Development of the Gustatory Solitary Nucleus

描述（由申请人提供）：脑干孤束核（rNST）的喙部是第一个中枢味觉中继。 rNST 接收来自面部和舌咽神经的初级传入投射，这些神经支配舌味蕾和乳头，并在反映周围组织的地形中集中终止。 rNST 细胞增殖和分化的时间过程和分子因素尚不清楚。鉴于有组织的味觉系统对于正常功能的重要性，长期目标是确定 rNST 如何在胚胎大鼠中建立。拟议的研究目标是确定： 孤束（ST）预测的发展；构成 rNST 的细胞成分增殖和分化的发育时间过程； rNST 神经元的生物物理特性和突触的发展；与 rNST 发育相关的特定分子因素；并且，操纵这些因子的表达以确定对 rNST 发育的影响。免疫荧光将用于确定分阶段胚胎中组成 rNST 的神经元和神经胶质细胞的发育时间过程。体外脑干切片制剂将用于确定神经元功能的发育。免疫组织化学和蛋白质印迹方法将用于鉴定对 NST 的增殖和分化以及整个发育阶段的细胞周期调节剂可能重要的 Sonic Hedgehog (Shh) 信号因子的表达。最后，外植体培养系统将用于分离脑干并在体外操纵位于 rNST 和周围的 Shh 信号分子。可行的假设是，在胚胎发育过程中，神经前体从第四脑室迁移并分化形成最早的rNST，其中神经元在神经胶质细胞之前分化； rNST 神经元在胚胎中具有功能，但特性发生变化；并且，Shh 信号传导调节 rNST 神经前体的增殖和分化时间以及 rNST 中新兴的神经生理功能。这些实验的结果将提供对 rNST 的早期建立、组织和功能的理解，并将证明参与该发育的可能分子途径。这将为理解味觉系统初级传入中继的形成提供重要基础，并为理解胚胎发生过程中发生环境操纵时该中继中发生的可塑性奠定基础。此外，该提案还解决了中枢神经系统神经模式形成过程中的基本问题。公共健康相关性：味觉指导所有脊椎动物的营养选择和摄取行为，由外周受体转导并直接传送到脑干，在脑干中与孤束喙核 (rNST) 中的其他感觉输入整合。了解接受这种感觉输入的孤核内神经元和神经胶质细胞的初始发育将有助于更全面地理解味觉信息如何整合到中枢神经系统中以告知饮食选择，这是维持生命的重要行为。确定 rNST 功能如何以及何时出现并在分子水平上受到调节对于理解味觉系统的可塑性至关重要。由于动物在出生时必须进食，因此 rNST 必须能够指导初始营养摄入。因此，表征控制 rNST 正常发育的因素对于了解母亲饮食、饮酒和药物如何影响 rNST 的神经发育和突触发生非常重要。

DESCRIPTION (provided by applicant): The rostral portion of the nucleus of the solitary tract (rNST) in the brainstem is the first central taste relay. The rNST receives primary afferent projections from facial and glossopharyngeal nerves, which innervate lingual taste buds and papillae and terminate centrally in a topography that reflects peripheral organization. The time course for and molecular factors involved in proliferation and differentiation of rNST cells are not well understood. Given the importance of an organized taste system for proper function, the long-term objective is to determine how the rNST is established in embryonic rat. The proposed research aims are to determine: the development of solitary tract (ST) projections; the developmental time course of proliferation and differentiation of cell components comprising the rNST; the development of biophysical properties and synapses of rNST neurons; specific molecular factors present in relation to rNST development; and, to manipulate the expression of these factors to determine effects on rNST development. Immunofluorescence will be used to identify the time course for development of neurons and glia that compose the rNST in staged embryos. An in vitro brainstem slice preparation will be used to determine development of neuron function. Immunohistochemical and Western blot approaches will be used to identify the expression of Sonic hedgehog (Shh) signaling factors potentially important for proliferation and differentiation in the NST, and cell cycle regulators across developmental stages. Finally, an explant culture system will be used to isolate the brainstem and manipulate in vitro Shh signaling molecules localized to the rNST and surround. The working hypotheses are that during embryonic development, neural precursors migrate from the fourth ventricle and differentiate to form the earliest rNST, where neurons differentiate before glia; that rNST neurons are functional, but with changing properties, in the embryo; and, that Shh signaling regulates the timing of proliferation and differentiation of rNST neural precursors and emerging neurophysiological function in rNST. Results of these experiments will provide an understanding of the early establishment, organization and function of the rNST and will demonstrate possible molecular pathways involved in that development. This will provide an important foundation for understanding the formation of the primary afferent relay of the gustatory system and lay the groundwork for understanding the demonstrated plasticity that occurs in this relay when environmental manipulations take place during embryogenesis. Furthermore, the proposal addresses essential questions in the process of neural patterning in the CNS. PUBLIC HEALTH RELEVANCE: Taste sensation, which guides nutritive choices and ingestive behaviors of all vertebrates, is transduced by peripheral receptors and conveyed directly to the brainstem where it is integrated with other sensory inputs in the rostral nucleus of the solitary tract (rNST). Knowledge of initial development of neurons and glia within the solitary nucleus that receive this sensory input will contribute to a more complete understanding of how taste information is integrated in the central nervous system to inform diet choices, a behavior important to sustain life. Determining how and when function emerges in rNST and is regulated at a molecular level will be essential in understanding plasticity of the taste system. Because animals have to feed at birth, the rNST must be functional to guide initial nutrient intake. It is therefore important to characterize factors controlling normal development of rNST to understand how maternal diet, alcohol consumption and drugs can influence neural development and synaptogenesis of the rNST.