INTRACELLULAR SIGNALING IN ENDOCRINE CELLS

GnRH和ET受体与磷脂酶C和D依赖的偶联 在垂体细胞中检测细胞内信号。促性腺激素释放激素诱导两者 最初和持续的IP3/DAG产生，而ET-1仅诱导 瞬时反应，可与GnRH诱导的DAG相媲美 蛋白激酶C耗竭细胞的反应。GnRH和ET诱导 磷脂酶C抑制剂可降低IP3/DAG和钙离子反应， U73122和新霉素。这两种磷脂酶抑制剂也降低了 激动剂诱导培养的垂体细胞释放黄体生成素。与之形成鲜明对比的是 激动剂对磷脂酶C、D活性的刺激作用 激活GnRH受体，但不激活ET受体。此外，GnRH-而不是ET- 在两者存在下，诱导的DAG形成减少 磷脂酶D抑制剂、乙醇和心得安与DAG的下降 GnRH诱导的c-fos表达受抑制 表情。我们还研究了它们之间的相互作用 受体介导的细胞内信号与质膜电 事件。结果表明，脑垂体细胞是可兴奋的，而且 每一个钙尖峰都是由钙进入在一次 动作电位(AP)。心动过速和钙振荡的频率 促性腺激素细胞受到去极化/超极化脉冲的调制。 使用这些和其他关于质膜通道特性的数据 在前面提供的促性腺激素细胞中，一个定量的数学模型 发展了对时空钙振荡的预测。模型 包含一组类似Hodgkin-Huxley的方程和 钙离子向细胞中心扩散。这表明AP诱导的 钙尖峰只在邻近的薄壳层中突出。 细胞表面。我们还发现，在促性腺激素释放激素刺激的细胞中， 在持续阶段期间的去极化脉冲可以引发瞬变 钙的上升与内源循环相似。此外，钙离子进入 在单个去偏振脉冲被发现移动的相位 随后的内源性钙振荡，而不影响其 频率两个连续的去偏振脉冲的应用显示 第二次脉冲引起的钙升高的大小取决于 两个连续脉冲之间经过的时间，表示每个脉冲 内源性或诱发的钙升高循环使钙释放 处于难治状态的促性腺激素的机制。电活动 可能在磷脂酶C诱导的调控中起重要作用 钙振荡，包括重置振荡器的时钟。

Coupling of GnRH and ET receptors with phospholipase C- and D-dependent intracellular signals was examined in pituitary cells. GnRH induced both initial and sustained IP3/DAG production, while ET-1 induced only a transient response, a profile that was comparable to the GnRH-induced DAG response in protein kinase C-depleted cells. GnRH- and ET-induced IP3/DAG and calcium responses were reduced by phospholipase C inhibitors, U73122 and neomycin. Those two phospholipase inhibitors also reduced agonist-induced LH release by cultured pituitary cells. In contrast to phospholipase C, phospholipase D activity was stimulated by agonist activation of GnRH but not ET receptors. Furthermore, GnRH- but not ET- induced DAG formation was reduced in the presence of the two phospholipase D inhibitors, ethanol and propanolol, and the fall in DAG production was accompanied by the inhibition of GnRH-induced c-fos expression. We have also examined the interactions between these receptor-mediated intracellular signals and plasma membrane electrical events. The results showed that pituitary cells are excitable and that each calcium spike is produced by the calcium entry during a single action potential (AP). The frequency of APs and calcium oscillations in gonadotrophs was modulated by depolarization/hyperpolarization pulses. Using these and other data on plasma membrane channel characterization in gonadotrophs provided earlier, a quantitative mathematical model predicting spatio-temporal calcium oscillations was developed. The model contains a set of Hodgkin-Huxley-like equations and the equation for calcium diffusion towards the cell center. It suggests that AP-induced calcium spiking is prominent only in a thin shell layer neighboring the cell surface. We also found that in GnRH-stimulated cells a brief depolarization pulse during the sustained phase can elicit a transient calcium rise similar to the endogenous cycle. In addition, calcium entry during a single depolarizing pulse was found to shift the phase of subsequent endogenous calcium oscillations, without affecting their frequency. The application of two consecutive depolarizing pulses showed that the size of the calcium rise evoked by the second pulse depended on the time lapsed between two consecutive pulses, indicating that each endogenous or evoked calcium rise cycle leaves the calcium release mechanism of the gonadotroph in a refractory state. Electrical activity may play an important role in the modulation of phospholipase C-induced calcium oscillations, including resetting the clock of the oscillator.