Yu K L, Rosenblum P M, Peter R E
Department of Zoology, University of Alberta, Edmonton, Canada.
Gen Comp Endocrinol. 1991 Feb;81(2):256-67. doi: 10.1016/0016-6480(91)90010-4.
In vitro release of gonadotropin releasing hormone (GnRH) from slices of the preoptic-anterior hypothalamic (P-AH) region and fragments of the pituitary of goldfish was studied using a static incubation system. Release of GnRH from both tissue preparations was stimulated by depolarizing concentrations of extracellular potassium ions (K+). Other putative secretagogues, calcium ionophore A23187 (1 microM), forskolin (100 microM), and prostaglandin E2 1 microM) also stimulated release of GnRH from both tissue preparations. Omission of Ca2+, or chelating the remaining remaining Ca2+ by EGTA (0.1 mM), abolished the release of GnRH stimulated by high K+ concentrations (60 mM), but did not reduce spontaneous release. Verapamil (1 microM), a voltage-sensitive calcium channel blocker, abolished the release of GnRH stimulated by high K+ or A21387 from both tissue preparations. The GnRH released in vitro from both the P-AH region and pituitary was concentrated by Sep-Pak and then separated by high-performance liquid chromatography. The major peak of the GnRH immunoreactivity was found to coelute with synthetic salmon GnRH [( Trp7,Leu8]-GnRH) and the minor peak with chicken GnRH-II [( Gln8]-GnRH). Dopamine (10 and 100 microM) inhibited GnRH release from both P-AH slices and pituitary fragments, while serotonin (1-100 microM) stimulated release from both. Norepinephrine (10-100 microM) stimulated GnRH release from P-AH slices but not from pituitary fragments. The results demonstrate that the release of GnRH from goldfish P-AH slices and pituitary fragments in vitro in response to various secretagogues and monoamines can be studied using a static incubation system.
利用静态孵育系统研究了金鱼视前区-下丘脑前部（P-AH）区域切片和垂体碎片中促性腺激素释放激素（GnRH）的体外释放情况。两种组织制剂中GnRH的释放均受到细胞外钾离子（K⁺）去极化浓度的刺激。其他假定的促分泌素，钙离子载体A23187（1微摩尔）、福斯高林（100微摩尔）和前列腺素E2（1微摩尔）也刺激了两种组织制剂中GnRH的释放。去除Ca²⁺，或用乙二醇双（2-氨基乙基醚）四乙酸（EGTA，0.1毫摩尔）螯合剩余的Ca²⁺，可消除高K⁺浓度（60毫摩尔）刺激的GnRH释放，但不降低自发释放。维拉帕米（1微摩尔），一种电压敏感的钙通道阻滞剂，可消除两种组织制剂中高K⁺或A21387刺激的GnRH释放。从P-AH区域和垂体体外释放的GnRH通过Sep-Pak柱浓缩，然后通过高效液相色谱分离。发现GnRH免疫反应性的主要峰与合成鲑鱼GnRH [（Trp7，Leu8）-GnRH]共洗脱，次要峰与鸡GnRH-II [（Gln8）-GnRH]共洗脱。多巴胺（10和100微摩尔）抑制P-AH切片和垂体碎片中GnRH的释放，而5-羟色胺（1-100微摩尔）刺激两者释放。去甲肾上腺素（10-100微摩尔）刺激P-AH切片中GnRH的释放，但不刺激垂体碎片中GnRH的释放。结果表明，利用静态孵育系统可以研究金鱼P-AH切片和垂体碎片中GnRH对各种促分泌素和单胺的体外释放情况。
