Lack of run-down of smooth muscle P2X receptor currents recorded with the amphotericin permeabilized patch technique, physiological and pharmacological characterization of the properties of mesenteric artery P2X receptor ion channels.
Immunoreactivity for P2X(1), P2X(4) and P2X(5) receptor subtypes was detected in the smooth muscle cell layer of second and third order rat mesenteric arteries immunoreactivity, for P2X(2), P2X(3), P2X(6) and P2X(7) receptors was below the level of detection in the smooth muscle layer. P2X receptor-mediated currents were recorded in patch clamp studies on acutely dissociated mesenteric artery smooth muscle cells. Purinergic agonists evoked transient inward currents that decayed rapidly in the continued presence of agonist (tau approximately 200 ms). Standard whole cell responses to repeated applications of agonist at 5 min intervals ran down. Run-down was unaffected by changes in extracellular calcium concentration, intracellular calcium buffering or the inclusion of ATP and GTP in the pipette solution. Run-down was overcome and reproducible responses to purinergic agonists were recorded using the amphotericin permeabilized patch recording configuration. The rank order of potency at the P2X receptor was ATP=2 methylthio ATP>alpha, beta-methylene ATP>CTP=l-beta,gamma-methylene ATP. Only ATP and 2meSATP were full agonists. The P2 receptor antagonists suramin and PPADS inhibited P2X receptor-mediated currents with IC(50)s of 4 microM and 70 nM respectively. These results provide further characterization of artery P2X receptors and demonstrate that the properties are dominated by a P2X(1)-like receptor phenotype. No evidence could be found for a phenotype corresponding to homomeric P2X(4) or P2X(5) receptors or to heteromeric P2X(1/5) receptors and the functional role of these receptors in arteries remains unclear.
Lewis
C J
CJ
Department of Cell Physiology & Pharmacology, Medical Sciences Building, University of Leicester, University Road, Leicester, LE1 9HN.
Evans
R J
RJ
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000666:Amphotericin B / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006160:Guanosine Triphosphate / Q000494:pharmacology; D007150:Immunohistochemistry; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology*; D008638:Mesenteric Arteries / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D009131:Muscle, Smooth, Vascular / Q000737:chemistry / Q000166:cytology / Q000502:physiology*; D018408:Patch-Clamp Techniques; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000032:analysis / Q000187:drug effects / Q000502:physiology*; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D058848:Receptors, Purinergic P2X4; D058485:Receptors, Purinergic P2X5; D058486:Receptors, Purinergic P2X7; D013498:Suramin / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology; D013997:Time Factors
D007473:Ion Channels; C546403:P2rx2 protein, rat; C546410:P2rx3 protein, rat; C548228:P2rx4 protein, rat; C546467:P2rx5 protein, rat; C546470:P2rx7 protein, rat; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D058848:Receptors, Purinergic P2X4; D058485:Receptors, Purinergic P2X5; D058486:Receptors, Purinergic P2X7; D013873:Thionucleotides; C110590:purinoceptor P2X6; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; D011732:Pyridoxal Phosphate; D013498:Suramin; D000666:Amphotericin B; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium; C008501:2-methylthio-ATP