AbstractIonotropic glutamate receptors (IGR), including NMDA, AMPA, and kainate receptors, are expressed in terminals with varied morphology in the superficial laminae (I–III) of the dorsal horn of the spinal cord. Some of these terminals can be identified as endings of primary afferents, whereas others establish symmetric synapses, suggesting that they may be γ‐aminobutyric acid (GABA)‐ergic. In the present study, we used confocal and electron microscopy of double immunostaining for GAD65, a marker for GABAergic terminals, and for subunits of IGRs to test directly whether IGRs are expressed in GABAergic terminals in laminae I–III of the dorsal horn. Although colocalization is hard to detect with confocal microscopy, electron microscopy reveals a substantial number of terminals immunoreactive for GAD65 also stained for IGRs. Among all GAD65‐immunoreactive terminals counted, 37% express the NMDA receptor subunit NR1; 28% are immunopositive using an antibody for the GluR2/4 subunits of the AMPA receptor; and 20–35% are immunopositive using antibodies for the kainate receptor subunits GluR5, GluR6/7, KA1, or KA2. Terminals immunoreactive for IGR subunits and GAD65 establish symmetric synapses onto dendrites and perikarya and can be presynaptic to primary afferent terminals within both type 1 and type 2 synaptic glomeruli. Activation of presynaptic IGR may reduce neurotransmitter release. As autoreceptors in terminals of Aδ and C afferent fibers in laminae I–III, presynaptic IGRs may play a role in inhibiting nociception. As heteroreceptors in GABAergic terminals in the same laminae, on the other hand, presynaptic IGRs may have an opposite role and even contribute to central sensitization and hyperalgesia. J. Comp. Neurol. 486:169–178, 2005. © 2005 Wiley‐Liss, Inc.