INCREASE IN CALCIUM IN SMOOTH-MUSCLE CELLS OF THE RABBIT BLADDER INDUCED BY ACETYLCHOLINE AND ATP

Cultured smooth muscle cells from rabbit urinary bladder were loaded w ith fura-2. Changes in intracellular Ca concentration [Ca-i(2+]) produ ced by acetylcholine (ACh) or adenosine triphosphate (ATP) were estima ted by measuring the fluorescence ratio F340/F380. Western blot analys is and immunohistochemical techniques showed that the cultured cells r etained alpha-smooth muscle actin. ATP produced a rapid but transient increase in [Ca2+](i) and ACh produced a delayed, prolonged increase. Application of ACh after ATP in Ca-free solution failed to elevate [Ca 2+](i) suggesting that both ACh and ATP release Ca2+ from the same int racellular stores. Following application of ACh but not ATP in Ca-free Krebs solution, reintroduction of Ca2+ produced elevation of [Ca2+](i ), indicating that ACh causes prolonged opening of channels in the mem brane. The sustained increase induced by ACh was abolished by nicardip ine (blocker of Ca2+ voltage dependent channel I-Ca(V)) or quinine (bl ocker of non-selective cation channels). Although the elevations to AC h or ATP were abolished by neomycin (an inhibitor of phospholipase C) the different time courses suggest that the mechanisms of release of C a2+ from intracellular stores or the pathway for refilling the stores is different. (C) 1998 Elsevier Science B.V. All rights reserved.