ATP Evokes Inward Currents in Corpus Cavernosum Myocytes

摘要

Introduction: Although adenosine triphosphate (ATP) has often been reported to relax the corpus cavernosum, this may be mediated by indirect effects, such as release of nitric oxide from the endothelium. Recent data suggest that P2X1 receptors may be up-regulated in diabetes, and these exert an anti-erectile effect by causing the corpus cavernosum smooth muscle cells (CCSMCs) to contract. However, to date, there is no functional evidence that ATP can directly stimulate CCSMC. Aims: This study aims to (i) to directly examine the effect of ATP on membrane currents in freshly isolated CCSMC, where influences of endothelium and other cells are absent; and (ii) to determine the receptor subtypes, ionic currents, and Ca2+ signals stimulated by ATP. Methods: CCSMCs were enzymatically dispersed from male New Zealand White rabbits for patch clamp recording and measurement of intracellular Ca2+ in fluo-4-loaded cells using spinning disk confocal microscopy. Main Outcome Measures: Patch clamp recordings were made of ATP-evoked membrane currents and spontaneous membrane currents. Spinning disk confocal imaging of intracellular Ca2+ was performed, and the response to ATP was recorded. Results: ATP evoked repeatable inward currents in CCSMC (1st application: -675±101pA; 2nd application: -694±120pA, N=9, P=0.77). ATP-induced currents were reduced by suramin from -380±121 to -124±37pA (N=8, P0.05), by α,β-methylene ATP from -755±235 to 139±49pA (N=5, P0.05), and by NF449 from -419±to -51±13pA (N=6, P0.05). In contrast, MRS2500, a P2Y1 antagonist, had no effect on ATP responses (control: -838±139pA; in MRS2500: -822±184pA, N=13, P=0.84) but blocked inward currents evoked by 2-MeSATP, a P2Y1,12,13agonist (control: -623±166pA; in MRS2500: -56±25pA, N=6, P0.05). The ATP-evoked inward current was unaffected by changing the transmembrane Cl- gradient but reversed in direction when extracellular Na+ was reduced, indicating that it was a cation current. Conclusions: ATP directly stimulates CCSMC by evoking a P2X-mediated cation current.