Glutamatergic and GABAergic Synapses in the Human Spinal Dorsal Horn Revealed With Immunohistochemistry

Abstract

Primary afferent neurons detect sensory stimuli in the periphery and transmit this information to the dorsal horn of the spinal cord, where it is processed by excitatory and inhibitory controls before being sent to the brain. Our understanding of the synaptic architecture of these spinal circuits in the rodent has been massively advanced using antibodies raised against scaffolding proteins Homer1 and gephyrin, which anchor glutamate and GABA receptors to the membrane, respectively. Few studies have attempted to visualize spinal cord synapses in human tissue, partly due to a lack of high-quality tissue with low postmortem intervals. In this study, we reveal both excitatory and inhibitory synapses at a high resolution in human lumbar spinal cord tissue using Homer1 and gephyrin immunolabeling and show that the basic organization of these proteins within the dorsal horn is similar to that in the rodent. Homer1+ puncta are highly colocalized with ionotropic glutamate receptors, and over 75% are in contact with a presynaptic axon terminal containing the vesicular glutamate transporter 2 (VGluT2). Similarly, most gephyrin+ profiles are coextensive with the GABA_A-α₂ subunit but fewer than 10% colocalize with Homer1+ puncta, confirming the specificity of these markers. Finally, we use Homer1 immunolabeling to demonstrate that primary afferents can form complex synaptic arrangements in human spinal cord. We conclude that these antibodies can be used as reliable tools for the study of human synaptic circuitry, and we have used them to reveal insight into the spinal connections underlying somatosensation that can be expanded upon in future studies.