Morales Sara, Camello Pedro J, Alcón Soledad, Salido Ginés M, Mawe Gary, Pozo María J
Deprtment of Physiology, University of Extremadura, Caceres, Spain.
Am J Physiol Gastrointest Liver Physiol. 2004 Jun;286(6):G1090-100. doi: 10.1152/ajpgi.00260.2003. Epub 2004 Jan 22.
We have evaluated the presence of capacitative Ca(2+) entry (CCE) in guinea pig gallbladder smooth muscle (GBSM), including a possible relation with activation of L-type Ca(2+) channels. Changes in cytosolic Ca(2+) concentration induced by Ca(2+) entry were assessed by digital microfluorometry in isolated, fura 2-loaded GBSM cells. Application of thapsigargin, a specific inhibitor of the Ca(2+) store pump, induced a transient Ca(2+) release followed by sustained entry of extracellular Ca(2+). Depletion of the stores with thapsigargin, cyclopiazonic acid, ryanodine and caffeine, high levels of the Ca(2+)-mobilizing hormone cholecystokinin octapeptide, or simple removal of external Ca(2+) resulted in a sustained increase in Ca(2+) entry on subsequent reapplication of Ca(2+). This entry was attenuated by 2-aminoethoxydiphenylborane, L-type Ca(2+) channel blockade, pinacidil, and Gd(3+). Accumulation of the voltage-sensitive dye 3,3'-dipentylcarbocyanine and direct intracellular recordings showed that depletion of the stores is sufficient for depolarization of the plasma membrane. Contractility studies in intact gallbladder muscle strips showed that CCE induced contractions. The CCE-evoked contraction was sensitive to 2-aminoethoxydiphenylborane, L-type Ca(2+) channel blockers, and Gd(3+). We conclude that, in GBSM, release of Ca(2+) from internal stores activates a CCE pathway and depolarizes plasma membrane, allowing coactivation of voltage-operated L-type Ca(2+) channels. This process may play a role in excitation-contraction coupling in GBSM.
我们评估了豚鼠胆囊平滑肌（GBSM）中容量性Ca(2+)内流（CCE）的存在情况，包括其与L型Ca(2+)通道激活之间的可能关系。通过数字显微荧光测定法，在分离的、负载fura 2的GBSM细胞中评估了由Ca(2+)内流引起的胞质Ca(2+)浓度变化。应用毒胡萝卜素（一种Ca(2+)储存泵的特异性抑制剂）可诱导Ca(2+)的瞬时释放，随后细胞外Ca(2+)持续内流。用毒胡萝卜素、环匹阿尼酸、ryanodine和咖啡因耗尽储存库，高水平的Ca(2+)动员激素胆囊收缩素八肽，或简单去除细胞外Ca(2+)，都会导致在随后重新添加Ca(2+)时Ca(2+)内流持续增加。这种内流会被2-氨基乙氧基二苯硼烷、L型Ca(2+)通道阻滞剂、吡那地尔和Gd(3+)减弱。电压敏感染料3,3'-二戊基羰花青的积累和直接细胞内记录表明，储存库的耗尽足以使质膜去极化。完整胆囊肌条的收缩性研究表明，CCE可诱导收缩。CCE诱发的收缩对2-氨基乙氧基二苯硼烷、L型Ca(2+)通道阻滞剂和Gd(3+)敏感。我们得出结论，在GBSM中，Ca(2+)从内部储存库的释放激活了CCE途径并使质膜去极化，从而使电压门控L型Ca(2+)通道共同激活。这一过程可能在GBSM的兴奋-收缩偶联中起作用。
