Properties of Ion Channels that Control Secretion

描述(申请人提供)：哺乳动物的肾上腺皮质分为外肾小球和内束，它们分泌不同的皮质类固醇激素，包括矿质皮质激素醛固酮和糖皮质激素。这些激素在调节电解质、水分和能量平衡方面起着至关重要的作用，因此对维持血压和血糖在正常范围内至关重要。皮质醇对血糖的精确控制至关重要，因为这种糖是大脑的主要能量来源。低血糖会迅速导致脑损伤和死亡。皮质类固醇的异常分泌会导致内分泌病理，包括库欣和艾迪生病。 在细胞水平上，肾上腺束状带(AZF)细胞的皮质醇分泌主要受脑垂体肽ACTH的控制，而球状带(AZG)细胞分泌的醛固酮主要受外周血管紧张素II(ALL)的影响。虽然已经鉴定了许多调节皮质类固醇分泌的生理刺激和细胞内信使，但将这些刺激与分泌联系起来的信号通路还不完全清楚。特别是，电活动和特定离子通道在皮质类固醇分泌中的作用尚未阐明。 牛AZF和AZG细胞表达几个离子通道，这些通道决定了它们的电学性质和与分泌有关的离子事件。K通道bTREK-1设定静息电位，被ATP激活，被ACTH和ALL抑制。因此，Trek-1通道在整合激素和代谢信号，并将这些信号耦合到去极化依赖的钙(Ca2+)进入和分泌中起着关键作用。 ACTH和ALL刺激的皮质醇和醛固酮的分泌依赖于钙依赖的动作电位的产生，已经开发了一个特定的模型。具体地说，ACTH和ALL对bTREK-1钾通道的抑制导致由相反的电压门控性钙电流和钾电流驱动的动作电位。这项提案中描述的实验验证了这一假说，并确定了控制肾上腺皮质离子通道功能和表达的信号通路。具体目标是：1)证实All通过钙和三磷酸腺苷依赖的信号通路抑制牛AZF和AZG细胞中的bTREK-1钾通道，并确定其分子机制。目的：1)证明bTREK-1通道设置牛AZF细胞的静息电位，并且ALL对这些通道的抑制与去极化依赖的醛固酮分泌有关；2)证明ACTH通过调节AZF细胞离子通道的功能和表达，对牛AZF细胞的电特性进行短期和长期控制，并研究其中涉及的分子机制；以及3)确定天然bTREK-1通道的ATP敏感性是否允许它们作为传感器发挥作用，其活性随血糖浓度和AZF细胞的代谢状态而变化。

DESCRIPTION (provided by applicant): The adrenal cortex of mammals is divided into an outer glomerulosa and inner fasciculata that secrete different corticosteroid hormones, including the mineralocorticoid aldosterone and the glucocorticoid cortisol. These hormones function critically in regulating electrolyte, water, and energy balance, and are therefore vital for maintaining blood pressure and plasma glucose within normal limits. Precise control of blood glucose by cortisol is essential because this sugar is the brain's primary energy source. Hypoglycemia rapidly leads to brain damage and death. Aberrant secretion of corticosteroids causes endocrine pathology, including Cushing's and Addison's diseases. At the cellular level, cortisol secretion by adrenal zona fasciculata (AZF) cells is controlled primarily by the pituitary peptide ACTH, while zona glomerulosa (AZG) cells secrete aldosterone in response to the peripheral peptide Angiotensin II (All). Although many of the physiological stimuli and intracellular messengers that regulate secretion of corticosteroids have been identified, the signaling pathways that link these stimuli to secretion are incompletely understood. In particular, the role of electrical activity and specific ion channels in the secretion of corticosteroids has not been clarified. Bovine AZF and AZG cells express several ion channels that determine their electrical properties and the ionic events involved in secretion. The K+ channel bTREK-1 sets the resting potential, is activated by ATP and inhibited by ACTH and All. Thus, TREK-1 channels act pivotally in integrating hormonal and metabolic signals and coupling these to depolarization-dependent calcium (Ca2+) entry and secretion. A specific model has been developed for ACTH- and All-stimulated cortisol and aldosterone secretion that depends on the generation of Ca2+-dependent action potentials. Specifically, the inhibition of bTREK-1 K+ channels by ACTH and All leads to action potentials driven by opposing voltage-gated Ca2+ and K+ currents. Experiments described in this proposal test this hypothesis and identify signaling pathways that control function and expression of adrenocortical ion channels. Specific Aims will be: 1) To demonstrate that All inhibits bTREK-1 K+ channels in bovine AZF and AZG cells by separate Ca2+ - and ATP hydrolysis-dependent signaling pathways, and to identify their molecular mechanisms. To demonstrate that bTREK-1 channels set the resting potential of bovine AZG cells and that inhibition of these channels by All is coupled to depolarization-dependent aldosterone secretion; 2) To demonstrate that ACTH exerts short and long term control over the electrical properties of bovine AZF cells by regulating both the function and expression of AZF cell ion channels and to characterize the molecular mechanisms involved; and 3) To determine whether the ATP sensitivity of native bTREK-1 channels allows them to function as sensors whose activity varies with the plasma glucose concentration and the metabolic state of the AZF cell.