title: Nitroblue tetrazolium blocks BK channels in cerebrovascular smooth muscle cell membranes.
abstract: The effects of p-nitroblue tetrazolium (NBT) on large conductance, calcium-activated potassium channels (BK channels) in enzymatically dispersed rat cerebrovascular smooth muscle cells (CVSMCs) were examined. Patch clamp methods were employed to record single BK channel currents from inside-out patches of CVMC membrane maintained at 21 - 23 degrees C. When applied to the cytoplasmic face of inside-out membrane patches (internally applied NBT), micromolar concentrations of NBT reversible reduced the mean open time of BK channels, without changing channel conductance. NBT altered the frequency distribution of BK channel open times from a two exponential to a single exponential form. In the absence of NBT, mean channel open time increased on membrane depolarization. In the presence of internally applied NBT, mean channel open became essentially independent of membrane potential. Internally applied NBT also reduced the mean closed time of BK channels when measured at membrane potentials in the range -80 mV to +20 mV. The combined effects of internal NBT on mean open and closed times resulted in the suppression of BK channel open probability when measured at positive membrane potentials. When applied to the external membrane face, micromolar concentrations of NBT reduced mean channel open time progressively as the membrane was hyperpolarized, and also reduced open probability at negative membrane potentials. A model is proposed in which NBT alters channel gating by binding to a site at or near to the cytoplasmic membrane face. Externally applied NBT suppressed BK channel open probability at concentrations which also inhibit nitric oxide synthase (NOS). Therefore, the potential role of potassium channel block in NBT actions previously attributed to NOS inhibition is discussed.
authors.affiliation: Department of Physiology, University of British Columbia, 2146 Health Sciences Mall, Vancouver, B.C. V6T 1Z3, Canada.
authors.forename: D
authors.initials: D
authors.lastname: Ye
authors.forename: J A
authors.initials: JA
authors.lastname: Pospisilik
authors.forename: D A
authors.initials: DA
authors.lastname: Mathers
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002536:Cerebral Arteries / Q000166:cytology / Q000187:drug effects; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects; D007700:Kinetics; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008564:Membrane Potentials / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D009580:Nitroblue Tetrazolium / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers; D024681:Potassium Channels, Calcium-Activated; D051381:Rats; D017208:Rats, Wistar
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D024681:Potassium Channels, Calcium-Activated; D009580:Nitroblue Tetrazolium