title: Direct block of native and cloned (Kir2.1) inward rectifier K+ channels by chloroethylclonidine.
abstract: 1. We have investigated the inhibition of inwardly rectifying potassium channels by the alpha-adrenergic agonist/antagonist chloroethylclonidine (CEC). We used two preparations; two-electrode voltage-clamp of rat isolated flexor digitorum brevis muscle and whole-cell patch-clamp of cell lines transfected with Kir2.1 (IRK1). 2. In skeletal muscle and at a membrane potential of -50 mV, chloroethylclonidine (CEC), an agonist at alpha2-adrenergic receptors and an antagonist at alpha1x-receptors, was found to inhibit the inward rectifier current with a Ki of 30 microM. 3. The inhibition of skeletal muscle inward rectifier current by CEC was not mimicked by clonidine, adrenaline or noradrenaline and was not sensitive to high concentrations of alpha1-(prazosin) or alpha2-(rauwolscine) antagonists. 4. The degree of current inhibition by CEC was found to vary with the membrane potential (approximately 70% block at -50 mV c.f. approximately 10% block at -190 mV). The kinetics of this voltage dependence were further investigated using recombinant inward rectifier K+ channels (Kir2.1) expressed in the MEL cell line. Using a two pulse protocol, we calculated the time constant for block to be approximately 8 s at 0 mV, and the rate of unblock was described by the relationship tau=exp((Vm+149)/22) s. 5. This block was effective when CEC was applied to either the inside or the outside of patch clamped cells, but ineffective when a polyamine binding site (aspartate 172) was mutated to asparagine. 6. The data suggest that the clonidine-like imidazoline compound, CEC, inhibits inward rectifier K+ channels independently of alpha-receptors by directly blocking the channel pore, possibly at an intracellular polyamine binding site.
authors.affiliation: Ion Channel Group, Department of Cell Physiology and Pharmacology, Leicester University, PO Box 138, Leicester LE1 9HN. R.Barret-Jolley@bham.ac.ujk
authors.forename: R
authors.initials: R
authors.lastname: Barrett-Jolley
authors.forename: C
authors.initials: C
authors.lastname: Dart
authors.forename: N B
authors.initials: NB
authors.lastname: Standen
mesh_terms: D000818:Animals; D003000:Clonidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D007700:Kinetics; D026902:Potassium Channel Blockers; D024661:Potassium Channels, Inwardly Rectifying; D051381:Rats; D011942:Receptors, Adrenergic, alpha / Q000502:physiology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors
chemical_list: D026902:Potassium Channel Blockers; D024661:Potassium Channels, Inwardly Rectifying; D011942:Receptors, Adrenergic, alpha; D011994:Recombinant Proteins; C051731:chlorethylclonidine; D003000:Clonidine