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 ionchannels

1 Immunoreactivity for P2X(1), P2X(4) and P2X(5) receptor subtypes was dete cted in the smooth muscle cell layer of second and third order rat mesenter ic arteries immunoreactivity, for P2X(2), P2X(3), P2X(6) and P2X(7) recepto rs was below the level of detection in the smooth muscle layer. 2 P2X receptor-mediated currents were recorded in patch clamp studies on ac utely dissociated mesenteric artery smooth muscle cells. Purinergic agonist s evoked transient inward currents that decayed rapidly in the continued pr esence of agonist (iota similar to 200 ms). Standard whole cell responses t o repeated applications of agonist at 5 min intervals ran down. Run-down wa s unaffected by changes in extracellular calcium concentration, intracellul ar calcium buffering or the inclusion of ATP and GTP in the pipette solutio n. 3 Run-down was overcome and reproducible responses to purinergic agonists w ere recorded using the amphotericin permeabilized patch recording configura tion. 4 The rank order of potency at the P2X receptor was ATP=2 methylthio ATP>al pha,beta -methylene ATP>CTP = 1-beta,gamma -methylene ATP. Only ATP and 2me SATP were full agonists. The P2 receptor antagonists suramin and PPADS inhi bited P2X receptor-mediated currents with IC(50)s of 4 muM and 70 nM respec tively. 5 These results provide further characterization of artery P2X receptors an d demonstrate that the properties are dominated by a P2X(1)-like receptor p henotype. No evidence could be found for a phenotype corresponding to homom eric P2X(4) or P2X(5) receptors or to heteromeric P2X(1/5) receptors and th e functional role of these receptors in arteries remains unclear.