Water transport between CNS compartments: functional and molecular interactions between aquaporins and ion channels

Neuroscience 168 (2010) 926 –940

REVIEW WATER TRANSPORT BETWEEN CNS COMPARTMENTS: FUNCTIONAL AND MOLECULAR INTERACTIONS BETWEEN AQUAPORINS AND ION CHANNELS V. BENFENATIa AND S. FERRONIb*

between water channels and ion channels in the pathogenesis of astroglia-related acute and chronic diseases of the CNS is also briefly discussed. © 2010 IBRO. Published by Elsevier Ltd. All rights reserved.

a

Istituto per lo Studio dei Materiali Nanostrutturati, ISMN, National Research Council, (CNR), Via Gobetti 101, 40129 Bologna, Italy b

Department of Human and General Physiology, University of Bologna, Via S. Donato 19/2, 40127 Bologna, Italy

Key words: brain homeostasis, potassium buffering, regulatory volume decrease, protein-protein interaction, water channels.

Abstract—The physiological ability of the mammalian CNS to integrate peripheral stimuli and to convey information to the body is tightly regulated by its capacity to preserve the ion composition and volume of the perineuronal milieu. It is well known that astroglial syncytium plays a crucial role in such process by controlling the homeostasis of ions and water through the selective transmembrane movement of inorganic and organic molecules and the equilibration of osmotic gradients. Astrocytes, in fact, by contacting neurons and cells lining the fluid-filled compartments, are in a strategic position to fulfill this role. They are endowed with ion and water channel proteins that are localized in specific plasma membrane domains facing diverse liquid spaces. Recent data in rodents have demonstrated that the precise dynamics of the astroglia-mediated homeostatic regulation of the CNS is dependent on the interactions between water channels and ion channels, and their anchoring with proteins that allow the formation of macromolecular complexes in specific cellular domains. Interplay can occur with or without direct molecular interactions suggesting the existence of different regulatory mechanisms. The importance of molecular and functional interactions is pinpointed by the numerous observations that as consequence of pathological insults leading to the derangement of ion and volume homeostasis the cell surface expression and/or polarized localization of these proteins is perturbed. Here, we critically discuss the experimental evidence concerning: (1) molecular and functional interplay of aquaporin 4, the major aquaporin protein in astroglial cells, with potassium and gap-junctional channels that are involved in extracellular potassium buffering. (2) the interactions of aquaporin 4 with chloride and calcium channels regulating cell volume homeostasis. The relevance of the crosstalk

Contents Astrocyte-mediated control of extracellular potassium homeostasis 928 Potassium channels implicated in the regulation of extracellular potassium concentration 928 Interactions between potassium channels and AQP4 in astroglia 929 Interplay between AQP4 and the gap junction protein Cx43 930 Brain volume homeostasis: interactions of AQP4 with ion channels controlling astrocyte regulatory volume decrease 930 Cell volume regulation in astrocytes 930 Astroglial ion channels involved in regulatory volume decrease 931 Interaction of AQP4 with VRAC 932 Intracellular calcium signalling and cell volume regulation in astrocytes: is there a role for transient receptor potential channels? 932 Conclusions and future perspectives 933 Acknowledgments 934 References 934

The regulation of water and ion balance is crucial for normal brain function. There is indication, in fact, that brain swelling occurring as consequence of several pathological states causes changes in electrolyte concentration and volume of the interstitial space that critically alter the physiological activity of the neuronal network (Chebabo et al., 1995; Hrabetová et al., 2002; Somjen, 2002). Accumulating evidence suggests that brain swelling is the result of the altered exchange processes of organic and inorganic molecules and modification of water transport between the fluid-filled compartments (microvasculature, ventricular and subarachnoid spaces) the astroglial syncytium and the interstitial space (see Kimelberg, 2004a). Astrocytes are the most abundant cell type in mammalian brain (Nedergaard et al., 2003). Their peculiar star-like shape is characterized by cell bodies from which several processes depart to enwrap synapses and brain microvessels (Fig. 1). Because of this spatial relationship astrocytes

*Corresponding author. Tel: ⫹39-051-209-5630; fax: ⫹39-051-209-5629. E-mail address: [email protected] (S. Ferroni). Abbreviations: AQP, aquaporin; [Ca2⫹]o, extracellular calcium concentration; [Ca2⫹]i, intracellular calcium concentration; CCE, capacitative calcium entry; Cx, connexin; DAPC, dystrophin-associated-protein-complex; IEM, immunogold electron microscopy; [K⫹]o, extracellular potassium concentration; [K⫹]i, intracellular potassium concentration; Kir channel, inwardly rectifying K⫹ channel; PLA2, phospholipase A2; RVD, regulatory volume decrease; siRNA, small interfering RNA; TRP, transient receptor potential; TRPV, transient receptor potential vanilloid-related; VRAC, volume-regulated anion channel.

0306-4522/10 $ - see front matter © 2010 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2009.12.017

926

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

927

Fig. 1. Hypothetical astroglial channel-mediated mechanisms involved in extracellular homeostasis. In rodent brain, astrocyte morphology is characterized by irregularly shaped cell body from which depart several leaflet like processes. Some process endfeet cover neuronal soma (blue arrows, a) and enwrap synapses (b). Others take contact with the surface of the brain or abut the blood vessels forming a perivascular sheet (c). Astrocytes are coupled by gap-junctions (d) that allow synchronization of the responses in distal cells of the astroglial syncytium. The astroglial architecture is functional to the homeostatic role of astrocytes, which is crucial for the maintenance of physiological neuronal activity. As result of action potential firing (1), neurotransmitters and ions are released in the perineuronal milieu (2). Astrocytes uptake the extracellularly accumulated potassium (K⫹) and glutamate (Glu⫺) in order to maintain their levels within the physiological range (3): Astroglial Na⫹/Glu⫺ co-transporter (GLT-1) is responsible (in conjunction with GLAST co-transporter) for Glu⫺ uptake (see Anderson and Swanson, 2000). Among the large variety of channels, pumps and transporters expressed by astrocytes, the inwardly rectifying K⫹ channel (Kir4.1) mediates the K⫹ influx at perineuronal endfeet. The excess in intracellular K⫹ is spatially redistributed via gap junctions through the syncytium (4) toward districts where extracellular K⫹ is low, or directly into the interstitial space apposed to fluid-filled cavities (Müller glial cells) (5). At the glia-vasculature interface K⫹ is likely secreted via Kir4.1 because the role in this process of large-conductance, calcium (Ca2⫹)-activated K⫹ (BKCa) channels and Kv channels mediating voltage-gated outward K⫹ currents that are expressed in vivo has not been demonstrated yet. During spatial buffering of K⫹, the transient osmotic gradient created upon K⫹ uptake, is rapidly counteracted by water flux. An exclusive role in osmotic water movement is mediated by AQP4, which is largely distributed at astroglial endfeet surrounding synapses and blood vessels. The intracellular chloride (Cl⫺) concentration, which is controlled by transporters, is critical to set the condition for the passive KCl uptake when operative. The role of Cl⫺ channels of the ClC family that have been described in astroglia in vitro and in situ is still unclear. Volume-regulated anion channels (VRACs) have been identified in vitro and might contribute to cell volume homeostasis. However, whether they are also expressed and active in vivo is unknown (?). The expression of the Ca2⫹ permeable channel named transient receptor potential vanilloid-related type 4 (TRPV4) has been reported both in vivo and in vitro but its functional role is uncertain. Pumps and transporters that contribute to the transmembrane movement of ions have not been included.

are in a critical position for regulating the volume and composition of the interstitial milieu. At the neuron–astrocyte interface the astroglial endfeet uptake and secrete ions, organic molecules, and water (for reviews, see Evanko et al., 2004; Simard and Nedergaard, 2004). In order to ensure ion and water homeostasis of the interstitial fluid opposite processes occur at the endfeet of astrocytes facing the vasculature or those of astroglia contacting the cells lining other fluid-filled compartments (Amiry-Moghaddam and Ottersen, 2003; Kofuji and New-

man, 2004; Nagelhus et al., 2004; Simard and Nedergaard, 2004). It is well known that astrocytes regulate the extracellular fluid homeostasis by using a variety of transmembrane proteins that drive passive and active transports of ions, organic osmolytes, and osmotically obliged water (Pasantes-Morales et al., 2006). By conceiving the astrocyte as a multifunctional unit regulating separated milieus, it is not surprising that ion channels and water channels display a polarized distribution (Fig. 1) and can co-localize

928

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

to form macromolecular complexes able to mediate specific cellular processes (Nielsen et al., 1997; Nagelhus et al., 1999; Amiry-Moghaddam and Ottersen, 2003). Here we will review the concepts on the role of astroglial cells in the control of ion and water homeostasis. We will critically analyze the information available on molecular and functional protein–protein interactions of the main astrocytic water channel, aquaporin 4 (AQP4), with ion channels that are relevant to ion and cell volume homeostasis. The results obtained in Müller glial cells, the main glial component of the retina, will also be reviewed.

ASTROCYTE-MEDIATED CONTROL OF EXTRACELLULAR POTASSIUM HOMEOSTASIS Potassium channels implicated in the regulation of extracellular potassium concentration In the CNS a basic notion for conceiving the information processing mediated by action potential firing is that the rise in extracellular K⫹ ([K⫹]o), which is observed following the re-polarization phase of action potential, is rapidly buffered in the interspaces narrowing synapses. This process avoids the ratio of extracellular versus intracellular K⫹ to reach the ceiling level that can compromise neuronal function. Neuron resting membrane potential, in fact, directly depends on the electrochemical equilibrium potential of K⫹, and failure of [K⫹]o regulation is known to be the cause of severe functional alterations of the neuronal network such as spreading depression, anoxic depolarization, and epileptiform activity (Somjen, 2002; Seifert et al., 2006). The idea that astroglial cells are crucial in the maintenance of [K⫹]o homeostasis found support from the seminal observations that glial cells possess an elevated permeability to K⫹ ions and have a membrane potential that depolarizes during elevated neuronal electrical activity closely following the Nernst equilibrium potential for K⫹ (Kuffler et al., 1966). Those results induced Orkand and colleagues to formulate a revolutionary theory on the role of these cells in the control of [K⫹]o homeostasis called K⫹ spatial buffering (Orkand et al., 1966). Basically, they proposed that in the astroglial syncytium local differences between membrane potential and K⫹ equilibrium potential allow the draining of K⫹ from extracellular compartments in which there is a transient [K⫹]o elevation to those where [K⫹]o is lower. Functional evidence of such a mechanism was also provided in Müller glial cells of the retina extending the concept to K⫹ siphoning (Newman et al., 1984). Path-clamp studies have provided molecular insights in the underlying channel proteins involved. Detailed biophysical analyses identified inwardly rectifying K⫹ (Kir) channels inhibited by barium ions (Ba2⫹) as key molecular players in the regulation of astroglial functions in vitro and in situ, and in Müller cells (Barres et al., 1990; Newman, 1993; Sontheimer and Waxman, 1993; Ransom and Sontheimer, 1995; MacFarlane and Sontheimer, 1997; Zhou and Kimelberg, 2000; Olsen et al., 2006; Kucheryavykh et al., 2007). Among the various Kir channels cloned so far (Kubo et al., 2005) at least five of these (Kir 2.1, 2.2, 2.3, 4.1, 5.1) have been identified in astrocytes (Schröder et al., 2002; Hibino

et al., 2004). A large body of evidence has demonstrated that Kir4.1 underlies the majority of the astrocytic Kir conductance (Takumi et al., 1995; Poopalasundaram et al., 2000; Li et al., 2001; Olsen et al., 2006; Butt and Kalsi, 2006). In Müller cells, expression of Kir2.1 accompanies that of Kir4.1 (Kofuji et al., 2002). The weakly-rectifying nature of Kir4.1 drives bidirectional movement of K⫹ in and out of the cell depending on the transmembrane K⫹ gradient, a property that allows the astroglial-mediated K⫹ release back into the extracellular space following neuronal activity (Newman, 1993). In the brain, Kir4.1 is well positioned to accomplish this role because it is enriched at the endfeet of astrocytic processes enwrapping synapses and facing blood vessels and pia mater (Higashi et al., 2001; Hibino et al., 2004) or perivascular processes in Müller glial cells (Nagelhus et al., 1999; Kofuji et al., 2002). Kir4.1 was found to heteromerize with Kir5.1 in mouse neocortex, whereas homomeric Kir4.1 was expressed by hippocampal astroglia (Hibino et al., 2004). The functional relevance of this differential channel composition is unknown. The prominent role of Kir4.1 in [K⫹]o buffering was demonstrated in astroglial cells of the ventral respiratory group, hippocampus and Müller cells by using Kir4.1 knockout mouse model (Kofuji et al., 2000; Neusch et al., 2006; Djukic et al., 2007). The same result was obtained by RNA interference (siRNA, small interfering RNA) approach to knockdown Kir4.1 in cultured astroglia (Kucheryavykh et al., 2007). It has to be emphasized that Kir4.1 was recently found to be also involved in growth control of glial cells (Higashimori and Sontheimer, 2007). The functional significance of Kir2.1 is still unknown, even though its localization at the endfeet of Müller glial cells contacting neurons suggests the implication in [K⫹]o clearance (Kofuji et al., 2002). As stated above the process of spatial redistribution of K⫹ in the astroglial syncytium by Kir channels assumes that these channels are compartmentalized in astrocytes. Such clustered pattern of expression suggests the presence of cellular microdomains formed of plasma membrane proteins arranged in multi-molecular functional complexes. Several transmembrane proteins, including ion channels, contain amino acid sequences that allow protein–protein interactions (Schulte, 2008). The analysis of Kir4.1 indicates that its C-terminus harbours a PDZ-binding motif for the interaction with proteins containing PDZ domains (Connors et al., 2004). Kir4.1 was found to colocalize with syntrophins, a group of proteins belonging to the dystrophin-associated-protein-complex (DAPC), and to form a macromolecular assembly with Dp71, ␤– dystroglycan, ␣-dystroglycan and ␣-syntrophin both in retinal Müller glial cells and astrocytes (Ishii et al., 1997; Guadagno and Moukhles, 2004; Hibino et al., 2004; Noël et al., 2005; Connors and Kofuji, 2006). Alfa-syntrophin was shown to be crucial for DAPC-Kir4.1 interaction in brain and retina, depicting a critical role of scaffolding proteins in [K⫹]o homeostasis (Connors et al., 2004; Connors and Kofuji, 2006). Two other anchoring proteins PSD95 and SAP97 were reported to co-localize with Kir4.1 in Müller cells,

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

thereby providing indirect evidence of their involvement in channel compartmentalization (Horio et al., 1997). It is worth noting that the mechanistic scheme of [K⫹]o buffering described above might not fit to all regions of the CNS owing to the fact that astroglial cells cannot be considered as a homogeneous cell population (McKhann et al., 1997; Walz, 2000; Zhou and Kimelberg, 2000; Wallraff et al., 2004). In the cortex, spatial mechanism seems to play a major role (Holthoff and Witte, 2000), but in other regions the situation may be different. Moreover, a passive K⫹ uptake mechanism accompanied by the influx of Cl⫺ has been proposed, even though its relevance to [K⫹]o clearance is still uncertain (for a review, see Walz, 1989) A Na⫹/K⫹ pump-mediated mechanism was reported to underlie [K⫹]o buffering in hippocampus (Xiong and Stringer, 2000; D’Ambrosio et al., 2002). Despite the wealth of data indicating that Kir4.1 current is the major component of the large astroglial K⫹ conductance throughout the CNS, in mature hippocampus there are differences in expression pattern between stratum lacunosum-moleculare and radiatum with the former displaying a higher level of Kir4.1 (Seifert et al., 2009). Because in stratum lacunosum-moleculare there is also the largest density of microvessels in the hippocampus (Shimada et al., 1992), spatial buffering is likely to be the dominant mechanism in stratum lacunosum-moleculare but could not operate in stratum radiatum. Importantly, recent work has provided evidence about the presence in stratum radiatum of K⫹ channels belonging to the two-pore domain channel family (TREK and TWIK subtypes) (Seifert et al., 2009; Zhou et al., 2009). These channels are open in a large range of membrane potentials, giving a substantial contribution to the high astrocyte K⫹ conductance, and hence might be involved in K⫹ uptake. It also remains to be ascertained the role in [K⫹]o homeostasis of outward K⫹ currents mediated by Kv and Ca2⫹-activated K⫹ channels that have been described in brain astrocytes both in vitro and in situ (Quandt and MacVicar, 1986; Tse et al., 1992; Kressin et al., 1995; Bordey and Sontheimer, 1999; Zhou and Kimelberg, 2000; Gebremedhin et al., 2003; Bekar et al., 2005; Filosa et al., 2006). Interactions between potassium channels and AQP4 in astroglia There are theoretical and experimental evidence that in the retina as well as in other regions of the CNS, the augment in [K⫹]o caused by neuronal activity produces a diminution in extracellular space owing to a transient increase in volume of astroglial endfeet mediated by the high rate of fluid accumulation accompanying K⫹ uptake (Ransom et al., 1985; Walz and Hinks, 1985; Holthoff and Witte, 2000; Østby et al., 2009). AQP4, which is the predominant water channel in astrocytes (see Amiry-Moghaddam and Ottersen, 2003), has been proposed as molecular partner of Kir4.1 in spatial buffering of K⫹ by facilitating the water movement through the plasma membrane (Nagelhus et al., 1999, 2004). In support of this hypothesis developmental studies revealed the overlapping of ontogenic expression of Kir4.1 and AQP4 both in astrocytes and Müller cells

929

(Wen et al., 1999; Wurm et al., 2006). Furthermore, confocal and immunogold electron microscopy (IEM) showed that AQP4 tightly co-localizes with Kir4.1 in Müller cells and in astrocyte endfeet of the neocortex in situ, exhibiting a patchy distribution in astrocytes facing blood vessels and pia (Nagelhus et al., 1999, 2004; Hibino et al., 2004). The peculiar distribution pattern suggested the presence of a molecular interplay between Kir4.1 and AQP4 (Nagelhus et al., 2004). This hypothesis found further support by co-immunoprecipitation experiments illustrating that AQP4 forms a macromolecular complex with Kir4.1 and proteins belonging to DAPC in Müller cells (Connors et al., 2004; Connors and Kofuji, 2006). Such co-localization was found in astrocytes and Müller cells in vitro when cultured in the presence of the basal lamina protein laminin with which DAPC interacts (Guadagno and Moukhles, 2004; Noël et al., 2005). Likewise, agrin, another protein of the basal lamina, promoted an increase in membrane-associated immunoreactivity of AQP4 in cultured astrocytes (Noell et al., 2007). The DAPC protein ␣-syntrophin was also shown to be involved in membrane anchoring of water channel AQP4 in situ (Neely et al., 2001; Amiry-Moghaddam et al., 2003a, 2004b). As additional evidence, IEM studies of ␣-syntrophin-null mice proved that in these animals there is a marked loss of AQP4 in perivascular membranes of astrocyte endfeet (Amiry-Moghaddam et al., 2003b), and in these mice a significant rearrangement of AQP4 distribution is observed in neocortex and cerebellum (AmiryMoghaddam et al., 2004b). Molecular interactions suggest that functional interplay between Kir4.1 and AQP4 is also occurring. Mice deficient of the ␣-syntrophin gene display a delay in [K⫹]o clearance caused by sustained neuronal activity (Amiry-Moghaddam et al., 2004a), even though in the cortex Kir4.1 distribution appears unchanged. Genetic ablation of AQP4 caused major susceptibility to seizures and slowed dynamics in [K⫹]o homeostasis (Binder et al., 2004, 2006; Binder and Steinhäuser, 2006). In models of cortical spreading depression the AQP4 null mice display altered capability of K⫹ uptake (Padmawar et al., 2005). Interestingly, co-localization is also lost in high grade human glioma in situ, in which AQP4 as well as Kir4.1 are mislocalized throughout the cells (Olsen and Sontheimer, 2004; Warth et al., 2005). Thus, alteration of this macromolecular complex and its functional impairment could be a phenotype of reactive response of glial cells to injuries or to other pathologies, thereby representing a clinical target for diagnostic and therapeutic interventions. Although in pathophysiological settings a link occurs between Kir4.1 and AQP4, more controversial are the data concerning the functional interactions in a more physiological context. Evidence emerging from comparative studies performed in freshly isolated retinal Müller cells and brain astrocytes from AQP4 knockout mice is not consistent with a functional interplay between AQP4 and Kir4.1 because genetic ablation of AQP4 did not alter Kir4.1 distribution and functional properties (Ruiz-Ederra et al., 2007; Zhang and Verkman, 2008). Moreover, in cell culture model of retinal and hippocampal astroglial cells, the water permeability of AQP4 was not modified in astrocytes in which

930

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

Kir4.1 was knocked down by siRNA or by pharmacological inhibition of Kir channels with Ba2⫹ (Zhang and Verkman, 2008). The reasons for these discrepancies remain to be ascertained but different approaches in cell manipulation might play a role. An alternative explanation could be that the two proteins do not belong to the same detergentresistant plasma membrane microdomains (lipid rafts). At the ultrastructural level the molecular interaction between two proteins might occur because of proximity of different membrane units devoted to [K⫹]o buffering and water homeostasis that might crosstalk to serve specific functions (Hibino and Kurachi, 2007). However, only a more detailed analysis of the direct dependence of astroglial Kir properties on AQP4 presence performed by in situ single cell electrophysiology and/or by means of K⫹ selective microelectrodes shall unravel the postulated functional interaction of AQP4 with Kir4.1. These studies should focus on properties of Kir conductance in astrocytes abutting blood vessels compared to those of Kir4.1 conductance of astroglial cells interposed at synaptic interspaces. Because astrocytes can be considered as multifunctional unit the role of Kir channel might be different in the diverse plasma membrane regions of a given cell. The hypothesis of distinct AQP4 and Kir4.1 domains is supported by recent evidence, provided in two different dystrophin null mice models, revealing that only perivascular pools of AQP4 co-localize with Dp71, the same dystrophin that binds Kir4.1 to DAPC macromolecular complex in the brain (Nicchia et al., 2008). There are also other K⫹ channels that may interact with AQP4 as in vivo studies about protein expression of Ca2⫹-activated K⫹ channels (rSlo) and the voltage-gated K⫹ channel Kv1.5 revealed that in rat they are highly expressed at astroglial endfeet ensheating microvasculature also rich in AQP4 (Roy et al., 1996; Price et al., 2002). Moreover, recent studies have provided evidence that astrocyte endfeet contain TREK and TWIK subtypes of the two-pore domain K⫹ channel family (Seifert et al., 2009; Zhou et al., 2009). Further studies are warranted to unravel whether these proteins are molecular partners of AQP4. Interplay between AQP4 and the gap junction protein Cx43 Astroglial cells are connected to each other by gap junctions (Fig. 1) that permit the intercellular passage of ions and small organic molecules such as inositol triphosphate (IP3) (Giaume and McCarthy, 1996). In astroglia gap-junctional coupling promotes biochemical cell-cell communication and non-synaptic electrical synchronization at distal cells (Rozental et al., 2000). Astrocytes express primarily Cx43 but also Cx30 and Cx26 (Nagy et al., 2004). Permeability and expression of gap junctions are differentially regulated under physiological and pathophysiological conditions. In cultured cortical astrocytes and in slices, gap junctional coupling is modulated by [K⫹]o elevation (Enkvist and McCarthy, 1994; Nagy and Li, 2000). Upon ischemic conditions, gap junctions allow free trafficking of death messengers in the astrocytic syncytium (Lin et al., 1998). The relationship of gap junction with neuronal isch-

emic damage is also supported by data revealing that the relatively unspecific gap junction inhibitor carbenoxolone (Benfenati et al., 2009; Ye et al., 2009) strongly reduced neuronal cell death under these conditions (de Pina-Benabou et al., 2005; Perez Velazquez et al., 2006). Gap junctions are involved in the propagation of spreading depression but little in that of Ca2⫹ waves (Nedergaard et al., 1995; Scemes et al., 1998). A contribution of gapjunction-mediated ion flow has been reported in the context of [K⫹]o buffering in hippocampal astrocytes using double knockout mouse for Cx30 and Cx43 (Wallraff et al., 2006). By analysis of Lucifer Yellow diffusion it has been demonstrated that Cx43 in vivo mediates fluxes of molecules specifically along the path of glial endfeet apposed to the vessel wall (Simard et al., 2003). The existence of functional interplay between Cxs and AQPs has been described (for a review, see Chanson et al., 2007). In lens fibers, expression of gap junction-forming protein was downregulated in mice lacking the major intrinsic protein (MIP), also known as AQP0 (Al-Ghoul et al., 2003). Immunoprecipitation and immunoblotting analyses from lens fiber lysates revealed that AQP0 directly associates with Cx45.6 and Cx56 at the early stages of embryonic chick lens development (Yu and Jiang, 2004). Several data indicate that in astrocytes a cooperative action between AQP4 and the astroglial gap junction protein Cx43 occurs in the regulation of water and [K⫹]o homeostasis. Cx43 is highly expressed in the process endfeet of astrocytes apposed to the large and medium-size vessels (Simard et al., 2003), the plasma membrane domain in which AQP4 is also enriched. The augment in [K⫹]o observed as result of the high neuronal activity during ischemia increased both the level of gap-junctional coupling and AQP4 expression (Ribeiro Mde et al., 2006). By contrast, downregulation of AQP4, Cx43 and Kir4.1 was reported in a mouse model of hyperammonemia, a process leading to the development of brain edema (Lichter-Konecki et al., 2008). Direct prove of the interactions between AQP4 and Cx43 has been obtained in mouse primary cultured astrocytes in which AQP4 knockdown by siRNA induced the downregulation of Cx43 and alteration in astrocyte cell-cell coupling (Nicchia et al., 2005).

BRAIN VOLUME HOMEOSTASIS: INTERACTIONS OF AQP4 WITH ION CHANNELS CONTROLLING ASTROCYTE REGULATORY VOLUME DECREASE Cell volume regulation in astrocytes The ability of mammalian cells to sense the osmotic changes in their environment and to regulate the cell volume in response to such changes assumes a particular relevance in the brain. In the CNS the extracellular fluid osmolality and the cell volume homeostasis are continuously challenged by the generation of local, transient osmotic microgradients of ions and molecules associated with the physiological neuronal activity (Simard and Nedergaard, 2004). The alteration in intracellular water content

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

931

Fig. 2. Astroglial swelling and possible routes promoting cell volume recovery. Astroglial cells can adjust their volume augmented as consequence of neuronal firing. At the endfeet K⫹, Na⫹ and Glu⫺ uptakes occur by means of Kir4.1, the Na⫹/Glu⫺ co-transporters and perhaps still elusive cation channels (?) (1). This increase in intracellular osmolytes is accompanied by osmotically driven water influx through diffusion and the water channel AQP4 (2). The mechanism underlying astroglial volume sensing, which initiates the series of events leading to cell volume recovery called regulatory volume decrease (RVD), remains to be elucidated unequivocally. Swelling-induced, intracellular Ca2⫹ elevation might be involved in osmotransduction (3) at least in vitro. In addition to TRP channels activated by Ca2⫹ release from intracellular stores that mediate CCE (not shown), other members of this superfamily able to promote swelling-induced Ca2⫹ entry are plausible molecular candidates for volume sensing. The osmo-sensitive Ca2⫹ channel TRPV4 is expressed by astrocytes in vitro and in vivo but its link to osmotransduction remains to be ascertained (?). The ensuing process of RVD is based on the extrusion of intracellular solutes paralleled by obliged water efflux. The volume-regulated anion channel (VRAC), by permeating inorganic (Cl⫺) and organic osmolytes such as taurine (Tau), and excitatory amino acids (EAA) has a critical role in RVD in vitro but its relevance in vivo is unclear. A contribution of K⫹ efflux is also likely but the specific volume-sensitive K⫹ channels involved remain to be identified (4). The osmolyte extrusion creates the gradient for water efflux and promotes RVD to recover the initial volume (5). Pumps and transporters that contribute to the transmembrane movement of ions have not been included.

affects signalling molecules and ion concentrations that can cause variations in cell activities and in cell-cell communication mechanisms. Because astrocytes are the cells principally involved in brain volume homeostasis, unravelling the mechanisms allowing these cells to respond to anisotonic environment would be of crucial importance. Ion channels, transporters and the water channel AQP4 play major roles in both the augment in astroglial cell volume and the associated mechanism of volume recovery termed regulatory volume decrease (RVD) (Kimelberg, 2005). A moderate, transient swelling of astrocyte endfeet occurs at active synapses as suggested by diminution of extracellular spaces (Dietzel et al., 1980; Sykova and Chvatal, 2000). Moreover, hypoxia/ischemia and traumatic injuries cause cerebral edema, which is accompanied by severe astrocyte swelling (cytotoxic edema) and impairment of RVD (Kimelberg, 2005). Astrocyte exposure to hypotonicity in vitro has provided some insights into RVD, which can be summarized to develop in three temporally distinct steps (Fig. 2); first, the formation of an intracellular osmolyte gradient causes cell swelling because of osmotically driven water inflow by passive diffusion also through the water channel AQP4. Second, swollen astrocyte senses volume change, which triggers a chain of biochemical reactions aimed at regulating volume homeostasis. Third, the effector mechanisms

of osmostransduction promote the release of inorganic and organic osmolytes and water, which lead to volume recovery. Among the astroglial mechanisms proposed for cell volume sensation, hypotonicity-induced elevation of intracellular calcium ([Ca2⫹]i) and cytoskeleton rearrangements have been envisaged to play a critical role (O’Connor and Kimelberg, 1993; Fischer et al., 1997; Mongin et al., 1999; Niggel et al., 2000; Mongin and Kimelberg, 2005). The relevance of [Ca2⫹]i dynamics as biochemical signal involved in osmotransduction is, however, still controversial because evidence against implication of [Ca2⫹]i rise in RVD has also been documented (Morales-Mulia et al., 1998; Quesada et al., 1999). Moreover, it is unclear whether Ca2⫹ per se or biochemical mechanisms depending on [Ca2⫹]i elevation are involved. Various kinases (p125FAK, p38, JNK, p56lck, p72syk and ERK1/ ERK2) and phospholipase A2 (PLA2) are also activated upon cell swelling in different cell types, but their exact contribution to astroglial RVD is still unknown (reviewed by Pasantes-Morales et al., 2000). Astroglial ion channels involved in regulatory volume decrease Over the last two decades all the attempts to identify the ionic mechanisms underlying the astroglial RVD have

932

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

been unsuccessful (for a review, see Pasantes-Morales et al., 2006). It is also unclear whether there is a unique process operating in different regions of the CNS or there are distinct molecular mechanisms. A recent report indicates that in response to ischemia astroglial cells display differential properties in cell volume adjustment revealing the presence of at least two functional populations in the cortex (Benesova et al., 2009). Future studies shall investigate whether this behaviour is the result of different expression/localization of plasma membrane proteins involved in cell volume homeostasis. There are several indications that at least in vitro the volume-regulated anion channels (VRACs) play a pivotal role in astroglial response to swelling by contributing to RVD (Walz and Mukerji, 1988; Sánchez-Olea et al., 1993; Parkerson and Sontheimer, 2003, 2004; Abdullaev et al., 2006; Kimelberg et al., 2006). Other studies in vitro and in vivo have revealed the presence of inwardly rectifying Cl⫺ channels of the ClC channel family that could be engaged in cell volume homeostasis (Ferroni et al., 1997; Sík et al., 2000; Makara et al., 2003). There is also evidence that channel-mediated K⫹ efflux plays a critical role in RVD, even though the K⫹ channels involved have not been identified yet (Pasantes-Morales et al., 1994; Vitarella et al., 1994; Quesada et al., 1999). The Kir4.1 channel was shown to be essential for swelling regulation at the endfeet of astrocytes of the spinal cord (Dibaj et al., 2007), but evidence for such mechanism also in other CNS regions is still lacking. Because of the possible role of [Ca2⫹]i-mediated signalling in osmotransduction controlling astroglial RVD the dissection of the molecular mechanisms underlying the swelling-induced [Ca2⫹]i elevation has become a high priority. In the following chapters we will review recent evidence about the interaction of AQP4 with Cl⫺ and Ca2⫹ channels that because of their features might be involved in cell volume regulation in astroglia. Interaction of AQP4 with VRAC In cultured astrocytes cell swelling activates VRAC current, which, in addition to promote transmembrane fluxes of inorganic anions, mediates the flow of small, osmotically active organic molecules such as taurine, the excitatory amino acids glutamate and aspartate, and perhaps also ATP (for a review, see Kimelberg et al., 2006). There are numerous in vitro and in vivo evidence that activation of VRACs in response to astrocyte swelling might exacerbate neuronal cell damage by an excitotoxic mechanism due to uncontrolled glutamate efflux (Rutledge et al., 1998; Phillis et al., 1998; Seki et al., 1999; Feustel et al., 2004). It has been reported that peroxynitrite and hydrogen peroxide, which are abundantly produced during ischemia and reperfusion, can regulate VRAC-mediated excitatory amino acid release via mechanisms involving src tyrosine kinase-activation and Ca2⫹/calmodulin-dependent protein kinase II (Haskew et al., 2002; Kimelberg, 2004b; Haskew-Layton et al., 2005). The modulation of astrocytic VRAC by Ca2⫹/ calmodulin-dependent protein kinase has also been described (Li et al., 2002; Olson et al., 2004). More recently, a role of VRAC-mediated neurotransmitter release from

astrocytes under physiological conditions has been proposed as nonsynaptic mechanism of astrocyte–neuron communication (Takano et al., 2005; Mulligan and MacVicar, 2006). Other studies have demonstrated the implication of VRACs in astroglial RVD in vitro (Parkerson and Sontheimer, 2003, 2004). The dysregulation of VRAC activity was suggested to be causally related to alterations in cell volume regulation that parallel the development of brain edema in stroke (Kimelberg, 2005). Thus, to define whether VRACs are localized in specific functional domains in astrocytes in vivo would be crucial for addressing their functional relevance. However, electrophysiological evidence about VRAC activity in astroglia in situ is lacking and the molecular identity of VRACs is still elusive (see Kimelberg et al., 2006). One of the difficulties in studying VRAC-mediated effects is the absence of selective channel blockers or activators. Notably, 4-(2-butyl-6,7-dichloro-2-cyclopentylindan-1on-5-yl)oxybutyric acid (DCPIB) was recently shown to have a certain degree of specificity suppressing astroglial VRAC current and reducing both the swelling-induced and ischemia-mediated release of glutamate (Decher et al., 2001; Abdullaev et al., 2006; Zhang et al., 2008; Haskew-Layton et al., 2008). It also remains to be ascertained unequivocally whether VRACs represent a single population of swelling-activated anion channels or they are different molecular entities that can be activated under different conditions (Liu et al., 2006). Physical interaction between VRAC and AQP4 cannot be explored directly and functional interplay has to be viewed also considering other interacting partners. We recently provided in vitro evidence in rat cultured astrocytes of the functional interaction between AQP4 and VRAC by demonstrating the downregulation of VRAC activity after the siRNA-mediated knockdown of AQP4 (Benfenati et al., 2007a). Although rearrangement of the actin cytoskeleton has previously been reported in AQP4-knockdown cultured astrocytes from rat, mouse and humans (Nicchia et al., 2005), VRAC downregulation in AQP4 knockdown rat astroglia did not depend on the polymerization of the actin cytoskeleton. This is noteworthy because it was previously shown that activation of astroglial VRAC was dependent on the state of polymerization of actin cytoskeleton that affects cell morphology (Lascola and Kraig, 1996; Lascola et al., 1998). The interplay between VRAC and AQP4 appears even more complex as inclusion of ATP in the patch-clamp pipette resulted in the restoration of hypotonic sensitivity of VRAC in AQP4 knockdown astrocytes, thereby suggesting the existence of an ATP-dependent mechanism of interaction (Benfenati et al., 2007a). Intracellular calcium signalling and cell volume regulation in astrocytes: is there a role for transient receptor potential channels? The full mechanistic understanding of RVD may require to define the pathways mediating [Ca2⫹]i elevation occurring in response to cell swelling. Astroglial RVD elicited by hypotonic stress was shown to be dependent on [Ca2⫹]i elevation and abolished upon diminution of extracellular

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

Ca2⫹ ([Ca2⫹]o) (Olson et al., 1990; O’Connor and Kimelberg, 1993). Moreover, VRACs, which are the main effector mechanism of astroglial RVD, were reported to be upregulated by activation of [Ca2⫹]i-dependent enzymatic pathways (Li et al., 2002; Haskew et al., 2002; HaskewLayton et al., 2005). The dissection of the relative contribution to the [Ca2⫹]i dynamics given by the Ca2⫹ release from internal stores and the Ca2⫹ influx through the plasma membrane is necessary for hypothesizing novel strategies to modulate RVD. Some members of the transient receptor potential channel superfamily (TRP) could be candidates to mediate swelling-induced [Ca2⫹]i rises (see Pasantes-Morales et al., 2006). The presence of transcripts for TRP proteins able to mediate capacitative Ca2⫹ entry (CCE) promoted by store depletion has been described in cultured astrocytes (Pizzo et al., 2001). The subtypes 1 and 4 of Canonical TRP channels (TRPC) have been identified in cultured astroglia (Song et al., 2005; Malarkey et al., 2008). TRPC4 was reported to co-localize with the scaffolding protein ZO-1, and TRPC4 lacking of the PDZ-interacting domain mislocalized in cultured astroglia (Song et al., 2005). Future analyses of the interactions between these proteins and AQP4 will provide insight into their relevance to RVD. There are indications that in various cell types the sustained Ca2⫹ entry observed in response to a variety of chemical and physical stimuli coupled to alteration of cell volume including membrane stretch, osmotic swelling or swelling-induced cytoskeletal modification is mediated by a specific subgroup of the TRP family called TRPV (Transient Receptor Potential Vanniloid related) (Liedtke et al., 2000; Muraki et al., 2003; Liedtke and Kim, 2005; Liedtke, 2006; Becker et al., 2005, 2009). In mammalians, the non-selective cation channels TRPV1, TRPV2 and TRPV4 have been identified as osmo- and mechano-sensitive TRPV channels delineating their implication in sensing and transduction mechanisms of osmotic stimuli (for reviews, see Mutai and Heller, 2003; Liedtke and Kim, 2005; Liedtke, 2006). TRPV4 is a polymodal channel, activated also by moderate heat (Guler et al., 2002; Watanabe et al., 2002a; Vriens et al., 2004) and endogenous signalling molecules such as anandamide and arachidonic acid (Watanabe et al., 2003). Different stimuli were shown to use distinct pathways to indirectly activate TRPV4 (Vriens et al., 2004; Wegierski et al., 2009). Cell swelling activation of TRPV4 has been described to depend upon PLA2 stimulation and formation of arachidonic acid (Vriens et al., 2004). Src-dependent phosphorylation of TRPV4 by swelling has also been reported (Xu et al., 2003; Wegierski et al., 2009). Hydrogen peroxide, and presumably oxidative stress, induced a strong upregulation of Src-dependent phosphorylation of TRPV4 channel (Wegierski et al., 2009). Interestingly, these molecules also modulate positively astroglial VRAC in vitro by activating the same enzymatic pathways (Haskew et al., 2002; Haskew-Layton et al., 2005). Several observations indicate that TRPV4 is critically involved in cell volume homeostasis and RVD in various cell types (Arniges et al., 2004; Becker et al., 2005, 2009;

933

Liu et al., 2006; Pan et al., 2008). We recently showed that TRPV4 is present in rat cortical astrocytes in vitro and in situ (Benfenati et al., 2007b). Exposure to the phorbol derivative 4-alpha-phorbol 12,13-didecanoate (4␣PDD), a selective activator of TRPV4 (Watanabe et al., 2002b), induced oscillatory [Ca2⫹]i signals in cultured cortical astrocytes as well as current response resembling those observed for endogenous TRPV4 in other cell types (Liedtke et al., 2000; Reiter et al., 2006). Hypotonicityinduced [Ca2⫹]i elevation was dependent on [Ca2⫹]o and was inhibited by ruthenium red, a non-specific blocker of TRPV4 (Vriens et al., 2009). Analysis of in situ expression revealed that TRPV4 is abundantly expressed in the membranes of astrocytic endfeet abutting the pial surface and the ependyma of the ventricular system, two plasma membrane domains also highly enriched in AQP4 (Nielsen et al., 1997). It has been reported that TRPV4 interacts with AQP5 in salivary gland and lung epithelia to regulate RVD (Liu et al., 2006; Sidhaye et al., 2006, 2008), and colocalizes with AQP2 in kidney cells (Tian et al., 2004). Whether the interplay between TRPV4 and AQP5 occurs at a functional level or depends on their physical interactions appears to be cell specific. In mouse lung epithelial cells, hypotonicityinduced reduction of AQP5 expression in the plasma membrane was tightly regulated by TRPV4 activation. This interaction was functional but did not require molecular crosstalk (Sidhaye et al., 2006, 2008). By contrast, in salivary glands TRPV4 and AQP5 co-immunoprecipitated and co-localized; the physical interaction was positively modulated by hypotonicity and was necessary for RVD (Liu et al., 2006). In salivary glands of AQP5 knockout mice the hypotonicity-induced [Ca2⫹]i transients and RVD mediated by TRPV4 were impaired and similar results were obtained upon expression of the N-terminus-deleted AQP5 (Liu et al., 2006). Preliminary data from our lab have evidenced that a molecular interaction between TRPV4 and AQP4 is present in astrocytes in vitro and in situ (Benfenati et al., 2008). TRPV4 and AQP4 co-localization was observed in astrocytic syncytia abutting the cortical surface or perivascular spaces, suggesting that TRPV4 and AQP4 complex could mediate the signalling cascade that causes astroglial release of osmolytes in response to osmotic stress. Collectively, these data confirm the role of AQPs in volume control as regulator of ion channel activity via mechanisms that are not dependent on their ability to mediate water flux. They also support the tenet that TRPV4 channel is a relevant partner of AQPs in cell volume homeostasis. Whether similar interplay occurs also with other members of TRP channel family is an issue that shall be addressed in further studies.

CONCLUSIONS AND FUTURE PERSPECTIVES Protein–protein interactions occur in a wide range of cellular processes and in the CNS they are fundamental for the regulation of diverse functions such as neurite outgrowth, synaptic formation and modulation, neurotransmission, signal transduction pathways and homeostasis

934

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

regulation (Blazer and Neubig, 2009). The expression pattern of protein channels in specific plasma membrane microdomains of polarized cells is considered to be essential for determining the correct cell functioning (Funke et al., 2005). The relevance of protein–protein interactions in the plasma membrane as possible sites of therapeutic intervention for diseases in which there is a pathogenetic change in cell volume is inferred by the wealth of data concerning altered protein membrane expression and mislocalization of ion channels and AQPs in several acute and chronic disorders of the CNS (Olsen and Sontheimer, 2004; Warth et al., 2005; Seifert et al., 2006; Habela et al., 2008). Investigating the interactions between AQPs and Cl⫺ channels would be of great interest in the context of malignant glioma. It is known, in fact, that cell volume regulation processes are crucial for tumor adhesion, migration and proliferation (Ullrich and Sontheimer, 1996; Ullrich et al., 1998; Soroceanu et al., 1999; Habela et al., 2009). Moreover, efflux via VRAC controls cell volume regulatory processes and has been linked to invasive migration of human glioma (Ransom et al., 2001; Ernest et al., 2005). It was recently demonstrated that the putative swelling-activated ClC-3 channel protein is expressed by glioma cells and is a critical regulator of malignant glioma cell cycle (Habela et al., 2008). The role of AQP4 and AQP1 in facilitating astroglial migration has been established in vitro and in vivo (Saadoun et al., 2002, 2005; McCoy and Sontheimer, 2007; Papadopoulos et al., 2008). The mechanisms whereby AQPs and ion channels enhance cell migration are still unclear. Changes in actin polymerization/depolymerization states are known to accompany volume changes (shrinkage and swelling) at lamellipodia of migrating cells and are crucial for tumor infiltrating processes (Saadoun et al., 2005; Papadopoulos et al., 2008). AQPs as well as volume sensitive Cl⫺ and K⫹ channels would facilitate transmembrane water and ionic fluxes to promote rapid osmotic changes at the edge of malignant cells. Reduction of Cx43 expression and gap junctional coupling has been observed in human gliomas and a marked rearrangement of AQP4 with loss of its perivascular localization has been described in the same cells (Soroceanu et al., 2001; Pu et al., 2004; Warth et al., 2005). It would be interesting to address the interactions of these two proteins in malignant glioma invasion. In this context, the identification of interacting partners of AQP9 in glioma could also be important because this AQP is upregulated in human glioblastoma (Warth et al., 2007). The identification of the functional and molecular interactions of AQP9 could also shed some light on its physiological role in normal astrocytes. In the brain AQP9 is expressed by astrocytes and some neurons (Badaut et al., 2004; Mylonakou et al., 2009). Enrichment of AQP9 was demonstrated in astrocyte processes in the periventricular region of parenchyma and in glia limitans at the interface with subarachnoid space, suggesting its contribution to water movement between cerebrospinal fluid and brain parenchyma (Badaut et al., 2001). The observation that astroglial expression of AQP9 is enhanced following focal ce-

rebral ischemia supports the view of its role in edema formation. It is worth noting, however, that the finding that AQP9 is also present in mitochondria (Amiry-Moghaddam et al., 2005) and transports energy substrates has led to the hypothesis that it could also play a role in cerebral energy metabolism even though these results are still controversial (Yang et al., 2006). Interactions between AQPs and ion channels may also be important when considering spinal cord. Recent in vivo studies provided evidence that following spinal cord injury astroglia display an upregulation of AQP4 and AQP1 (Nesic et al., 2006, 2008). Whereas the role of increased level of AQP4 was associated to the development of spinal cord edema, AQP1 upregulation was not affected by hypertonicity suggesting that it may have additional roles (Nesic et al., 2008). Thus, addressing AQP1 interactions may help to define its functional relevance more accurately. Another fundamental question that shall be addressed is the role of protein–protein interactions of AQPs in genetic and transcriptional diseases of the CNS which manifest with altered expression and/or localization of ion channels (for reviews see Waxman, 2001; Camerino et al., 2008). The potential role of this interplay in the regulation of cell viability modulated by changes in cell volume that lead to apoptotic cell death also remains unexplored (Jessica Chen et al., 2008). This lack of information is partly due to the fact that the set of anchoring proteins regulating the molecular and functional interactions is not fully delineated yet. Dystroglycanopathies due to genetic defective dystroglycan protein complex are the only diseases that have been found to be associated to mislocalization of AQPs and the interacting ion channels in astroglia (Nico et al., 2003, 2004). Much work remains to be done but strategies based on the use of complementary molecular and cellular approaches such as proteomic and yeast twohybrid system analyses, combined with fluorescence resonance energy transfer (FRET) and total internal reflection fluorescence (TIRF) determinations using the multiphoton fluorescence lifetime imaging microscopy (FLIM) technique (for reviews see Miller and Stagljar, 2004; Llères et al., 2007; Smyth and Shaw, 2008) will certainly prove to be of great help to screen potential interactors of AQPs that could be verified in vivo upon conditional gene manipulation in astroglia (Slezak et al., 2007). Acknowledgments—We are grateful to the members of the lab and to Helena Pivonkova (Department of Cellular Neurophysiology, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic) for comments on the manuscript. Supported by the Italian Ministry of Education, University and Research (SF), and BIMORE project, Marie Curie Network, 6th Framework Programme of the European Union (VB).

REFERENCES Abdullaev IF, Rudkouskaya A, Schools GP, Kimelberg HK, Mongin AA (2006) Pharmacological comparison of swelling activated excitatory amino acid release and Cl⫺ currents in rat cultured astrocytes. J Physiol 572:677– 689.

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940 Al-Ghoul KJ, Kirk T, Kuszak AJ, Zoltoski RK, Shiels A, Kuszak JR (2003) Lens structure in MIP-deficient mice. Anat Rec A Discov Mol Cell Evol Biol 273:714 –730. Amiry-Moghaddam M, Ottersen OP (2003) The molecular basis of water transport in the brain. Nat Rev Neurosci 4:991–1001. Amiry-Moghaddam M, Otsuka T, Hurn PD, et al (2003a) An alphasyntrophin-dependent pool of AQP4 in astroglial end-feet confers bidirectional water flow between blood and brain. Proc Natl Acad Sci U S A 100:2106 –2111. Amiry-Moghaddam M, Williamson A, Palomba M, et al (2003b) Delayed K⫹ clearance associated with aquaporin-4 mislocalization. Proc Natl Acad Sci U S A 100:13615–13620. Amiry-Moghaddam M, Xue R, Haug FM, Neely JD, Bhardwaj A, Agre P, Adams ME, Froehner SC, Mori S, Ottersen OP (2004a) Alphasyntrophin deletion removes the perivascular but not endothelial pool of aquaporin-4 at the blood-brain barrier and delays the development of brain edema in an experimental model of acute hyponatremia. FASEB J 18:542–544. Amiry-Moghaddam M, Frydenlund DS, Ottersen OP (2004b) Anchoring of aquaporin-4 in brain: molecular mechanism and implication for the physiology of water transport. Neuroscience 129:905–913. Amiry-Moghaddam M, Lindland H, Zelenin S, Roberg BA, Gundersen BB, Petersen P, Rinvik E, Torgner IA, Ottersen OP (2005) Brain mitochondria contain aquaporin water channels: evidence for the expression of a short AQP9 isoform in the inner mitochondrial membrane. FASEB J 19:1459 –1467. Anderson CM, Swanson RA (2000) Astrocyte glutamate transport: review of properties, regulation, and physiological functions. Glia 32:1–14. Arniges M, Vázquez E, Fernández-Fernández JM, Valverde MA (2004) Swelling-activated Ca2⫹ entry via TRPV4 channel is defective in cystic fibrosis airway epithelia. J Biol Chem 279: 54062–54068. Badaut J, Hirt L, Granziera C, Bogousslavsky J, Magistretti PJ, Regli L (2001) Astrocyte-specific expression of aquaporin-9 in mouse brain is increased after transient focal cerebral ischemia. J Cereb Blood Flow Metab 21:477– 482. Badaut J, Petit JM, Brunet JF, Magistretti PJ, Charriaut-Marlangue C, Regli L (2004) Distribution of Aquaporin 9 in the adult rat brain: preferential expression in catecholaminergic neurons and in glial cells. Neuroscience 128:27–38. Barres BA, Koroshetz WJ, Chun LL, Corey DP (1990) Ion channel expression by white matter glia: the type-1 astrocyte. Neuron 5:527–544. Becker D, Bereiter-Hahn J, Jendrach M (2009) Functional interaction of the cation channel transient receptor potential vanilloid 4 (TRPV4) and actin in volume regulation. Eur J Cell Biol 88:141– 152. Becker D, Blase C, Bereiter-Hahn J, Jendrach M (2005) TRPV4 exhibits a functional role in cell-volume regulation. J Cell Sci 118: 2435–2440. Bekar LK, Loewen ME, Cao K, Sun X, Leis J, Wang R, Forsyth GW, Walz W (2005) Complex expression and localization of inactivating Kv channels in cultured hippocampal astrocytes. J Neurophysiol 93:1699 –1709. Benesova J, Hock M, Butenko O, Prajerova I, Anderova M, Chvatal A (2009) Quantification of astrocyte volume changes during ischemia in situ reveals two populations of astrocytes in the cortex of GFAP/ EGFP mice. J Neurosci Res 87:96 –111. Benfenati V, Nicchia GP, Svelto M, Rapisarda C, Frigeri A, Ferroni S (2007a) Functional down-regulation of volume-regulated anion channels in AQP4 knockdown cultured rat cortical astrocytes. J Neurochem 100:87–104. Benfenati V, Amiry-Moghaddam M, Caprini M, Mylonakou MN, Rapisarda C, Ottersen OP, Ferroni S (2007b) Expression and functional characterization of transient receptor potential vanilloid-related channel 4 (TRPV4) in rat cortical astrocytes. Neuroscience 148:876 – 892.

935

Benfenati V, Dovizio M, Caprini M, Mylonakou MN, Ferroni S, Ottersen OP, Amiry-Moghaddam M (2008) AQP4 and TRPV4 forms a molecular complex in astrocytes. Poster Presented at: 6th Federation of European Neuroscience Societies (FENS) Forum; July 14; Oslo, Norway. Abstracts 130:2. Benfenati V, Caprini M, Nicchia GP, Rossi A, Dovizio M, Cervetto C, Nobile M, Ferroni S (2009) Carbenoxolone inhibits volume-regulated anion conductance in cultured rat cortical astroglia. Channels 3:323–336. Binder DK, Oshio K, Ma T, Verkman AS, Manley GT (2004) Increased seizure threshold in mice lacking aquaporin-4 water channels. Neuroreport 15:259 –262. Binder DK, Steinhäuser C (2006) Functional changes in astroglial cells in epilepsy. Glia 54:358 –368. Binder DK, Yao X, Zador Z, Sick TJ, Verkman AS, Manley GT (2006) Increased seizure duration and slowed potassium kinetics in mice lacking aquaporin-4 water channels. Glia 53:631– 636. Blazer LL, Neubig RR (2009) Small molecule protein-protein interaction inhibitors as CNS therapeutic agents: current progress and future hurdles. Neuropsychopharmacology 34:126 –141. Bordey A, Sontheimer H (1999) Differential inhibition of glial K(⫹) currents by 4-AP. J Neurophysiol 82:3476 –3487. Butt AM, Kalsi A (2006) Inwardly rectifying potassium channels (Kir) in central nervous system glia: a special role for Kir4.1 in glial functions. J Cell Mol Med 10:33– 44. Camerino DC, Desaphy JF, Tricarico D, Pierno S, Liantonio A (2008) Therapeutic approaches to ion channel diseases. Adv Genet 64:81–145. Chanson M, Kotsias BA, Peracchia C, O’Grady SM (2007) Interactions of connexins with other membrane channels and transporters. Prog Biophys Mol Biol 94:233–244. Chebabo SR, Hester MA, Jing J, Aitken PG, Somjen GG (1995) Interstitial space, electrical resistance and ion concentrations during hypotonia of rat hippocampal slices. J Physiol 487:685– 697. Connors NC, Kofuji P (2006) Potassium channel Kir 4.1 macromolecular complex in retinal glial cells. Glia 53:124 –131. Connors NC, Adams ME, Froehner SC, Kofuji P (2004) The potassium channel Kir4.1 associates with the dystrophin-glycoprotein complex via alpha-syntrophin in glia. J Biol Chem 279:28387–28392. D’Ambrosio R, Gordon DS, Winn HR (2002) Differential role of KIR channel and Na(⫹)/K(⫹)-pump in the regulation of extracellular K(⫹) in rat hippocampus. J Neurophysiol 87:87–102. Decher N, Lang HJ, Nilius B, Brüggemann A, Busch AE, Steinmeyer K (2001) DCPIB is a novel selective blocker of I(Cl,swell) and prevents swelling-induced shortening of guinea-pig atrial action potential duration. Br J Pharmacol 134:1467–1479. de Pina-Benabou MH, Szostak V, Kyrozis A, Rempe D, Uziel D, Urban-Maldonado M, Benabou S, Spray DC, Federoff HJ, Stanton PK, Rozental R (2005) Blockade of gap junctions in vivo provides neuroprotection after perinatal global ischemia. Stroke 36:2232– 2257. Dibaj P, Kaiser M, Hirrlinger J, Kirchhoff F, Neusch C (2007) Kir4.1 channels regulate swelling of astroglial processes in experimental spinal cord edema. J Neurochem 103:2620 –2628. Dietzel I, Heinemann U, Hofmeier G, Lux HD (1980) Transient changes in the size of the extracellular space in the sensorimotor cortex of cats in relation to stimulus-induced changes in potassium concentration. Exp Brain Res 40:432– 439. Djukic B, Casper KB, Philpot BD, Chin LS, McCarthy KD (2007) Conditional knock-out of Kir4.1 leads to glial membrane depolarization, inhibition of potassium and glutamate uptake, and enhanced short-term synaptic potentiation. J Neurosci 27:11354 – 11365. Enkvist MO, McCarthy KD (1994) Astroglial gap junction communication is increased by treatment with either glutamate or high K⫹ concentration. J Neurochem 62:489 – 495. Ernest NJ, Weaver AK, Van Duyn LB, Sontheimer HW (2005) Relative contribution of chloride channels and transporters to regulatory

936

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

volume decrease in human glioma cells. Am J Physiol Cell Physiol 288:C1451–C1460. Evanko DS, Zhang Q, Zorec R, Haydon PG (2004) Defining pathways of loss and secretion of chemical messengers from astrocytes. Glia 47:233–240. Ferroni S, Marchini C, Nobile M, Rapisarda C (1997) Characterization of an inwardly rectifying chloride conductance expressed by cultured rat cortical astrocytes. Glia 21:217–227. Feustel PJ, Jin Y, Kimelberg HK (2004) Volume-regulated anion channels are the predominant contributors to release of excitatory amino acids in the ischemic cortical penumbra. Stroke 35:1164 – 1168. Filosa JA, Bonev AD, Straub SV, Meredith AL, Wilkerson MK, Aldrich RW, Nelson MT (2006) Local potassium signaling couples neuronal activity to vasodilation in the brain. Nat Neurosci 9:1397–1403. Fischer R, Schliess F, Haussinger D (1997) Characterization of the hypo-osmolarity-induced Ca2⫹ response in cultured rat astrocytes. Glia 20:51–58. Funke L, Dakoji S, Bredt DS (2005) Membrane-associated guanylate kinases regulate adhesion and plasticity at cell junctions. Annu Rev Biochem 74:219 –245. Gebremedhin D, Yamaura K, Zhang C, Bylund J, Koehler RC, Harder DR (2003) Metabotropic glutamate receptor activation enhances the activities of two types of Ca2⫹-activated K⫹ channels in rat hippocampal astrocytes. J Neurosci 23:1678 –1687. Giaume C, McCarthy KD (1996) Control of gap-junctional communication in astrocytic networks. Trends Neurosci 19:319 –325. Guadagno E, Moukhles H (2004) Laminin-induced aggregation of the inwardly rectifying potassium channel, Kir4.1, and the waterpermeable channel, AQP4, via a dystroglycan-containing complex in astrocytes. Glia 47:138 –149. Guler AD, Lee H, Iida T, Shimizu I, Tominaga M, Caterina M (2002) Heat-evoked activation of the ion channel, TRPV4. J Neurosci 22:6408 – 6414. Habela CW, Ernest NJ, Swindall AF, Sontheimer H (2009) Chloride accumulation drives volume dynamics underlying cell proliferation and migration. J Neurophysiol 101:750 –757. Habela CW, Olsen ML, Sontheimer H (2008) ClC3 is a critical regulator of the cell cycle in normal and malignant glial cells. J Neurosci 28:9205–9217. Haskew RE, Mongin AA, Kimelberg HK (2002) Peroxynitrite enhances astrocytic volume-sensitive excitatory amino acid release via a src tyrosine kinase-dependent mechanism. J Neurochem 82:903–912. Haskew-Layton RE, Mongin AA, Kimelberg HK (2005) Hydrogen peroxide potentiates volume-sensitive excitatory amino acid release via a mechanism involving Ca2⫹/calmodulin-dependent protein kinase II. J Biol Chem 280:3548 –3554. Haskew-Layton RE, Rudkouskaya A, Jin Y, Feustel PJ, Kimelberg HK, Mongin AA (2008) Two distinct modes of hypoosmotic mediuminduced release of excitatory amino acids and taurine in the rat brain in vivo. PLoS ONE 3:e3543. Hibino H, Fujita A, Iwai K, Yamada M, Kurachi Y (2004) Differential assembly of inwardly rectifying K⫹ channel subunits, Kir4.1 and Kir5.1, in brain astrocytes. J Biol Chem 279:44065– 44073. Hibino H, Kurachi Y (2007) Distinct detergent-resistant membrane microdomains (lipid rafts) respectively harvest K(⫹) and water transport systems in brain astroglia. Eur J Neurosci 26:2539 – 2555. Higashi K, Fujita A, Inanobe A, Tanemoto M, Doi K, Kubo T, Kurachi Y (2001) An inwardly rectifying K(⫹) channel, Kir4.1, expressed in astrocytes surrounds synapses and blood vessels in brain. Am J Physiol Cell Physiol 281:C922–C931. Higashimori H, Sontheimer H (2007) Role of Kir4.1 channels in growth control of glia. Glia 55:1668 –1679. Holthoff K, Witte OW (2000) Directed spatial potassium redistribution in rat neocortex. Glia 29:288 –292. Horio Y, Hibino H, Inanobe A, et al (1997) Clustering and enhanced activity of an inwardly rectifying potassium channel, Kir4.1, by an

anchoring protein, PSD-95/SAP90. J Biol Chem 272:12885– 12888. Hrabetová S, Chen KC, Masri D, Nicholson C (2002) Water compartmentalization and spread of ischemic injury in thick-slice ischemia model. J Cereb Blood Flow Metab 22:80 – 88. Ishii M, Horio Y, Tada Y, Hibino H, Inanobe A, Ito M, Yamada M, Gotow T, Uchiyama Y, Kurachi Y (1997) Expression and clustered distribution of an inwardly rectifying potassium channel, KAB-2/Kir4.1, on mammalian retinal Müller cell membrane: their regulation by insulin and laminin signals. J Neurosci 17:7725–7735. Jessica CM, Sepramaniam S, Armugam A, Shyan Choy M, Manikandan J, Melendez AJ, Jeyaseelan K, Sang Cheung N (2008) Water and ion channels: crucial in the initiation and progression of apoptosis in central nervous system? Curr Neuropharmacol 6:102– 116. Kimelberg HK (2004a) Water homeostasis in the brain: basic concepts. Neuroscience 129:851– 860. Kimelberg HK (2004b) Increased release of excitatory amino acids by the actions of ATP and peroxynitrite on volume-regulated anion channels (VRACs) in astrocytes. Neurochem Int 45:511–519. Kimelberg HK (2005) Astrocytic swelling in cerebral ischemia as a possible cause of injury and target for therapy. Glia 50:389 –397. Kimelberg HK, Macvicar BA, Sontheimer H (2006) Anion channels in astrocytes: biophysics, pharmacology, and function. Glia 54:747– 757. Kofuji P, Ceelen P, Zahs KR, Surbeck LW, Lester HA, Newman EA (2000) Genetic inactivation of an inwardly rectifying potassium channel (Kir4.1 subunit) in mice: phenotypic impact in retina. J Neurosci 20:5733–5740. Kofuji P, Biedermann B, Siddharthan V, Raap M, Iandiev I, Milenkovic I, Thomzig A, Veh RW, Bringmann A, Reichenbach A (2002) Kir potassium channel subunit expression in retinal glial cells: implications for spatial potassium buffering. Glia 39:292–303. Kofuji P, Newman EA (2004) Potassium buffering in the central nervous system. Neuroscience 129:1045–1056. Kressin K, Kuprijanova E, Jabs R, Seifert G, Steinhäuser C (1995) Developmental regulation of Na⫹ and K⫹ conductances in glial cells of mouse hippocampal brain slices. Glia 15:173–187. Kubo Y, Adelman JP, Clapham DE, Jan LY, Karschin A, Kurachi Y, Lazdunski M, Nichols CG, Seino S, Vandenberg CA (2005) International Union of Pharmacology. LIV. Nomenclature and molecular relationships of inwardly rectifying potassium channels. Pharmacol Rev 57:509 –526. Kucheryavykh YV, Kucheryavykh LY, Nichols CG, Maldonado HM, Baksi K, Reichenbach A, Skatchkov SN, Eaton MJ (2007) Downregulation of Kir4.1 inward rectifying potassium channel subunits by RNAi impairs potassium transfer and glutamate uptake by cultured cortical astrocytes. Glia 55:274 –281. Kuffler SW, Nicholls JG, Orkand RK (1966) Physiological properties of glial cells in the central nervous system of amphibia. J Neurophysiol 29:768 –787. Lascola CD, Kraig RP (1996) Whole-cell chloride currents in rat astrocytes accompany changes in cell morphology. J Neurosci 16: 2532–2545. Lascola CD, Nelson DJ, Kraig RP (1998) Cytoskeletal actin gates a Clchannel in neocortical astrocytes. J Neurosci 18:1679 –1692. Li G, Liu Y, Olson JE (2002) Calcium/calmodulin-modulated chloride and taurine conductances in cultured rat astrocytes. Brain Res 925:1– 8. Li L, Head V, Timpe LC (2001) Identification of an inward rectifier potassium channel gene expressed in mouse cortical astrocytes. Glia 33:57–71. Lichter-Konecki U, Mangin JM, Gordish-Dressman H, Hoffman EP, Gallo V (2008) Gene expression profiling of astrocytes from hyperammonemic mice reveals altered pathways for water and potassium homeostasis in vivo. Glia 56:365–377.

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940 Liedtke W (2006) Transient receptor potential vanilloid channels functioning in transduction of osmotic stimuli. J Endocrinol 191:515–523. Liedtke W, Choe Y, Marti-Renom MA, Bell AM, Denis CS, Sali A, Hudspeth AJ, Friedman JM, Heller S (2000) Vanilloid receptorrelated osmotically activated channel (VR-OAC), a candidate vertebrate osmoreceptor. Cell 103:525–535. Liedtke W, Kim C (2005) Functionality of the TRPV subfamily of TRP ion channels: add mechano-TRP and osmo-TRP to the lexicon! Cell Mol Life Sci 62:2985–3001. Lin JH, Weigel H, Cotrina ML, Liu S, Bueno E, Hansen AJ, Hansen TW, Goldman S, Nedergaard M (1998) Gap-junction-mediated propagation and amplification of cell injury. Nat Neurosci 1:494 – 500. Liu HT, Tashmukhamedov BA, Inoue H, Okada Y, Sabirov RZ (2006) Roles of two types of anion channels in glutamate release from mouse astrocytes under ischemic or osmotic stress. Glia 54: 343–357. Liu X, Bandyopadhyay B, Nakamoto T, Singh B, Liedtke W, Melvin JE, Ambudkar I (2006) A role for AQP5 in activation of TRPV4 by hypotonicity: concerted involvement of AQP5 and TRPV4 in regulation of cell volume recovery. J Biol Chem 281:15485–15495. Llères D, Swift S, Lamond AI (2007) Detecting protein-protein interactions in vivo with FRET using multiphoton fluorescence lifetime imaging microscopy (FLIM). Curr Protoc Cytom Chapter 12:Unit 12.10. MacFarlane SN, Sontheimer H (1997) Electrophysiological changes that accompany reactive gliosis in vitro. J Neurosci 17:7316 –7329. Makara JK, Rappert A, Matthias K, Steinhäuser C, Spät A, Kettenmann H (2003) Astrocytes from mouse brain slices express ClC2-mediated Cl- currents regulated during development and after injury. Mol Cell Neurosci 23:521–530. Malarkey EB, Ni Y, Parpura V (2008) Ca2⫹ entry through TRPC1 channels contributes to intracellular Ca2⫹ dynamics and consequent glutamate release from rat astrocytes. Glia 56:821– 835. McCoy E, Sontheimer H (2007) Expression and function of water channels (aquaporins) in migrating malignant astrocytes. Glia 55:1034 –1043. McKhann GM 2nd, D’Ambrosio R, Janigro D (1997) Heterogeneity of astrocyte resting membrane potentials and intercellular coupling revealed by whole-cell and gramicidin-perforated patch recordings from cultured neocortical and hippocampal slice astrocytes. J Neurosci 17:6850 – 6863. Miller J, Stagljar I (2004) Using the yeast two-hybrid system to identify interacting proteins. Methods Mol Biol 261:247–262. Mongin AA, Cai Z, Kimelberg HK (1999) Volume-dependent taurine release from cultured astrocytes requires permissive [Ca2⫹]i and calmodulin. Am J Physiol 277:823– 832. Mongin AA, Kimelberg HK (2005) ATP regulates anion channel-mediated organic osmolyte release from cultured rat astrocytes via multiple Ca2⫹-sensitive mechanisms. Am J Physiol Cell Physiol 288:C204 –C213. Morales-Mulia S, Vaca L, Hernandez-Cruz A, Pasantes-Morales H (1998) Osmotic swelling-induced changes in cytosolic calcium do not affect regulatory volume decrease in rat cultured suspended cerebellar astrocytes. J Neurochem 71:2330 –2338. Mulligan SJ, MacVicar BA (2006) VRACs CARVe a path for novel mechanisms of communication in the CNS. Sci STKE 357:pe42. Muraki K, Iwata Y, Katanosaka Y, Ito T, Ohya S, Shigekawa M, Imaizumi Y (2003) TRPV2 is a component of osmotically sensitive cation channels in murine aortic myocytes. Circ Res 93:829 – 838. Mutai H, Heller S (2003) Vertebrate and invertebrate TRPV-like mechanoreceptors. Cell Calcium 33:471– 478. Mylonakou MN, Petersen PH, Rinvik E, Rojek A, Valdimarsdottir E, Zelenin S, Zeuthen T, Nielsen S, Ottersen OP, Amiry-Moghaddam M (2009) Analysis of mice with targeted deletion of AQP9 gene provides conclusive evidence for expression of AQP9 in neurons. J Neurosci Res 87:1310 –1322.

937

Nagelhus EA, Horio Y, Inanobe A, Fujita A, Haug FM, Nielsen S, Kurachi Y, Ottersen OP (1999) Immunogold evidence suggests that coupling of K⫹ siphoning and water transport in rat retinal Müller cells is mediated by a coenrichment of Kir4.1 and AQP4 in specific membrane domains. Glia 26:47–54. Nagelhus EA, Mathiisen TM, Ottersen OP (2004) Aquaporin-4 in the central nervous system: cellular and subcellular distribution and coexpression with Kir4.1. Neuroscience 129:905–913. Nagy JI, Li WE (2000) A brain slice model for in vitro analyses of astrocytic gap junction and connexin 43 regulation: actions of ischemia, glutamate and elevated potassium. Eur J Neurosci 12:4567– 4572. Nagy JI, Dudek FE, Rash JE (2004) Update on connexins and gap junctions in neurons and glia in the mammalian nervous system. Brain Res Brain Res Rev 47:191–215. Nedergaard M, Cooper AJ, Goldman SA (1995) Gap junctions are required for the propagation of spreading depression. J Neurobiol 28:433– 444. Nedergaard M, Ransom B, Goldman SA (2003) New roles for astrocytes: redefining the functional architecture of the brain. Trends Neurosci 26:523–530. Neely JD, Amiry-Moghaddam M, Ottersen OP, Froehner SC, Agre P, Adams ME (2001) Syntrophin-dependent expression and localization of aquaporin-4 water channel protein. Proc Natl Acad Sci U S A 98:14108 –14113. Nesic O, Lee J, Unabia GC, Johnson K, Ye Z, Vergara L, Hulsebosch CE, Perez-Polo JR (2008) Aquaporin 1—a novel player in spinal cord injury. J Neurochem 105:628 – 640. Nesic O, Lee J, Ye Z, Unabia GC, Rafati D, Hulsebosch CE, PerezPolo JR (2006) Acute and chronic changes in aquaporin 4 expression after spinal cord injury. Neuroscience 143:779 –792. Neusch C, Papadopoulos N, Müller M, Maletzki I, Winter SM, Hirrlinger J, Handschuh M, Bähr M, Richter DW, Kirchhoff F, Hülsmann S (2006) Lack of the Kir4.1 channel subunit abolishes K⫹ buffering properties of astrocytes in the ventral respiratory group: impact on extracellular K⫹ regulation. J Neurophysiol 95:1843–1852. Newman EA, Frambach DA, Odette LL (1984) Control of extracellular potassium levels by retinal glial cell K⫹ siphoning. Science 225: 1174 –1175. Newman EA (1993) Inward-rectifying potassium channels in retinal glial (Müller) cells. J Neurosci 13:3333–3345. Nicchia GP, Rossi A, Nudel U, Svelto M, Frigeri A (2008) Dystrophindependent and -independent AQP4 pools are expressed in the mouse brain. Glia 56:869 – 876. Nicchia GP, Srinivas M, Li W, Brosnan CF, Frigeri A, Spray DC (2005) New possible roles for aquaporin-4 in astrocytes: cell cytoskeleton and functional relationship with connexin 43. FASEB J 19:1674 – 1676. Nico B, Frigeri A, Nicchia GP, Corsi P, Ribatti D, Quondamatteo F, Herken R, Girolamo F, Marzullo A, Svelto M, Roncali L (2003) Severe alterations of endothelial and glial cells in the blood-brain barrier of dystrophic mdx mice. Glia 42:235–251. Nico B, Paola Nicchia G, Frigeri A, Corsi P, Mangieri D, Ribatti D, Svelto M, Roncali L (2004) Altered blood-brain barrier development in dystrophic MDX mice. Neuroscience 125:921–935. Nielsen S, Nagelhus EA, Amiry-Moghaddam M, Bourque C, Agre P, Ottersen OP (1997) Specialized membrane domains for water transport in glial cells: high-resolution immunogold cytochemistry of aquaporin-4 in rat brain. J Neurosci 17:171–180. Niggel J, Sigurdson W, Sachs F (2000) Mechanically induced calcium movements in astrocytes, bovine aortic endothelial cells and C6 glioma cells. J Membr Biol 174:121–134. Noël G, Belda M, Guadagno E, Micoud J, Klöcker N, Moukhles H (2005) Dystroglycan and Kir4.1 coclustering in retinal Müller glia is regulated by laminin-1 and requires the PDZ-ligand domain of Kir4.1. J Neurochem 94:691–702. Noell S, Fallier-Becker P, Beyer C, Kröger S, Mack AF, Wolburg H (2007) Effects of agrin on the expression and distribution of the

938

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

water channel protein aquaporin-4 and volume regulation in cultured astrocytes. Eur J Neurosci 26:2109 –2118. O’Connor ER, Kimelberg HK (1993) Role of calcium in astrocyte volume regulation and in the release of ions and amino acids. J Neurosci 13:2638 –2650. Olsen ML, Sontheimer H (2004) Mislocalization of Kir channels in malignant glia. Glia 46:63–73. Olsen ML, Higashimori H, Campbell SL, Hablitz JJ, Sontheimer H (2006) Functional expression of Kir4.1 channels in spinal cord astrocytes. Glia 53:516 –528. Olson JE, Fleischhacker D, Murray WB, Holtzman D (1990) Control of astrocyte volume by intracellular and extracellular Ca2⫹. Glia 3:405– 412. Olson JE, Li GZ, Wang L, Lu L (2004) Volume-regulated anion conductance in cultured rat cerebral astrocytes requires calmodulin activity. Glia 46:391– 401. Orkand RK, Nicholls JG, Kuffler SW (1966) Effect of nerve impulses on the membrane potential of glial cells in the central nervous system of amphibia. J Neurophysiol 29:788 – 806. Østby I, Øyehaug L, Einevoll GT, Nagelhus EA, Plahte E, Zeuthen T, Lloyd CM, Ottersen OP, Omholt SW (2009) Astrocytic mechanisms explaining neural-activity-induced shrinkage of extraneuronal space. PLoS Comput Biol 5:e1000272. Padmawar P, Yao X, Bloch O, Manley GT, Verkman AS (2005) K⫹ waves in brain cortex visualized using a long-wavelength K⫹sensing fluorescent indicator. Nat Methods 2:825– 827. Pan Z, Yang H, Mergler S, Liu H, Tachado SD, Zhang F, Kao WW, Koziel H, Pleyer U, Reinach PS (2008) Dependence of regulatory volume decrease on transient receptor potential vanilloid 4 (TRPV4) expression in human corneal epithelial cells. Cell Calcium 44:374 –385. Papadopoulos MC, Saadoun S, Verkman AS (2008) Aquaporins and cell migration. Pflugers Arch 456:693–700. Parkerson KA, Sontheimer H (2003) Contribution of chloride channels to volume regulation of cortical astrocytes. Am J Physiol Cell Physiol 284:1460 –1467. Parkerson KA, Sontheimer H (2004) Biophysical and pharmacological characterization of hypotonically activated chloride currents in cortical astrocytes. Glia 46:419 – 436. Pasantes-Morales H, Cardin V, Tuz K (2000) Signaling events during swelling and regulatory volume decrease. Neurochem Res 25: 1301–1314. Pasantes-Morales H, Murray RA, Lilja L, Morán J (1994) Regulatory volume decrease in cultured astrocytes. I. Potassium- and chloride-activated permeability. Am J Physiol 266:C165–C171. Pasantes-Morales H, Lezama RA, Ramos-Mandujano G, Tuz KL (2006) Mechanisms of cell volume regulation in hypo-osmolality. Am J Med 119:S4 –S11. Perez Velazquez JL, Kokarovtseva L, Sarbaziha R, Jeyapalan Z, Leshchenko Y (2006) Role of gap junctional coupling in astrocytic networks in the determination of global ischaemia-induced oxidative stress and hippocampal damage. Eur J Neurosci 23:1–10. Phillis JW, Song D, O’Regan MH (1998) Tamoxifen, a chloride channel blocker, reduces glutamate and aspartate release from the ischemic cerebral cortex. Brain Res 780:352–355. Pizzo P, Burgo A, Pozzan T, Fasolato C (2001) Role of capacitative calcium entry on glutamate-induced calcium influx in type-I rat cortical astrocytes. J Neurochem 79:98 –109. Poopalasundaram S, Knott C, Shamotienko OG, Foran PG, Dolly JO, Ghiani CA, Gallo V, Wilkin GP (2000) Glial heterogeneity in expression of the inwardly rectifying K(⫹) channel, Kir4.1, in adult rat CNS. Glia 30:362–372. Price DL, Ludwig JW, Mi H, Schwarz TL, Ellisman MH (2002) Distribution of rSlo Ca2⫹-activated K⫹ channels in rat astrocyte perivascular endfeet. Brai Res 956:183–193. Pu P, Xia Z, Yu S, Huang Q (2004) Altered expression of Cx43 in astrocytic tumors. Clin Neurol Neurosurg 107:49 –54.

Quandt FN, MacVicar BA (1986) Calcium activated potassium channels in cultured astrocytes. Neuroscience 19:29 – 41. Quesada O, Ordaz B, Morales-Mulia S, Pasantes-Morales H (1999) Influence of Ca2⫹ on K⫹ efflux during regulatory volume decrease in cultured astrocytes. J Neurosci Res 57:350 –358. Ransom BR, Yamate CL, Connors BW (1985) Activity-dependent shrinkage of extracellular space in rat optic nerve: a developmental study. J Neurosci 5:532–535. Ransom CB, O’Neal JT, Sontheimer H (2001) Volume-activated chloride currents contribute to the resting conductance and invasive migration of human glioma cells. J Neurosci 21:7674 –7683. Ransom CB, Sontheimer H (1995) Biophysical and pharmacological characterization of inwardly rectifying K⫹ currents in rat spinal cord astrocytes. J Neurophysiol 73:333–346. Reiter B, Kraft R, Gunzel D, Zeissig S, Schulzke JD, Fromm M, Harteneck C (2006) TRPV4-mediated regulation of epithelial permeability. FASEB J 20:1802–1812. Ribeiro Mde C, Hirt L, Bogousslavsky J, Regli L, Badaut J (2006) Time course of aquaporin expression after transient focal cerebral ischemia in mice. J Neurosci Res 83:1231–1240. Roy ML, Saal D, Perney T, Sontheimer H, Waxman SG, Kaczmarek LK (1996) Manipulation of the delayed rectifier Kv1.5 potassium channel in glial cells by antisense oligodeoxynucleotides. Glia 18:177–184. Rozental R, Giaume C, Spray DC (2000) Gap junctions in the nervous system. Brain Res Brain Res Rev 32:11–15. Ruiz-Ederra J, Zhang H, Verkman AS (2007) Evidence against functional interaction between aquaporin-4 water channels and Kir4.1 potassium channels in retinal Müller cells. J Biol Chem 282: 21866 –21872. Rutledge EM, Aschner M, Kimelberg HK (1998) Pharmacological characterization of swelling-induced D-[3H] aspartate release from primary astrocyte cultures. Am J Physiol 274:1511–1520. Saadoun S, Papadopoulos MC, Davies DC, Krishna S, Bell BA (2002) Aquaporin-4 expression is increased in oedematous human brain tumours. J Neurol Neurosurg Psychiatry 72:262–265. Saadoun S, Papadopoulos MC, Watanabe H, Yan D, Manley GT, Verkman AS (2005) Involvement of aquaporin-4 in astroglial cell migration and glial scar formation. J Cell Sci 118:5691–5698. Sánchez-Olea R, Peña C, Morán J, Pasantes-Morales H (1993) Inhibition of volume regulation and efflux of osmoregulatory amino acids by blockers of Cl- transport in cultured astrocytes. Neurosci Lett 156:141–144. Scemes E, Dermietzel R, Spray DC (1998) Calcium waves between astrocytes from Cx43 knockout mice. Glia 24:65–73. Schröder W, Seifert G, Hüttmann K, Hinterkeuser S, Steinhäuser C (2002) AMPA receptor-mediated modulation of inward rectifier K⫹ channels in astrocytes of mouse hippocampus. Mol Cell Neurosci 19:447– 458. Schulte U (2008) Protein-protein interactions and subunit composition of ion channels. CNS Neurol Disord Drug Targets 7:172–186. Seifert G, Schilling K, Steinhäuser C (2006) Astrocyte dysfunction in neurological disorders: a molecular perspective. Nat Rev Neurosci 7:194 –206. Seifert G, Hüttmann K, Binder DK, Hartmann C, Wyczynski A, Neusch C, Steinhäuser C (2009) Analysis of astroglial K⫹ channel expression in the developing hippocampus reveals a predominant role of the Kir4.1 subunit. J Neurosci 29:7474 –7488. Seki Y, Feustel PJ, Keller RW Jr, Trammer BI, Kimelberg HK (1999) Inhibition of ischemia-induced glutamate release in rat striatum by dihydrokinate and an anion channel blocker. Stroke 30:433– 440. Shimada M, Akagi N, Goto H, Watanabe H, Nakanishi M, Hirose Y, Watanabe M (1992) Microvessel and astroglial cell densities in the mouse hippocampus. J Anat 180:89 –95. Sidhaye VK, Guler AD, Schweitzer KS, D’Alessio F, Caterina MJ, King LS (2006) Transient receptor potential vanilloid 4 regulates aqua-

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940 porin-5 abundance under hypotonic conditions. Proc Natl Acad Sci U S A 103:4747– 4752. Sidhaye VK, Schweitzer KS, Caterina MJ, Shimoda L, King LS (2008) Shear stress regulates aquaporin-5 and airway epithelial barrier function. Proc Natl Acad Sci U S A 105:3345–3350. Sík A, Smith RL, Freund TF (2000) Distribution of chloride channel-2immunoreactive neuronal and astrocytic processes in the hippocampus. Neuroscience 101:51– 65. Simard M, Arcuino G, Takano T, Liu QS, Nedergaard M (2003) Signaling at the gliovascular interface. J Neurosci 23:9254 –9262. Simard M, Nedergaard M (2004) The neurobiology of glia in the context of water and ion homeostasis. Neuroscience 129:877– 896. Slezak M, Göritz C, Niemiec A, Frisén J, Chambon P, Metzger D, Pfrieger FW (2007) Transgenic mice for conditional gene manipulation in astroglial cells. Glia 55:1565–1576. Smyth JW, Shaw RM (2008) Visualizing ion channel dynamics at the plasma membrane. Heart Rhythm 5:S7–S11. Somjen GG (2002) Ion regulation in the brain: implications for pathophysiology. Neuroscientist 8:254 –267. Song X, Zhao Y, Narcisse L, Duffy H, Kress Y, Lee S, Brosnan CF (2005) Canonical transient receptor potential channel 4 (TRPC4) co-localizes with the scaffolding protein ZO-1 in human fetal astrocytes in culture. Glia 49:418 – 429. Sontheimer H, Waxman SG (1993) Expression of voltage-activated ion channels by astrocytes and oligodendrocytes in the hippocampal slice. J Neurophysiol 70:1863–1873. Soroceanu L, Manning TJ Jr, Sontheimer H (1999) Modulation of glioma cell migration and invasion using CI(⫺) and K(⫹) ion channel blockers. J Neurosci 19:5942–5954. Soroceanu L, Manning TJ Jr, Sontheimer H (2001) Reduced expression of connexin-43 and functional gap junction coupling in human gliomas. Glia 33:107–117. Sykova E, Chvatal A (2000) Glial cells and volume transmission in the CNS. Neurochem Int 36:397– 409. Takano T, Kang J, Jaiswal JK, Simon SM, Lin JH, Yu Y, Li Y, Yang J, Dienel G, Zielke HR, Nedergaard M (2005) Receptor-mediated glutamate release from volume sensitive channels in astrocytes. Proc Natl Acad Sci U S A 102:16466 –16471. Takumi T, Ishii T, Horio Y, et al (1995) A novel ATP-dependent inward rectifier potassium channel expressed predominantly in glial cells. J Biol Chem 270:16339 –16346. Tian W, Salanova M, Xu H, Lindsley JN, Oyama TT, Anderson S, Bachmann S, Cohen DM (2004) Renal expression of osmotically responsive cation channel TRPV4 is restricted to water-impermeant nephron segments. Am J Physiol Renal Physiol 287: F17–F24. Tse FW, Fraser DD, Duffy S, MacVicar BA (1992) Voltage-activated K⫹ currents in acutely isolated hippocampal astrocytes. J Neurosci 12:1781–1788. Ullrich N, Bordey A, Gillespie GY, Sontheimer H (1998) Expression of voltage-activated chloride currents in acute slices of human gliomas. Neuroscience 83:1161–1173. Ullrich N, Sontheimer H (1996) Biophysical and pharmacological characterization of chloride currents in human astrocytoma cells. Am J Physiol 270:C1511–C1521. Vitarella D, DiRisio DJ, Kimelberg HK, Aschner M (1994) Potassium and taurine release are highly correlated with regulatory volume decrease in neonatal primary rat astrocyte cultures. J Neurochem 63:1143–1149. Vriens J, Appendino G, Nilius B (2009) Pharmacology of vanilloid transient receptor potential cation channels. Mol Pharmacol 25: 1263–1279. Vriens J, Watanabe H, Janssens A, Droogmans G, Voets T, Nilius B (2004) Cell swelling, heat, and chemical agonists use distinct pathways for the activation of the cation channel TRPV4. Proc Natl Acad Sci U S A 101:396 – 401.

939

Wallraff A, Odermatt B, Willecke K, Steinhäuser C (2004) Distinct types of astroglial cells in the hippocampus differ in gap junction coupling. Glia 48:36 – 43. Wallraff A, Köhling R, Heinemann U, Theis M, Willecke K, Steinhäuser C (2006) The impact of astrocytic gap junctional coupling on potassium buffering in the hippocampus. J Neurosci 26:5438 –5447. Walz W (1989) Role of glial cells in the regulation of the brain ion microenvironment. Prog Neurobiol 33:309 –333. Walz W (2000) Controversy surrounding the existence of discrete functional classes of astrocytes in adult gray matter. Glia 31: 95–103. Walz W, Hinks EC (1985) Carrier-mediated KCl accumulation accompanied by water movements is involved in the control of physiological K⫹ levels by astrocytes. Brain Res 343:44 –51. Walz W, Mukerji S (1988) KCl movements during potassium-induced cytotoxic swelling of cultured astrocytes. Exp Neurol 99:17–29. Warth A, Mittelbronn M, Hülper P, Erdlenbruch B, Wolburg H (2007) Expression of the water channel protein aquaporin-9 in malignant brain tumors. Appl Immunohistochem Mol Morphol 15:193–198. Warth A, Mittelbronn M, Wolburg H (2005) Redistribution of the water channel protein aquaporin-4 and the K⫹ channel protein Kir4.1 differs in low- and high-grade human brain tumors. Acta Neuropathol 109:418 – 426. Watanabe H, Vriens J, Suh SH, Benham CD, Droogmans G, Nilius B (2002a) Heat-evoked activation of TRPV4 channels in a HEK293 cell expression system and in native mouse aorta endothelial cells. J Biol Chem 277:47044 – 47051. Watanabe H, Davis JB, Smart D, Jerman JC, Smith GD, Hayes P, Vriens J, Cairns W, Wissenbach U, Prenen J, Flockerzi V, Droogmans G, Benham CD, Nilius B (2002b) Activation of TRPV4 channels (hVRL-2/mTRP12) by phorbol derivatives. J Biol Chem 277:13569 –13577. Watanabe H, Vriens J, Prenen J, Droogmans G, Voets T, Nilius B (2003) Anandamide and arachidonic acid use epoxyeicosatrienoic acids to activate TRPV4 channels. Nature 424:434 – 438. Waxman SG (2001) Transcriptional channelopathies: an emerging class of disorders. Nat Rev Neurosci 2:652– 659. Wegierski T, Lewandrowski U, Müller B, Sickmann A, Walz G (2009) Tyrosine phosphorylation modulates the activity of TRPV4 in response to defined stimuli. J Biol Chem 284:2923–2933. Wen H, Nagelhus EA, Amiry-Moghaddam M, Agre P, Ottersen OP, Nielsen S (1999) Ontogeny of water transport in rat brain: postnatal expression of the aquaporin-4 water channel. Eur J Neurosci 11:935–945. Wurm A, Pannike T, Iandiev I, Wiedemann P, Reichenbach A, Brigmann A (2006) The developmental expression of K⫹ channels in retinal glial cells is associated with a decrease of osmotic cell swelling. Glia 54:411– 423. Xiong ZQ, Stringer JL (2000) Sodium pump activity, not glial spatial buffering, clears potassium after epileptiform activity induced in the dentate gyrus. J Neurophysiol 83:1443–1451. Xu H, Zhao H, Tian W, Yoshida K, Roullet JB, Cohen DM (2003) Regulation of a transient receptor potential (TRP) channel by tyrosine phosphorylation. SRC family kinase-dependent tyrosine phosphorylation of TRPV on TYR-253 mediates its response to hypotonic stress. J Biol Chem 278:11520 –11527. Yang B, Zhao D, Verkman AS (2006) Evidence against functionally significant aquaporin expression in mitochondria. J Biol Chem 281:16202–16206. Ye ZC, Oberheim N, Kettenmann H, Ransom BR (2009) Pharmacological “cross-inhibition” of connexin hemichannels and swelling activated anion channels. Glia 57:258 –269. Yu XS, Jiang JX (2004) Interaction of major intrinsic protein (aquaporin-0) with fiber connexins in lens development. J Cell Sci 117:871– 880. Zhang H, Verkman AS (2008) Aquaporin-4 independent Kir4.1 K⫹ channel function in brain glial cells. Mol Cell Neurosci 37:1–10.

940

V. Benfenati and S. Ferroni / Neuroscience 168 (2010) 926 –940

Zhang Y, Zhang H, Feustel PJ, Kimelberg HK (2008) DCPIB, a specific inhibitor of volume regulated anion channels (VRACs), reduces infarct size in MCAO and the release of glutamate in the ischemic cortical penumbra. Exp Neurol 210:514 –520. Zhou M, Kimelberg HK (2000) Freshly isolated astrocytes from rat hippocampus show two distinct current patterns and diffe-

rent [K(⫹)](o) uptake capabilities. J Neurophysiol 84:2746 – 2757. Zhou M, Xu G, Xie M, Zhang X, Schools GP, Ma L, Kimelberg HK, Chen H (2009) TWIK-1 and TREK-1 are potassium channels contributing significantly to astrocyte passive conductance in rat hippocampal slices. J Neurosci 29:8551– 8564.

(Accepted 5 December 2009) (Available online 22 December 2009)