Postsynaptic autophosphorylation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) at Thr286/287 is crucial for the induction of long-term potentiation at many glutamatergic synapses, and has also been implicated in the persistence of synaptic potentiation. However, the availability of CaMKII phosphorylated at Thr286/287 at individual glutamatergic synapses in vivo is unclear. We used post-embedding immunogold labelling to quantitatively analyse the ultrastructural localization of CaMKII phosphorylated at Thr286/287 (pCaMKII) at synapses formed by presumed nociceptive and low-threshold mechanosensitive primary afferent nerve endings in laminae I–IV of rat spinal cord. Immunogold labelling was enriched in the postsynaptic densities of such synapses, consistent with observations in pre-embedding immunoperoxidase-stained dorsal horn. Presynaptic axoplasm also exhibited sparse immunogold labelling, in peptidergic terminals partly associated with dense core vesicles. Analysis of single or serial pCaMKII-immunolabelled sections indicated that the large majority of synapses formed either by presumed peptidergic or non-peptidergic nociceptive primary afferent terminals in laminae I–II of the spinal cord, or by presumed low-threshold mechanosensitive primary afferent terminals in laminae IIi–IV, contained pCaMKII in their postsynaptic density. However, the postsynaptic levels of pCaMKII immunolabelling at low-threshold primary afferent synapses were only ∼ 50% of those at nociceptive synapses. These results suggest that constitutively autophosphorylated CaMKII in the postsynaptic density is a common characteristic of glutamatergic synapses, thus potentially contributing to maintenance of synaptic efficacy. Furthermore, pCaMKII appears to be differentially regulated between high- and low-threshold primary afferent synapses, possibly reflecting different susceptibility to synaptic plasticity between these afferent pathways.