リズム運動パターン生成 Rhythmic Motor Pattern Generation

P1-1-78 ショウジョウバエ幼虫の運動回路におけるGABA作動性神経細胞の機能解析 Identification and functional analysis of a class of local GABAergic interneurons in the Drosophila larval motor circuits ○伏木彬1, 高坂洋史1, 能瀬聡直1,2 ○Akira Fushiki1, Hiroshi Kohsaka1, Akinao Nose1,2 東京大学大学院　新領域創成科学研究科　複雑理工学専攻1, 東京大学大学院　理学系研究科　物理学専攻2 Dept of Complexity Sci and Eng, Grad Sch of Frontier Sci, Univ of Tokyo, Japan1, Dept of Physics, Grad Sch of Science, Univ of Tokyo, Japan2 The balance between excitatory and inhibitory neurons is believed to be critical for rhythmic animal movements. Yet, little is known about the mechanisms of how excitatory and inhibitory neurons interact to generate appropriate motor outputs in the developing and mature nervous system. We are trying to address this question using the motor circuits of Drosophila larvae as a model. By using GAL4/UAS system and GCaMP-based calcium imaging, we identified a class of GABAergic interneurons that show propagating activity patterns corresponding to the wave of muscle contractions during larval locomotion. These neurons are segmentally repeated (one neuron per hemisegment) and are located in the dorsolateral areas of the ventral nerve cord. Channelrhodopsin-2 (ChR2)-mediated activation of these neurons in the 3rd instar larvae induced acute paralysis of muscles in the abdominal segments and ceased locomotion, suggesting that these neurons act to inhibit motor outputs. When we blocked their neurotransmitter release throughout embryonic and larval periods by expression of tetanus toxin light chain (TeTxLc), the larval locomotion was severely defected: there was a dramatic decrease in the frequency and speed of peristalsis. In contrast, acute inhibition with halorhodopsin (NpHR) or archaerhodopsin (Arch) in the 3rd instar larvae did not alter the mobility of larvae. These results may suggest that the activity of these neurons is essential during the development and/or maintenance of the neural circuits that generate appropriate larval locomotion. To further pursue this possibility, we are currently studying the effects of temporal inhibition of these neurons in specific stages of embryonic and larval life. We are also trying to identify upstream and downstream neurons by reconstruction from electron microscope (EM) image data (in collaboration with Dr. Albert Cardona in Janelia Farm Research Campus).

P1-1-79 光遺伝学と電気生理学手法によるショウジョウバエ幼虫の蠕動運動を制御する神経回路の解明 Optogenetic and electrophysiological dissection of Drosophila neural networks that regulate larval locomotion ○高木俊輔1, 高坂洋史2, 風間北斗3, 能瀬聡直1,2 ○Shunsuke Takagi1, Hiroshi Kohsaka2, Hokto Kazama3, Akinao Nose1,2 東京大院・理学系・物理学1, 東京大院・新領域・複雑理工学2, 理研・脳科学総合研究センター3 Dept. of Phys., Grad. Sch. of Sci., Univ. of Tokyo1, Dept. of Comp. Sci. and Eng., Grad. Sch. of Front. Sci., Univ. of Tokyo2, RIKEN, Inst. of Brain Sci.3 Understanding how motor patterns are regulated by neural circuits remains a major goal in neuroscience. We address this question by using, as a model, the neural circuits that regulate coordinated peristaltic movement of Drosophila larvae. We have recently identified a class of segmentally arrayed local interneurons, termed PMSIs (period-positive median segmental interneurons). When the function of PMSIs is temporally inhibited, the speed of larval crawling is greatly reduced. PMSIs form putative synaptic endings with motor neurons (MNs) and induce relaxation of muscles when activated by light. These results suggest that PMSIs regulate the speed of larval locomotion by locally inhibiting the activity of MNs. In this study, we further explored the role of PMSIs in the regulation of motor activity by combining optogenetics and electrophysiology. We expressed channelrhodopsin2 (ChR2) in PMSIs and temporarily activated these neurons while recording the activity of MNs by a patch electrode. The membrane potential and current were recorded from MNs in dissected larvae, in a whole-cell current-clamp and voltage-clamp mode, respectively. Current injections delivered via the recording pipette evoked trains of action potentials in MNs in a control situation. When PMSIs were optically activated during the current injection, however, the firing rate of some MNs was dramatically decreased with their resting potential hyperpolarized. The results provide direct evidence that these interneurons send inhibitory signals to motor neurons. We are currently studying if response to PMSIs activation differs among distinct types of motor neurons, by injecting dye from the electrode during and after the recording, and identifying the type of motor neurons according to the visualized morphology.

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