Pharmacological evaluation of the role of cytochrome P450 in intracellular calcium signalling in rat pancreatic acinar cells.
We have investigated whether the cytochrome P450 system is involved in Ca(2+) signalling in rat pancreatic acinar cells. Intracellular free [Ca(2+)] ([Ca(2+)](i)) was measured in collagenase-isolated cells using fura-2 microspectrofluorimetry and imaging. The imidazole P450 inhibitor ketoconazole (5 - 50 microM) inhibited [Ca(2+)](i) oscillations induced by cholecystokinin octapeptide (CCK). However, ketoconazole also raised baseline [Ca(2+)](i) when applied in the absence of CCK. These effects were mimicked by 5 - 50 microM SKF96365, an imidazole widely used as an inhibitor of Ca(2+) entry. The non-imidazole P450 inhibitor proadifen (SKF525A) inhibited CCK-induced [Ca(2+)](i) oscillations at a concentration of 10 - 50 microM. Proadifen alone caused intracellular Ca(2+) release at 25 or 50 microM, but not at 10 microM. Octadecynoic acid and 1-aminobenzotriazole, structurally-unrelated non-imidazole P450 inhibitors, did not alter baseline [Ca(2+)](i) or CCK-evoked oscillations. We compared cumulative CCK dose-response relationship in control cells and in cells where P450 had been induced by prior injection of animals with beta-naphthoflavone. Only minor differences were apparent, with induced cells showing some decrease in responsiveness at moderate and higher concentration of CCK (30 pM - 3 nM). Direct assessment of depletion-activated Ca(2+) entry showed no clear differences between control and induced cells. In conclusion, we could find no compelling evidence for a role of P450 in controlling Ca(2+) signalling generally, or Ca(2+) entry in particular, in pancreatic acinar cells. Induction of P450 is therefore probably toxic to acinar cells via a Ca(2+)-independent mechanism.
Bruce
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JI
School of Biological Sciences, University of Manchester, G38 Stopford Building, Oxford Road, Manchester, M13 9PT, UK.
Elliott
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AC
D000818:Animals; D020013:Calcium Signaling; D003577:Cytochrome P-450 Enzyme System / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004790:Enzyme Induction; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007654:Ketoconazole / Q000494:pharmacology; D008297:Male; D010179:Pancreas / Q000378:metabolism*; D011335:Proadifen / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012844:Sincalide / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology
D005231:Fatty Acids, Unsaturated; D007093:Imidazoles; D007211:Indoles; C047008:17-octadecynoic acid; D019284:Thapsigargin; D003577:Cytochrome P-450 Enzyme System; D011335:Proadifen; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D012844:Sincalide; D007654:Ketoconazole; C000543:cyclopiazonic acid