Chronic exposure to TPA depletes PKCalpha and augments Ca-dependent insulin secretion from cultured rat islets

The insulin secretory responses of rat islets to glucose (15 mM), 12-O-tetradecanoylphorbol13-acetate (TPA; 500 nM), and potassium (30 mM) were determined fromperifused islets cultured for 22-24 h in CMRL-1066 medium (controlcultured) or islets cultured in the additional presence of 500 nM TPA.Islet content of protein kinase C  (PKC) and serine and threoninephosphoprotein patterns were also monitored after the culture period.Compared with freshly isolated islets, culturing alone had no adverseeffect on the capacity of TPA or 30 mM potassium to stimulate secretionor on the islet content of PKC. In agreement with previous studies, culturing in TPA reduced the islet content of immunoreactive PKC by>95% and abolished the capacity of the phorbol ester to stimulate secretion during a subsequent dynamic perifusion. Culturing in TPAslightly improved the insulin secretory response to 15 mM glucosecompared with control-cultured islets; however, sustained rates of 15 mM glucose-induced secretion from these islets were significantly lessthan the responses of freshly isolated islets. Islets cultured in TPAresponded to 30 mM potassium with a markedly amplified insulinsecretory response that was abolished by nitrendipine. Enhancedphosphorylation of several islet proteins was also observed inTPA-cultured islets compared with control-cultured islets. Thesefindings demonstrate that culturing alone impairs glucose-induced secretion, a response that is improved but still subnormal compared with freshly isolated islet responses, if TPA is included in the culture medium. Sustained phosphorylation of several islet proteins inTPA-cultured islets may account, at least in part, for augmented calcium-dependent secretion.