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Spatiotemporal analysis of glutamatergic neurotransmission
e94
Abstracts / Neuroscience Research 71S (2011) e46–e107
cific inhibitory synapses in the basal nucleus (BA), but not lateral nucleus, of the amygdala. The synapses, termed invaginating synapses, consisted of conventional symmetrical contact and unique perisynaptic invagination of nerve terminals into perikarya. At invaginating synapses, DGL␣ was preferentially recruited to concave somatic membrane of postsynaptic pyramidal neurons, while invaginating presynaptic terminals highly expressed CB1 , MGL, and CCK. No such molecular convergence was seen for flat perisomatic synapses made by parvalbumin-positive interneurons. On the other hand, DGL␣ and CB1 were expressed weakly at axo-spinous excitatory synapses. Consistent with these morphological data, thresholds for DGL␣mediated depolarization-induced retrograde suppression were much lower for inhibitory synapses than for excitatory synapses in BA pyramidal neurons. Moreover, depolarization-induced suppression was readily saturated for inhibition, but never for excitation. These findings suggest that perisomatic inhibition by invaginating synapses is a key target of 2-AG-mediated control of the excitability of BA pyramidal neurons. doi:10.1016/j.neures.2011.07.401
O4-E-3-2 Lognormal sparse connectivity generates intrinsic noise optimal for information processing in cortical networks Jun-nosuke Teramae 1,2 , Yasuhiro Tsubo 1 , Tomoki Fukai 1,3 1
BSI, RIKEN, Saitama, Japan Saitama, Japan
2
PRESTO, JST, Saitama, Japan
3
CREST, JST,
In the absence of sensory stimulation, cortical circuits generate intrinsic irregular activity. There has been much recent interest in the genesis and functional roles of such spontaneous activity or noise in the brain. However, the computational rule governing the generation and functional significance of noise is unknown. Here, we present a biologically plausible mechanism to generate intrinsic noise and to utilize it for information processing in recurrent neural networks. In a recurrent neural network model, we show that a simple synaptic transformation, a lognormal scaling of recurrent excitatory potentials (EPSPs) results in spontaneous noise generation. This synaptic connectivity coordinates asynchronous sparse firing, spike sequence generation and routing, UP state maintenance, and excitatory-inhibitory balance. Our model for the first time links these seemingly unrelated observations within a self-consistent framework for network dynamics. We also show that the lognormal scaling prevented the network from falling into a seizure-like state in response to external stimuli and information about external stimuli simultaneously travel along multiple pathways without much interference. This implies that the cortex processes information in parallel, but not in a densely distributed manner (PnDP). Remarkably, the lognormal scaling creates functional asymmetries between dense/weak and sparse/strong synaptic inputs that both maximizes and modulates the flow of information in the network. Dense/weak synapses in lognormal-connected circuits are intrinsically optimal for information processing by strong synapses. To validate our modeling results of the synaptic dualism, we perform dynamic clamp recordings from cortical neurons to induce lognormally distributed EPSPs in the membrane dynamics. Our results may have important implications for understanding and conceptualizing the computational principles underlying neural circuit function. Research fund: KAKENHI 20700304 (to J.T.), 22700323 (to Y.T.) and 22115013 (to T.F.). doi:10.1016/j.neures.2011.07.402
O4-E-3-3 Spatiotemporal analysis of glutamatergic neurotransmission Hirokazu Sakamoto , Shigeyuki Namiki, Kenzo Hirose Dept. Neurobiol., Grad. Sch. of Med., Tokyo Univ., Tokyo, Japan Neurotransmitter release at synapses is fundamental to information processing in brain, yet little is known about its spatiotemporal dynamics at the level of single synapses. Here, we visualized glutamate, a major excitatory neurotransmitter, released from individual presynaptic terminals with single quantum resolution using a fluorescent glutamate sensor. Our analysis shows that the number of releasable vesicles and the release probability were regulated by cell-specific and synapse-specific mechanisms, and that their heterogeneity had different impacts on spatiotemporal dynamics of glutamate release. Furthermore, we identified the molecular mechanisms underlying synapse-specific regulation of these presynaptic properties. Research fund: Global COE, SRPBS.
O4-E-3-4 Cis complex of NB-2/contactin-5 and amyloid precursor-like protein 1 (APLP1) is localized at the presynaptic sites Yasushi Shimoda 1 , Fumiya Koseki 1 , Masaki Itoh 1 , Kyohei Osada 1 , Manabu Toyoshima 1 , Kazutada Watanabe 1,2 1
Dept Bioeng, Nagaoka Univ Tech, Nagaoka, Japan 2 Nagaoka Nat Coll Tech, Nagaoka, Japan NB-2/contactin-5, a GPI-anchored glycoprotein belonging to the immunoglobulin superfamily, is expressed at glutamatergic synapses in the central auditory system. Deficiency of NB-2 causes a deficit of synapse formation and induces apoptosis in the auditory neurons. However, the molecular mechanism underlying NB-2-mediated synapse formation remains unknown. It was recently reported that NB-2 interacts with amyloid precursor-like protein 1 (APLP1), a member of amyloid precursor protein family known to be involved in synapse formation and function. In this study, we investigated the physiological significance of interaction between NB-2 and APLP1. First, we performed pull-down assay, cell surface binding assay and immunoprecipitation to confirm the interaction of NB-2 with APLP1. Next, in situ hybridization and immunohistochemical analysis demonstrated that NB-2 and APLP1 are likely to co-localize in the cerebral cortex, the hippocampus and the auditory system, suggesting that NB-2 and APLP1 may interact in these area. Immunofluorescence of cultured hippocampal neurons showed that both NB-2 and APLP1 signals overlapped with synapsin 1 signals, suggesting that NB-2 and APLP1 are co-localized at the presynaptic sites. To biochemically examine their co-localization on the presynaptic membrane, we isolated the pre- and postsynaptic membrane fractions from the synaptosomal fraction of the cerebral cortex. Western blotting revealed that both NB-2 and APLP1 were enriched in the presynaptic fraction. NB-2 and APLP1 were co-immunoprecipitated from the lysate of HEK293T cells co-expressing NB-2 and APLP1, but not from the lysate of mixed culture of cells expressing NB-2 with cells expressing APLP1, implying that NB-2 and APLP1 interact on the membrane in cis manner. Taken together, our results suggest that NB-2 forms cis-complex with APLP1 on the presynaptic membrane. There is a possibility that NB-2 might regulate processing of APLP1 to control synapse formation. Research fund: KAKENHI (18300120). doi:10.1016/j.neures.2011.07.404
O4-F-1-1 Identification of a critical genetic determinant of midbrain commissural neurons Yasuyuki Inamata , Ryuichi Shirasaki Grad Sch Frontier Biosci., Osaka Univ., Suita, Japan During development, the construction of proper neuronal circuits depends upon the fidelity with which growing axons select specific pathways to reach their target cells. Although studies of developing spinal cord have provided insights into basic strategies of neuronal fate specification, the knowledge and understanding of the genetic programs for wiring the nervous system are still limited. To address this question, we have been studying molecular programs that direct the navigation of commissural axons that cross the floor plate (FP) at the ventral midline of the CNS. Previously, we have shown that dorsal commissural neurons from the spinal cord rostrally to the midbrain share axon guidance mechanisms in the context of netrin/DCC signaling (Shirasaki et al., 1996, Neuron). This raised the possibility that genetic programs upstream of the axon guidance mechanisms are also shared. However, this assumption did not hold true, for the bHLH transcription factor Atoh1 that acts as a determinant of dI1 dorsal commissural neurons is absent in the CNS rostral to the midbrain/hindbrain boundary. This prompted us to search for the missing determinant in the midbrain. In the current study, we show in mice that the midbrain commissural neurons also express Lhx2 and Robo3, components of which are indispensable for midline crossing caudal to the midbrain. Intriguingly, in gain-of-function experiments using in vivo electroporation, an ectopic expression of Dbx1 induces the generation of commissural neurons. Conversely, loss-of-function of endogenous Dbx1 with a dominant-negative form of Dbx1 results in an opposite phenotype. These results indicate that Dbx1 is a critical intrinsic determinant of midbrain commissural neurons in vivo. Our results also suggest that Atoh1and Dbx1-initiated transcriptional cascades possess molecular programs for expression of similar guidance effectors. Research fund: Grant-in-Aid for Young Scientists (A) and (S) (18680028, 21670002). doi:10.1016/j.neures.2011.07.405
doi:10.1016/j.neures.2011.07.403
Abstracts / Neuroscience Research 71S (2011) e46–e107
cific inhibitory synapses in the basal nucleus (BA), but not lateral nucleus, of the amygdala. The synapses, termed invaginating synapses, consisted of conventional symmetrical contact and unique perisynaptic invagination of nerve terminals into perikarya. At invaginating synapses, DGL␣ was preferentially recruited to concave somatic membrane of postsynaptic pyramidal neurons, while invaginating presynaptic terminals highly expressed CB1 , MGL, and CCK. No such molecular convergence was seen for flat perisomatic synapses made by parvalbumin-positive interneurons. On the other hand, DGL␣ and CB1 were expressed weakly at axo-spinous excitatory synapses. Consistent with these morphological data, thresholds for DGL␣mediated depolarization-induced retrograde suppression were much lower for inhibitory synapses than for excitatory synapses in BA pyramidal neurons. Moreover, depolarization-induced suppression was readily saturated for inhibition, but never for excitation. These findings suggest that perisomatic inhibition by invaginating synapses is a key target of 2-AG-mediated control of the excitability of BA pyramidal neurons. doi:10.1016/j.neures.2011.07.401
O4-E-3-2 Lognormal sparse connectivity generates intrinsic noise optimal for information processing in cortical networks Jun-nosuke Teramae 1,2 , Yasuhiro Tsubo 1 , Tomoki Fukai 1,3 1
BSI, RIKEN, Saitama, Japan Saitama, Japan
2
PRESTO, JST, Saitama, Japan
3
CREST, JST,
In the absence of sensory stimulation, cortical circuits generate intrinsic irregular activity. There has been much recent interest in the genesis and functional roles of such spontaneous activity or noise in the brain. However, the computational rule governing the generation and functional significance of noise is unknown. Here, we present a biologically plausible mechanism to generate intrinsic noise and to utilize it for information processing in recurrent neural networks. In a recurrent neural network model, we show that a simple synaptic transformation, a lognormal scaling of recurrent excitatory potentials (EPSPs) results in spontaneous noise generation. This synaptic connectivity coordinates asynchronous sparse firing, spike sequence generation and routing, UP state maintenance, and excitatory-inhibitory balance. Our model for the first time links these seemingly unrelated observations within a self-consistent framework for network dynamics. We also show that the lognormal scaling prevented the network from falling into a seizure-like state in response to external stimuli and information about external stimuli simultaneously travel along multiple pathways without much interference. This implies that the cortex processes information in parallel, but not in a densely distributed manner (PnDP). Remarkably, the lognormal scaling creates functional asymmetries between dense/weak and sparse/strong synaptic inputs that both maximizes and modulates the flow of information in the network. Dense/weak synapses in lognormal-connected circuits are intrinsically optimal for information processing by strong synapses. To validate our modeling results of the synaptic dualism, we perform dynamic clamp recordings from cortical neurons to induce lognormally distributed EPSPs in the membrane dynamics. Our results may have important implications for understanding and conceptualizing the computational principles underlying neural circuit function. Research fund: KAKENHI 20700304 (to J.T.), 22700323 (to Y.T.) and 22115013 (to T.F.). doi:10.1016/j.neures.2011.07.402
O4-E-3-3 Spatiotemporal analysis of glutamatergic neurotransmission Hirokazu Sakamoto , Shigeyuki Namiki, Kenzo Hirose Dept. Neurobiol., Grad. Sch. of Med., Tokyo Univ., Tokyo, Japan Neurotransmitter release at synapses is fundamental to information processing in brain, yet little is known about its spatiotemporal dynamics at the level of single synapses. Here, we visualized glutamate, a major excitatory neurotransmitter, released from individual presynaptic terminals with single quantum resolution using a fluorescent glutamate sensor. Our analysis shows that the number of releasable vesicles and the release probability were regulated by cell-specific and synapse-specific mechanisms, and that their heterogeneity had different impacts on spatiotemporal dynamics of glutamate release. Furthermore, we identified the molecular mechanisms underlying synapse-specific regulation of these presynaptic properties. Research fund: Global COE, SRPBS.
O4-E-3-4 Cis complex of NB-2/contactin-5 and amyloid precursor-like protein 1 (APLP1) is localized at the presynaptic sites Yasushi Shimoda 1 , Fumiya Koseki 1 , Masaki Itoh 1 , Kyohei Osada 1 , Manabu Toyoshima 1 , Kazutada Watanabe 1,2 1
Dept Bioeng, Nagaoka Univ Tech, Nagaoka, Japan 2 Nagaoka Nat Coll Tech, Nagaoka, Japan NB-2/contactin-5, a GPI-anchored glycoprotein belonging to the immunoglobulin superfamily, is expressed at glutamatergic synapses in the central auditory system. Deficiency of NB-2 causes a deficit of synapse formation and induces apoptosis in the auditory neurons. However, the molecular mechanism underlying NB-2-mediated synapse formation remains unknown. It was recently reported that NB-2 interacts with amyloid precursor-like protein 1 (APLP1), a member of amyloid precursor protein family known to be involved in synapse formation and function. In this study, we investigated the physiological significance of interaction between NB-2 and APLP1. First, we performed pull-down assay, cell surface binding assay and immunoprecipitation to confirm the interaction of NB-2 with APLP1. Next, in situ hybridization and immunohistochemical analysis demonstrated that NB-2 and APLP1 are likely to co-localize in the cerebral cortex, the hippocampus and the auditory system, suggesting that NB-2 and APLP1 may interact in these area. Immunofluorescence of cultured hippocampal neurons showed that both NB-2 and APLP1 signals overlapped with synapsin 1 signals, suggesting that NB-2 and APLP1 are co-localized at the presynaptic sites. To biochemically examine their co-localization on the presynaptic membrane, we isolated the pre- and postsynaptic membrane fractions from the synaptosomal fraction of the cerebral cortex. Western blotting revealed that both NB-2 and APLP1 were enriched in the presynaptic fraction. NB-2 and APLP1 were co-immunoprecipitated from the lysate of HEK293T cells co-expressing NB-2 and APLP1, but not from the lysate of mixed culture of cells expressing NB-2 with cells expressing APLP1, implying that NB-2 and APLP1 interact on the membrane in cis manner. Taken together, our results suggest that NB-2 forms cis-complex with APLP1 on the presynaptic membrane. There is a possibility that NB-2 might regulate processing of APLP1 to control synapse formation. Research fund: KAKENHI (18300120). doi:10.1016/j.neures.2011.07.404
O4-F-1-1 Identification of a critical genetic determinant of midbrain commissural neurons Yasuyuki Inamata , Ryuichi Shirasaki Grad Sch Frontier Biosci., Osaka Univ., Suita, Japan During development, the construction of proper neuronal circuits depends upon the fidelity with which growing axons select specific pathways to reach their target cells. Although studies of developing spinal cord have provided insights into basic strategies of neuronal fate specification, the knowledge and understanding of the genetic programs for wiring the nervous system are still limited. To address this question, we have been studying molecular programs that direct the navigation of commissural axons that cross the floor plate (FP) at the ventral midline of the CNS. Previously, we have shown that dorsal commissural neurons from the spinal cord rostrally to the midbrain share axon guidance mechanisms in the context of netrin/DCC signaling (Shirasaki et al., 1996, Neuron). This raised the possibility that genetic programs upstream of the axon guidance mechanisms are also shared. However, this assumption did not hold true, for the bHLH transcription factor Atoh1 that acts as a determinant of dI1 dorsal commissural neurons is absent in the CNS rostral to the midbrain/hindbrain boundary. This prompted us to search for the missing determinant in the midbrain. In the current study, we show in mice that the midbrain commissural neurons also express Lhx2 and Robo3, components of which are indispensable for midline crossing caudal to the midbrain. Intriguingly, in gain-of-function experiments using in vivo electroporation, an ectopic expression of Dbx1 induces the generation of commissural neurons. Conversely, loss-of-function of endogenous Dbx1 with a dominant-negative form of Dbx1 results in an opposite phenotype. These results indicate that Dbx1 is a critical intrinsic determinant of midbrain commissural neurons in vivo. Our results also suggest that Atoh1and Dbx1-initiated transcriptional cascades possess molecular programs for expression of similar guidance effectors. Research fund: Grant-in-Aid for Young Scientists (A) and (S) (18680028, 21670002). doi:10.1016/j.neures.2011.07.405
doi:10.1016/j.neures.2011.07.403