Moreover, a pure population of KARs was initially isolated from DRG neurons that are likely to be C fiber nociceptors (Huettner, 1990). Molecular and electrophysiological characterization of these neurons led us to conclude that these DRG KARs are made up of heteromeric GluK1 and GluK5 subunits (Sommer et al., 1992; Bahn et al., 1994; Rozas et al., 2003), and since glutamate-induced currents are lost in GluK1-deficient mice, this appears to be the only iGluR expressed in these cells (Mulle et al., 2000; Rozas et al., 2003). Although pain perception cannot be properly considered a disease, persistent or recurrent

Neuron

Review Maturation of synaptic networks during development

Kainate Receptors

Postsynaptic depolarization at some excitatory synapses

Enhancement of neuronal excitability

Presynaptic modulation of Glu and GABA release

betw Balance

een ex

citation

and inhi

apeutic trials appear to have been abandoned (see Bhangoo and Swanson, 2013 and references therein). Thus, the genetic linkage of KAR subunits to diseases are extremely illustrative as to the diseases that may be influenced or triggered by KARs, represent promising lines for further studies into their mechanistic causes. However, much work remains to be done before definitive conclusions can be drawn regarding the exact roles of KARs in brain disease.

bition

Activity of Neuronal Circuits

Brain Performance

Figure 5. Diagram Illustrating How KARs Could Influence Cognitive Functions by Modifying Key Functional Features of Neuronal and Circuit Activity Any alteration in the regulation of these activities, including circuit maturation during development, may provoke sufficient disequilibrium as to lead to a disease state.

pain is associated to a number of disorders of distinct origins and pathophysiological bases, including neuropathic pain. Initial support for the involvement of KARs in pain transmission came from the fact that several KAR antagonists possess analgesic activity in a number of animal models of pain. For instance, SYM 2081 increases the latency of escape in the hot plate and chronic constriction injury tests, presumably acting as a functional antagonist (Sutton et al., 1999), whereas the antagonist of GluK1-containing receptors, LY382884, decreases the frequency of paw licking induced by the subcutaneous injection of formalin (Simmons et al., 1998). In keeping with these results, the ablation of Grik1 gene mitigates pain-associated behavior (Ko et al., 2005; see Bhangoo and Swanson, 2013 for a review and references therein). Interestingly, the activation of primary afferent sensory fibers produces a kainate receptor-mediated EPSC on the dorsal horn neurons (Li et al., 1999). As in the CNS, these synaptic responses are characterized by slow onset and decay time constants. A remarkable feature of these KAR-mediated EPSCs is that they can only be elicited upon nerve stimulation at intensities strong enough to activate the high-threshold Ad and C fibers. This feature raises the possibility that KARs may be exclusively involved in nociceptive transmission at this level, a hypothesis that received significant support when opiate agonists were shown to reduce the amplitude of the KAR-mediated EPSC in dorsal horn neurons (Li et al., 1999). In addition, this receptor subtype is also expressed by trigeminal neurons (Sahara et al., 1997) and KARs are generally expressed along nociceptive pathways, from DRG neurons to the cortex (see Wu et al., 2007 for a review). The strong indications that GluK1 antagonists modulate pain perception have led to several clinical trials to validate KARs as therapeutic targets for pain treatment (reviewed by Bhangoo and Swanson, 2013). While some of these demonstrated certain efficacy, and positive results were reported in phases I and II for migraine, postoperative pain, and analogous cases, these ther-

Perspective It is now apparent that KARs play different roles in synaptic transmission and its modulation to those fulfilled by other glutamate receptors, such as AMPARs and NMDARs (see Figure 5), yet there is still much to be learned concerning the role of KARs in brain performance. Several functional implications directly emanate from the role played by KARs as ion channel forming receptors at synapses, including a role in short- and long-term synaptic plasticity. New and unexpected roles for KARs come from their capacity to signal though noncanonical metabotropic pathway. Although the importance of both signaling modes has been demonstrated in neuronal physiology, it is unclear which may be more relevant and under which circumstances, something we hope will be revealed in the near future. Similarly, it remains unclear which subunits may be responsible for coupling to G proteins and how an ion channel couples to and activates a G protein. These questions, relevant to fully understand KARs, await further advances. It is also necessary to more strictly examine the role of KARs in brain disease, as indicated by the linkage of SNPs and mutations in KAR encoding genes to several devastating diseases, such as schizophrenia and bipolar disorders, the most promising syndromes linked to KAR malfunction. Such studies should benefit from the already abundant information of the roles played by KARs in synaptic physiology, and the availability of KO and transgenic models will be particularly beneficial in this enterprise. Nevertheless, new models are still to be developed. These experiments will reveal how KARs participate in normal behavior and whether they are suitable targets for therapeutic interventions. The plethora of proteins able to interact with KARs, some of them demonstrated to be true ancillary proteins, opens a new field of research to analyze their role not only in pacing affinity and channel gating but also in the polarized trafficking of these receptors. How do they get into the presynaptic terminals? How do they get into the synaptic spines? Is there a specific role for abundant extrasynaptic KARs? Are all these proteinprotein interactions regulated by neuronal activity or any other functional factors? In summary, after 20 years of research following their functional identification in CNS neurons, KARs remain vaguely defined entities. There is a lot of information available but understanding the functions of KARs still lags behind that of other glutamate receptors and a comprehensive model is still lacking. The potential of these receptors as targets for new therapeutic interventions is extensive and could well represent just the tip of an iceberg. The detailed study of currently available KAR-deficient mice and the development of new animal models (e.g., conditional KOs and mice overexpressing KARs) should fuel Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 305

Neuron

Review progress in this area, perhaps unraveling how these receptors may more efficiently serve as therapeutic targets. ACKNOWLEDGMENTS The authors’ research is supported by grants to J.L. from the Spanish MICINN (BFU2011-24084), CONSOLIDER (CSD2007-00023), and Prometeo/2011/ 086. J.M.M. was supported by a PhD fellowship from the Portuguese Fundac¸a˜o para a Cieˆncia e a Tecnologia. Discussion and input from the members of J.L.’s laboratory are warmly acknowledged. REFERENCES Agrawal, S.G., and Evans, R.H. (1986). The primary afferent depolarizing action of kainate in the rat. Br. J. Pharmacol. 87, 345–355. Ahmad, Y., Bhatia, M.S., Mediratta, P.K., Sharma, K.K., Negi, H., Chosdol, K., and Sinha, S. (2009). Association between the ionotropic glutamate receptor kainate3 (GRIK3) Ser310Ala polymorphism and schizophrenia in the Indian population. World J. Biol. Psychiatry 10, 330–333. Ali, A.B., Rossier, J., Staiger, J.F., and Audinat, E. (2001). Kainate receptors regulate unitary IPSCs elicited in pyramidal cells by fast-spiking interneurons in the neocortex. J. Neurosci. 21, 2992–2999. Artinian, J., Peret, A., Marti, G., Epsztein, J., and Cre´pel, V. (2011). Synaptic kainate receptors in interplay with INaP shift the sparse firing of dentate granule cells to a sustained rhythmic mode in temporal lobe epilepsy. J. Neurosci. 31, 10811–10818. Bah, J., Quach, H., Ebstein, R.P., Segman, R.H., Melke, J., Jamain, S., Rietschel, M., Modai, I., Kanas, K., Karni, O., et al. (2004). Maternal transmission disequilibrium of the glutamate receptor GRIK2 in schizophrenia. Neuroreport 15, 1987–1991. Bahn, S., Volk, B., and Wisden, W. (1994). Kainate receptor gene expression in the developing rat brain. J. Neurosci. 14, 5525–5547. Barberis, A., Sachidhanandam, S., and Mulle, C. (2008). GluR6/KA2 kainate receptors mediate slow-deactivating currents. J. Neurosci. 28, 6402–6406. Begni, S., Popoli, M., Moraschi, S., Bignotti, S., Tura, G.B., and Gennarelli, M. (2002). Association between the ionotropic glutamate receptor kainate 3 (GRIK3) ser310ala polymorphism and schizophrenia. Mol. Psychiatry 7, 416–418. Benes, F.M., Todtenkopf, M.S., and Kostoulakos, P. (2001). GluR5,6,7 subunit immunoreactivity on apical pyramidal cell dendrites in hippocampus of schizophrenics and manic depressives. Hippocampus 11, 482–491. Beneyto, M., Kristiansen, L.V., Oni-Orisan, A., McCullumsmith, R.E., and Meador-Woodruff, J.H. (2007). Abnormal glutamate receptor expression in the medial temporal lobe in schizophrenia and mood disorders. Neuropsychopharmacology 32, 1888–1902.

Braga, M.F.M., Aroniadou-Anderjaska, V., and Li, H. (2004). The physiological role of kainate receptors in the amygdala. Mol. Neurobiol. 30, 127–141. Braga, M.F., Aroniadou-Anderjaska, V., Li, H., and Rogawski, M.A. (2009). Topiramate reduces excitability in the basolateral amygdala by selectively inhibiting GluK1 (GluR5) kainate receptors on interneurons and positively modulating GABAA receptors on principal neurons. J. Pharmacol. Exp. Ther. 330, 558–566. Bureau, I., Dieudonne, S., Coussen, F., and Mulle, C. (2000). Kainate receptormediated synaptic currents in cerebellar Golgi cells are not shaped by diffusion of glutamate. Proc. Natl. Acad. Sci. USA 97, 6838–6843. Caiati, M.D., Sivakumaran, S., and Cherubini, E. (2010). In the developing rat hippocampus, endogenous activation of presynaptic kainate receptors reduces GABA release from mossy fiber terminals. J. Neurosci. 30, 1750–1759. Carta, M., Opazo, P., Veran, J., Athane´, A., Choquet, D., Coussen, F., and Mulle, C. (2013). CaMKII-dependent phosphorylation of GluK5 mediates plasticity of kainate receptors. EMBO J. 32, 496–510. Castillo, P.E., Malenka, R.C., and Nicoll, R.A. (1997). Kainate receptors mediate a slow postsynaptic current in hippocampal CA3 neurons. Nature 388, 182–186. Catches, J.S., Xu, J., and Contractor, A. (2012). Genetic ablation of the GluK4 kainate receptor subunit causes anxiolytic and antidepressant-like behavior in mice. Behav. Brain Res. 228, 406–414. Chamberlain, S.E.L., Gonza´lez-Gonza´lez, I.M., Wilkinson, K.A., Konopacki, F.A., Kantamneni, S., Henley, J.M., and Mellor, J.R. (2012). SUMOylation and phosphorylation of GluK2 regulate kainate receptor trafficking and synaptic plasticity. Nat. Neurosci. 15, 845–852. Chamberlain, S.E.L., Sadowski, J.H.L.P., Teles-Grilo Ruivo, L.M., Atherton, L.A., and Mellor, J.R. (2013). Long-term depression of synaptic kainate receptors reduces excitability by relieving inhibition of the slow afterhyperpolarization. J. Neurosci. 33, 9536–9545. Chattopadhyay, B., Ghosh, S., Gangopadhyay, P.K., Das, S.K., Roy, T., Sinha, K.K., Jha, D.K., Mukherjee, S.C., Chakraborty, A., Singhal, B.S., et al. (2003). Modulation of age at onset in Huntington’s disease and spinocerebellar ataxia type 2 patients originated from eastern India. Neurosci. Lett. 345, 93–96. Chittajallu, R., Vignes, M., Dev, K.K., Barnes, J.M., Collingridge, G.L., and Henley, J.M. (1996). Regulation of glutamate release by presynaptic kainate receptors in the hippocampus. Nature 379, 78–81. Christensen, J.K., Paternain, A.V., Selak, S., Ahring, P.K., and Lerma, J. (2004). A mosaic of functional kainate receptors in hippocampal interneurons. J. Neurosci. 24, 8986–8993. Clarke, V.R., Ballyk, B.A., Hoo, K.H., Mandelzys, A., Pellizzari, A., Bath, C.P., Thomas, J., Sharpe, E.F., Davies, C.H., Ornstein, P.L., et al. (1997). A hippocampal GluR5 kainate receptor regulating inhibitory synaptic transmission. Nature 389, 599–603.

Bhangoo, S.K., and Swanson, G.T. (2013). Kainate receptor signaling in pain pathways. Mol. Pharmacol. 83, 307–315.

Clarke, V.R., Collingridge, G.L., Lauri, S.E., and Taira, T. (2012). Synaptic kainate receptors in CA1 interneurons gate the threshold of theta-frequencyinduced long-term potentiation. J. Neurosci. 32, 18215–18226.

Blackwood, D.H.R., Pickard, B.J., Thomson, P.A., Evans, K.L., Porteous, D.J., and Muir, W.J. (2007). Are some genetic risk factors common to schizophrenia, bipolar disorder and depression? Evidence from DISC1, GRIK4 and NRG1. Neurotox. Res. 11, 73–83.

Contractor, A., Swanson, G., and Heinemann, S.F. (2001). Kainate receptors are involved in short- and long-term plasticity at mossy fiber synapses in the hippocampus. Neuron 29, 209–216.

Blackwood, D.H.R., Thiagarajah, T., Malloy, P., Pickard, B.S., and Muir, W.J. (2008). Chromosome abnormalities, mental retardation and the search for genes in bipolar disorder and schizophrenia. Neurotox. Res. 14, 113–120. Bonfardin, V.D.J., Fossat, P., Theodosis, D.T., and Oliet, S.H.R. (2010). Gliadependent switch of kainate receptor presynaptic action. J. Neurosci. 30, 985–995. Bowie, D. (2002). External anions and cations distinguish between AMPA and kainate receptor gating mechanisms. J. Physiol. 539, 725–733. Bowie, D. (2010). Ion-dependent gating of kainate receptors. J. Physiol. 588, 67–81.

306 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

Contractor, A., Mulle, C., and Swanson, G.T. (2011). Kainate receptors coming of age: milestones of two decades of research. Trends Neurosci. 34, 154–163. Copits, B.A., and Swanson, G.T. (2012). Dancing partners at the synapse: auxiliary subunits that shape kainate receptor function. Nat. Rev. Neurosci. 13, 675–686. Copits, B.A., and Swanson, G.T. (2013a). Lateral thinking: CaMKII uncouples kainate receptors from mossy fibre synapses. EMBO J. 32, 487–489. Copits, B.A., and Swanson, G.T. (2013b). Kainate receptor post-translational modifications differentially regulate association with 4.1N to control activitydependent receptor endocytosis. J. Biol. Chem. 288, 8952–8965.

Neuron

Review Copits, B.A., Robbins, J.S., Frausto, S., and Swanson, G.T. (2011). Synaptic targeting and functional modulation of GluK1 kainate receptors by the auxiliary neuropilin and tolloid-like (NETO) proteins. J. Neurosci. 31, 7334–7340. Cossart, R., Esclapez, M., Hirsch, J.C., Bernard, C., and Ben-Ari, Y. (1998). GluR5 kainate receptor activation in interneurons increases tonic inhibition of pyramidal cells. Nat. Neurosci. 1, 470–478. Cossart, R., Tyzio, R., Dinocourt, C., Esclapez, M., Hirsch, J.C., Ben-Ari, Y., and Bernard, C. (2001). Presynaptic kainate receptors that enhance the release of GABA on CA1 hippocampal interneurons. Neuron 29, 497–508. Coussen, F., and Mulle, C. (2006). Kainate receptor-interacting proteins and membrane trafficking. Biochem. Soc. Trans. 34, 927–930. Coussen, F., Perrais, D., Jaskolski, F., Sachidhanandam, S., Normand, E., Bockaert, J., Marin, P., and Mulle, C. (2005). Co-assembly of two GluR6 kainate receptor splice variants within a functional protein complex. Neuron 47, 555–566. Cunha, R.A., Constantino, M.D., and Ribeiro, J.A. (1997). Inhibition of [3H] gamma-aminobutyric acid release by kainate receptor activation in rat hippocampal synaptosomes. Eur. J. Pharmacol. 323, 167–172. Cunha, R.A., Malva, J.O., and Ribeiro, J.A. (1999). Kainate receptors coupled to G(i)/G(o) proteins in the rat hippocampus. Mol. Pharmacol. 56, 429–433. Cunha, R.A., Malva, J.O., and Ribeiro, J.A. (2000). Pertussis toxin prevents presynaptic inhibition by kainate receptors of rat hippocampal [(3)H]GABA release. FEBS Lett. 469, 159–162. Daw, M.I., Pelkey, K.A., Chittajallu, R., and McBain, C.J. (2010). Presynaptic kainate receptor activation preserves asynchronous GABA release despite the reduction in synchronous release from hippocampal cholecystokinin interneurons. J. Neurosci. 30, 11202–11209. De Waard, M., Liu, H., Walker, D., Scott, V.E., Gurnett, C.A., and Campbell, K.P. (1997). Direct binding of G-protein betagamma complex to voltagedependent calcium channels. Nature 385, 446–450. Delaney, A.J., and Jahr, C.E. (2002). Kainate receptors differentially regulate release at two parallel fiber synapses. Neuron 36, 475–482. Diguet, E., Fernagut, P.-O., Normand, E., Centelles, L., Mulle, C., and Tison, F. (2004). Experimental basis for the putative role of GluR6/kainate glutamate receptor subunit in Huntington’s disease natural history. Neurobiol. Dis. 15, 667–675. Djurovic, S., Ka¨hler, A.K., Kulle, B., Jo¨nsson, E.G., Agartz, I., Le Hellard, S., Hall, H., Jakobsen, K.D., Hansen, T., Melle, I., et al. (2009). A possible association between schizophrenia and GRIK3 polymorphisms in a multicenter sample of Scandinavian origin (SCOPE). Schizophr. Res. 107, 242–248. Dracheva, S., Byne, W., Chin, B., and Haroutunian, V. (2008). Ionotropic glutamate receptor mRNA expression in the human thalamus: absence of change in schizophrenia. Brain Res. 1214, 23–34. Dutta, S., Das, S., Guhathakurta, S., Sen, B., Sinha, S., Chatterjee, A., Ghosh, S., Ahmed, S., Ghosh, S., and Usha, R. (2007). Glutamate receptor 6 gene (GluR6 or GRIK2) polymorphisms in the Indian population: a genetic association study on autism spectrum disorder. Cell. Mol. Neurobiol. 27, 1035–1047. Epsztein, J., Represa, A., Jorquera, I., Ben-Ari, Y., and Cre´pel, V. (2005). Recurrent mossy fibers establish aberrant kainate receptor-operated synapses on granule cells from epileptic rats. J. Neurosci. 25, 8229–8239. Fernandes, H.B., Catches, J.S., Petralia, R.S., Copits, B.A., Xu, J., Russell, T.A., Swanson, G.T., and Contractor, A. (2009). High-affinity kainate receptor subunits are necessary for ionotropic but not metabotropic signaling. Neuron 63, 818–829. Fisahn, A., Heinemann, S.F., and McBain, C.J. (2005). The kainate receptor subunit GluR6 mediates metabotropic regulation of the slow and medium AHP currents in mouse hippocampal neurones. J. Physiol. 562, 199–203. Fisher, J.L., and Mott, D.D. (2012). The auxiliary subunits Neto1 and Neto2 reduce voltage-dependent inhibition of recombinant kainate receptors. J. Neurosci. 32, 12928–12933. Fisher, J.L., and Mott, D.D. (2013). Modulation of homomeric and heteromeric kainate receptors by the auxiliary subunit Neto1. J. Physiol. 2013, 24.

Frankle, W.G., Lerma, J., and Laruelle, M. (2003). The synaptic hypothesis of schizophrenia. Neuron 39, 205–216. Freitag, C.M. (2007). The genetics of autistic disorders and its clinical relevance: a review of the literature. Mol. Psychiatry 12, 2–22. Frerking, M., and Ohliger-Frerking, P. (2002). AMPA receptors and kainate receptors encode different features of afferent activity. J. Neurosci. 22, 7434– 7443. Frerking, M., Malenka, R.C., and Nicoll, R.A. (1998). Synaptic activation of kainate receptors on hippocampal interneurons. Nat. Neurosci. 1, 479–486. Frerking, M., Schmitz, D., Zhou, Q., Johansen, J., and Nicoll, R.A. (2001). Kainate receptors depress excitatory synaptic transmission at CA3—>CA1 synapses in the hippocampus via a direct presynaptic action. J. Neurosci. 21, 2958–2966. Furukawa, H., Singh, S.K., Mancusso, R., and Gouaux, E. (2005). Subunit arrangement and function in NMDA receptors. Nature 438, 185–192. Garcia, E.P., Mehta, S., Blair, L.A., Wells, D.G., Shang, J., Fukushima, T., Fallon, J.R., Garner, C.C., and Marshall, J. (1998). SAP90 binds and clusters kainate receptors causing incomplete desensitization. Neuron 21, 727–739. Ge´cz, J., Barnett, S., Liu, J., Hollway, G., Donnelly, A., Eyre, H., Eshkevari, H.S., Baltazar, R., Grunn, A., Nagaraja, R., et al. (1999). Characterization of the human glutamate receptor subunit 3 gene (GRIA3), a candidate for bipolar disorder and nonspecific X-linked mental retardation. Genomics 62, 356–368. Gelsomino, G., Menna, E., Antonucci, F., Rodighiero, S., Riganti, L., Mulle, C., Benfenati, F., Valtorta, F., Verderio, C., and Matteoli, M. (2013). Kainate induces mobilization of synaptic vesicles at the growth cone through the activation of protein kinase A. Cereb. Cortex 23, 531–541. Goda, Y., and Davis, G.W. (2003). Mechanisms of synapse assembly and disassembly. Neuron 40, 243–264. Goldin, M., Epsztein, J., Jorquera, I., Represa, A., Ben-Ari, Y., Cre´pel, V., and Cossart, R. (2007). Synaptic kainate receptors tune oriens-lacunosum moleculare interneurons to operate at theta frequency. J. Neurosci. 27, 9560–9572. Gouaux, E. (2004). Structure and function of AMPA receptors. J. Physiol. 554, 249–253. Grabauskas, G., Lancaster, B., O’Connor, V., and Wheal, H.V. (2007). Protein kinase signalling requirements for metabotropic action of kainate receptors in rat CA1 pyramidal neurones. J. Physiol. 579, 363–373. Gregor, P., Gaston, S.M., Yang, X.L., O’Regan, J.P., Rosen, D.R., Tanzi, R.E., Patterson, D., Haines, J.L., Horvitz, H.R., Uhl, G.R., et al. (1994). Genetic and physical mapping of the GLUR5 glutamate receptor gene on human chromosome 21. Hum. Genet. 94, 565–570. Hirbec, H., Francis, J.C., Lauri, S.E., Braithwaite, S.P., Coussen, F., Mulle, C., Dev, K.K., Coutinho, V., Meyer, G., Isaac, J.T.R., et al. (2003). Rapid and differential regulation of AMPA and kainate receptors at hippocampal mossy fibre synapses by PICK1 and GRIP. Neuron 37, 625–638. Huettner, J.E. (1990). Glutamate receptor channels in rat DRG neurons: activation by kainate and quisqualate and blockade of desensitization by Con A. Neuron 5, 255–266. Ivakine, E.A., Acton, B.A., Mahadevan, V., Ormond, J., Tang, M., Pressey, J.C., Huang, M.Y., Ng, D., Delpire, E., Salter, M.W., et al. (2013). Neto2 is a KCC2 interacting protein required for neuronal Cl- regulation in hippocampal neurons. Proc. Natl. Acad. Sci. USA 110, 3561–3566. Izzi, C., Barbon, A., Kretz, R., Sander, T., and Barlati, S. (2002). Sequencing of the GRIK1 gene in patients with juvenile absence epilepsy does not reveal mutations affecting receptor structure. Am. J. Med. Genet. 114, 354–359. Jamain, S., Betancur, C., Quach, H., Philippe, A., Fellous, M., Giros, B., Gillberg, C., Leboyer, M., and Bourgeron, T.; Paris Autism Research International Sibpair (PARIS) Study. (2002). Linkage and association of the glutamate receptor 6 gene with autism. Mol. Psychiatry 7, 302–310. Jane, D.E., Lodge, D., and Collingridge, G.L. (2009). Kainate receptors: pharmacology, function and therapeutic potential. Neuropharmacology 56, 90–113.

Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 307

Neuron

Review Jaskolski, F., Coussen, F., and Mulle, C. (2005). Subcellular localization and trafficking of kainate receptors. Trends Pharmacol. Sci. 26, 20–26. Jiang, L., Xu, J., Nedergaard, M., and Kang, J. (2001). A kainate receptor increases the efficacy of GABAergic synapses. Neuron 30, 503–513. Jin, X.-T., and Smith, Y. (2007). Activation of presynaptic kainate receptors suppresses GABAergic synaptic transmission in the rat globus pallidus. Neuroscience 149, 338–349. Kamiya, H., and Ozawa, S. (1998). Kainate receptor-mediated inhibition of presynaptic Ca2+ influx and EPSP in area CA1 of the rat hippocampus. J. Physiol. 509, 833–845. Kamiya, H., and Ozawa, S. (2000). Kainate receptor-mediated presynaptic inhibition at the mouse hippocampal mossy fibre synapse. J. Physiol. 523, 653–665.

Lee, J.-H., Lee, J.-M., Ramos, E.M., Gillis, T., Mysore, J.S., Kishikawa, S., Hadzi, T., Hendricks, A.E., Hayden, M.R., Morrison, P.J., et al.; Registry Study of the European Huntington’s Disease Network; Huntington Study Group COHORT project. (2012). TAA repeat variation in the GRIK2 gene does not influence age at onset in Huntington’s disease. Biochem. Biophys. Res. Commun. 424, 404–408. Lerma, J. (1997). Kainate reveals its targets. Neuron 19, 1155–1158. Lerma, J. (2003). Roles and rules of kainate receptors in synaptic transmission. Nat. Rev. Neurosci. 4, 481–495. Lerma, J. (2006). Kainate receptor physiology. Curr. Opin. Pharmacol. 6, 89–97. Lerma, J. (2011). Net(o) excitement for kainate receptors. Nat. Neurosci. 14, 808–810.

Kamiya, H., Ozawa, S., and Manabe, T. (2002). Kainate receptor-dependent short-term plasticity of presynaptic Ca2+ influx at the hippocampal mossy fiber synapses. J. Neurosci. 22, 9237–9243.

Lerma, J., Paternain, A.V., Naranjo, J.R., and Mellstro¨m, B. (1993). Functional kainate-selective glutamate receptors in cultured hippocampal neurons. Proc. Natl. Acad. Sci. USA 90, 11688–11692.

Khalilov, I., Hirsch, J., Cossart, R., and Ben-Ari, Y. (2002). Paradoxical antiepileptic effects of a GluR5 agonist of kainate receptors. J. Neurophysiol. 88, 523–527.

Lerma, J., Morales, M., Vicente, M.A., and Herreras, O. (1997). Glutamate receptors of the kainate type and synaptic transmission. Trends Neurosci. 20, 9–12.

Kidd, F.L., and Isaac, J.T. (1999). Developmental and activity-dependent regulation of kainate receptors at thalamocortical synapses. Nature 400, 569–573.

Lerma, J., Paternain, A.V., Rodrı´guez-Moreno, A., and Lo´pez-Garcı´a, J.C. (2001). Molecular physiology of kainate receptors. Physiol. Rev. 81, 971–998.

Kilic, G., Ismail Kucukali, C., Orhan, N., Ozkok, E., Zengin, A., Aydin, M., and Kara, I. (2010). Are GRIK3 (T928G) gene variants in schizophrenia patients different from those in their first-degree relatives? Psychiatry Res. 175, 43–46.

Li, H., and Rogawski, M.A. (1998). GluR5 kainate receptor mediated synaptic transmission in rat basolateral amygdala in vitro. Neuropharmacology 37, 1279–1286.

Knight, H.M., Walker, R., James, R., Porteous, D.J., Muir, W.J., Blackwood, D.H.R., and Pickard, B.S. (2012). GRIK4/KA1 protein expression in human brain and correlation with bipolar disorder risk variant status. Am. J. Med. Genet. B. Neuropsychiatr. Genet. 159B, 21–29.

Li, P., Wilding, T.J., Kim, S.J., Calejesan, A.A., Huettner, J.E., and Zhuo, M. (1999). Kainate-receptor-mediated sensory synaptic transmission in mammalian spinal cord. Nature 397, 161–164.

Ko, S., Zhao, M.G., Toyoda, H., Qiu, C.S., and Zhuo, M. (2005). Altered behavioral responses to noxious stimuli and fear in glutamate receptor 5 (GluR5)- or GluR6-deficient mice. J. Neurosci. 25, 977–984.

Li, Z., He, Z., Tang, W., Tang, R., Huang, K., Xu, Z., Xu, Y., Li, L., Li, X., Feng, G., et al. (2008). No genetic association between polymorphisms in the kainatetype glutamate receptor gene, GRIK4, and schizophrenia in the Chinese population. Prog. Neuropsychopharmacol. Biol. Psychiatry 32, 876–880.

Kwon, H.-B., and Castillo, P.E. (2008). Role of glutamate autoreceptors at hippocampal mossy fiber synapses. Neuron 60, 1082–1094. Laezza, F., Wilding, T.J., Sequeira, S., Coussen, F., Zhang, X.Z., Hill-Robinson, R., Mulle, C., Huettner, J.E., and Craig, A.M. (2007). KRIP6: a novel BTB/kelch protein regulating function of kainate receptors. Mol. Cell. Neurosci. 34, 539–550. Laezza, F., Wilding, T.J., Sequeira, S., Craig, A.M., and Huettner, J.E. (2008). The BTB/kelch protein, KRIP6, modulates the interaction of PICK1 with GluR6 kainate receptors. Neuropharmacology 55, 1131–1139. Lancaster, B., and Adams, P.R. (1986). Calcium-dependent current generating the afterhyperpolarization of hippocampal neurons. J. Neurophysiol. 55, 1268– 1282.

Li, J.-M., Zeng, Y.-J., Peng, F., Li, L., Yang, T.-H., Hong, Z., Lei, D., Chen, Z., and Zhou, D. (2010). Aberrant glutamate receptor 5 expression in temporal lobe epilepsy lesions. Brain Res. 1311, 166–174. Liu, Q.S., Patrylo, P.R., Gao, X.B., and van den Pol, A.N. (1999). Kainate acts at presynaptic receptors to increase GABA release from hypothalamic neurons. J. Neurophysiol. 82, 1059–1062. Lourenc¸o, J., Cannich, A., Carta, M., Coussen, F., Mulle, C., and Marsicano, G. (2010). Synaptic activation of kainate receptors gates presynaptic CB(1) signaling at GABAergic synapses. Nat. Neurosci. 13, 197–204. Lourenc¸o, J., Matias, I., Marsicano, G., and Mulle, C. (2011). Pharmacological activation of kainate receptors drives endocannabinoid mobilization. J. Neurosci. 31, 3243–3248.

Lanore, F., Labrousse, V.F., Szabo, Z., Normand, E., Blanchet, C., and Mulle, C. (2012). Deficits in morphofunctional maturation of hippocampal mossy fiber synapses in a mouse model of intellectual disability. J. Neurosci. 32, 17882– 17893.

Lowry, E.R., Kruyer, A., Norris, E.H., Cederroth, C.R., and Strickland, S. (2013). The GluK4 kainate receptor subunit regulates memory, mood, and excitotoxic neurodegeneration. Neuroscience 235, 215–225.

Lauri, S.E., Bortolotto, Z.A., Bleakman, D., Ornstein, P.L., Lodge, D., Isaac, J.T., and Collingridge, G.L. (2001). A critical role of a facilitatory presynaptic kainate receptor in mossy fiber LTP. Neuron 32, 697–709.

Lucarini, N., Verrotti, A., Napolioni, V., Bosco, G., and Curatolo, P. (2007). Genetic polymorphisms and idiopathic generalized epilepsies. Pediatr. Neurol. 37, 157–164.

Lauri, S.E., Bortolotto, Z.A., Nistico, R., Bleakman, D., Ornstein, P.L., Lodge, D., Isaac, J.T.R., and Collingridge, G.L. (2003). A role for Ca2+ stores in kainate receptor-dependent synaptic facilitation and LTP at mossy fiber synapses in the hippocampus. Neuron 39, 327–341.

MacDonald, M.E., Vonsattel, J.P., Shrinidhi, J., Couropmitree, N.N., Cupples, L.A., Bird, E.D., Gusella, J.F., and Myers, R.H. (1999). Evidence for the GluR6 gene associated with younger onset age of Huntington’s disease. Neurology 53, 1330–1332.

Lauri, S.E., Segerstra˚le, M., Vesikansa, A., Maingret, F., Mulle, C., Collingridge, G.L., Isaac, J.T., and Taira, T. (2005). Endogenous activation of kainate receptors regulates glutamate release and network activity in the developing hippocampus. J. Neurosci. 25, 4473–4484.

Maingret, F., Lauri, S.E., Taira, T., and Isaac, J.T. (2005). Profound regulation of neonatal CA1 rat hippocampal GABAergic transmission by functionally distinct kainate receptor populations. J. Physiol. 567, 131–142.

Lauri, S.E., Vesikansa, A., Segerstra˚le, M., Collingridge, G.L., Isaac, J.T.R., and Taira, T. (2006). Functional maturation of CA1 synapses involves activity-dependent loss of tonic kainate receptor-mediated inhibition of glutamate release. Neuron 50, 415–429.

308 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

Mathew, S.S., Pozzo-Miller, L., and Hablitz, J.J. (2008). Kainate modulates presynaptic GABA release from two vesicle pools. J. Neurosci. 28, 725–731. Matute, C. (2011). Therapeutic potential of kainate receptors. CNS Neurosci. Ther. 17, 661–669.

Neuron

Review Mayer, M.L. (2005). Crystal structures of the GluR5 and GluR6 ligand binding cores: molecular mechanisms underlying kainate receptor selectivity. Neuron 45, 539–552. McQueen, M.B., Devlin, B., Faraone, S.V., Nimgaonkar, V.L., Sklar, P., Smoller, J.W., Abou Jamra, R., Albus, M., Bacanu, S.-A., Baron, M., et al. (2005). Combined analysis from eleven linkage studies of bipolar disorder provides strong evidence of susceptibility loci on chromosomes 6q and 8q. Am. J. Hum. Genet. 77, 582–595. Meador-Woodruff, J.H., Davis, K.L., and Haroutunian, V. (2001). Abnormal kainate receptor expression in prefrontal cortex in schizophrenia. Neuropsychopharmacology 24, 545–552. Melyan, Z., Wheal, H.V., and Lancaster, B. (2002). Metabotropic-mediated kainate receptor regulation of IsAHP and excitability in pyramidal cells. Neuron 34, 107–114.

type glutamate receptor gene, GRIK4, in schizophrenia and bipolar disorder. Mol. Psychiatry 11, 847–857. Pickard, B.S., Knight, H.M., Hamilton, R.S., Soares, D.C., Walker, R., Boyd, J.K.F., Machell, J., Maclean, A., McGhee, K.A., Condie, A., et al. (2008). A common variant in the 3’UTR of the GRIK4 glutamate receptor gene affects transcript abundance and protects against bipolar disorder. Proc. Natl. Acad. Sci. USA 105, 14940–14945. Pinheiro, P., and Mulle, C. (2006). Kainate receptors. Cell Tissue Res. 326, 457–482. Pinheiro, P.S., Perrais, D., Coussen, F., Barhanin, J., Bettler, B., Mann, J.R., Malva, J.O., Heinemann, S.F., and Mulle, C. (2007). GluR7 is an essential subunit of presynaptic kainate autoreceptors at hippocampal mossy fiber synapses. Proc. Natl. Acad. Sci. USA 104, 12181–12186.

Melyan, Z., Lancaster, B., and Wheal, H.V. (2004). Metabotropic regulation of intrinsic excitability by synaptic activation of kainate receptors. J. Neurosci. 24, 4530–4534.

Pinheiro, P.S., Lanore, F., Veran, J., Artinian, J., Blanchet, C., Cre´pel, V., Perrais, D., and Mulle, C. (2013). Selective block of postsynaptic kainate receptors reveals their function at hippocampal mossy fiber synapses. Cereb. Cortex 23, 323–331.

Min, M.Y., Melyan, Z., and Kullmann, D.M. (1999). Synaptically released glutamate reduces gamma-aminobutyric acid (GABA)ergic inhibition in the hippocampus via kainate receptors. Proc. Natl. Acad. Sci. USA 96, 9932–9937.

Plested, A.J.R., and Mayer, M.L. (2007). Structure and mechanism of kainate receptor modulation by anions. Neuron 53, 829–841.

Mondin, M., Carta, M., Normand, E., Mulle, C., and Coussen, F. (2010). Profilin II regulates the exocytosis of kainate glutamate receptors. J. Biol. Chem. 285, 40060–40071. Motazacker, M.M., Rost, B.R., Hucho, T., Garshasbi, M., Kahrizi, K., Ullmann, R., Abedini, S.S., Nieh, S.E., Amini, S.H., Goswami, C., et al. (2007). A defect in the ionotropic glutamate receptor 6 gene (GRIK2) is associated with autosomal recessive mental retardation. Am. J. Hum. Genet. 81, 792–798. Mulle, C., Sailer, A., Pe´rez-Otan˜o, I., Dickinson-Anson, H., Castillo, P.E., Bureau, I., Maron, C., Gage, F.H., Mann, J.R., Bettler, B., and Heinemann, S.F. (1998). Altered synaptic physiology and reduced susceptibility to kainate-induced seizures in GluR6-deficient mice. Nature 392, 601–605. Mulle, C., Sailer, A., Swanson, G.T., Brana, C., O’Gorman, S., Bettler, B., and Heinemann, S.F. (2000). Subunit composition of kainate receptors in hippocampal interneurons. Neuron 28, 475–484. Nakashiba, T., Young, J.Z., McHugh, T.J., Buhl, D.L., and Tonegawa, S. (2008). Transgenic inhibition of synaptic transmission reveals role of CA3 output in hippocampal learning. Science 319, 1260–1264. Negrete-Dı´az, J.V., Sihra, T.S., Delgado-Garcı´a, J.M., and Rodrı´guez-Moreno, A. (2006). Kainate receptor-mediated inhibition of glutamate release involves protein kinase A in the mouse hippocampus. J. Neurophysiol. 96, 1829–1837. Ng, D., Pitcher, G.M., Szilard, R.K., Sertie´, A., Kanisek, M., Clapcote, S.J., Lipina, T., Kalia, L.V., Joo, D., McKerlie, C., et al. (2009). Neto1 is a novel CUB-domain NMDA receptor-interacting protein required for synaptic plasticity and learning. PLoS Biol. 7, e41. Paddock, S., Laje, G., Charney, D., Rush, A.J., Wilson, A.F., Sorant, A.J.M., Lipsky, R., Wisniewski, S.R., Manji, H., and McMahon, F.J. (2007). Association of GRIK4 with outcome of antidepressant treatment in the STAR*D cohort. Am. J. Psychiatry 164, 1181–1188. Paternain, A.V., Morales, M., and Lerma, J. (1995). Selective antagonism of AMPA receptors unmasks kainate receptor-mediated responses in hippocampal neurons. Neuron 14, 185–189. Paternain, A.V., Cohen, A., Stern-Bach, Y., and Lerma, J. (2003). A role for extracellular Na+ in the channel gating of native and recombinant kainate receptors. J. Neurosci. 23, 8641–8648.

Plested, A.J.R., Vijayan, R., Biggin, P.C., and Mayer, M.L. (2008). Molecular basis of kainate receptor modulation by sodium. Neuron 58, 720–735. Porsolt, R.D., Bertin, A., and Jalfre, M. (1977). Behavioral despair in mice: a primary screening test for antidepressants. Arch. Int. Pharmacodyn. Ther. 229, 327–336. Porter, R.H., Eastwood, S.L., and Harrison, P.J. (1997). Distribution of kainate receptor subunit mRNAs in human hippocampus, neocortex and cerebellum, and bilateral reduction of hippocampal GluR6 and KA2 transcripts in schizophrenia. Brain Res. 751, 217–231. Ripke, S., Sanders, A.R., Kendler, K.S., Levinson, D.F., Sklar, P., Holmans, P.A., Lin, D.-Y., Duan, J., Ophoff, R.A., Andreassen, O.A., et al.; Schizophrenia Psychiatric Genome-Wide Association Study (GWAS) Consortium. (2011). Genome-wide association study identifies five new schizophrenia loci. Nat. Genet. 43, 969–976. Rivera, R., Rozas, J.L., and Lerma, J. (2007). PKC-dependent autoregulation of membrane kainate receptors. EMBO J. 26, 4359–4367. Rodrigues, R.J., and Lerma, J. (2012). Metabotropic signaling by kainate receptors. WIREs: Membrane Transport and Signaling 1, 399–410. Rodrı´guez-Moreno, A., and Lerma, J. (1998). Kainate receptor modulation of GABA release involves a metabotropic function. Neuron 20, 1211–1218. Rodrı´guez-Moreno, A., Herreras, O., and Lerma, J. (1997). Kainate receptors presynaptically downregulate GABAergic inhibition in the rat hippocampus. Neuron 19, 893–901. Rodrı´guez-Moreno, A., Lo´pez-Garcı´a, J.C., and Lerma, J. (2000). Two populations of kainate receptors with separate signaling mechanisms in hippocampal interneurons. Proc. Natl. Acad. Sci. USA 97, 1293–1298. Rozas, J.L., Paternain, A.V., and Lerma, J. (2003). Noncanonical signaling by ionotropic kainate receptors. Neuron 39, 543–553. Rubinsztein, D.C., Leggo, J., Chiano, M., Dodge, A., Norbury, G., Rosser, E., and Craufurd, D. (1997). Genotypes at the GluR6 kainate receptor locus are associated with variation in the age of onset of Huntington disease. Proc. Natl. Acad. Sci. USA 94, 3872–3876.

Perkinton, M.S., and Sihra, T.S. (1999). A high-affinity presynaptic kainate-type glutamate receptor facilitates glutamate exocytosis from cerebral cortex nerve terminals (synaptosomes). Neuroscience 90, 1281–1292.

Ruiz, A., Sachidhanandam, S., Utvik, J.K., Coussen, F., and Mulle, C. (2005). Distinct subunits in heteromeric kainate receptors mediate ionotropic and metabotropic function at hippocampal mossy fiber synapses. J. Neurosci. 25, 11710–11718.

Perrais, D., Pinheiro, P.S., Jane, D.E., and Mulle, C. (2009). Antagonism of recombinant and native GluK3-containing kainate receptors. Neuropharmacology 56, 131–140.

Sachidhanandam, S., Blanchet, C., Jeantet, Y., Cho, Y.H., and Mulle, C. (2009). Kainate receptors act as conditional amplifiers of spike transmission at hippocampal mossy fiber synapses. J. Neurosci. 29, 5000–5008.

Pickard, B.S., Malloy, M.P., Christoforou, A., Thomson, P.A., Evans, K.L., Morris, S.W., Hampson, M., Porteous, D.J., Blackwood, D.H., and Muir, W.J. (2006). Cytogenetic and genetic evidence supports a role for the kainate-

Sahara, Y., Noro, N., Iida, Y., Soma, K., and Nakamura, Y. (1997). Glutamate receptor subunits GluR5 and KA-2 are coexpressed in rat trigeminal ganglion neurons. J. Neurosci. 17, 6611–6620.

Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 309

Neuron

Review Salinas, G.D., Blair, L.A.C., Needleman, L.A., Gonzales, J.D., Chen, Y., Li, M., Singer, J.D., and Marshall, J. (2006). Actinfilin is a Cul3 substrate adaptor, linking GluR6 kainate receptor subunits to the ubiquitin-proteasome pathway. J. Biol. Chem. 281, 40164–40173.

Sklar, P., Ripke, S., Scott, L.J., Andreassen, O.A., Cichon, S., Craddock, N., Edenberg, H.J., Nurnberger, J.I., Rietschel, M., Blackwood, D., et al.; Psychiatric GWAS Consortium Bipolar Disorder Working Group. (2011). Large-scale genome-wide association analysis of bipolar disorder identifies a new susceptibility locus near ODZ4. Nat. Genet. 43, 977–983.

Sallert, M., Malkki, H., Segerstra˚le, M., Taira, T., and Lauri, S.E. (2007). Effects of the kainate receptor agonist ATPA on glutamatergic synaptic transmission and plasticity during early postnatal development. Neuropharmacology 52, 1354–1365.

Sloviter, R.S., and Damiano, B.P. (1981). On the relationship between kainic acid-induced epileptiform activity and hippocampal neuronal damage. Neuropharmacology 20, 1003–1011.

Salmen, B., Beed, P.S., Ozdogan, T., Maier, N., Johenning, F.W., Winterer, J., Breustedt, J., and Schmitz, D. (2012). GluK1 inhibits calcium dependent and independent transmitter release at associational/commissural synapses in area CA3 of the hippocampus. Hippocampus 22, 57–68.

Smolders, I., Bortolotto, Z.A., Clarke, V.R.J., Warre, R., Khan, G.M., O’Neill, M.J., Ornstein, P.L., Bleakman, D., Ogden, A., Weiss, B., et al. (2002). Antagonists of GLU(K5)-containing kainate receptors prevent pilocarpine-induced limbic seizures. Nat. Neurosci. 5, 796–804.

Sander, T., Hildmann, T., Kretz, R., Fu¨rst, R., Sailer, U., Bauer, G., Schmitz, B., Beck-Mannagetta, G., Wienker, T.F., and Janz, D. (1997). Allelic association of juvenile absence epilepsy with a GluR5 kainate receptor gene (GRIK1) polymorphism. Am. J. Med. Genet. 74, 416–421.

Snell, R.G., MacMillan, J.C., Cheadle, J.P., Fenton, I., Lazarou, L.P., Davies, P., MacDonald, M.E., Gusella, J.F., Harper, P.S., and Shaw, D.J. (1993). Relationship between trinucleotide repeat expansion and phenotypic variation in Huntington’s disease. Nat. Genet. 4, 393–397.

Scarr, E., Beneyto, M., Meador-Woodruff, J.H., and Dean, B. (2005). Cortical glutamatergic markers in schizophrenia. Neuropsychopharmacology 30, 1521–1531.

Sokolov, B.P. (1998). Expression of NMDAR1, GluR1, GluR7, and KA1 glutamate receptor mRNAs is decreased in frontal cortex of ‘‘neuroleptic-free’’ schizophrenics: evidence on reversible up-regulation by typical neuroleptics. J. Neurochem. 71, 2454–2464.

Schiffer, H.H., and Heinemann, S.F. (2007). Association of the human kainate receptor GluR7 gene (GRIK3) with recurrent major depressive disorder. Am. J. Med. Genet. B. Neuropsychiatr. Genet. 144B, 20–26. Schiffer, H.H., Swanson, G.T., Masliah, E., and Heinemann, S.F. (2000). Unequal expression of allelic kainate receptor GluR7 mRNAs in human brains. J. Neurosci. 20, 9025–9033. Schmitz, D., Frerking, M., and Nicoll, R.A. (2000). Synaptic activation of presynaptic kainate receptors on hippocampal mossy fiber synapses. Neuron 27, 327–338. Schmitz, D., Mellor, J., and Nicoll, R.A. (2001). Presynaptic kainate receptor mediation of frequency facilitation at hippocampal mossy fiber synapses. Science 291, 1972–1976. Schmitz, D., Mellor, J., Breustedt, J., and Nicoll, R.A. (2003). Presynaptic kainate receptors impart an associative property to hippocampal mossy fiber long-term potentiation. Nat. Neurosci. 6, 1058–1063. Scott, R., Lalic, T., Kullmann, D.M., Capogna, M., and Rusakov, D.A. (2008). Target-cell specificity of kainate autoreceptor and Ca2+-store-dependent short-term plasticity at hippocampal mossy fiber synapses. J. Neurosci. 28, 13139–13149.

Sommer, B., Burnashev, N., Verdoorn, T.A., Keina¨nen, K., Sakmann, B., and Seeburg, P.H. (1992). A glutamate receptor channel with high affinity for domoate and kainate. EMBO J. 11, 1651–1656. Soto, D., Coombs, I.D., Kelly, L., Farrant, M., and Cull-Candy, S.G. (2007). Stargazin attenuates intracellular polyamine block of calcium-permeable AMPA receptors. Nat. Neurosci. 10, 1260–1267. Straub, C., Hunt, D.L., Yamasaki, M., Kim, K.S., Watanabe, M., Castillo, P.E., and Tomita, S. (2011a). Distinct functions of kainate receptors in the brain are determined by the auxiliary subunit Neto1. Nat. Neurosci. 14, 866–873. Straub, C., Zhang, W., and Howe, J.R. (2011b). Neto2 modulation of kainate receptors with different subunit compositions. J. Neurosci. 31, 8078–8082. Sutton, J.L., Maccecchini, M.L., and Kajander, K.C. (1999). The kainate receptor antagonist 2S,4R-4-methylglutamate attenuates mechanical allodynia and thermal hyperalgesia in a rat model of nerve injury. Neuroscience 91, 283–292. Szatmari, P., Maziade, M., Zwaigenbaum, L., Me´rette, C., Roy, M.A., Joober, R., and Palmour, R. (2007). Informative phenotypes for genetic studies of psychiatric disorders. Am. J. Med. Genet. B. Neuropsychiatr. Genet. 144B, 581–588.

Segerstra˚le, M., Juuri, J., Lanore, F., Piepponen, P., Lauri, S.E., Mulle, C., and Taira, T. (2010). High firing rate of neonatal hippocampal interneurons is caused by attenuation of afterhyperpolarizing potassium currents by tonically active kainate receptors. J. Neurosci. 30, 6507–6514.

Tang, M., Pelkey, K.A., Ng, D., Ivakine, E., McBain, C.J., Salter, M.W., and McInnes, R.R. (2011). Neto1 is an auxiliary subunit of native synaptic kainate receptors. J. Neurosci. 31, 10009–10018.

Selak, S., Paternain, A.V., Aller, M.I., Pico´, E., Rivera, R., and Lerma, J. (2009). A role for SNAP25 in internalization of kainate receptors and synaptic plasticity. Neuron 63, 357–371.

Tang, M., Ivakine, E., Mahadevan, V., Salter, M.W., and McInnes, R.R. (2012). Neto2 interacts with the scaffolding protein GRIP and regulates synaptic abundance of kainate receptors. PLoS ONE 7, e51433.

Shaltiel, G., Maeng, S., Malkesman, O., Pearson, B., Schloesser, R.J., Tragon, T., Rogawski, M., Gasior, M., Luckenbaugh, D., Chen, G., and Manji, H.K. (2008). Evidence for the involvement of the kainate receptor subunit GluR6 (GRIK2) in mediating behavioral displays related to behavioral symptoms of mania. Mol. Psychiatry 13, 858–872.

Tashiro, A., Dunaevsky, A., Blazeski, R., Mason, C.A., and Yuste, R. (2003). Bidirectional regulation of hippocampal mossy fiber filopodial motility by kainate receptors: a two-step model of synaptogenesis. Neuron 38, 773–784.

Shibata, H., Shibata, A., Ninomiya, H., Tashiro, N., and Fukumaki, Y. (2002). Association study of polymorphisms in the GluR6 kainate receptor gene (GRIK2) with schizophrenia. Psychiatry Res. 113, 59–67. Shibata, H., Aramaki, T., Sakai, M., Ninomiya, H., Tashiro, N., Iwata, N., Ozaki, N., and Fukumaki, Y. (2006). Association study of polymorphisms in the GluR7, KA1 and KA2 kainate receptor genes (GRIK3, GRIK4, GRIK5) with schizophrenia. Psychiatry Res. 141, 39–51. Shuang, M., Liu, J., Jia, M.X., Yang, J.Z., Wu, S.P., Gong, X.H., Ling, Y.S., Ruan, Y., Yang, X.L., and Zhang, D. (2004). Family-based association study between autism and glutamate receptor 6 gene in Chinese Han trios. Am. J. Med. Genet. B. Neuropsychiatr. Genet. 131B, 48–50. Simmons, R.M., Li, D.L., Hoo, K.H., Deverill, M., Ornstein, P.L., and Iyengar, S. (1998). Kainate GluR5 receptor subtype mediates the nociceptive response to formalin in the rat. Neuropharmacology 37, 25–36.

310 Neuron 80, October 16, 2013 ª2013 Elsevier Inc.

Tomita, S., and Castillo, P.E. (2012). Neto1 and Neto2: auxiliary subunits that determine key properties of native kainate receptors. J. Physiol. 590, 2217– 2223. Vesikansa, A., Sakha, P., Kuja-Panula, J., Molchanova, S., Rivera, C., Huttunen, H.J., Rauvala, H., Taira, T., and Lauri, S.E. (2012). Expression of GluK1c underlies the developmental switch in presynaptic kainate receptor function. Sci Rep 2, 310. Vignes, M., and Collingridge, G.L. (1997). The synaptic activation of kainate receptors. Nature 388, 179–182. Vignes, M., Clarke, V.R., Parry, M.J., Bleakman, D., Lodge, D., Ornstein, P.L., and Collingridge, G.L. (1998). The GluR5 subtype of kainate receptor regulates excitatory synaptic transmission in areas CA1 and CA3 of the rat hippocampus. Neuropharmacology 37, 1269–1277. Vincent, P., and Mulle, C. (2009). Kainate receptors in epilepsy and excitotoxicity. Neuroscience 158, 309–323.

Neuron

Review Vivithanaporn, P., Yan, S., and Swanson, G.T. (2006). Intracellular trafficking of KA2 kainate receptors mediated by interactions with coatomer protein complex I (COPI) and 14-3-3 chaperone systems. J. Biol. Chem. 281, 15475–15484. Wang, R., Mellem, J.E., Jensen, M., Brockie, P.J., Walker, C.S., Hoerndli, F.J., Hauth, L., Madsen, D.M., and Maricq, A.V. (2012). The SOL-2/Neto auxiliary protein modulates the function of AMPA-subtype ionotropic glutamate receptors. Neuron 75, 838–850. Wilding, T.J., and Huettner, J.E. (1995). Differential antagonism of alphaamino-3-hydroxy-5-methyl-4- isoxazolepropionic acid-preferring and kainate-preferring receptors by 2,3-benzodiazepines. Mol. Pharmacol. 47, 582–587. Wilson, G.M., Flibotte, S., Chopra, V., Melnyk, B.L., Honer, W.G., and Holt, R.A. (2006). DNA copy-number analysis in bipolar disorder and schizophrenia reveals aberrations in genes involved in glutamate signaling. Hum. Mol. Genet. 15, 743–749. Wisden, W., and Seeburg, P.H. (1993). A complex mosaic of high-affinity kainate receptors in rat brain. J. Neurosci. 13, 3582–3598.

Wu, L.J., Ko, S.W., and Zhuo, M. (2007). Kainate receptors and pain: from dorsal root ganglion to the anterior cingulate cortex. Curr. Pharm. Des. 13, 1597–1605. Yan, D., Yamasaki, M., Straub, C., Watanabe, M., and Tomita, S. (2013). Homeostatic control of synaptic transmission by distinct glutamate receptors. Neuron 78, 687–699. Zhang, L.I., and Poo, M.-M. (2001). Electrical activity and development of neural circuits. Nat. Neurosci. Suppl. 4, 1207–1214. Zhang, W., St-Gelais, F., Grabner, C.P., Trinidad, J.C., Sumioka, A., MorimotoTomita, M., Kim, K.S., Straub, C., Burlingame, A.L., Howe, J.R., and Tomita, S. (2009). A transmembrane accessory subunit that modulates kainate-type glutamate receptors. Neuron 61, 385–396. Zhang, Z., Wang, D., Sun, T., Xu, J., Chiang, H.C., Shin, W., and Wu, L.G. (2013). The SNARE proteins SNAP25 and synaptobrevin are involved in endocytosis at hippocampal synapses. J. Neurosci. 33, 9169–9175. Ziegra, C.J., Willard, J.M., and Oswald, R.E. (1992). Coupling of a purified goldfish brain kainate receptor with a pertussis toxin-sensitive G protein. Proc. Natl. Acad. Sci. USA 89, 4134–4138.

Neuron 80, October 16, 2013 ª2013 Elsevier Inc. 311