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 Ca2+- free or Ca2+-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 Ca2+-independent activation of the plasma membrane Ca2+ pump by Ca2+ mobilizing agonists.
|Original language||English (US)|
|Number of pages||7|
|Journal||Journal of Biological Chemistry|
|State||Published - Jan 1 1992|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology