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1 Department of Physiology, University of Extremadura, Caceres, Spain
2 Department of Anatomy and Neurobiology, University of Vermont, Burlington, Vermont, USA
* To whom correspondence should be addressed. E-mail: mjpozo{at}unex.es.
We have evaluated the presence of capacitative calcium entry (CCE) in guinea pig gallbladder smooth muscle (GBSM), including a possible relationship with activation of L-type Ca2+ channels. Changes in cytosolic Ca2+ concentration ([Ca2+]i) induced by Ca2+ entry were assessed by digital microfluorimetry in isolated, fura-2 loaded GBSM cells. Application of thapsigargin, a specific inhibitor of the Ca2+ store pump, induced a transient Ca2+ release followed by sustained entry of extracellular Ca2+ ions. Depletion of the stores with thapsigargin, cyclopiazonic acid (CPA), ryanodine and caffeine, high levels of the Ca2+-mobilizing hormone CCK, or by simple removal of external calcium, resulted in a sustained increase in Ca2+ entry upon subsequent reapplication of Ca2+. This entry was attenuated by 2-APB, L-type Ca2+ channel blockade, pinacidil and by Gd 3+ ions. Both accumulation of the voltage-sensitive dye DiOC5(3) and direct intracellular recordings showed that depletion of the stores is sufficient for depolarization of plasma membrane. Contractility studies in intact gallbladder muscle strips showed that capacitative calcium entry induced contractions. The CCE-evoked contraction was sensitive to 2-APB, L-type blockers and Gd3+. We conclude that in GBSM release of [Ca2+]i from internal stores activates a capacitative Ca2+ entry pathway and depolarizes plasma membrane, allowing coactivation of voltage-operated L type Ca2+ channels. This process may play a role in excitation-contraction coupling in GBSM.
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