Caffeine inhibits the agonist-evoked cytosolic Ca2+ signal in mouse pancreatic acinar cells by blocking inositol trisphosphate production.
The inhibitory effects of caffeine on receptor-activated cytosolic Ca2+ signal generation in isolated mouse pancreatic acinar cells were investigated. Using the ability of caffeine to quench Indo-1 fluorescence we measured simultaneously the free intracellular Ca2+ concentration ([Ca2+]i) and the intracellular caffeine concentration ([caffeine]i). We also measured inositol 1,4,5-trisphosphate (InsP3) production with a radioreceptor assay. When caffeine was added to the extracellular solution during a sustained receptor-activated increase in [Ca2+]i, [caffeine]i rose to its steady level within a few seconds. This was accompanied by a decrease of [Ca2+]i, which started only after [caffeine]i had reached an apparent threshold concentration (about 2 mM in the case of 0.5 microM acetylcholine (ACh) stimulation). Above this [caffeine]i level there was a linear relationship between [caffeine]i and [Ca2+]i. Throughout the caffeine exposure [Ca2+]i remained at a steady low level. Following removal of caffeine from the bath, [caffeine]i decreased to zero within seconds. There was no significant increase in [Ca2+]i until [caffeine]i had been reduced to the threshold level (about 2 mM at 0.5 microM ACh). Caffeine inhibited Ca2+ signals evoked by ACh, cholecystokinin, and ATP and also inhibited signals generated in the absence of external Ca2+. Caffeine application had the same effect as removal of agonist allowing recovery from apparent desensitization. Caffeine inhibited the agonist-evoked production of InsP3 in a dose-dependent manner. Our results demonstrate the acute and reversible dose-dependent inhibition of agonist-evoked cytosolic Ca2+ signal generation due to rapid intracellular caffeine accumulation and washout. The inhibition can be explained by the reduction of agonist-evoked InsP3 production.
Toescu
E C
EC
Medical Research Council Secretory Control Research Group, Liverpool University, United Kingdom.
O'Neill
S C
SC
Petersen
O H
OH
Eisner
D A
DA
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D007211:Indoles; D015544:Inositol 1,4,5-Trisphosphate / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D051379:Mice; D010179:Pancreas / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*
D005456:Fluorescent Dyes; D007211:Indoles; D002110:Caffeine; D015544:Inositol 1,4,5-Trisphosphate; C048960:indo-1; D000109:Acetylcholine; D002118:Calcium