Chromaffin cells of the adrenal medulla have an important role in the sympathetic stress response. They secrete catecholamines and other hormones into the bloodstream upon stimulation by the neurotransmitter pituitary adenylate cyclase-activating polypeptide (PACAP). PACAP causes a long-lasting and robust secretory response from chromaffin cells. However, the cellular mechanisms by which PACAP causes secretion remain unclear. Our previous work showed that the secretory response to PACAP relies on signaling through phospholipase C epsilon (PLCε). The objective of this study was to clarify the role of signaling events downstream of PLCε. Here, it is demonstrated that a brief exposure of chromaffin cells to PACAP caused diacylglycerol (DAG) production—a process that was dependent on PLCε activity. DAG then activated protein kinase C (PKC), prompting its redistribution to the plasma membrane. PKC activation was important for the increases in cytosolic Ca2+ and exocytosis that were evoked by PACAP. Indeed, pharmacological inhibition of PKC with NPC 15437, a competitive inhibitor of DAG binding, significantly disrupted the secretory response. NPC 15437 application also eliminated PACAP-stimulated effects on the readily releasable pool size, the Ca2+ sensitivity of granule fusion, and the voltage dependence of Ca2+ channel activation. Quantitative PCR revealed PKCβ, PKCε, and PKCμ to be highly expressed in the mouse chromaffin cell. Genetic knockdown of PKCβ and PKCε disrupted PACAP-evoked secretion, while knockdown of PKCμ had no measurable effect. This study highlights important roles for PKC signaling in a highly regulated pathway for exocytosis that is stimulated by PACAP.

中文翻译：

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PKC 信号传导在嗜铬细胞胞吐作用中的作用。


肾上腺髓质的嗜铬细胞在交感神经应激反应中起重要作用。它们在神经递质垂体腺苷酸环化酶激活多肽 （PACAP） 的刺激下将儿茶酚胺和其他激素分泌到血液中。PACAP 引起嗜铬细胞持久而强大的分泌反应。然而，PACAP 引起分泌的细胞机制仍不清楚。我们之前的工作表明，对 PACAP 的分泌反应依赖于通过磷脂酶 C ε （PLCε） 的信号传导。本研究的目的是阐明 PLCε 下游信号转导事件的作用。在这里，证明嗜铬细胞短暂暴露于 PACAP 会导致甘油二酯 （DAG） 的产生 - 这一过程依赖于 PLCε 活性。然后 DAG 激活蛋白激酶 C （PKC），促使其重新分布到质膜。PKC 激活对于 PACAP 诱发的胞质 Ca2+ 和胞吐作用的增加很重要。事实上，NPC 15437（一种 DAG 结合的竞争性抑制剂）对 PKC 的药理学抑制显着破坏了分泌反应。NPC 15437 的应用还消除了 PACAP 刺激对易于释放的池大小、颗粒融合的 Ca2+ 敏感性以及 Ca2+ 通道激活的电压依赖性的影响。定量 PCR 显示 PKCβ 、 PKCε 和 PKCμ 在小鼠嗜铬细胞中高表达。PKCβ 和 PKCε 的基因敲低破坏了 PACAP 诱发的分泌，而 PKCμ 的敲低没有可测量的效果。本研究强调了 PKC 信号转导在 PACAP 刺激的高度调节的胞吐作用途径中的重要作用。