Group III metabotropic glutamate receptors (mGluRs) are involved in nociceptive transmission in the spinal cord. However, the cellular mechanism underlying the modulation of synaptic transmission from nociceptive primary afferents to dorsal horn neurons by group III mGluRs has yet to be explored. In this study, we used transgenic mice expressing enhanced green fluorescent protein (EGFP) under the control of the glutamate decarboxylase (GAD) 65 promoter to identify specific subpopulations of GABAergic inhibitory interneurons. By GABA immunolabeling, we confirmed the majority of GAD65-EGFP-expressing neurons were GABAergic. Because GAD65-EGFP-expressing neurons have not been examined in detail before, we first investigated the physiological properties of GAD65-EGFP- and non-EGFP-expressing neurons in substantia gelatinosa (SG) of the spinal dorsal horn. Membrane properties, such as the resting membrane potential, membrane capacitance, action potential threshold, and action potential height, differed significantly between these two groups of neurons. Most EGFP-expressing neurons displayed a tonic firing pattern (73% of recorded neurons) and received monosynaptic Aδ and/or C primary afferent inputs (85% of recorded neurons). In contrast, we observed a delayed firing pattern in 53% of non-EGFP-expressing neurons. After identifying the physiological properties of EGFP-expressing neurons, we tested the effects of group III mGluRs on synaptic transmission pharmacologically. A group III mGluR agonist, L-AP4, attenuated Aδ fiber-evoked synaptic transmission but did not affect C fiber-evoked synaptic transmission to EGFP-expressing neurons. Similar primary afferent-specific inhibition by L-AP4 was also observed in non-EGFP-expressing neurons. Moreover, Aδ fiber-evoked synaptic transmission was suppressed by a selective mGluR7 agonist, AMN082. These results suggest that modulation of the synaptic transmission from primary afferents to SG neurons by group III mGluR agonist is specific to the type of nociceptive primary afferents but not to the type of target neurons.