TOP ＞ シンポジウム

シンポジウム Principle and breakdown of higher brain function by decoding intracellular signaling 3S4-1 Optogenetic manipulation and imaging of CaMKII-Rho GTPase signaling pathway during synaptic plasticity Murakoshi Hideji National Institute for Physiological Sciences Ca2+/Calmodulin-dependent kinase II（CaMKII）is one of the most important signaling molecules for synaptic plasticity underlying learning and memory. Here, we developed a photo-activatable CaMKII（paCaMKII）. Light-induced spine specific CaMKII activation successfully induced the structural plasticity, which suggesting that CaMKII activation is sufficient for the plasticity. In addition, we imaged the activity of Rho GTPases, RhoA and Cdc42, by using 2pFLIM and found that these molecules are activated via CaMKII. Furthermore, since the loss of function assay suggests that RhoA and Cdc42 works for triggering and maintaining the structural plasticity, respectively, these molecules may cooperatively work for establishing the spine structural plasticity. 3S4-2 Reward action of dopamine on the structural plasticity of dendritic spines Yagishita Sho1，Kasai Haruo1,2 1Lab. of Structural Phys. Med., Univ. of Tokyo，2CREST Animal behaviors are reinforced by subsequent rewards following within a narrow time window. Such reward signals are primarily coded by dopamine, which modulates the synaptic connections of medium spiny neurons in the striatum. However, it has been difficult to understand why dopamine reinforces preceding, but not subsequent, sensorimotor events if dopamine always activates downstream molecules such as protein-kinase A（PKA）. In acute slices of mouse brain, we selectively stimulated dopaminergic and glutamatergic inputs on D1R-MSNs by optogenetic stimulation of dopaminergic fibers and two-photon uncaging of caged-glutamates paired with APs（STDP）. We found that dopamine markedly potentiated spine enlargement, but this only occurred within a narrow time window（0.3-2 s）closely following STDP, which is consistent with behavioral conditioning findings. FRET imaging of Ca2+/calmodulin-dependent protein kinase II（CaMKII）and protein-kinase A（PKA）revealed that the sequence detection involved molecular signaling upstream of PKA activation：Sufficient generation of cAMP for PKA activation occurred only when spikes preceded dopamine to prime adenylyl-cyclase（AC1）, otherwise cAMP was effectively removed by a potent phosphodiesterase（PDE）activity in thin distal dendrites of MSNs due to a large surface-to-volume ratio. Therefore, PKA was activated only within the specific timing for reinforcement, which then activated CaMKII through the dopamine- and cAMP-regulated phosphoprotein 32 kDa（DARPP-32）. Thus, these intracellular mechanisms can explain the reward action of dopamine. We are now studying how D1R-MSNs are involved in a Pavlovian conditioning task using head-fixed mice. 3S4-3 Phospho-proteomic analysis enables discovery of reward signals Nagai Taku1,3，Amano Mutsuki2,3，Yamada Kiyofumi1,3，Kaibuchi Kozo2,3 1Dept. Neuropsychopharmacol. Hosp. Pharm., Nagoya Univ. Grad. Sch. Med.，2Dept. Cell Pharmacol., Nagoya Univ. Grad. Sch. Med.，3SRPBS It is well known that dopamine（DA）is necessary for motor function, motivation, working memory and the reward system. The principal target of DA is medium spiny neurons（MSNs）, which are divided into two distinct classes, DA type 1 receptor（D1R）-or type 2 receptor-expressing neurons（D1R-MSNs or D2R-MSNs, respectively）, within the striatum. D1R-MSNs in the striatum form direct projections to the substantia nigra pars reticulata, whereas D2R-MSNs form indirect projections to the substantia nigra pars reticulata via the pallidum and subthalamic nuclei. These two pathways control the dynamic balance in the basal ganglia-thalamocortical circuit. DA is believed to regulate membrane properties acting through D1R-protein kinase A（PKA）for controlling reward-related behaviors. However, how PKA regulates the D1R-MSN excitability and reward-related behaviors remains largely unknown. To elucidate PKA-dependent reward signaling in D1R-MSNs, we performed phospho-proteomic analyses to comprehensively identify the PKA substrates downstream of D1Rs in the striatum. We also attempted to characterize neurochemical and behavioral significance of these phosphorylated substrates. We will introduce our recent findings in this symposium. 3S4-4 Genetic manipulation of memory engram Matsuo Naoki Dept. of Mol. and Behav. Neurosci., Grad. Sch. of Med., Osaka Univ. It is a fundamental question how memories are represented in the brain. A prevailing hypothesis suggests that memory is encoded by a cooperative activity of specific subset group of neurons. However, identifying these neurons supporting a given memory is challenging because these neuronal ensembles are likely sparsely distributed within the brain. To circumvent this difficulty, we have previously developed a transgenic system in mice that allows us to manipulate neurons activated during a relevant behavior. In the system, the expression of a given transgene is regulated by neuronal activity via the promoter of c-fos gene, whose expression is rapidly and transiently induced in response to neuronal activity, and is also dependent on a tetracycline inducible expression system. I will introduce approaches trying to manipulate the memory engram to address many unanswered questions about memory. I will also discuss about the possibility to manipulate a molecular activity in those neuronal ensembles. 3S4-5 Computational modeling of dopaminergic actions on striatal medium spiny neurons Yoshimoto Junichiro，Yukinawa Naoto，Nakano Takashi Neural Computation Unit, Okinawa Institute of Science and Technology Graduate University The striatum is the input structure of the basal ganglia and involved in various aspects of reward evaluation and incentive-based learning. Since it was suggested that dopamine encodes the reward prediction error, the neural substrates of this type of learning has often been discussed in the context of reinforcement learning theory, in which the dopamine-dependent corticostriatal plasticity is thought to be a critical element for linking sensory inputs and motor actions via reward signals, resulting in goal-directed behavior. How can this plasticity be interpreted as an outcome of intracellular molecular signaling cascades? What dysfunction of molecular signaling could lead to the breakdown of the plasticity? What extracellular signal could modulate the plasticity? In this talk, we present our studies to seek for possible answers of these questions from a viewpoint of“dynamical systems.” The majority of neurons in the striatum is constituted by medium spiny neurons（MSNs）, whose population is divided into two subpopulations：one expresses dopamine D1-like receptors（D1Rs）and the other subpopulation expresses dopamine D2-like receptors（D2Rs）. In the first half, we present a kinetic signal transduction model of D1R-expressing MSN constructed based on existing literature and database, and show that the bistability of the positive feedback loop constituted by PKA, PP2A and DARPP-32（pThr75）could play an important role in reverting long-term depression to long-term potentiation in dopamine-dependent plasticity.In the second half, we present our on-going study on a kinetic signal transduction model of D2R-expressing MSN, which is based on observations that dopamine exerts pharmacological actions opposite to that of D1R-expressing MSN and counteracts adenosine signaling, which facilitates PKA activity via adenosine A2A-like receptors（A2ARs）. The model predicts that the bidirectional plasticity of the MSN could be maintained through the balance between dopamine and adenosine signals. 3S4-6 Comparative genetic analysis of autism and schizophrenia：Focus on rare variants Aleksic Branko，Kushima Itaru，Ozaki Norio Nagoya University, Graduate School of Medicine, Department of Psychiatry There is strong evidence that genetic factors make substantial contributions to the etiology of autism spectrum disorder（ASD）and schizophrenia, with heritability estimates being at least 80%. In recent years new molecular genetics findings, particularly from the application of genome-wide association studies（GWASs）, have implicated risk factors for ASD and schizophrenia, and have suggested the possibility of a genetic overlap between them. Earlier, we conducted low resolution copy number variation（CNV）screening using affimetrix 5.0 array in order to catalog CNVs that may increase the schizophrenia susceptibility in the Japanese population. The current study is an extension of previous project. For the CNV detection, we are using high resolution comparative genomic hybridization array（aCGH）with 720k probes and 4,000 bps resolution. In addition we are conducting whole genome next generation whole genome sequencing for the patients in whom large psychiatric CNVs have been detected.Besides the known large CNVs that are previously reported to be associated with schizophrenia we found hundreds of small to medium size novel, exon disrupting sequence variations in more than 10% of patients with developmental disorders. Many of those are in functionally relevant protein domains, with the potential to affect physiological function of the affected gene productIn summary, these findings point to the number of genomic variants that may be relevant to the pathoetiology of schizophrenia and ASD were below the detection threshold of last generation CNV typing technologies. In addition, much future work is required, and this work should not be constrained by current categorical diagnostic systems. Such studies should explore the relationship of genes and genetic risk factors to symptomatology across current diagnostic categories.