Activation of the plasma membrane Ca2+ pump during agonist stimulation of pancreatic acini. |
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Authors: | B X Zhang H Zhao P Loessberg S Muallem |
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Institution: | Department of Physiology, University of Texas Southwestern Medical Center, Dallas 75235-9040. |
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Abstract: | The role of internal stores and plasma membrane Ca2+ pumps in controlling Ca2+]i during agonist stimulation and their regulation by agonists are not well understood. We report here measurements of intracellular (Ca2+]i) and extracellular (Ca2+]o) Ca2+ concentrations in agonist-stimulated pancreatic acini in an effort to directly address these questions. Stimulation of acini suspended in Ca(2+)-free or Ca(2+)-containing medium with Ca2+ mobilizing agonists resulted in a typical transient increase in Ca2+]i. Thapsigargin, a specific inhibitor of internal Ca2+ pumps, inhibited the rate of Ca2+]i reduction after agonist stimulation by approximately 40%. Under the same conditions, thapsigargin had no effect on the rate of the unidirectional Ca2+ efflux across the plasma membrane as revealed by measurements of Ca2+]o. These findings suggest that internal Ca2+ pumps actively remove Ca2+ from the cytosol during continued agonist stimulation. The correlation between the reduction in Ca2+]i and the increase in Ca2+]o showed that Ca2+ efflux from cells stimulated with agonist and thapsigargin represent Ca2+ efflux across the plasma membrane. Inhibition of cells exposed to agonist and thapsigargin with a specific antagonist sharply reduced the rates of the Ca2+]i decrease and the accompanied Ca2+]o increase. Hence, at comparable Ca2+]i, Ca2+ efflux from stimulated cells was about 3-fold faster than that from resting cells, indicating that agonists directly activate the plasma membrane Ca2+ pump. To study the role of Ca2+]i increase in plasma membrane Ca2+ pump activation the acini were loaded with 1,2-bis-(2-aminophenoxyethane-N,N,N',N')-tetraacetic acid (BAPTA), and Ca2+]o was measured during agonist stimulation. Surprisingly, although BAPTA completely prevented the increase in Ca2+]i, Ca2+ efflux rate was reduced by only 34%. These findings provide the first evidence for Ca(2+)-independent activation of the plasma membrane Ca2+ pump by Ca2+ mobilizing agonists. |
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