IONIC MECHANISMS RELATED TO SECRETION IN PITUITARY CELLS

细胞膜电活动与胞吐的功能耦合 从内分泌细胞释放激素是 内分泌腺分泌生理学。 在垂体的情况下， 激素分泌、参与的膜离子通道和分子 将刺激性和抑制性促分泌素受体与 这些离子通道现在才被发现。 这个任务很复杂 由于存在分泌一种以上激素的细胞亚型 以及多个分泌调节因子之间的相互作用。 许多 关键的促分泌素受体属于七个超家族 跨膜结构域受体，其抑制配体结合进入 通过鸟嘌呤核苷酸结合蛋白（G蛋白）的细胞作用。 在这些受体中，有多巴胺受体（D2型）， 催乳素分泌的调节剂，以及控制 抑制生长激素释放。 这一目标的主要目的是 本研究采用膜片钳电生理方法， 离子通道功能的变化是这些作用的基础， 和其他激动剂对培养的单个垂体细胞的作用，以及 阐明耦合机制，共同的和独特的，调节 分泌亚型中的兴奋-分泌偶联。 正常大鼠前额叶 垂体细胞将使用反向溶血斑进行鉴定 测定（RHPA）和新开发的方法，顺序细胞免疫印迹 明确区分细胞类型和定量的SCIBA分析 单细胞水平分泌的变化。 此外，我们将 检查正常和突变多巴胺受体的细胞系， 与信号转导途径的其他元件重组。 关键 要解决的问题是：（1）G蛋白偶联D2的身份 离子通道的受体，（2）参与这种偶联的机制， (3)受体偶联特异性的结构决定因素， G蛋白;（4）D_2受体相对免疫的性质， 脱敏，（5）离子通道和离子通道的确切作用 浓度变化调节激素分泌。 这 研究将使我们更清楚地了解 调节垂体细胞的激素分泌。 此外，共同 垂体和脑多巴胺受体亚型同一性 表明这些研究可能提供了机械的见解， 这些神经递质在情感障碍和神经系统疾病中的作用 调节植物的功能。

The functional coupling between membrane electrical events and exocytotic release of hormones from endocrine cells is a key element in the physiology of secretion in endocrine glands. In the case of pituitary hormone secretion the membrane ion channels involved and the molecular mechanisms coupling stimulatory and inhibitory secretogogue receptors to these ion channels are only now being revealed. This task is complicated due to the presence of cellular subtypes secreting more than one hormone and interactions between more than one regulator of secretion. Many of the key secretagogue receptors belong to the superfamily of seven membrane spanning domain receptors that transduce ligand binding into cellular action via guanine nucleotide binding proteins (G-proteins). Among such receptors are those for dopamine (D2 type), the primary regulator of prolactin secretion, and somatostatin which governs inhibition of growth hormone release. The principle objective of this study is to employ patch clamp electrophysiological methods to examine the changes in ion channel function which underlie the actions of these and other agonists on individual pituitary cells in culture, and to elucidate the coupling mechanisms, both common and unique, which regulate excitation-secretion coupling in secretory subtypes. Normal rat anterior pituitary cells will be identified using the reverse hemolytic plaque assay (RHPA) and a newly developed method, the sequential cell immunoblot assay (SCIBA) to unambiguously discriminate cell types and quantify changes in secretion at the single cell level. In addition we will examine cell lines in which normal and mutated dopamine receptors are recombined with other elements of the signal transduction pathway. Key issues to be addressed are (1) the identity of G-proteins coupling D2 receptors to ion channels, (2) the mechanisms involved in this coupling, (3) the structural determinants of coupling specificity of receptors and G-proteins, (4) the nature of the relative immunity of D2 receptors to desensitization, and (5) the precise role of ion channel and ion concentration changes in the regulation of hormone secretion. This research will lead to a clearer understanding of the events underlying regulated hormone secretion in pituitary cells. In addition, the common identity of dopamine receptor subtypes in both pituitary and brain suggest that these studies may provide mechanistic insight into the actions of these neurotransmitters in affective disorders and neural regulation of vegetative functions.