Secretin internalization and adenosine 3',5'-monophosphate levels in pancreatic acinar cells.
The internalization of [125I]secretin in pancreatic acinar cells was evaluated by differentiation of surface-bound and internalized radioligand with an acidified glycine buffer. The amount of surface-bound radioligand was 2-fold higher at 37 C than at 4 C between 15 and 60 min; internalized radioactivity was more than 10-fold greater at 37 C than at 4 C during the same time period. The effects of chloroquine, dithiothreitol (DTT), carbonyl cyanide m-chlorophenyl-hydrazone (CCCP), and dansylcadaverine on the binding and internalization of secretin were then evaluated. Chloroquine (0.1 mM), a lysosomotropic agent, did not affect secretin radioligand binding to its pancreatic receptor, while DTT, a sulfhydryl reducing agent, significantly lowered binding by more than 90% from 15-60 min. The metabolic inhibitor CCCP, however, significantly enhanced binding of the secretin radioligand in both the surface and the internalized pools. Surface binding was 1.6- to 3.3-fold greater from 15-60 min (P less than or equal to 0.01) in acinar cells exposed to CCCP than in untreated controls, while internalized radioactivity increased from 2.8- to 1.4-fold above the control value (P less than or equal to 0.01) at the same times. Dansylcadaverine, an inhibitor of receptor recycling, reduced internalized radioligand by 40% after 30 min of binding. The effects of these chemical agents on cAMP production were also considered. cAMP production was significantly reduced by DTT, CCCP, and dansylcadaverine at secretin concentrations of 0.1, 3.0, and 10 nM, respectively. This study demonstrates that 1) pancreatic acinar cells rapidly internalize [125I]secretin; 2) internalization of secretin does not enhance cAMP production; and 3) disulfide linkages are important for secretin receptor activity.
Izzo
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RS
Department of Medicine, Nassau County Medical Center, East Meadow, New York 11554.
Chen
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AI
Pellecchia
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Praissman
M
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D000249:Adenosine Monophosphate / Q000378:metabolism*; D000818:Animals; D001414:Bacitracin / Q000494:pharmacology; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002738:Chloroquine / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D010179:Pancreas / Q000166:cytology / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled; D011964:Receptors, Gastrointestinal Hormone / Q000378:metabolism; D012633:Secretin / Q000378:metabolism*
D043562:Receptors, G-Protein-Coupled; D011964:Receptors, Gastrointestinal Hormone; C029937:secretin receptor; D012633:Secretin; D001414:Bacitracin; D000249:Adenosine Monophosphate; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D002738:Chloroquine; D000242:Cyclic AMP; C008542:monodansylcadaverine; D002103:Cadaverine; D004229:Dithiothreitol