Dynamics of calcium clearance in mouse pancreatic beta-cells.
Pancreatic beta-cells maintain glucose homeostasis by their regulated Ca(2+)-dependent secretion of insulin. Several cellular mechanisms control intracellular Ca(2+) levels, but their relative significance in mouse beta-cells is not fully known. We used photometry to measure the dynamics of cytosolic Ca(2+) ([Ca(2+)](i)) clearance after brief, depolarization-induced Ca(2+) entry. Treatment with thapsigargin or cyclopiazonic acid, inhibitors of the sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA) pumps, nearly doubled the peak and slowed the decay of the depolarization-induced Ca(2+) transients. The remaining thapsigargin-insensitive decay was slowed further by inhibition of the plasma membrane Ca(2+)-ATPase (PMCA) and plasma membrane Na(+)/Ca(2+) exchanger (NCX) via alkalization of the bath solution, by adding lanthanum, or by substitution of Na(+) with Li(+). Mitochondrial Ca(2+) uptake contributed little to clearance in thapsigargin-pretreated cells. Together, the SERCA, PMCA, and NCX transport mechanisms accounted for 89 to 97% of clearance in normal solutions. We developed a quantitative model for the dynamic role of removal mechanisms over a wide range of [Ca(2+)](i). According to our model, 50 to 64% of initial Ca(2+) removal is via the SERCA pump, whereas the NCX contributes 21-30% of the extrusion at high [Ca(2+)](i), and the PMCA contributes 21-27% at low [Ca(2+)](i).
Chen
Liangyi
L
Department of Physiology & Biophysics, School of Medicine, University of Washington, Seattle 98195-7290, USA. lychen@u.washington.edu
Koh
Duk-Su
DS
Hille
Bertil
B
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000187:drug effects / Q000502:physiology*; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008657:Metabolic Clearance Rate; D051379:Mice; D008807:Mice, Inbred BALB C; D008954:Models, Biological; D011189:Potassium Chloride / Q000494:pharmacology; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
D011189:Potassium Chloride; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium