A quantitative study of neurons which express neurokinin-1 or somatostatin sst2a receptor in rat spinal dorsal horn.

The neurokinin-1 and somatostatin sst2a receptors have both been identified on spinal cord neurons. In this study we have estimated the proportions of neurons in different parts of the spinal cord which express these receptors, by using a monoclonal antibody against a neuronal nuclear protein named NeuN and combining the optical disector method with confocal microscopy. The NeuN antibody was initially tested on over 3200 neurons identified with antisera against a variety of compounds, including neuropeptides, enzymes and receptors, and also on astrocytes and oligodendrocytes. All of the neurons, but none of the glial cells that were examined possessed NeuN-immunoreactivity, which suggests that NeuN is a reliable marker for all spinal cord neurons. We found that approximately 45% of neurons in lamina I, 23-29% of those in laminae IV-VI and 18% in lamina X possessed the neurokinin-1 receptor, while the receptor was present on a smaller proportion of neurons in laminae II and III (6% and 11%, respectively). Thirteen percent of lamina I neurons and 15% of those in lamina II expressed the sst2a receptor. To provide further information about the types of neuron which possess the sst2a receptor, we searched for possible co-existence with the neurokinin-1 receptor as well as with GABA and glycine. sst2a and neurokinin-1 receptors were not co-localized on neurons in laminae I and II. All of the sst2a-immunoreactive neurons examined were also GABA-immunoreactive, and 83.5% were glycine-immunoreactive, indicating that the receptor is located on inhibitory neurons in the superficial dorsal horn. These results demonstrate the proportions of neurons in each region of the spinal cord which can be directly activated by substance P or somatostatin acting through these receptors. Levels of receptors can change in pathological states, and this method could be used to determine whether or not these changes involve alterations in the number of neurons which express receptors. In addition, the method can be used to estimate the sizes of neurochemically-defined populations of spinal cord neurons.