Sodium fluoride stimulates exocytosis at a late site of calcium interaction in stimulus-secretion coupling: studies with the RINm5F beta cell line.
In the insulin-secreting beta cell line RINm5F, sodium fluoride stimulated exocytosis in a concentration (5-15 mM)- and temperature-dependent manner. Depletion of aluminum with the chelator deferoxamine or addition of aluminum to the buffer failed to affect the NaF-stimulated insulin release. This suggests that stimulation of heterotrimeric G proteins or inhibition of phosphatases or other enzymes by fluoroaluminate, an analog of the phosphate moiety, is not involved in the insulinotropic action of NaF. Removal of extracellular Ca2+ suppressed the NaF-stimulated insulin release. However, nitrendipine, a blocker of L-type voltage-dependent Ca2+ channels, did not inhibit the NaF-stimulated insulin release and NaF did not cause any changes in the cytosolic free calcium concentration ([Ca2+]i). Decreasing [Ca2+]i with thapsigargin or increasing [Ca2+]i with ionomycin or a depolarizing concentration of KCl resulted in suppression or enhancement of NaF-stimulated insulin release, respectively. Furthermore, NaF enhanced Ca(2+)-induced insulin release in electrically permeabilized RINm5F cells. These findings indicate that the effect of NaF on exocytosis is dependent on [Ca2+]i, although NaF itself does not change [Ca2+]i. Inhibitors of protein kinase C, such as staurosporine and bisindolylmaleimide, in concentrations sufficient to block the effects of phorbol esters, did not attenuate the NaF-stimulated insulin release. Neither cellular cAMP content nor [3H]arachidonic acid release was increased by NaF. NaF-stimulated insulin release was synergistically enhanced by the activation of protein kinases A and C. Finally, trifluoperazine, an inhibitor of calmodulin and other Ca(2+)-binding proteins, inhibited the insulinotropic action of NaF in a concentration-dependent manner. Trifluoperazine (50 microM) and W-7 (100 microM) nullified the 10 mM NaF-stimulated insulin release. It is concluded that NaF evokes exocytosis by a novel mechanism of sensitization to Ca2+, possibly on a Ca(2+)-responsive protein that is sensitive to trifluoperazine and W-7, leading to exocytosis. Protein kinases A and C also act at this site or at a more distal point.
Komatsu
M
M
Department of Pharmacology, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853.
McDermott
A M
AM
Sharp
G W
GW
D000535:Aluminum / Q000502:physiology; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D003676:Deferoxamine / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects* / Q000502:physiology; D005110:Extracellular Space / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D015759:Ionomycin / Q000494:pharmacology; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism*; D007700:Kinetics; D008555:Melitten / Q000494:pharmacology; D009568:Nitrendipine / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D012636:Secretory Rate / Q000187:drug effects; D012969:Sodium Fluoride / Q000494:pharmacology*; D013268:Stimulation, Chemical; D013449:Sulfonamides / Q000494:pharmacology; D013696:Temperature; D013729:Terpenes / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019284:Thapsigargin; D014268:Trifluoperazine / Q000494:pharmacology
D002135:Calcium-Binding Proteins; D007328:Insulin; D013449:Sulfonamides; D013729:Terpenes; D005576:Colforsin; D008555:Melitten; D014268:Trifluoperazine; D016718:Arachidonic Acid; D015759:Ionomycin; C017967:W 7; D011189:Potassium Chloride; D019284:Thapsigargin; D012969:Sodium Fluoride; D009568:Nitrendipine; D000535:Aluminum; D000242:Cyclic AMP; D011493:Protein Kinase C; D003676:Deferoxamine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium