title: Large-conductance voltage- and Ca2+-activated K+ channel regulation by protein kinase C in guinea pig urinary bladder smooth muscle.
abstract: Large-conductance voltage- and Ca(2+)-activated K(+) (BK) channels are critical regulators of detrusor smooth muscle (DSM) excitability and contractility. PKC modulates the contraction of DSM and BK channel activity in non-DSM cells; however, the cellular mechanism regulating the PKC-BK channel interaction in DSM remains unknown. We provide a novel mechanistic insight into BK channel regulation by PKC in DSM. We used patch-clamp electrophysiology, live-cell Ca(2+) imaging, and functional studies of DSM contractility to elucidate BK channel regulation by PKC at cellular and tissue levels. Voltage-clamp experiments showed that pharmacological activation of PKC with PMA inhibited the spontaneous transient BK currents in native freshly isolated guinea pig DSM cells. Current-clamp recordings revealed that PMA significantly depolarized DSM membrane potential and inhibited the spontaneous transient hyperpolarizations in DSM cells. The PMA inhibitory effects on DSM membrane potential were completely abolished by the selective BK channel inhibitor paxilline. Activation of PKC with PMA did not affect the amplitude of the voltage-step-induced whole cell steady-state BK current or the single BK channel open probability (recorded in cell-attached mode) upon inhibition of all major Ca(2+) sources for BK channel activation with thapsigargin, ryanodine, and nifedipine. PKC activation with PMA elevated intracellular Ca(2+) levels in DSM cells and increased spontaneous phasic and nerve-evoked contractions of DSM isolated strips. Our results support the concept that PKC activation leads to a reduction of BK channel activity in DSM via a Ca(2+)-dependent mechanism, thus increasing DSM contractility.
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina.
authors.forename: Kiril L
authors.initials: KL
authors.lastname: Hristov
authors.forename: Amy C
authors.initials: AC
authors.lastname: Smith
authors.forename: Shankar P
authors.initials: SP
authors.lastname: Parajuli
authors.forename: John
authors.initials: J
authors.lastname: Malysz
authors.forename: Georgi V
authors.initials: GV
authors.lastname: Petkov
mesh_terms: D000818:Animals; D020013:Calcium Signaling; D004558:Electric Stimulation; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D005071:Evoked Potentials; D006168:Guinea Pigs; D015640:Ion Channel Gating; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology* / Q000294:innervation; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism*; D026902:Potassium Channel Blockers / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D013997:Time Factors; D001743:Urinary Bladder / Q000187:drug effects / Q000201:enzymology* / Q000294:innervation
chemical_list: D020536:Enzyme Activators; D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D011493:Protein Kinase C; D011188:Potassium
keywords: detrusor; phorbol 12-myristate 13-acetate; ryanodine receptors