Garrido-Gracia J C, Bellido C, Aguilar R, Sánchez-Criado J E
Department of Cell Biology, Physiology and Immunology, University of Córdoba, Avda. Menéndez Pidal s/n, 14004 Córdoba, Spain.
J Physiol Biochem. 2006 Mar;62(1):35-42. doi: 10.1007/BF03165804.
In the absence of progesterone (P), the anti-P at the receptor RU486 reduces basal and GnRH-stimulated LH secretion both in vivo and in vitro, demonstrating the existence of a ligand-independent activation of progesterone receptor (LIAPR). The aim of the present study was to determine which component of the intracellular LH secretory pathway activated by GnRH is responsible for LIAPR. To do this, anterior pituitary dispersed cells from female rats in proestrus, cultured in the presence of 17beta-estradiol, were incubated with activators or inhibitors of PKC, cAMP-PKA signalling pathways or intracellular calcium (Ca2+) traffic, in the presence or absence of RU486. Results showed that RU486 reduced both GnRH- and the PKC activator PMA-induced LH secretion. In GnRH-stimulated cells incubated with the PKC inhibitor BIS-I or treated with PMA "overnight", RU486 had no effect on reduced LH secretion, nor on stimulated LH secretion elicited by the Ca2+ ionophore ionomycin. Moreover, when GnRH- or PMA-treated cells were co-incubated with 1 microM of the L-type Ca2+ channel blocker nifedipine or the intracellular Ca2+ chelator BAPTA-AM, RU486 potentiated the expected inhibition of these drugs on LH secretion. Activation (forskolin, 8-Br-cAMP) or inhibition (MDL-12,330A) of the cAMP-PKA signalling cascade affected neither the GnRH- and PMA-induced increase of LH secretion nor the reduction of LH secretion due to RU486. Taken together, the data point to the existence of a Ca2+ -independent PKC-PR cross-talk mechanism as part of the intracellular signalling of GnRH-stimulated LH secretion.
在缺乏孕酮（P）的情况下，受体拮抗剂RU486在体内和体外均能降低基础状态下以及GnRH刺激后的促黄体生成素（LH）分泌，这表明存在一种不依赖配体的孕酮受体激活（LIAPR）现象。本研究的目的是确定GnRH激活的细胞内LH分泌途径中的哪个成分负责LIAPR。为此，将处于动情前期的雌性大鼠的垂体前叶分散细胞在17β-雌二醇存在的情况下进行培养，然后在有或无RU486的情况下，与蛋白激酶C（PKC）、环磷酸腺苷-蛋白激酶A（cAMP-PKA）信号通路或细胞内钙（Ca2+）转运的激活剂或抑制剂一起孵育。结果显示，RU486降低了GnRH和PKC激活剂佛波酯（PMA）诱导的LH分泌。在用PKC抑制剂双吲哚马来酰胺-I（BIS-I）孵育或“过夜”用PMA处理的GnRH刺激细胞中，RU486对降低的LH分泌没有影响，对钙离子载体离子霉素刺激的LH分泌也没有影响。此外，当用GnRH或PMA处理的细胞与1微摩尔的L型钙通道阻滞剂硝苯地平或细胞内钙螯合剂1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸-乙酰甲酯（BAPTA-AM）共同孵育时，RU486增强了这些药物对LH分泌的预期抑制作用。cAMP-PKA信号级联的激活（福斯高林、8-溴环磷酸腺苷）或抑制（MDL-12,330A）既不影响GnRH和PMA诱导的LH分泌增加，也不影响由于RU486导致的LH分泌减少。综上所述，数据表明存在一种不依赖Ca2+的PKC-孕酮受体相互作用机制，作为GnRH刺激LH分泌的细胞内信号传导的一部分。
