ATPgammaS, a nonhydrolyzable ATP analog, was found to dose-dependently generate an inward current at a holding potential of -70 mV (EC(50)=43 microM) in lamina IX neurons of rat spinal cord slices using the whole-cell patch-clamp technique. This inward current had an extrapolated reversal potential of -9 mV and was resistant to the Na(+)-channel blocker tetrodotoxin, glutamate-receptor antagonists or nominally Ca(2+)-free medium. ATP gamma S also increased the frequency and amplitude of glutamatergic spontaneous excitatory postsynaptic current (sEPSC); this action was dose-dependent and sensitive to tetrodotoxin. Unlike ATP gamma S, the P2X-receptor agonist, BzATP or alpha,beta-methylene ATP, did not change holding currents, but the current response produced by ATP gamma S disappeared in the presence of the P2-receptor antagonist PPADS. The sEPSC frequency and amplitude increase was observed with alpha,beta-methylene ATP, but not with the P2Y-receptor agonist, 2-methylthio ADP, UTP or UDP. The current response by ATP gamma S was suppressed by the addition of GDP beta S into the patch-pipette solution. As for ATP gamma S, 2-methylthio ADP produced an inward current, while UTP and UDP had no effect on holding currents. The P2Y(1)-receptor antagonist MRS2179 inhibited the ATP gamma S-induced inward current, but did not affect the sEPSC frequency and amplitude increase produced by ATP gamma S. These data indicate that extracellular ATP increases the excitability of lamina IX neurons by membrane depolarization (probably through non-selective cation-channel activation) and spontaneous excitatory transmission enhancement, which may be mediated by P2Y(1) and P2X receptors, respectively. This finding supports the idea that purinergic receptor antagonists provide a therapy for spinal cord injury.