Comprehensive understanding of the multifunctional GnRH peptidergic nervous system

TN-GnRH神经元的传感器输入：环境变化冲击动物的感觉器官，然后由大脑的感觉处理机制处理。在这里，我们分析了从感觉处理脑区产生的TN-GnRH神经元的突触输入。我们获得的电生理证据表明，对TN-GnRH神经元的去极化GABA能突触输入的发生，这可能在感觉输入到神经调节TN-GnRH神经元的传递中起重要作用。我们新开发了一种培养分离的TN-GnRH神经元的方法，这将使我们能够在二维原代培养系统中重建GnRH神经元的三维结构。我们计划使用这种新的制备的同时记录细胞内钙动力学（钙成像），神经活动（电生理学），和胞吐GnRH释放（碳纤维安培法和膜电容测量）。生成和分析的转基因青鳉鱼：我们首先生成了一个转基因青鳉线，其中三个形态以及功能不同的GnRH神经元表达GFP荧光蛋白。使用这样的转基因青鳉鱼，gnrh 2-GFP转基因青鳉鱼，我们可以记录自发电活动的gnrh 2神经元的第一次在脊椎动物的大脑。我们发现，它们表现出类似于神经调节TN-GnRH神经元的自发规则起搏活动，这可能表明自发规则起搏活动可能代表神经调节功能。我们还成功地记录了转gnrh 1-GFP基因青鳉的促垂体性GnRH神经元（gneh 1神经元）的电活动，其表现出与神经调节性gnrh 2和gnrh 3神经元完全不同的不规则自发活动。

Sensor inputs to the TN-GnRH neurons: The environmental changes impinge upon the sensory organs of the animals and then are processed by the sensory processing mechanisms of the brain. Here, we analyzed the synaptic inputs to the TN-GnRH neurons that arise from the sensory processing brain areas. We obtained electrophysiological evidence to show the occurrence of depolarizing GABAergic synaptic inputs to the TN-GnRH neurons, which may play an important role in the transmission of sensory inputs to the neuromodulatory TN-GnRH neurons.Isolated primary culture of TN-GnRH neurons : We newly developed a method to culture isolated TN-GnRH neurons, which should enable us to reconstitute three dimensional structures of the GnRH neurons in a two-dimensional primary culture system. We plan to use this new preparation for the simultaneous recording of intracellular calcium dynamics (calcium imaging), neural-activities (electrophysiology), and exocytotic GnRH release (carbon fiber amperometry and membrane capacitance measurement).Generation and analysis of transgenic medaka fish: We first generated a transgenic medaka line in which three morphologically as well as functionally different GnRH neurons express GFP fluorescent proteins. Using one of such transgenic medaka fish, gnrh2-GFP transgenic medaka fish, we could record spontaneous electrical activities of the gnrh2 neurons for the first time in the vertebrate brains. We showed that they exhibit spontaneous regular pacemaker activities similar to the neuromodulatory TN-GnRH neurons, which may suggest that the spontaneous regular pacemaker activity may represent neuromodulatory functions. We also succeeded in recording the electrical activities of the hypophysiotropic GnRH neurons (gneh1 neurons) from gnrh1-GFP transgenic medaka, which showed irregular spontaneous activities quite different from the neuromodulatory gnrh2 and gnrh3 neurons.