Hideki Miwa

Functional contributions of synaptically localized NR2B subunits of the NMDA receptor to synaptic transmission and long-term potentiation in the adult mouse CNS.

The NMDA‐type glutamate receptor is a heteromeric complex composed of the NR1 and at least one of the NR2 subunits. Switching from the NR2B to the NR2A subunit is thought to underlie functional alteration of the NMDA receptor during synaptic maturation, and it is generally believed that it results in preferential localization of NR2A subunits on the synaptic site and that of NR2B subunits on the extracellular site in the mature brain. It has also been proposed that activation of the NR2A and NR2B subunits results in long‐term potentiation (LTP) and long‐term depression (LTD), respectively. Furthermore, recent reports suggest that synaptic and extrasynaptic receptors may have distinct roles in synaptic plasticity as well as in gene expression associated with neuronal death. Here, we have investigated whether NR2B subunit‐containing receptors are present and functional at mature synapses in the lateral nucleus of the amygdala (LA) and the CA1 region of the hippocampus, comparing their properties between the two brain regions. We have found, in contrast to the above hypotheses, that the NR2B subunit significantly contributes to synaptic transmission as well as LTP induction. Furthermore, its contribution is greater in the LA than in the CA1 region, and biophysical properties of NMDA receptors and the NR2B/NR2A ratio are different between the two brain regions. These results indicate that NR2B subunit‐containing NMDA receptors accumulate on the synaptic site and are responsible for the unique properties of synaptic function and plasticity in the amygdala.

Reduced axonal localization of a Caps2 splice variant impairs axonal release of BDNF and causes autistic-like behavior in mice

Ca2+-dependent activator protein for secretion 2 (CAPS2 or CADPS2) potently promotes the release of brain-derived neurotrophic factor (BDNF). A rare splicing form of CAPS2 with deletion of exon3 (dex3) was identified to be overrepresented in some patients with autism. Here, we generated Caps2-dex3 mice and verified a severe impairment in axonal Caps2-dex3 localization, contributing to a reduction in BDNF release from axons. In addition, circuit connectivity, measured by spine and interneuron density, was diminished globally. The collective effect of reduced axonal BDNF release during development was a striking and selective repertoire of deficits in social- and anxiety-related behaviors. Together, these findings represent a unique mouse model of a molecular mechanism linking BDNF-mediated coordination of brain development to autism-related behaviors and patient genotype.

Glutamate Decarboxylase 67 Deficiency in a Subset of GABAergic Neurons Induces Schizophrenia-Related Phenotypes

Decreased expression of the GABA synthetic enzyme glutamate decarboxylase 67 (GAD67) in a subset of GABAergic neurons, including parvalbumin (PV)-expressing neurons, has been observed in postmortem brain studies of schizophrenics and in animal models of schizophrenia. However, it is unclear whether and how the perturbations of GAD67-mediated GABA synthesis and signaling contribute to the pathogenesis of schizophrenia. To address this issue, we generated the mice lacking GAD67 primarily in PV neurons and characterized them with focus on schizophrenia-related parameters. We found that heterozygous mutant mice exhibited schizophrenia-related behavioral abnormalities such as deficits in prepulse inhibition, MK-801 sensitivity, and social memory. Furthermore, we observed reduced inhibitory synaptic transmission, altered properties of NMDA receptor-mediated synaptic responses in pyramidal neurons, and increased spine density in hippocampal CA1 apical dendrites, suggesting a possible link between GAD67 deficiency and disturbed glutamatergic excitatory synaptic functions in schizophrenia. Thus, our results indicate that the mice heterozygous for GAD67 deficiency primarily in PV neurons share several neurochemical and behavioral abnormalities with schizophrenia, offering a novel tool for addressing the underlying pathophysiology of schizophrenia.

Contribution of parvalbumin and somatostatin-expressing GABAergic neurons to slow oscillations and the balance in beta-gamma oscillations across cortical layers

Cortical interneurons are classified into several subtypes that contribute to cortical oscillatory activity. Parvalbumin (PV)-expressing cells, a type of inhibitory interneuron, are involved in the gamma oscillations of local field potentials (LFPs). Under ketamine-xylazine anesthesia or sleep, mammalian cortical circuits exhibit slow oscillations in which the active-up state and silent-down state alternate at ~1 Hz. The up state is composed of various high-frequency oscillations, including gamma oscillations. However, it is unclear how PV cells and somatostatin (SOM) cells contribute to the slow oscillations and the high-frequency oscillations nested in the up state. To address these questions, we used mice lacking glutamate decarboxylase 67, primarily in PV cells (PV-GAD67 mice) or in SOM cells (SOM-GAD67 mice). We then compared LFPs between PV-GAD67 mice and SOM-GAD67 mice. PV cells target the proximal regions of pyramidal cells, whereas SOM cells are dendrite-preferring interneurons. We found that the up state was shortened in duration in the PV-GAD67 mice, but tended to be longer in SOM-GAD67 mice. Firing rate tended to increase in PV-GAD67 mice, but tended to decrease in SOM-GAD67 mice. We also found that delta oscillations tended to increase in SOM-GAD67 mice, but tended to decrease in PV-GAD67 mice. Current source density and wavelet analyses were performed to determine the depth profiles of various high-frequency oscillations. High gamma and ripple (60–200 Hz) power decreased in the neocortical upper layers specifically in PV-GAD67 mice, but not in SOM-GAD67. In addition, beta power (15–30 Hz) increased in the deep layers, specifically in PV-GAD67 mice. These results suggest that PV cells play important roles in persistence of the up state and in the balance between gamma and beta bands across cortical layers, whereas SOM and PV cells may make an asymmetric contribution to regulate up-state and delta oscillations.

Parvalbumin-positive neuron-specific GAD67 knockout mice exhibit schizophrenia-like phenotypes

P3-s11 Gene expression analysis of hippocampi from alpha-CaMKII heterozygous knockout (KO) mice and Schnurri-2 KO mice using public microarray databases Hironori K. Nakamura 1 , Keizo Takao 2,3, Keiko Toyama 1,2, Tsuyoshi Takaji 4, Shunsuke Ishii 5, Tsuyoshi Miyakawa 1,2,3 1 Div. of Sys. Med. Sci., Inst. for Comprehensive Med. Sci., Fujita Health Univ., Toyoake, Japan 2 CREST, JST, Saitama, Japan 3 NIPS, Okazaki, Japan 4 Inst. for Develop. Res., Aichi Human Service Center, Kasugai, Japan 5 RIKEN Tsukuba Inst., Tsukuba, Japan

Immunohistochemical characterization of calcium-binding protein-containing GABAergic neurons in the lateral amygdala using vesicular GABA transporter (VGAT)-Venus transgenic mice

The cerebral cortex consists of multiple neuron subtypes, which exhibit diverse modulation patterns in response to neurotransmitters including noradrenaline and acetylcholine. To examine whether cholinergic effects on unitary inhibitory postsynaptic currents (uIPSCs) are differentially regulated by acetylcholine receptors depending on their presynaptic and postsynaptic cell subtypes, we performed multiple whole-cell patch clamp recording from layer V GABAergic interneurons and pyramidal cells in acute slices containing the insular cortex. In fast spiking (FS) to pyramidal cell synapses, carbachol (CCh, 10 M) invariably decreased uIPSC amplitude by 51.3 ± 6.7% (mean ± S.E.M.) which accompanied the increases in coefficient of variation of the 1st uIPSC amplitude and paired-pulse ratio of 2nd to 1st uIPSC amplitude (PPR). CCh-induced uIPSC suppression was dose-dependent and blocked by 100 M atropine, a muscarinic receptor antagonist. The similar cholinergic suppression was observed in non-FS to pyramidal cell synapses. In contrast, inhibitory connections composed of presynaptic FS and postsynaptic FS/non-FS cells showed heterogeneous effects on uIPSC amplitude by CCh. In 40.7% pairs, uIPSCs were suppressed by 29.5 ± 5.0%, while in 37.0% pairs uIPSCs were increased by 59.9 ± 11.7%. Non-FS to FS/non-FS synapses also often showed CCh-induced uIPSC facilitation by 28.5 ± 11.9% in 47.1% pairs, whereas 41.2% pairs showed suppression of uIPSC amplitude by 46.0 ± 11.6%. Both in presynaptic FS and non-FS to postsynaptic FS/non-FS connections that showed facilitation of uIPSCs PPR, was larger than that in synapses showing CCh-induced suppression, suggesting that CCh tends to facilitate GABA release in interneuron-interneuron synapses with lower release probability. These results suggest that acetylcholine receptors differently regulate GABAergic synaptic transmission depending on postsynaptic interneuron subtype.

The roles of vesicular GABA transporter (VGAT) in quantal release of GABA

P1-a26 Motilin has a modulatory action on the postsynaptic GABA receptors specifically in the excitatory medial vestibular nuclear neurons Hiroshi Todaka 1 , Tetsuya Tatsukawa 1, Yuchio Yanagawa 3, Katsuei Shibuki 2, Soichi Nagao 1 1 RIKEN, BSI, Lab. for Motor Learning Control, Saitama, Japan 2 Department of Neurophysiology, Brain Research Institute, Niigata University, Niigata, Japan 3 Department of Genetic and Behavioral Neuroscience, Gunma University, Gunma, Japan

Analysis of long-lasting hyperpolarization mediated by GABAb receptors in the lateral amygdala of mice slice preparation using voltage-sensitive dye imaging

Emotional disorders are quite often accompanied with various psychiatric diseases and stress disorders, and it is an urgent problem to overcome these disorders for many peoples. Mice and rats exhibit profound emotional disorders such as elevated anxiety after having a status epilepticus (SE) by pilocarpine injection, however, its precise mechanism is not yet clarified. In order to know the pathogenic origin of the emotional disorders, we studied the changes in behavior and brain structures of mice after inducing SE with various conditions. In the present study, we used well-established behavior tests for anxiety, including a light and dark preference tests, an open field test, and an elevated plus maze test. Marked facilitation of anxiety-like behavior were observed in the mice which induced SE for 4.5 h as early as 4-6 days after SE. This emotional change was very stable and still observed 8 months after SE. It is known that SE induced mossy fiber sprouting of granule cells in dentate gyrus as well as a loss of parvalbumin-positive inter neurons in the cerebral cortex and hippocampus. Since mossy fiber sprouting was not observed on 6 days after SE by immunohistochemistry with antineuropeptide Y antibody, the sprouting was unlikely to be a pathogenic change for the emotional disorder. The mice in which SE was terminated at 1 hour by a phenobarbital administration also exhibited marked facilitation of anxiety-like behaviors 2 days after SE, however, this change was reversible and the mice showed a normal behavior 6 days after SE. These results suggested that there were at least two different mechanisms for the appearance of emotional disorder after pilocarpine-induced SE, one was reversible and another was irreversible, and the duration of SE was critical for the selection of mechanisms.

Postsynaptic modulation of glutamatergic responses by Adenosine A1 receptors in the hippocampal CA1 region

Ropinirole is a non-ergot dopamine agonist used for Parkinson disease. Although the non-ergot drugs have little adverse effects on gastrointestinal tracts compared to ergot-type drugs, they occasionally show serious side effects such as sleep attacks and daytime sleepiness. Because orexin neurons in the hypothalamus play a key role in maintaining arousal state, we examined the effect of ropinirole on orexin neurons maintained in hypothalamic slice culture. Ropinirole at concentrations over 100 M induced a decrease in the number of orexin-immunoreactive neurons but not MCH-immunoreactive neurons in a concentration-dependent manner. This decrease was reversible after washout of ropinirole and was significantly attenuated by a dopamine D2 antagonist (sulpiride) or a serotonin 5-HT1A antagonist (pindolol). Thus, depletion of orexin may contribute to the adverse effect of ropinirole such as sleep attacks and daytime sleepiness during the treatment of Parkinson disease.