The dorsal root reflex (DRR) is an antidromic action potential originating in the spinal cord that propagates toward the periphery. Given that both GABA A and 5-HT 3 receptors are involved in the generation of DRRs and stimulation of the periaqueductal gray (PAG) can induce the release of GABA and serotonin within the spinal cord, we investigated the modulation of DRRs by the PAG descending system. The central end of the cut left L5 dorsal root in adult Sprague–Dawley rats was tested with single fiber recording. Stimulating electrodes were placed in the PAG, sciatic nerve, or transcutaneously across hindpaws. Fifty-seven DRRs were recorded for the effect of PAG stimulation in 19 rats, and 51 DRRs from 26 rats and nine DRRs from seven rats were recorded for an effect of ipsilateral and contralateral peripheral stimulation, respectively. The results were expressed as a percentage of the number of DRRs over the number of stimuli. PAG stimulation at 0.2, 0.5, 5, 20, and 50 Hz produced ratio’s of 113.16±9.84, 114.54±12.22, 24.6±3.23, 17.77±4.76, and 12.62±3.44 (%), respectively. Stimulation at ipsilateral peripheral nerve evoked DRRs of 103.26±8.93, 95.27±10.57, 37.66±7.55, 11.32±4.96, and 5.32±3.82 (%), respectively. Stimulation of the contralateral peripheral nerve evoked DRRs of 90.88±15.59, 44.30±10.77, 6.29±1.63, 0.45±0.19, and 0.29±0.15 (%), respectively. Transection at the thoracic spinal level completely eliminated PAG-induced DRRs. In conclusion, both PAG and peripheral stimulation produced DRRs in a frequency dependent manner. Stimulus intensity has no significant effect on DRRs.