Phosphorylation of 3 particulate proteins in rat pancreatic acini in response to vasoactive intestinal peptide (VIP), secretin and cholecystokinin (CCK-8).
Rat pancreatic acini were preincubated with 0.4 mM 32Pi for 45 min at 37 degrees C, then exposed for 15 min to VIP, secretin or CCK-8. The incubation was terminated with a stop solution and a fraction rich in mitochondria and zymogen granules was separated from a microsome-rich fraction by differential centrifugation. After heating in the presence of SDS, beta-mercaptoethanol was added and the pattern of equivalent amounts of 32P-labelled proteins was examined by autoradiography of SDS-PAGE gels. VIP, secretin, and CCK-8 stimulated the phosphorylation of a Mr=33 K microsomal protein and that of two proteins of Mr=21 K and Mr=25 K mostly present in a fraction rich in mitochondria and zymogen granules. Stimulations were dose-dependent, the highest stimulant concentrations tested allowing 2- to 3-fold increases of phosphorylation over basal. When 1 nM CCK-8 was used simultaneously with 1 microM VIP, the cyclic AMP levels attained and the pattern of protein phosphorylation were similar to those obtained with VIP alone, and there was a potentiation of amylase secretion; when a supra-maximal 0.1 microM CCK-8 concentration was added, the VIP-induced elevation in cyclic AMP levels and the phosphorylation of the Mr=21 K and Mr=25 K proteins were partially antagonized, and no potentiation any more of secretion occurred. To conclude the in vitro phosphorylation of three particulate proteins (Mr=33 K, 25 K, and 21 K) was similarly increased in rat pancreatic acini in response to secretin and VIP (acting through cyclic AMP) and to CCK-8 (acting mostly through Ca2+).
D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D010179:Pancreas / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010750:Phosphoproteins / Q000302:isolation & purification; D010766:Phosphorylation; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D012633:Secretin / Q000494:pharmacology*; D012844:Sincalide / Q000494:pharmacology*; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology*
D010750:Phosphoproteins; D011506:Proteins; D012633:Secretin; D014660:Vasoactive Intestinal Peptide; D000242:Cyclic AMP; D012844:Sincalide