LHRH PULSATILE RELEASE IN CULTURED LHRH NEURONS FROM EMBRYONIC OLFACTORY PLACODE

胚胎嗅觉板培养的 LHRH 神经元中的 LHRH 脉冲释放

• 批准号: 6116442
• 负责人: Ei Terasawa-Grilley
• 金额: $ 5.34万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6116442
• 关键词: animal tissue; bioimaging /biomedical imaging; clinical research; endocrine gland /system; growth /development; microscopy; mother /infant health care; nervous system; women's health

OBJECTIVE To understand the mechanism of LHRH pulse generation, LHRH release pattern and the role of calcium in neurosecretion will be investigated in cultured LHRH neurons. Previously we have described primary LHRH cell cultures derived from the embryonic olfactory placode. The first study showed that cultured LHRH neurons release the decapeptide into media. It was found that LHRH cells release LHRH in a pulsatile manner at approximately 50-minute intervals. Further, LHRH release was stimulated by depolarization with high K+ and the Na+ channel opener, veratridine. However, while the Na+ channel blocker, tetrodotoxin (TTX) suppressed the effects of veratridine, TTX did not alter the effects of high K+. The second study examined the role of extracellular and intracellular Ca2+ in LHRH release. The results are summarized as follows 1) Exposing the cells to a low Ca2+ (20 nM) buffer solution suppressed LHRH release, while exposure to a normal Ca2+ solution (1.25 mM) maintained pulsatile LHRH release; 2) LHRH release from cultured LHRH cells was stimulated by the voltage sensitive L-type Ca2+ channel agonist, Bay K 8644 (10 5M), while it was suppressed by th e L-type Ca2+ channel blocker, nifedipine (1 5M), but not by the N-type channel blocker, (-conotoxin GVIA (1 5M); 3) The intracellular Ca2+ stimulant, ryanodine (1 5M) stimulated LHRH release, while the intracellular Ca2+ transporting ATPase antagonist, thapsigargin (1 and 10 5M), did not yield consistent results; 4) Carbonyl cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP, 1 (M), a mitochondrial Ca2+ mobilizer, stimulated LHRH release, while ruthenium red, a mitochondrial Ca2+ uptake inhibitor, did not induce consistent results. These results indicate that 1) the presence of extracellular Ca2+ is essential for LHRH neurosecretion, 2) Ca2+ enters the cell via L-type channels, but not N-type channels, and 3) mobilization of intracellular Ca2+ from inositol 1,4,5-triphosphate (IP3)-sensitive stores as well as mitochondrial stores, appears to contribute to LHRH release in these cells. FUTURE DIRECTIONS We will examine if LHRH release is correlated with intracellular calcium oscillations. KEY WORDS LHRH neurons, pulsatility, monkey embryos, LHRH release in vitro, intracellular Ca2+ FUNDING NIH HD15433 & RR00167 PUBLICATIONS Terasawa, E. 1998. Cellular mechanism of pulsatile LHRH release. Gen. Comp. Endocrinol. 112 283-295. [J]

目的了解促黄体生成素(LHRH)脉冲产生机制。 LHRH的释放模式和钙在神经分泌中的作用将是 在培养的LHRH神经元上进行了研究。之前我们已经描述过 胚胎嗅觉来源的原代LHRH细胞培养 普洛伊德。第一项研究表明，培养的LHRH神经元释放 将十肽带入媒体。研究发现，LHRH细胞释放LHRH 以脉动的方式以大约50分钟的间隔。此外， 高K+和Na+去极化刺激LHRH释放 通道开通剂，藜芦碱。然而，虽然钠离子通道阻滞剂， 河豚毒素(TTX)可抑制藜芦碱的作用，但不能 改变高K+的影响。第二项研究考察了 LHRH释放中的细胞外和细胞内钙离子。结果是 总结如下：1)细胞暴露在低钙(20 NM)环境中 缓冲液抑制LHRH的释放，而暴露于正常的 钙离子溶液(1.25 mM)维持LHRH的脉动释放； 电压刺激培养的LHRH细胞释放 敏感的L型钙通道激动剂，Bay K8644(10.5M)，而它 被L类钙通道阻滞剂硝苯地平(1.5M)抑制， 但不是由N型通道阻滞剂，(-芋螺毒素GVIA(1 5M)；3) 细胞内钙离子刺激剂兰尼定(1.5 M)刺激LHRH 释放，而细胞内钙转运ATPase拮抗剂， Thapsigarin(1和10 5M)，结果不一致；4) 对三氟甲氧基苯肼(FCCP)，1(M)，a 线粒体钙激动剂，刺激LHRH释放，而Ru 线粒体钙摄取抑制剂RED不能诱导 结果。这些结果表明：1)细胞外的存在 钙离子是促性腺激素释放激素神经分泌所必需的，2)钙离子通过 L经络，而不是N型经络；3)动员 1，4，5-三磷酸肌醇(IP3)敏感的细胞内钙离子 储存和线粒体储存似乎对LHRH有贡献 在这些细胞中释放。未来方向我们将研究LHRH是否 释放与细胞内钙振荡有关。钥匙 LHRH神经元，搏动性，猴胚胎，LHRH释放 NIH HD15433和RR00167出版物的体外、细胞内钙离子资助 Terasawa，E.1998。LHRH脉动性释放的细胞机制。 上将薪酬内分泌激素。112 283-295。[J]