Intercellular Communication in Retinal Development

描述（由申请人提供）：神经元的输出不仅受兴奋的影响，还受神经递质，γ -氨基丁酸（GABA）和甘氨酸介导的抑制的大小和时间的影响。在许多神经系统疾病中，当兴奋不能通过抑制得到适当控制时，神经回路就会出现功能障碍。我们对细胞相互作用的理解是组装和维持适当的抑制性连接远远落后于我们对兴奋电路的了解。在这个项目中，我们建议显著推进对控制轴突兴奋性神经递质释放的抑制性连接的发育和功能维持的认识。我们将重点关注视网膜双极细胞轴突末端的抑制性突触。这些神经元对于将视觉信号从光感受器传递到视网膜神经节细胞至关重要。来自这些细胞的信息传递是由至少两种不同类型的抑制突触形成的。在Aim 1中，我们将遗传标记调节抑制作用的离子性GABAA和GABAC受体到相同的轴突上，但具有不同的动力学。我们将使用相关荧光成像和连续块面扫描电子显微镜来绘制不同突触类型的连接模式。利用GABA受体亚基特异性条件敲除小鼠，我们将确定携带α 1 GABAA受体亚基的成熟GABA能突触是否需要在发育过程中短暂丰富的α 3亚基的存在。在Aim 2中，我们将采用成像、电生理分析和抑制传递紊乱的突变小鼠来确定神经传递在双极细胞轴突上建立适当的抑制性突触类型组合中的作用。在Aim 3中，我们将区分用于调节双极细胞中不同细胞区室（轴突与树突）中gaba能突触的途径。我们将使用分子和遗传方法来改变细胞内氯化物通量

DESCRIPTION (provided by applicant): The output of a neuron is shaped not only by excitation but also by the magnitude and timing of inhibition mediated by the neurotransmitters, gamma-aminobutyric acid (GABA) and glycine. Circuits become dysfunctional as found in many diseases of the nervous system when excitation is not controlled properly by inhibition. Our understanding of the cellular interactions that assemble and maintain appropriate inhibitory connections lags far behind our knowledge about excitatory circuits. In this project, we propose to significantly advance knowledge of the development and functional maintenance of inhibitory connections that control the release of excitatory neurotransmitters from axons. We will focus on inhibitory synapses on the axon terminals of retinal bipolar cells. These neurons are essential for relaying visual signals from photoreceptors to the retinal ganglion cells. Transmission from these cells is shaped by at least two different types of inhibitory synapses. In Aim 1, we will genetically label ionotropic GABAA and GABAC receptors that regulate inhibition onto the same axon, but with different kinetics. We will use correlative fluorescence imaging and serial block face scanning electron microscopy to map connectivity patterns of the different synapse types. Using GABA receptor subunit specific conditional knockout mice, we will determine whether mature GABAergic synapses bearing alpha1 GABAA receptor subunits require the presence of alpha3 subunits that are transiently abundant during development. In Aim 2, we will employ imaging, electrophysiological assays, and mutant mice with perturbed inhibitory transmission to ascertain the role of neurotransmission in establishing the appropriate combination of inhibitory synapse types on bipolar cell axons. In Aim 3, we will distinguish the pathways that are employed to regulate GABAergic synapses in distinct cell compartments, axon versus dendrite, of bipolar cells. We will use molecular and genetic approaches to alter intracellular chloride flux specifically in these neurons. Our results will significantly advance understanding of the cellular mechanisms that regulate the development and maintenance of presynaptic inhibition across circuits that act in parallel to process sensory signals.