Baes M, Vale W W
Clayton Foundation Laboratories for Peptide Biology, Salk Institute, La Jolla, California 92037.
Endocrinology. 1989 Jan;124(1):104-10. doi: 10.1210/endo-124-1-104.
The regulation of GRF secretion was studied using a fetal rat hypothalamic cell culture system. The cells were subjected to short term release experiments on days 10-18 after plating, and GRF secretion was assessed by RIA. The identity of GRF immunoreactivity in the incubation medium was confirmed by reverse phase liquid chromatographic analysis. Depolarization of the cells with 56 mM K+ evoked a 4-fold increase in basal GRF release. When cultures were pretreated for 6 days with the adenylate cyclase activator forskolin, basal GRF release was augmented in subsequent release experiments to levels 2-fold greater than those in the control cultures. In nonpretreated cultures, forskolin (1-100 microM) and the protein kinase C activator phorbol 12-myristate 13-acetate (10 nM-1 microM), stimulated basal GRF release in a dose-dependent fashion. The Ca2+ channel blocker verapamil (100 microM) significantly inhibited the GRF response to both forskolin and phorbol 12-myristate 13-acetate. The gamma-aminobutyric acid (GABA) agonist muscimol (0.1-10 microM) inhibited forskolin-stimulated, but not K+ stimulated, GRF release in a dose-dependent manner. This inhibition was reversed by the GABA antagonists bicuculline and picrotoxinin. Muscimol (10 microM) slightly suppressed basal GRF release. The present findings suggest that GRF secretion can be evoked by agents known to increase intracellular cAMP levels or activate protein kinase-C. They also support a role for GABA in the inhibitory control of GRF secretion.
利用胎鼠下丘脑细胞培养系统研究了生长激素释放因子（GRF）分泌的调节机制。细胞在接种后第10 - 18天进行短期释放实验，采用放射免疫分析法（RIA）评估GRF的分泌情况。通过反相液相色谱分析确认了培养液中GRF免疫反应性的特征。用56 mM K⁺使细胞去极化，可使基础GRF释放增加4倍。当培养物用腺苷酸环化酶激活剂福斯可林预处理6天时，在随后的释放实验中基础GRF释放增加，达到比对照培养物高2倍的水平。在未预处理的培养物中，福斯可林（1 - 100 microM）和蛋白激酶C激活剂佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯（10 nM - 1 microM）以剂量依赖方式刺激基础GRF释放。钙通道阻滞剂维拉帕米（100 microM）显著抑制GRF对福斯可林和佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯的反应。γ-氨基丁酸（GABA）激动剂蝇蕈醇（0.1 - 10 microM）以剂量依赖方式抑制福斯可林刺激的GRF释放，但不抑制K⁺刺激的GRF释放。GABA拮抗剂荷包牡丹碱和苦味毒可逆转这种抑制作用。蝇蕈醇（10 microM）略微抑制基础GRF释放。目前的研究结果表明，已知能增加细胞内cAMP水平或激活蛋白激酶C的物质可诱发GRF分泌。这些结果也支持GABA在GRF分泌的抑制性控制中发挥作用。
