Impulses in primary afferent nerve fibers may produce short-or long-lasting modifications in spinal nociception. Here we have identified a robust long-term depression (LTD) of synaptic transmission in substantia gelatinosa neurons that can be induced by low-frequency stimulation of primary afferent A␦-fibers. Synaptic transmission between dorsal root afferents and neurons in the substantia gelatinosa of the spinal cord dorsal horn was examined by intracellular recording in a transverse slice dorsal root preparation of rat spinal cord. Conditioning stimulation of dorsal roots with 900 pulses given at 1 Hz (10 V, 0.1 msec) produced LTD of EPSP amplitudes in substantia gelatinosa neurons to 41 Ϯ 10% of control that lasted for at least 2 hr. When A-and C-fibers were recruited, conditioning stimulation was as effective as A-fiber stimulation alone. After LTD, synaptic strength could be increased to its original level by applying a second, high-frequency tetanic stimulus to the dorsal root, indicating that LTD is reversible and not attributable to damage of individual synapses. Bath application of the GABA A receptor antagonist bicuculline and glycine receptor antagonist strychnine did not affect LTD. When NMDA receptors were blocked by bath application of D-2-amino-5-phosphonovaleric acid, LTD was abolished or strongly reduced. Loading substantia gelatinosa neurons with Ca 2ϩ chelator BAPTA also blocked or reduced LTD. After incubation of slices with calyculin A, a selective and membrane permeable inhibitor of protein phosphatases 1 and 2A, LTD was not attenuated. We propose that this form of LTD may be relevant for long-lasting segmental antinociception after afferent stimulation.