Srivastava R K, Van der Kraak G
Department of Zoology, University of Guelph, Ontario, Canada.
Gen Comp Endocrinol. 1994 Feb;93(2):181-91. doi: 10.1006/gcen.1994.1021.
For the present study, calcium ionophore A23187 and phorbol 12-myristate 13-acetate (PMA) were used to investigate the possible roles that changes in intracellular calcium content and protein kinase C activation play in steroid production by goldfish vitellogenic ovarian follicles (0.2-0.7 mm in diameter) incubated in vitro. While largely ineffective when tested alone, PMA and A23187 exhibit synergism leading to increased testosterone and 17 beta-estradiol production. Production of both steroids was also increased by the addition of either human chorionic gonadotropin (hCG) or forskolin, a direct activator of adenylate cyclase. However, hCG- and forskolin-stimulated testosterone and 17 beta-estradiol production was attenuated by PMA and A23187 either alone or in combination. Both drugs blocked the stimulatory actions of hCG on adenosine 3',5'-cyclic monophosphate (cAMP) formation. PMA also influenced steroid production at sites distal to cAMP formation as it blocked the actions of dibutyryl cAMP a permeant analog of cAMP. This action was prior to cholesterol side-chain cleavage as PMA had no effect on the metabolism of 25-hydroxycholesterol to testosterone or 17 beta-estradiol. By comparison, A23187 had no effects on either dbcAMP- or 25-hydroxycholesterol-stimulated steroid production. Separation of vitellogenic and prematurational, full-grown (0.9-1.1 mm in diameter) ovarian follicles from the same fish lead to the demonstration of opposing actions of A23187 in the two follicle types. Whereas A23187 inhibited hCG-stimulated steroid production in vitellogenic follicles, it had an additive effect on the stimulatory action of hCG on testosterone production by prematurational, full-grown ovarian follicles. In conclusion, these studies on vitellogenic ovarian follicles demonstrate specific regulatory actions of calcium and protein kinase C at multiple steps in the steroidogenic cascade and that ovarian responsiveness to calcium changes during the course of follicle development in the goldfish.
在本研究中，使用钙离子载体A23187和佛波醇12 - 肉豆蔻酸酯13 - 乙酸酯（PMA）来研究细胞内钙含量变化和蛋白激酶C激活在体外培养的金鱼卵黄生成期卵巢卵泡（直径0.2 - 0.7毫米）类固醇生成中可能发挥的作用。单独测试时，PMA和A23187大多无效，但二者表现出协同作用，导致睾酮和17β - 雌二醇生成增加。添加人绒毛膜促性腺激素（hCG）或腺苷酸环化酶的直接激活剂福斯高林也会增加这两种类固醇的生成。然而，单独或联合使用PMA和A23187会减弱hCG和福斯高林刺激的睾酮和17β - 雌二醇生成。两种药物均阻断了hCG对3',5'-环磷酸腺苷（cAMP）形成的刺激作用。PMA还影响cAMP形成位点远端的类固醇生成，因为它阻断了二丁酰cAMP（一种cAMP的通透类似物）的作用。此作用发生在胆固醇侧链裂解之前，因为PMA对25 - 羟基胆固醇向睾酮或17β - 雌二醇的代谢没有影响。相比之下，A23187对dbcAMP或25 - 羟基胆固醇刺激的类固醇生成均无影响。从同一条鱼中分离出卵黄生成期和早熟的、完全成熟的（直径0.9 - 1.1毫米）卵巢卵泡，结果表明A23187在两种卵泡类型中具有相反的作用。在卵黄生成期卵泡中，A23187抑制hCG刺激的类固醇生成，而在早熟的、完全成熟的卵巢卵泡中，它对hCG刺激的睾酮生成具有相加作用。总之，这些关于卵黄生成期卵巢卵泡的研究表明，钙和蛋白激酶C在类固醇生成级联反应的多个步骤中具有特定的调节作用，并且在金鱼卵泡发育过程中卵巢对钙变化具有反应性。
