Dual Action of Protein Kinase C Activation in the Regulation of Insulin Release by Muscarinic Agonist from Rat Insulinoma Cell Line (RINr)*

Abstract

The role of protein kinase C in muscarinic agonist-induced insulin release from rat insulinoma cells was investigated. The dose-dependent stimulation of insulin secretion by carbamylcholine (carbachol) was associated with dose-dependent increase in the release of 3H-inositolphosphates from prelabeled rat insulinoma cell line (RINr) cells. After preincubation with 32P-orthophosphates, carbachol also evoked a rapid decrease in 32P-labeling of phosphatidylinositol-4,5-bisphophate with concomitant increase in 32P-labeling of phosphatidic acid. Furthermore, carbachol significantly increased membrane-associated protein kinase C activity with a simultaneous decrease of its activity in cytosol. Although phorbol-12,13-dibutyrate (PDBu), a protein kinase C activator, also stimulated insulin release, insulin secretion induced by concomitant administration of carbachol and PDBu was clearly less than the level expected on the basis of an additive action. Moreover, PDBu significantly inhibited inositolphospholipid turnover stimulated by carbachol. Finally, PDBu inhibited the binding of 3H-scopolamine binding revealed that PDBu decreased the number of muscarinic receptors without altering its affinity. These findings suggest that activation of protein kinase C not only mediates muscarinic stimulation of insulin secretion from RINr cells but also operates a negative feedback mechanism in a signal transduction system, at least in part, via down-regulation of muscarinic receptors.

Translocation of protein kinase C in rat islets of Langerhans Effects of a phorbol ester, carbachol and glucose

“…The cholinergic muscarinic agonist, CCh, promoted translocation of PKC from cytosol to membranes in intact islets, as has also been recently reported in studies using rat insulinoma cells [16] and, in a preliminary report, a hamster cell line [ 181. These observations concur with the known effects of cholinergic agonists to promote DAG formation in islet membranes [19], and intact islets [20], and suggest that the DAG thus formed is involved in the activation and translocation of PKC.…”

“…In common with many other cell types [ 13-161, exposing intact islets to a phorbol ester produced a detectable shift of PKC activity to the membrane fraction. Despite the relatively high concentrations of phorbol esters used in our experiments, the extent of PKC translocation in islets was less than that reported in other tissues [13,14], including a rat insulinoma cell line [16]. This appears to be a real difference between tissues, since we were able to show considerably more translocation of PKC activity in parallel experiments using rat anterior pituitary tissue (46% of total in membranes in pituitary vs 24% in islets; Persaud and Jones, unpublished).…”

Processing and secretion of insulin-related peptides in an insulinoma cell line

“…These secretagogues have been shown to potentiate the stimulation of insulin secretion by glucose in primary and transformed pancreatic islet cells (Arkammar et al, 1989;Hill et al, 1987;Praz et al, 1983;Rajan et al, 1989;Yamatani et al, 1988). The initial content of immunoreactive IRP in the cells was 1193 ± 55 U/10 5 cells.…”

The diverse roles of protein kinase C in pancreatic β-cell function

“…cPKCs are the most strongly implicated isoforms [13]. However, there is also good evidence that PKC activation acts as a feedback inhibitor in the G q signalling pathway [11,[14][15][16][17]. It would not be surprising if this effect were mediated by activation of different PKC isoforms to those responsible for the predominant stimulation of secretion, although this has not been investigated extensively.…”