Insulin-like growth factor-I-enhanced secretion is abolished in protein kinase C-deficient chromaffin cells.
Previous studies have demonstrated that bovine chromaffin cells cultured in medium with 10 nM insulin-like growth factor-I (IGF-I) secrete about twofold more catecholamine when exposed to secretory stimuli than do cells cultured without IGF-I. The purpose of this study was to determine whether protein kinase C (PKC) is involved in the effect of IGF-I on secretion from these cells. PKC was down-regulated in the cells by 16-18 h of treatment with beta-phorbol didecanoate (beta-PDD; 100 nM). Such treatment had no effect on high-K(+)-stimulated secretion from cells cultured without IGF-I; however, secretion from cells cultured with IGF-I was reduced to a level comparable to that in cells cultured without the peptide. The inactive isomer, alpha-PDD (100 nM), had no effect on secretion from untreated or IGF-I-treated chromaffin cells. The effect of beta-PDD was time and concentration dependent, with 100 nM beta-PDD producing a maximal effect in 8-10 h. In situ PKC activity measured in permeabilized cells treated with PMA (300 nM) was decreased by approximately 40% by 10 h and was reduced to almost basal levels by 18 h. Immunoblotting experiments demonstrated that both alpha- and epsilon-PKC were lost from the cells with time courses similar to that seen in the in situ PKC assay. Overnight treatment with the PKC inhibitor H7 (100 microM) prevented the enhanced secretion normally seen in IGF-I-treated cells, whereas HA1004 had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)
Penberthy
W T
WT
Department of Biochemistry, College of Medicine, University of Tennessee, Memphis 38163.
Dahmer
M K
MK
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000313:Adrenal Medulla / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D000595:Amino Acid Sequence; D000704:Analysis of Variance; D000818:Animals; D001921:Brain / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology*; D010879:Piperazines / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D051381:Rats; D013379:Substrate Specificity; D013449:Sulfonamides; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002395:Catecholamines; D007546:Isoquinolines; D010446:Peptide Fragments; D010703:Phorbol Esters; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; C008876:phorbol-12,13-didecanoate; D007334:Insulin-Like Growth Factor I; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; C043204:N-(2-guanidinoethyl)-5-isoquinolinesulfonamide; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium