ATP activated currents were studied in Leydig cells of mice using the patch-clamp technique. Whole-cell currents were rapidly activating and slowly desensitizing (55 % decrement from the peak value, upon exposure to 100 µM ATP for 60 s), requiring 3 minutes of washout to recover 100 % of the response. The concentration-response relationships for ATP, ATPS and 2MeSATP were described by the Hill equation with a K_D of 44µM, 110 µM and 643 µM, respectively, and n equal 2. The order of efficacy of agonists was: ATPATPS>2MeSATP>BzATP. MeATP, GTP, UTP, cAMP and adenosine were ineffective. Suramin and PPADS blocked the responses in a concentration dependent manner. The ATP activated currents were dependent on the extracellular pH, being maximal at pH 6.5 and decreased with both acidification and alkalinization (apparent pKs of 5.9 and 7.4, respectively). The whole-cell current-voltage relationship showed inward rectification and reversed near 0 mV. Experiments performed in bi-ionic conditions for measurement of reversal potentials showed that this channel is highly permeable to calcium (P_Ca/P_Na = 5.32), but not so to chloride (P_Cl/P_Na = 0.03) or NMDG (P_NMDG/P_Na = 0.09). Unitary currents, recorded in outside-out patches, had a chord conductance of 27 pS (between -90 and -50 mV) and were inward rectifying. The average current passing through the excised patch decreased with time (= 13 s), resembling desensitization of the macroscopic current. These findings indicate that the ATP receptor present in Leydig cells shows properties most similar to that of cloned homomeric P2X₂.