シンポジウム（Symposium）

Symposium A new era of neuroscience with zebrafish シンポジウムゼブラフィッシュが拓く神経科学

7月26日（金）15:10～15:30　第7会場（朱鷺メッセ　2F　201B） 2S07a-1 ゼブラフィッシュ小脳神経回路の発生と機能 Masahiko Hibi（日比正彦）^1,2，Tsubasa Itoh（伊藤翼）¹，Shinnosuke Yura（由良信之介）¹，Koji Matsuda（松田光司）^1,2，Takashi Shimizu（清水貴史）^1,2 ¹名古屋大学大学院理学研究科生命理学専攻 ²名古屋大学生物機能開発利用研究センター The cerebellum plays an important role in some forms of skillful movements, motor learning, and cognitive functions. Structure and development of the cerebellar neural circuits are generally conserved among vertebrates. Purkinje cells (PCs) and the neurons in the inferior olivary nuclei (IO neuros) in the hindbrain compose the cerebellar neural circuity. Previous studies revealed that PCs and the IO neurons are derived from neuronal progenitors that express the proneural gene ptf1a in the ventricular zone (VZ) of the hindbrain (rhombomere, r1-7). Although the cell fate of these neurons is known to be controlled by gradients of Fgf (high at rhombomere [r] 1 and 4) and retinoic acid (RA) signals (high at r7), it remains elusive what factors function downstream of these signals to generate the different neurons from the progenitors. We addressed this issue by using zebrafish as a model. We found that the homeobox gene gsx2 is expressed in the VZ of the r7 and required for differentiation of the IO neurons. As the gsx2 expression is regulated by the Fgf and RA signals, gsx2 functions downstream of the rostro-caudal positional information to control specification of the IO neurons. We found differentiating or differentiated PCs express genes encoding transcriptional activators Foxp1b and Foxp4, and transcriptional corepressors Skor1b and Skor2. foxp1b;foxp4 double mutant larvae showed a strong reduction in PCs. skor1b;skor2 double mutant larvae completely lacked PCs but instead showed a marked increase in granule cells (GCs), indicating that Foxp1b and Foxp4, and Skor1b and Skor2 function redundantly as key transcriptional regulators in the initial step of PC differentiation. Foxp1b and Foxp4 positively regulate expression of genes required for the PC differentiation, whereas Skor1b and Skor2 suppress the GC fate by repressing the GC genes. Our findings shed light on the mechanism by which transcriptional regulators control differentiation of different type of neurons from the same progenitors for the cerebellar neural circuit formation. In addition, we shall discuss new optotenetic tools to decipher functions of the cerebellar neural circuits in zebrafish.		7月26日（金）15:30～15:50　第7会場（朱鷺メッセ　2F　201B） 2S07a-2 脊髄V1ニューロンは、ロコモーション中の運動ニューロンの選択的動員パターンの生成に必須の役割を果たす Shin-ichi Higashishima（東島真一），Yukiko Kimura（木村有希子）自然科学研究機構生命創成探究センター Vertebrates can produce movements of widely varying strength and speed by using rhythmic networks of neurons located in the spinal cord. A leading model for determining the recruitment patterns of motoneurons (MNs) during speed/strength changes is the ""size principle. According to this principle, the pool of active cells steadily increases in size with progressive increases in the force and speed of movement. It is assumed that MNs that are bigger in size and innervate fast-type muscles are added to smaller MNs that innervate slow-type muscle as the speed/strength of movement increases. Indeed, in larval zebrafish, it has been found that MNs of larger size are recruited only during strong/fast movements. Similar phenomena were also found in a class of excitatory premotor interneurons (V2a neurons). Studies in larval zebrafish, however, have also shown that recruitment patterns did not perfectly follow a simple adding rule. Slow-type MNs as well as slow-type V2a neurons were found to actually be inactive during stronger/faster movements, suggesting that, with increasing speed/strength of movements, deactivation of slow-type MNs and interneurons occurs. Unlike fish, skeletal muscles in mammals consist of mixed fibers with slow- and fast-type muscle fibers intermingled, which makes it more difficult to accurately examine the activities of slow- and fast-type muscles separately. Nonetheless, several lines of evidence suggest that preferential recruitment of faster muscle fibers during rapid contractions occur. Importantly, however, the neuronal basis for the silencing of slow-component neurons during fast/strong movements has remained largely unknown in any vertebrate species. Here, we performed functional analyses of spinal V1 neurons by selectively killing them in larval zebrafish, revealing two functions of V1 neurons. The first is the long-proposed role of V1 neurons: they play an important role in shortening the cycle period during swimming by providing in-phase inhibition. The second is that V1 neurons play an important role in the selection of active sets of neurons. We showed that strong inhibitory inputs coming from V1 neurons play a crucial role in suppressing the activities of slow-type V2a and motor neurons, and, consequently, of slow muscles during fast swimming. Our results thus highlight the critical role of spinal inhibitory neurons for silencing slow-component neurons during fast movements.

7月26日（金）15:50～16:10　第7会場（朱鷺メッセ　2F　201B） 2S07a-3 オプティックフロー情報処理に関わる神経回路の解剖学的基盤 Fumi Kubo（久保郁）国立遺伝研 Animals use global image motion to actively stabilize their position by compensatory body movements, such as the optomotor response. In zebrafish, the main visual area that process optic flow information is the pretectum. Previous studies have shown that pretectal neurons distinguish different optic flow patterns, such as rotation and translation, to drive appropriate compensatory behaviors. To elucidate critical neuroanatomical features that underlie this sensorimotor transformation, we have combined functional imaging and morphological reconstruction of single cells. Synaptic terminals of direction-selective retinal ganglion cells (DS-RGCs) are located within retinal arborization field 5 (AF5) in the pretectum, where they meet the dendrites of pretectal neurons that show "simple" tuning to monocular optic flow. Translation-selective neurons, which respond selectively to optic flow in the same direction for both eyes, are intermingled with "simple" cells and share neuropil, but do not receive inputs from DS-RGCs. Furthermore, we used a recently compiled atlas of single neurons in larval zebrafish to reveal projection patterns of pretectal neurons. We found that mutually exclusive populations of pretectal projection neurons innervate either the reticular formation or the cerebellum, which in turn control optomotor behavior. These results suggest that local computations in a defined pretectal circuit transform optic flow signals into motor commands that drive optomotor behavior.		7月26日（金）16:10～16:30　第7会場（朱鷺メッセ　2F　201B） 2S07a-4 嗅覚記憶と意欲行動の神経回路メカニズム Nobuhiko Miyasaka（宮坂信彦）¹，Yoshihiro Yoshihara（吉原良浩）^1,2 ¹理研CBS システム分子行動学 ²理研CBS 花王連携セ One of the most important roles of the olfactory system is to learn and memorize odor cues associated with food, allowing animals to efficiently find food for survival. However, its underlying circuit mechanisms are not fully understood. Here we develop a simple behavioral paradigm for appetitive olfactory conditioning in zebrafish and analyze brain regions that are activated after learning. Administration of a synthetic odorant, morpholine, into a test tank does not elicit obvious behavioral changes in naive zebrafish, whereas repeated pairing of the odorant with food results in odorant-evoked attraction of zebrafish to the odorant source and/or food ports before supplying food. In situ hybridization analyses following the probe trial reveal that the number of c-fos-positive cells in a specific subnucleus of the thalamus, whose mammalian counterpart is still unknown, is significantly higher in the paired conditioning group than that in the unpaired control group, although the numbers of c-fos-positive cells in the posterior and medial zones of dorsal telencephalic area (Dp and Dm, homologs of the piriform cortex and the pallial amygdala in mammals, respectively) are unchanged between the two groups. Furthermore, upon transferring naive zebrafish from their home tank to a test tank, c-fos-positive cells increases in number in the thalamic subnucleus. These results imply that activation of the specific subnucleus of the thalamus could correlate with a brain state common to motivated behaviors including appetitive behavior evoked by a reinforced odorant cue and exploratory behavior in a novel environment. Further experiments are ongoing to clarify the molecular expression profile, neuroanatomy and function of the c-fos-positive neurons in the thalamic subnucleus.

7月26日（金）16:30～16:50　第7会場（朱鷺メッセ　2F　201B） 2S07a-5 ゼブラフィッシュ成魚における予測誤差による行動選択機構 Makio Torigoe（鳥越万紀夫）¹，Tanvir Islam（Islam Tanvir）^1,4，Hisaya Kakinuma（柿沼久哉）^1,4，Chi Chung Alan Fung（Fung Alan Chi Chung）²，Takuya Isomura（磯村拓哉）³，Hideaki Shimazaki（島崎秀昭）³，Tazu Aoki（青木田鶴）¹，Tomoki Fukai（深井朋樹）²，Hitoshi Okamoto（岡本仁）^1,4 ¹理研CBS 意思決定回路動態 ²理研CBS 神経情報・脳計算 ³理研CBS 数理脳科学 ⁴理研CBS 花王連携セ Capacity to predict the desirable future and to plan the proper behavioral strategy to realize it has been regarded as the inherent functions of the higher vertebrate brain in the decision making. However, adult zebrafish has recently drawn attention as a model animal for the study of decision making due to its capability of various adaptive behaviors and the conservation of the basic telencephalic structure which is contributing to the decision making. In this study, we aimed at directly addressing this process by establishing the closed-loop virtual reality system for the head-tethered adult zebrafish with the 2-photon calcium imaging system. The adult zebrafish harboring G-CaMP7 in the excitatory neurons were trained to perform visual-based active and passive avoidance tasks and simultaneously the neural activities were imaged in the cellular level. The Non-negative Matrix Factorization analysis revealed the one ensemble of neurons whose activities were suppressed by the recognized backward movement of the landscape, and the other ensemble suppressed by perceiving red goal color after learning. These results suggest that these ensembles encode the errors from the predictions of each perceptual state which is favorable to achieve the active avoidance, i.e. perceiving the visual flow of backward movement of the landscape and perceiving the red goal color. To test this, we artificially enhanced these prediction errors by introducing the open-loop condition in which fish could not swim forward and could not reach the goal however hard fish beat the tail. In this condition, the activities of the two ensembles elevated and fish behaved to correct errors. In addition, we changed the goal color to different colors as soon as reaching the goal. We observe the increased activity of another ensemble when fish perceived the different colors of the goal. Taken together, at the onset of the trial, learned fish conceives two future conditions as the favorable status on its way to the safe goal, i.e. one with the backwardly moving landscape and the other with the color of the safe goal. The two different neural ensembles monitor the discrepancy between these predictions and the perceived real external status. Once fish reaches the goal, another ensemble is set to work to monitor whether fish keeps staying in the safe goal. These results suggest that fish sets the behavioral strategy to actively realize these predictions, and fish's capability of prediction of future.		7月26日（金）16:50～17:10　第7会場（朱鷺メッセ　2F　201B） 2S07a-6 ゼブラフィッシュにおける扁桃体および海馬機能の研究 Koichi Kawakami（川上浩一）^1,2 ¹国立遺伝研遺伝形質発生遺伝 ²総研大院遺伝学 The amygdala and hippocampus are two major components of the mammalian brains and perform crucial roles in the processing of emotional memory and episodic and spatial memory, respectively. In teleost, the medial and lateral zone of the dorsal telencephalon (Dm and Dl) have been postulated to be homologs of the mammalian amygdala and hippocampus based on neuroanatomical and functional studies. However, both Dm and Dl are broad areas in the dorsal telencephalon and the neural circuitries mediating the amygdalar and hippocampal functions have yet to be explored. Here we identified the neuronal populations that are essential for emotional learning and episodic and spatial learning by a genetic approach in zebrafish. We performed large-scale gene trap and enhancer trap screens and generated transgenic fish that expressed Gal4FF, a synthetic Gal4 transcription activator, in specific regions and neuronal circuits in the brain. Then we crossed these brain-specific Gal4FF transgenic fish lines with UAS-neurotoxin lines to inhibit the activity of the Gal4FF-expressing neurons, and analyzed behaviors of the double transgenic fish. We found that, when the activity of a subpopulation of neurons in the Dm was inhibited, the fish showed deficits in the emotional learning (fear conditioning) paradigm. Furthermore, we found that, when the activity of a subpopulation of neurons in the Dl was inhibited, the fish showed deficits in the episodic (trace fear conditioning) and spatial learning paradigms. Thus, we propose these neuronal populations are functional equivalents of the mammalian amygdala and hippocampus, respectively. We identified projections of these neuronal circuits by making serial cross sections and also by preparing cleared brains with the Scale method and light-sheet microscopy. Our study provides a basis for understanding essential neuronal circuits mediating evolutionarily conserved behaviors in vertebrates.