Development of the Gustatory Solitary Nucleus

描述(申请人提供)：脑干中孤束核(RNST)的吻侧部分是第一个中央味觉传递。RNST从面神经和舌咽神经接受初级传入投射，这些神经支配舌味蕾和舌乳头，并在反映周围组织的地形图中央终止。RNST细胞增殖和分化的时间进程和分子因素尚不清楚。鉴于有组织的味觉系统对正常功能的重要性，长期目标是确定RNST是如何在胚胎大鼠中建立的。拟议的研究目的是确定：孤束投射的发展；组成RNST的细胞成分的增殖和分化的发育时间进程；RNST神经元的生物物理特性和突触的发展；与RNST发育有关的特定分子因素；以及，操纵这些因素的表达，以确定对RNST发育的影响。免疫荧光将被用来确定组成RNST的神经元和神经胶质细胞在分期胚胎中的发育过程。一种体外脑干切片制备将用于确定神经元功能的发展。免疫组织化学和Western印迹方法将被用来确定对NST的增殖和分化具有潜在重要性的Sonic hedgehog(Shh)信号因子的表达，以及不同发育阶段的细胞周期调节因子。最后，将使用外植体培养系统分离脑干，并在体外操纵定位于RNST及其周围的Shh信号分子。工作假设是，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，其中神经元在胶质细胞之前分化；RNST神经元在胚胎中具有功能，但具有变化的性质；Shh信号调控RNST神经前体的增殖和分化以及RNST中新出现的神经生理功能。这些实验的结果将提供对RNST早期建立、组织和功能的理解，并将展示参与这一发展的可能的分子途径。这将为理解味觉系统初级传入中继器的形成提供重要的基础，并为理解胚胎发生过程中环境操纵时在该中继器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。与公共健康相关：味觉指导所有脊椎动物的营养选择和摄食行为，由外周受体传递并直接传送到脑干，在那里它与孤束嘴核(RNST)的其他感觉输入整合在一起。了解接受这种感觉输入的孤束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，从而为饮食选择提供信息，这是维持生命的重要行为。确定RNST中功能如何以及何时出现并在分子水平上受到调控，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，RNST必须具有指导初始营养摄入的功能。因此，确定控制RNST正常发育的因素，以了解母亲的饮食、饮酒和药物如何影响RNST的神经发育和突触发生是重要的。

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.