Activation of small conductance Ca(2+)-dependent K+ channels by purinergic agonists in smooth muscle cells of the mouse ileum.
1. Whole-cell and single-channel K+ currents were recorded at room temperature (22-24 degrees C), from smooth muscle cells enzymatically dispersed from the mouse ileum, using variations of the patch-clamp technique. 2. Net outward K+ currents recorded through amphotericin-B-perforated patches in response to step depolarizations positive to -50 mV from a holding potential of -80 mV were decreased by up to 70% by external apamin (0.5 microM). Apamin-sensitive whole-cell currents were also recorded from cells perfused internally with 150 nM Ca2+ but not from cells perfused internally with 85 nM Ca2+. 3. Three types of non-inactivating Ca(2+)-sensitive K+ channels were identified in cell-attached and excised patches under an asymmetrical K+ gradient: (i) large conductance (BKCa; approximately 200 pS) channels blocked by 2 mM external TEA; (ii) intermediate conductance (IKCa; approximately 39 pS) channels blocked by 2 mM external TEA and inhibited by external apamin (0.5 microM); and (iii) small conductance (SKCa; approximately 10 pS) channels that were not blocked by 5 mM external TEA but were sensitive to extracellular apamin (0.5 microM). 4. The TEA-resistant SKCa channels were activated by an increase in [Ca2+]i with an EC50 of 1.5 microM and a Hill coefficient of 1.3. 5. P2 purinoceptor agonists 2-methylthioATP (2-MeSATP), 2-chloroATP and ATP (10-50 microM) increased an apamin-sensitive whole-cell outward K+ current. Extrapatch application of 2-MeSATP (20-100 microM) stimulated the apamin-sensitive IKCa and SKCa channels and activated an apamin-sensitive steady outward current at 0 mV. 6. Smooth muscle cells from the mouse ileum possess two apamin-sensitive K+ channels (IKCa and SKCa); of these, the IKCa channels are TEA sensitive while the SKCa channels are TEA resistant. These channels, along with an apamin-sensitive but TEA-resistant steady outward current, may mediate membrane hyperpolarization elicited by purinergic agonists.
Vogalis
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Centre for Swallowing and Motility Disorders, Brockton/West Roxbury Veterans Administration Medical Center, MA 02132, USA. vogalis@brockton.va.gov
Goyal
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RK
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001030:Apamin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D007082:Ileum / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology; D015640:Ion Channel Gating / Q000502:physiology; D051379:Mice; D009130:Muscle, Smooth / Q000737:chemistry* / Q000166:cytology / Q000187:drug effects; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000502:physiology*; D058913:Purinergic Agonists; D019789:Tetraethylammonium / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology
D015221:Potassium Channels; D058913:Purinergic Agonists; D013873:Thionucleotides; D001030:Apamin; C006979:2-chloro-ATP; D019789:Tetraethylammonium; D000255:Adenosine Triphosphate; D002118:Calcium; C008501:2-methylthio-ATP