title: Paxilline inhibition of the alpha-subunit of the high-conductance calcium-activated potassium channel.
abstract: High conductance calcium-activated (maxi-K) channels are potently blocked by a family of indole diterpenes that includes paxilline. Paxilline stimulates binding of charybdotoxin (ChTX) to maxi-K channels in vascular smooth muscle and blocks these channels in electrophysiological experiments (Knaus et al., 1994b). These results suggested that paxilline blocked maxi-K channels at a site distance from the ChTX binding site located near the external entrance to the pore. Here we have examined block of the cloned alpha subunit (slo) of the maxi-K channel in excised membrane patches after internal application of paxilline. Paxilline caused a reversible inhibition of channel currents with slow washout kinetics. In the presence of 10 muM intracellular calcium, paxilline blocked currents elicited by brief voltage pulses with a Ki of 1.9 nM and a Hill coefficient near one. Changing the internal calcium by the fold caused a two to three fold change in the Ki for paxilline block, with less block occurring at high calcium concentrations. Paxilline reduced the maximum of the conductance-voltage relation in a calcium-sensitive manner with less block occurring at high calcium concentrations, and caused a 20 mV depolarizing shift in the midpoint for channel opening. The time-course of relief of paxilline block by elevated calcium was more rapid than washout of paxilline suggesting an allosteric interaction between calcium and paxilline.
authors.affiliation: Department of Membrane Biochemistry and Biophysics, Merck Research Labs, Rahway, NJ 07065, USA.
authors.forename: M
authors.initials: M
authors.lastname: Sanchez
authors.forename: O B
authors.initials: OB
authors.lastname: McManus
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D007211:Indoles / Q000494:pharmacology*; D015640:Ion Channel Gating / Q000187:drug effects; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000096:biosynthesis / Q000378:metabolism; D024681:Potassium Channels, Calcium-Activated; D013997:Time Factors; D014981:Xenopus
chemical_list: D007211:Indoles; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024681:Potassium Channels, Calcium-Activated; C048220:paxilline; D002118:Calcium