cAMP-Mediated Stabilization of Fusion Pores in Cultured Rat Pituitary Lactotrophs
cAMP-Mediated Stabilization of Fusion Pores in Cultured Rat Pituitary Lactotrophs
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DOI：
10.1523/jneurosci.5351-12.2013
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发表时间：
2013-05-01
影响因子：
5.3
通讯作者：
Zorec, Robert
中科院分区：
文献类型：
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作者：
Calejo, Ana Isabel;Jorgacevski, Jernej;Zorec, Robert
受调节的胞吐作用介导激素和递质的释放。该过程的最后一步是囊泡和质膜之间的合并以及融合孔的形成。一旦形成，最初稳定且狭窄的融合孔可能会可逆地变宽（瞬时胞吐作用）或完全打开（完全融合胞吐作用）。胞吐作用通常是由胞质钙活性升高触发的。然而，其他第二信使，例如 cAMP，据报道可以调节分泌。 cAMP 影响不同融合孔状态之间转变的方式仍不清楚。在这里，激素释放研究表明，在毛喉素（一种腺苷酸环化酶激活剂）和膜渗透性 cAMP 类似物 (dbcAMP) 的刺激下，分离的大鼠泌乳素细胞中催乳素的释放表现出双相浓度依赖性。尽管在较低浓度（2-10 μM forskolin 和 2.5-5 mM dbcAMP）下，这些药物刺激催乳素释放，但在较高浓度（50 μM forskolin 和 10-15 mM dbcAMP）下测量到抑制作用。通过使用高分辨率电容 (C-m) 测量，我们记录了 C-m 的离散增加，这代表基本的胞吐事件。 cAMP 的升高使完全融合事件的频率保持不变，同时增加了瞬态事件的频率。通过增加的融合孔电导和延长的融合孔停留时间来测量，它们表现出更宽的融合孔。 dbcAMP 提高了观察到瞬时融合孔有节奏地重新打开的概率。总之，cAMP 介导的宽融合孔的稳定阻止囊泡进入胞吐作用的完全融合阶段，这阻碍了囊泡内容物在高 cAMP 浓度下的排出。
Regulated exocytosis mediates the release of hormones and transmitters. The last step of this process is represented by the merger between the vesicle and the plasma membranes, and the formation of a fusion pore. Once formed, the initially stable and narrow fusion pore may reversibly widen (transient exocytosis) or fully open (full-fusion exocytosis). Exocytosis is typically triggered by an elevation in cytosolic calcium activity. However, other second messengers, such as cAMP, have been reported to modulate secretion. The way in which cAMP influences the transitions between different fusion pore states remains unclear. Here, hormone release studies show that prolactin release from isolated rat lactotrophs stimulated by forskolin, an activator of adenylyl cyclases, and by membrane-permeable cAMP analog (dbcAMP), exhibit a biphasic concentration dependency. Although at lower concentrations (2-10 mu M forskolin and 2.5-5 mM dbcAMP) these agents stimulate prolactin release, an inhibition is measured at higher concentrations (50 mu M forskolin and 10-15 mM dbcAMP). By using high-resolution capacitance (C-m) measurements, we recorded discrete increases in C-m, which represent elementary exocytic events. An elevation of cAMP leaves the frequency of full-fusion events unchanged while increasing the frequency of transient events. These exhibited a wider fusion pore as measured by increased fusion pore conductance and a prolonged fusion pore dwell time. The probability of observing rhythmic reopening of transient fusion pores was elevated by dbcAMP. In conclusion, cAMP-mediated stabilization of wide fusion pores prevents vesicles from proceeding to the full-fusion stage of exocytosis, which hinders vesicle content discharge at high cAMP concentrations.