Effect of Thapsigargin on Cytosolic Ca²⁺ Level and Amylase Release in Rat Parotid Acinar Cells

Abstract

At concentrations >0.01 μM, thapsigargin (ThG) dose-dependently caused an increase in cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) in rat parotid acinar cells, as measured by the fluorescent Ca²⁺-indicator fura-2. In the absence of extracellular Ca²⁺, a transient increase in [Ca²⁺]_i by ThG was observed, and subsequent addition of carbachol (CCh) did not produce a further [Ca²⁺]_i response, suggesting that ThG released Ca²⁺ from the CCh-sensitive intracellular Ca²⁺ pool. Since ThG did not stimulate formation of inositol phosphates, the ThG-induced Ca²⁺ mobilization is independent of phosphoinositide breakdown. High concentrations (>0.1 μM) of ThG induced amylase release from rat parotide acini, but the effect was very poor as compared with that of CCh or the protein kinase C activator, PMA(phorbol 12-myristate 13-acetate). Combined addition of ThG and PMA modestly potentiated amylase release induced by PMA alone. These results support the view that amylase release by muscarinic stimulation is mediated mainly by activation of protein kinase C rather than a rise in [Ca²⁺]_i although Ca²⁺ may modulate the secretory response.