The effects of nitric oxide-containing solution and different nitric oxide donors were investigated on spontaneously active neurons using extracellular recording technique in areas of rat spinal cord slices where high levels of nitric oxide synthase are present. In lamina X, 93% of all neurons investigated (n = 84) increased their firing rate and 2% decreased it by superfusion with the nitric oxide donor sodium nitroprusside. In contrast, 49% of all neurons in laminae I and II (n = 90) were inhibited and only 28% were activated. Both effects were due to the postsynaptic action of sodium nitroprusside, because they could still be observed in medium containing 0.3 mM Ca2+ and 9 mM Mg2+, known to block synaptic transmission. Application of 8-bromo-cyclic-GMP caused an excitation of every neuron which was excited by sodium nitroprusside and an inhibition of every cell which was inhibited by sodium nitroprusside (n = 25). This effect was different from the effect of 8-bromo-cyclic-AMP, which mimicked only the excitatory, but not the inhibitory response of sodium nitroprusside. These results provide evidence that nitric oxide in the spinal cord can directly cause an excitation or an inhibition of the electrical activity of spinal neurons. Another, more general conclusion from our results is that the nitric oxide-induced production of cyclic-GMP alone does not allow any prediction about an excitatory or inhibitory effect on the neuronal activity, which has to be determined separately.