Matozaki T
Second Department of Internal Medicine, Kobe University School of Medicine, Japan.
Nihon Naibunpi Gakkai Zasshi. 1988 Oct 20;64(10):1051-64. doi: 10.1507/endocrine1927.64.10_1051.
To clarify the possible role of a guanine nucleotide regulatory protein in the signal transducing system activated by cholecystokinin (CCK), the actions of CCK on rat pancreatic acini were compared with those of NaF, which is a well-known activator of stimulatory and inhibitory guanine nucleotide regulatory proteins. Guanine nucleotides reduced the binding of 125I-CCK-octapeptide (CCK8) to acinar cell membranes, with the rank order of potency being guanyl-5'-yl imidodiphosphate (Gpp(NH)p) greater than GTP greater than GDP greater than GMP. Scatchard analysis of labeled CCK binding revealed that the decrease in CCK binding caused by Gpp(NH)p was due to the decrease in an affinity constant of CCK for its receptors with no significant change in the maximal binding capacity. When acini were incubated with increasing concentrations of either CCK8 or NaF, maximal stimulation of amylase release occurred at 100 pM CCK8 or 10 mM NaF, respectively, and supramaximal concentrations of CCK8 or NaF caused its submaximal stimulation. Further, CCK and NaF similarly stimulated the hydrolysis of polyphosphoinositide in acini and the release of Ca2+ from the intracellular Ca2+ store into the cytoplasm although there was a lag period prior to any detectable stimulation by NaF. The doses of NaF necessary for Ca2+ mobilization and inositol phosphate generation were nearly the same, with a maximal stimulation at 20 mM NaF. NaF, at concentrations up to 20 mM, a supramaximal concentration for amylase release, produced no significant change in the cellular cyclic AMP level. In addition, 10 mM NaF potentiated the amylase release stimulated by VIP, a well-known secretagogue which functions via cyclic AMP, suggesting that the stimulatory effects of NaF are independent of cellular cyclic AMP. Gpp(NH)p also activated the hydrolysis of polyphosphoinositide in a cell-free pancreatic acinar cell membrane preparation, with a half-maximal and a maximal stimulation at 1 microM and 10 microM, respectively. Furthermore, the effects of submaximal concentrations of CCK8 on polyphosphoinositide hydrolysis were markedly potentiated in the presence of 100 microM GTP, which alone was ineffective. These results, therefore, strongly suggest that pancreatic CCK receptors may be coupled to the activation of polyphosphoinositide breakdown by a guanine nucleotide regulatory protein.
为阐明鸟嘌呤核苷酸调节蛋白在胆囊收缩素（CCK）激活的信号转导系统中可能的作用，将CCK对大鼠胰腺腺泡的作用与NaF的作用进行了比较，NaF是一种众所周知的刺激性和抑制性鸟嘌呤核苷酸调节蛋白激活剂。鸟嘌呤核苷酸降低了125I-CCK-八肽（CCK8）与腺泡细胞膜的结合，其效力顺序为鸟苷-5'-基亚氨基二磷酸（Gpp(NH)p）＞GTP＞GDP＞GMP。对标记的CCK结合进行Scatchard分析表明，Gpp(NH)p引起的CCK结合减少是由于CCK与其受体的亲和常数降低，而最大结合容量无显著变化。当腺泡与浓度不断增加的CCK8或NaF孵育时，淀粉酶释放的最大刺激分别在100 pM CCK8或10 mM NaF时出现，而CCK8或NaF的超最大浓度导致其亚最大刺激。此外，CCK和NaF同样刺激腺泡中多磷酸肌醇的水解以及Ca2+从细胞内Ca2+储存库释放到细胞质中，尽管在NaF产生任何可检测到的刺激之前有一个延迟期。Ca2+动员和肌醇磷酸生成所需的NaF剂量几乎相同，在20 mM NaF时达到最大刺激。高达20 mM的NaF浓度（淀粉酶释放的超最大浓度）对细胞内的环磷酸腺苷水平没有显著影响。此外，10 mM NaF增强了由VIP刺激的淀粉酶释放，VIP是一种通过环磷酸腺苷起作用的众所周知的促分泌素，这表明NaF的刺激作用独立于细胞内的环磷酸腺苷。Gpp(NH)p在无细胞的胰腺腺泡细胞膜制剂中也激活了多磷酸肌醇的水解，在1 μM和10 μM时分别达到半最大刺激和最大刺激。此外，在100 μM GTP存在的情况下，亚最大浓度的CCK8对多磷酸肌醇水解的作用明显增强，而单独的GTP无效。因此，这些结果强烈表明胰腺CCK受体可能通过鸟嘌呤核苷酸调节蛋白与多磷酸肌醇分解的激活相偶联。
