PROTEIN-KINASE A MEDIATES ACTIVATION OF ATP-SENSITIVE K-MUSCLE( CURRENTS BY CGRP IN GALLBLADDER SMOOTH)

The signal transduction mechanisms underlying the activation of ATP-se nsitive potassium (K-ATP) current by calcitonin gene-related peptide ( CGRP) in gallbladder smooth muscle were examined with intracellular mi croelectrode recording and whole cell patch-clamp techniques. In the i ntact gallbladder preparation, the adenylyl cyclase activator forskoli n hyperpolarized the membrane potential and abolished spontaneous acti on potentials. This response was inhibited by the K-ATP channel blocke r glibenclamide. CGRP (10 nM), forskolin (10 mu M), the membrane-perme able adenosine 3',5'-cyclic monophosphate (cAMP) analogue adenosine 3' ,5'cyclic monophosphothioate (Sp-cAMP[S]; 500 mu M), and the catalytic subunit of protein kinase A (100 U/ml) activated glibenclamide-sensit ive currents in enzymatically dissociated gallbladder smooth muscle ce lls. CGRP activation of potassium currents was prevented by dialysis o f the cell cytoplasm with guanosine 5'-O-(2-thiodiphosphate) (5 mM) or a specific peptide inhibitor of protein kinase A (2.3 mu M). Okadaic acid (5 mu M), a phosphatase inhibitor, slowed the deactivation of the K-ATP current, following removal of CGRP. The results of this study i ndicate that CGRP hyperpolarizes gallbladder smooth muscle by elevatio n of cAMP and subsequent stimulation of protein kinase A.