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  • Title: Muscarinic suppression of Ca2+ current in smooth muscle cells of the guinea-pig urinary bladder.
    Author: Yoshino M, Yabu H.
    Journal: Exp Physiol; 1995 Jul; 80(4):575-87. PubMed ID: 7576597.
    Abstract:
    The suppressive action of carbachol (CCh) on the Ca2+ current (ICa) in smooth muscle cells of the guinea-pig urinary bladder was investigated using the whole-cell patch clamp technique. Bath application of 10 microM CCh reduced the amplitude of ICa by 92 +/- 3.8% (n = 9). Adding 1 microM atropine to the bath completely blocked the action of CCh, indicating that the suppressive action of CCh on ICa is mediated by the activation of muscarinic receptors. Intracellular perfusion of the non-hydrolysable GTP analogue, guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; 200 microM) mimicked the effects of CCh. Sustained suppression of ICa was observed when GTP gamma S was present in the cytoplasm. Intracellular perfusion of inositol 1,4,5-trisphosphate (InsP 3; 20 microM) also suppressed ICa; its effect was not sustained but transient. The protein kinase C activator, phorbol 12,13-dibutyrate (PDBu), however, could not mimic the effects of CCh on ICa. When intracellular Ca2+ was strongly buffered by the Ca2+ chelator EGTA (20 mM) in the patch pipette, the sustained suppression of ICa was abolished. Inclusion of 3 mg/ml heparin, a blocker of InsP3-induced Ca2+ release, in the patch pipette reduced the degree of sustained ICa suppression by 43.2 +/- 1.9% (n = 7). Adding thapsigargin (TG), a sarcoplasmic reticulum Ca2+-ATPase inhibitor, to a wash solution reduced the recovery of ICa by about 50%, suggesting that approximately half of the ICa suppression induced by CCh is due to Ca2+ release from TG-sensitive internal Ca2+ stores. From these results it appears that CCh suppresses ICa via two independent mechanisms: (1) Ca(2+)-mediated inactivation of the Ca2+ channel, which is caused by Ca2+ release from InsP3- and TG-sensitive internal stores, and (2) a GTP-binding protein-mediated mechanism, which requires intracellular Ca2+.
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