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The Physiological Role of Calmodulin in Rat Parotid Amylase Release

钙调蛋白在大鼠腮腺淀粉酶释放中的生理作用

基本信息

批准号：
63570876
负责人：
TOJYO Yosuke
金额：
$ 1.15万
依托单位：
Higashi-Nippon-Gakuen University, School of Dentistry
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1988
资助国家：
日本
起止时间：
1988 至 1989
项目状态：
已结题

项目摘要

Using dispersed rat parotid cells, the effects of three calmodulin antagonists trifluoperazine (TFP), W-7 and W-5, on amylase release and acinar cell structure were examined. TFP and W-7 strongly inhibited both isoproterenol (ISO)- or DBcAMP-stimulated amylase release at a concentration of 50 or 100 muM, while W-5, a weak calmodulin antagonist, had a little effect. Cyclic AMP level was markedly elevated by ISO even in the presence of TFP and W-7.Electron micrographs demonstrated that treatment of parotid cells with either TFP or W-7 caused a loss of luminal microvilli and surface folds. When cells were stimulated by ISO in the presence of TFP or W-7, the enlarged lumina did not recover to their original size and the discharged secretory material was retained in the lumina. The structural changes were similar, in part, to those evoked by cytochalasin D, a microfilament disrupting agent. W-5 affected the acinar cell structure only a little.Amylase release induced by ISO or DBcAMP was significantly enhanced by pretreatment with Ca^<2+>. Depletion of intracellular Ca^<2+> by pre-treatment with the Ca^<2+>-chelators EGTA and BAPTA-AM reduced the amylase release.The effects of ISO on cytosolic free Ca^<2+>([Ca^<2+>]i)were examined using the fluorescent Ca^<2+>-indicator fura-2. At concentrations up to 1 mM, ISO caused a rapid increase in [Ca^<2+>]i, while 1 muM ISO, which stimulates the maximum amylase release, had little effect. Since the alpha-adrenoceptor antagonist phentolamine blocked the ISO-induced increasg+in [Ca^<2+>]i better than the beta-adrenoceptor antagonist, the increase in [Ca^<2+>]i is due to an activation of alpha-adrenoceptors rather than beta-adrenoceptors.This study suggests that calmodulin is involved in the exocytosis of parotid amylase through the regulation of the cytoskeletal system. Calmodulin may be functional even at resting levels of [Ca^<2+>]i.

采用大鼠腮腺细胞，研究了三种钙调素拮抗剂三氟拉嗪（TFP）W-7和W-5对腺泡细胞淀粉酶释放和结构的影响。TFP和W-7在50或100 μ M的浓度下强烈抑制异丙肾上腺素（ISO）或DBcAMP刺激的淀粉酶释放，而弱钙调素拮抗剂W-5的作用很小。电镜观察表明，TFP或W-7处理腮腺细胞后，腮腺细胞腔微绒毛和表面皱褶消失。当细胞在TFP或W-7存在下被ISO刺激时，扩大的管腔不能恢复到其原始大小，并且排出的分泌物质保留在管腔中。的结构变化是相似的，部分，引起的细胞松弛素D，微丝破坏剂。W-5对腺泡细胞结构影响不大，Ca^<2+>预处理可显著增强ISO或DBcAMP诱导的淀粉酶释放。用Ca^2+螯合剂EGTA和BAPTA-AM预处理细胞内Ca^2+，使其耗竭，淀粉酶释放减少，用荧光Ca^2+指示剂fura-2检测ISO对胞浆游离Ca^2+（[Ca^2+]i）的影响。当浓度达到1 mM时，ISO引起[Ca^<2+>]i的快速增加，而1 μ M ISO刺激淀粉酶的最大释放，几乎没有影响。由于α-肾上腺素能受体拮抗剂酚妥拉明比β-肾上腺素能受体拮抗剂能更好地阻断ISO引起的[Ca ^<2 +>]i的增加，[Ca^<2+]i的增加是由于α-肾上腺素能受体而不是β-肾上腺素能受体的激活。钙调蛋白甚至在[Ca^<2+>]i的静息水平下也可能是功能性的。