Liu T C, Jackson G L
Endocrinology. 1986 Jul;119(1):236-43. doi: 10.1210/endo-119-1-236.
We determined the role of microfilaments in regulating LH synthesis (translation or glycosylation) and release from cultured rat anterior pituitary cells under basal and GnRH-stimulated conditions. Cells were pretreated for 2 h with microfilament-disrupting drugs, cytochalasin B (CB; 2 and 20 microM) or cytochalasin D (CD; 1 and 10 microM). LH synthesis and release were measured after 4 h of incubation with or without 1 nM GnRH and drugs. LH translation and glycosylation were monitored by measuring the incorporation of [14C]alanine and [3H]glucosamine, respectively, into total (cell and medium) immunoprecipitable LH. Immunoreactive LH (IRLH) in medium and cells was measured by RIA. GnRH at 1 nM significantly (P less than 0.01) increased the release of IRLH and total [3H]LH (glycosylation), but had no effect on total [14C]LH (translation), uptake, or incorporation of precursors into total protein. Neither CB (2 and 20 microM) nor CD (10 microM) altered basal or GnRH-stimulated IRHL release. Neither drug altered basal medium concentrations of [3H]LH or [14C]LH. In contrast, both CB and CD reduced (P less than 0.01) GnRH-stimulated [3H]LH in the medium and total system (LH glycosylation). CB reduced (P less than 0.01) [3H]glucosamine uptake, total [3H]protein synthesis, and basal level of total [3H]LH, while CD had no effects on these parameters. Thus, CD exerted a more specific inhibitory effect on GnRH-stimulated LH glycosylation than CB. CB (2 and 20 microM) increased (P less than 0.01), while CD (10 microM) decreased (P less than 0.01) [14C]alanine uptake, total [14C]LH, and [14C]protein under both basal and GnRH-stimulated conditions. These results demonstrated that while the cytochalasins did not inhibit either basal or GnRH-stimulated IRLH release, they did inhibit GnRH-stimulated LH glycosylation, although the effect of CB was due partially to reduced [3H]glucosamine uptake. Integrity of microfilaments appears to be important for GnRH-enhanced LH glycosylation, but not for GnRH-enhanced LH release.
我们确定了微丝在基础条件和促性腺激素释放激素（GnRH）刺激条件下对培养的大鼠垂体前叶细胞中促黄体生成素（LH）合成（翻译或糖基化）及释放的调节作用。细胞先用微丝破坏药物细胞松弛素B（CB；2和20微摩尔）或细胞松弛素D（CD；1和10微摩尔）预处理2小时。在有或无1纳摩尔GnRH及药物存在的情况下孵育4小时后，测量LH的合成和释放。通过分别测量[14C]丙氨酸和[3H]葡糖胺掺入总的（细胞和培养基）可免疫沉淀LH中的量，来监测LH的翻译和糖基化。通过放射免疫分析（RIA）测量培养基和细胞中的免疫反应性LH（IRLH）。1纳摩尔的GnRH显著（P<0.01）增加了IRLH的释放及总的[3H]LH（糖基化），但对总的[14C]LH（翻译）、摄取或前体掺入总蛋白没有影响。2微摩尔和20微摩尔的CB以及10微摩尔的CD均未改变基础或GnRH刺激的IRHL释放。两种药物均未改变培养基中[3H]LH或[14C]LH的基础浓度。相反，CB和CD均降低了（P<0.01）GnRH刺激的培养基和整个系统中的[3H]LH（LH糖基化）。CB降低了（P<0.01）[3H]葡糖胺摄取、总的[3H]蛋白合成以及总的[3H]LH的基础水平，而CD对这些参数没有影响。因此，与CB相比，CD对GnRH刺激的LH糖基化具有更特异性的抑制作用。在基础和GnRH刺激条件下，2微摩尔和微摩尔的CB增加了（P<0.01）[14C]丙氨酸摄取、总的[14C]LH和[14C]蛋白，而10微摩尔的CD降低了（P<0.01）它们。这些结果表明，虽然细胞松弛素不抑制基础或GnRH刺激的IRLH释放，但它们确实抑制GnRH刺激的LH糖基化，尽管CB的作用部分归因于[3H]葡糖胺摄取的减少。微丝的完整性似乎对GnRH增强的LH糖基化很重要，但对GnRH增强的LH释放不重要。
