Koshiyama H, Tashjian A H
Laboratory of Toxicology, Harvard School of Public Health, Boston, Massachusetts 02115.
Endocrinology. 1991 Jun;128(6):2715-22. doi: 10.1210/endo-128-6-2715.
In GH4C1 rat pituitary cells, a GTP-binding protein appears to be involved in signal transduction between the TRH receptor and phospholipase C. In certain other cell types, another role for GTP has been reported, namely regulation of Ca2+ translocation from one intracellular pool to another. Using digitonin-permeabilized GH4C1 cells, we have investigated whether an analogous process occurs in pituitary cells. In permeabilized GH4C1 cells, TRH, inositol 1,4,5-trisphosphate (IP3), and nonhydrolyzable GTP analogs guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) and 5'-guanylyl imidodiphosphate each increased free Ca2+ concentration [( Ca2+]). Unlike several other systems, GTP did not increase [Ca2+]. Guanosine 5'-O-(2-thiodiphosphate) inhibited Ca2+ release induced by both TRH and GTP gamma S. Heparin abolished IP3-induced Ca2+ release but did not prevent Ca2+ release induced by TRH or GTP gamma S, suggesting a mechanism for their actions that did not depend solely on IP3 production. Neomycin inhibited GTP gamma S-induced Ca2+ release, but it did not prevent TRH- or IP3-induced Ca2+ release. In the absence of ATP, GTP gamma S did not elevate [Ca2+], although TRH and IP3 did, suggesting that ATP-dependent sequestration of Ca2+ was necessary for the action of GTP gamma S in this system, but not for TRH and IP3. Repeated additions of IP3 resulted in an attenuation of the response to IP3- GTP gamma S, which itself increased [Ca2+] after IP3 attenuation, restored the attenuated Ca2+ response to IP3. We conclude that, in permeabilized GH4C1 cells, GTP gamma S as well as TRH cause intracellular Ca2+ release; however, their mechanisms of action are, at least in part, distinct. Furthermore, the IP3-depletable Ca2+ pool can be refilled from a GTP gamma S-sensitive compartment via Ca2+ transport through the cytosol.
在GH4C1大鼠垂体细胞中，一种GTP结合蛋白似乎参与了促甲状腺激素释放激素（TRH）受体与磷脂酶C之间的信号转导。在某些其他细胞类型中，已报道GTP的另一个作用，即调节Ca2+从一个细胞内池转运到另一个细胞内池。利用洋地黄皂苷通透的GH4C1细胞，我们研究了垂体细胞中是否发生类似过程。在通透的GH4C1细胞中，TRH、肌醇1,4,5-三磷酸（IP3）以及不可水解的GTP类似物鸟苷5'-O-(3-硫代三磷酸)（GTPγS）和5'-鸟苷酰亚胺二磷酸均增加了游离Ca2+浓度[Ca2+]。与其他几个系统不同，GTP并未增加[Ca2+]。鸟苷5'-O-(2-硫代二磷酸)抑制TRH和GTPγS诱导的Ca2+释放。肝素消除了IP3诱导的Ca2+释放，但并未阻止TRH或GTPγS诱导的Ca2+释放，这表明它们的作用机制并非仅依赖于IP3的产生。新霉素抑制GTPγS诱导的Ca2+释放，但不阻止TRH或IP3诱导的Ca2+释放。在没有ATP的情况下，GTPγS不会升高[Ca2+]，尽管TRH和IP3会，这表明ATP依赖的Ca2+螯合对于该系统中GTPγS的作用是必要的，但对TRH和IP3则不是。重复添加IP3导致对IP3-GTPγS的反应减弱，而GTPγS本身在IP3反应减弱后增加[Ca2+]，恢复了对IP3减弱的Ca2+反应。我们得出结论，在通透的GH4C1细胞中，GTPγS以及TRH均引起细胞内Ca2+释放；然而，它们的作用机制至少部分是不同的。此外，可被IP3耗尽的Ca2+池可通过Ca2+经细胞质的转运从对GTPγS敏感的区室重新填充。
