鯉魚腦下腺“促性腺激素”釋放研究

Abstract

摘要： 本實驗之目的為1．利用離體（in vitro）鯉魚腦下腺灌流培養系統探討正腎上腺素（norepinephrine, NE)。促性腺激素釋放激素（gonadotropin-releasing hormone, GnRH）及多巴胺（dopamine, DA）對腦下腺之直接作用方式為刺激或抑制，且更進一步探討adrenergicα及β二型受體與NE對鯉魚“促性腺激素”(carp gonadotropin, cGtH）釋放作用影響之相關性。2．利用高敏感度之“液相層析-電化學偵檢法”(LC-EC)觀察鯉魚腦下腺中NE及DA之含量，藉以瞭解NE及DA在鯉魚活體內是否具有生理意義。 在動態離體腦下腺培養系統中，1 uM至100 uM之NE可直接刺激鯉魚腦下腺釋放cGtH，其刺激效果在性腺處於發育初期及發育中期之鯉魚腦下腺較顯著，而在性腺處退化狀態之鯉魚腦下腺其效果則不佳，即NE之作用會受到季節變化及性腺成熟度之影響。Adrenergic α及β型受體在cGtH釋放上之作用乃借α型拮抗藥物”prazosin”及β型拮抗藥物”propranolol”分別探討之。以NE與拮抗藥物共同處理鯉魚腦下腺時，100 uM之prazosin只稍微影響NE對cGtH釋放之刺激效界，propranolol則會完全抑制NE之刺激效果，即β型受體可能參與NE在鯉魚腦下腺刺激cGtH分泌之作用。 本實驗亦分別利用人工合成之“鮭魚促性腺激素釋放激素”(saloon gonadotropin-releasing hormone, sGnRH）及“促性腺激素釋放激素類似物”(gonadotropin-releasing hormone analogue, GnRH-A)探討GnRH對cGtH釋放之調節作用。結果sGnRH 及GnRH-A均可刺激cGtH之分泌但其刺激效果不穩定。 DA對cGtH釋放之抑制性作用，乃藉DA拮抗藥物“haloperidol”對鯉魚腦下腺之影響而得。10uM之haloperidol可使cGtH基礎釋放量提高；利用10uM之haloperidol與luM GnRH共同處理鯉魚腦下腺時，haloperidol可提高sGnRH及 GnRH-A對cGtH釋放之刺激效果。 以高效液相層析-電化學偵檢法於不同月份偵測鯉魚腦下腺中NE及DA之含量發現：NE在每個鯉魚腦下腺之總含量最低為315pg，最高為2032pg; DA之總含量則最低為248pg，最高為3000Pg。在不同月份中均可測到NE及DA，其量由生殖季前至生殖季後有逐漸上升之趨勢。由NE及DA持續。明顯存在之現象顯示：NE及DA在鯉魚生殖上應具有重要之生理意義。 綜合本實驗利用不同激素或藥物，處理離體鯉魚腦下腺之效用，及以高效液相層析-電化學偵檢法偵測鯉魚腦下腺內NE及DA含量之結果顯示：(1)NE極可能經由β型受體而直接刺激鯉魚腦下腺分泌cGtH，且NE在鯉魚生殖機制上具有生理意義。(2)GnRH對cGtH之釋放具有刺激效果；(3)DA會持續抑制鯉魚腦下腺自發性及GnRH引發之cGtH釋放。(4）鯉魚本身所具之GnRH與sGnRH，二者之結構可能有極大的差異。

Abstract: Release of radioimmunoassayable common carp gonadotropin(cGtH) from freshly prepared common carp pituitary fragments was measured using a column superfusion system to investigate the effects of norepinephrine(NE), dopamine(DA), salmon gonadotropin-releasing hormone(sGnRH) and gonadotropin-releasing hormone analogue(GnRH-A) on the regulation of cGtH secretion. Contents of NE and DA in the pituitary and hypothalamus were also measured using Liquid chromatography-Electrochemical detector. Treatment of pituitary fragments from carp with GSI less than 25% with 1 uM, 10 uM and 100 UN NE increased the amount of cGtH released into the perifusate, and the effects of NE were blocked by a specificβantagonist, propranolol. But on regressing gonads NE could not elicit significant effects. Perifusion with 10 uM DA did not significantly decreased the amount of spontaneous cGtH release but treatment with a DA antagonist, haloperidol, potentiated the cGtH spontaneous secretion and GnRH-induced cGtH release. Treatment of pituitary fragments with sGnRH or GnRH-A alone did not consistently stimulate cGtH secretion from the carp pituitary. Detectable amounts of NE and DA within the pituitary could be measured from 1989 November to 1990 April and the contents of NE and DA both increased progressively during this period. Those contents of NE were between 315 pg to 2032 pg per pituitary and those contents of DA were between 248 pg to 3000 pg per pituitary. These results provided in vitro evidence for direct NE stimulation of cGtH release by the adrenergicβreceptor and the extent of NE effect was affected by seasonal factor. In addition, the existence of detectable amounts of NE within carp pituitary strongly suggested a possible physiological role of NE in the regulation of carp reproduction. The present study showed that administration of haloperidol, a dopamine D2 antagonist, potentiated cGtH release from the carp pituitary. In addition, tissue content of DA in pituitary increased instead of decreasing when entering the breeding season. These results suggests that DA is continuously present and tonically exerts its inhibitory effect on cGtH secretion.