Thapsigargin-induced Ca2+ mobilization in acutely isolated mouse lacrimal acinar cells is dependent on a basal level of Ins(1,4,5)P3 and is inhibited by heparin.
The tumour-promoting agent thapsigargin has been shown to inhibit the microsomal Ca(2+)-ATPase and cause Ca2+ mobilization in a variety of cell types including exocrine acinar cells [Bird, Obie and Putney (1992) J. Biol. Chem. 267, 18382-18386]. When applied to acutely isolated lacrimal acinar cells, thapsigargin caused a slow biphasic activation of both the Ca(2+)-dependent K+ and Cl- currents measured using the whole-cell patch-clamp technique. If the only action of thapsigargin is to inhibit sequestration into Ca2+ pools, then Ca2+ mobilization following exposure to thapsigargin indicates that there is a significant 'leak' of Ca2+ into the cytoplasm, which is normally countered by Ca(2+)-ATPase activity. In the present study, we introduced the Ins(1,4,5)P3 receptor antagonist heparin (200 micrograms/ml) into lacrimal acinar cells via the patch-clamp pipette. Following a 5 min preincubation in the presence of heparin, neither acetylcholine (1 microM) nor thapsigargin (1 microM) caused any significant increase in either Ca(2+)-dependent current. Caffeine has been shown to suppress basal Ins(1,4,5)P3 levels in exocrine acinar cells [Toescu, O'Neill, Petersen and Eisner (1992) J. Biol. Chem. 267, 23467-23470]. Preincubation with caffeine (10 mM) also inhibited the response to subsequent exposure to thapsigargin. These data suggest that, in acutely isolated lacrimal cells, the source of the Ca2+ leak which gives rise to Ca2+ mobilization following inhibition of Ca2+ re-uptake by thapsigargin is Ca2+ release, from Ins(1,4,5)P3-dependent Ca2+ pools, caused by resting Ins(1,4,5)P3 levels.
Smith
P M
PM
Physiological Laboratory, University of Liverpool, U.K.
Gallacher
D V
DV
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D018118:Chloride Channels / Q000187:drug effects; D006493:Heparin / Q000494:pharmacology*; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D007765:Lacrimal Apparatus / Q000166:cytology / Q000378:metabolism*; D008297:Male; D051379:Mice; D015221:Potassium Channels / Q000187:drug effects; D013729:Terpenes / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D019284:Thapsigargin
D018118:Chloride Channels; D015221:Potassium Channels; D013729:Terpenes; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D006493:Heparin; D002118:Calcium