Zawalich W S, Zawalich K C
Yale University School of Nursing, New Haven, Connecticut 06536-0740.
Endocrinology. 1990 May;126(5):2307-12. doi: 10.1210/endo-126-5-2307.
Isolated rat islets of Langerhans were incubated for 2 h in a myo-[2-3H]inositol-containing solution to label their phosphoinositides. Also included during this labeling period was forskolin (0.1-5 microM), a compound established to elevate islet cAMP levels. These islets were subsequently perifused, and their insulin secretory responses to 20 mM glucose or 1 microM of the phorbol ester 12-O-tetradecanoyl phorbol-13-acetate (TPA) were assessed. Determined in parallel with secretion were [3H] inositol efflux patterns and, at the termination of the perifusion, labeled inositol phosphate accumulation. The following major observations were made. 1) Forskolin had no deleterious effect on the total amount of [3H]inositol incorporated by the islets during the labeling period. 2) However, labeling in forskolin resulted in subsequent dose-dependent decreases in 20 mM glucose-induced insulin secretion, [3H]inositol efflux and inositol phosphate accumulation. 3) Inclusion of the diacylglycerol (DAG) kinase inhibitor monooleoylglycerol (50 microM) restored to a significant degree glucose-induced release from forskolin-desensitized islets. 4) Pretreatment with 5 microM forskolin had no deleterious effect on TPA-induced insulin release. 5) Prior exposure to forskolin also impaired phosphoinositide hydrolysis in response to cholecystokinin stimulation. 6) Similar to forskolin, labeling in isobutylmethylxanthine (1 mM) reduced in a parallel fashion islet [3H]inositol efflux and insulin secretion in response to 20 mM glucose stimulation. These findings demonstrate that prior chronic elevation of islet cAMP levels suppresses the activation of phospholipase-C in response to subsequent stimulation. Defective insulin secretory responsiveness of these islets appears to be the result of impaired generation of phosphoinositide-derived second messenger molecules, particularly DAG. By substituting for DAG, however, TPA circumvents this biochemical lesion and evokes a normal insulin secretory response from forskolin-pretreated islets.
将分离的大鼠胰岛在含肌醇-[2-³H]的溶液中孵育2小时，以标记其磷酸肌醇。在该标记期间还加入了福斯可林（0.1 - 5微摩尔），这是一种能提高胰岛环磷酸腺苷（cAMP）水平的化合物。随后对这些胰岛进行灌流，并评估它们对20毫摩尔葡萄糖或1微摩尔佛波酯12 - O - 十四酰佛波醇 - 13 - 乙酸酯（TPA）的胰岛素分泌反应。与分泌同时测定的是[³H]肌醇流出模式，并在灌流结束时测定标记的肌醇磷酸积累。得出以下主要观察结果。1）福斯可林对标记期间胰岛摄取的[³H]肌醇总量没有有害影响。2）然而，在福斯可林存在下进行标记导致随后20毫摩尔葡萄糖诱导的胰岛素分泌、[³H]肌醇流出和肌醇磷酸积累呈剂量依赖性降低。3）加入二酰基甘油（DAG）激酶抑制剂单油酰甘油（50微摩尔）在很大程度上恢复了福斯可林脱敏胰岛对葡萄糖诱导的释放。4）用5微摩尔福斯可林预处理对TPA诱导的胰岛素释放没有有害影响。5）预先暴露于福斯可林也损害了胆囊收缩素刺激引起的磷酸肌醇水解。6）与福斯可林相似，在异丁基甲基黄嘌呤（1毫摩尔）存在下进行标记以平行方式降低了胰岛对20毫摩尔葡萄糖刺激的[³H]肌醇流出和胰岛素分泌。这些发现表明，预先长期升高胰岛cAMP水平会抑制后续刺激引起的磷脂酶 - C的激活。这些胰岛胰岛素分泌反应缺陷似乎是磷酸肌醇衍生的第二信使分子，特别是DAG生成受损的结果。然而，通过替代DAG，TPA绕过了这种生化损伤，并从福斯可林预处理的胰岛中引发正常的胰岛素分泌反应。
