Elsevier

Neuroscience

Volume 153, Issue 2, 2 May 2008, Pages 507-517
Neuroscience

Pain mechanism
Neurochemical characterization of neuronal populations expressing protein kinase C gamma isoform in the spinal cord and gracile nucleus of the rat

https://doi.org/10.1016/j.neuroscience.2008.01.082Get rights and content

Abstract

Protein kinase C gamma (PKCγ) is widely distributed throughout the CNS and is thought to play a role in long term hyper-excitability in nociceptive neurones. Here, we provide the first report of PKCγ cells in the dorsal column nuclei of the adult rat. Retrograde labeling of PKCγ cells from the thalamus with choleragenoid revealed that 25% of the PKCγ positive gracile cells projected to the thalamus. Further, we have characterized the distribution of PKCγ within gracile nucleus in terms of colocalization with various neurotransmitter receptors or enzymes and calcium binding proteins, and compared this with PKCγ colocalization in cells of laminae I–III of the spinal cord. We show that approximately 90% of the PKCγ cells in the gracile nucleus and 60% in the dorsal horn were immuno-positive for the AMPA receptor subunit glutamate 2/3 (GluR2/3). Little coexpression was seen with neurokinin 1 receptor, nitric oxide synthase (NOS) and the AMPA receptor subunit GluR1, markers of distinct neuronal subpopulations. In the spinal cord, a quarter of PKCγ cells expressed calbindin, but very few cells did so in the gracile nucleus.

Electrical stimulation at c-fiber strength of the normal or injured sciatic nerve was used to induce c-fos as a marker of postsynaptic activation in the spinal cord and gracile nucleus. Quantitative analysis of the number of PKCγ positive gracile cells that expressed also c-fos increased from none to 24% after injury, indicating an alteration in the sensory activation pattern in these neurones after injury. C-fos was not induced in inner lamina II following c-fiber electrical stimulation of the intact or axotomized sciatic nerve, indicating no such plasticity at the spinal cord level. As dorsal column nuclei cells may contribute to allodynia after peripheral nerve injury, pharmacological modulation of PKCγ activity may therefore be a possible way to ameliorate neuropathic pain after peripheral nerve injury.

Section snippets

Experimental procedures

All animal procedures were carried out in accordance with the UK Home Office Scientific Procedures Act (1986). Efforts were made to minimize the number of animals used and their suffering. A total of 24 rats were used in the present study. Ten adult Wistar rats were overdosed with sodium pentobarbital and perfused through the ascending aorta with 50 ml saline followed by 400 ml 4% paraformaldehyde in 0.1 M phosphate buffer at pH 7.4. The brainstem region containing the dorsal column nuclei and

General distribution of PKCγ in the medulla

PKCγ cells have been reported in the medullary dorsal horn of the spinal trigeminal nucleus and in the cochlear nucleus (Li et al 2001a, Li et al 2001b, Tanaka and Saito 1992). We also observed immunopositive neurons within these regions but also within the para-trigeminal and lateral reticular nuclei. Rostral to the obex of the medulla, especially strong terminal labeling was seen in the vestibular nuclei. Positive fibers were also seen in the corticospinal and trigeminal tracts and the medial

Discussion

Our study demonstrates the following new findings with regard to PKCγ neurons: Firstly, we have identified and characterized a population of neurons within the gracile nucleus in terms of neurotransmitter receptors and shown that a proportion project to the thalamus. Secondly, we show that some of these cells are capable of eliciting c-fos expression after peripheral nerve injury using c-fiber strength electrical stimulation. Thirdly, in the spinal cord, we have further characterized the PKCγ

Conclusion

In conclusion, this study has furthered the knowledge of PKCγ expressing cells by showing that more than 80% of spinal cord neurons expressed either the GluR1 or GluR2/3 AMPA receptor subunits, with a differential lamina distribution. Although PKCγ cells are known to play a role in chronic pain conditions little is known about the direct synaptic contacts to these cells. We suggest that glutamatergic input is likely to be very important, either from primary afferents or from local excitatory

Acknowledgments

This work was supported by the Wellcome Trust. A.S.H. was also the recipient of a Wellcome Trust Summer Vacation Scholarship.

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