The activation of urocortin immunoreactive neurons in the Edinger-Westphal nucleus following acute pain stress in rats

Urocortin, a member of the corticotropin releasing factor (CRF) peptide family, has a 45% sequence identity to CRF. Urocortin is ten-times more potent than CRF in increasing cAMP in cells expressing the CRF₂ receptor, therefore it was postulated to be an endogenous ligand for this receptor. Urocortin possesses the biological activity of CRF, and by activating the CRF₂ receptors, it can directly affect autonomic functions and play an important role in modifying the efferent components of endocrine, immune and behavioral responses to stress. Although urocortin's distribution in the rat brain has been described, with the most abundant urocortin-ir perikarya present in the Edinger-Westphal nucleus (E-WN), little is known about the physiological significance of brain urocortin. Since immediate early gene expression is seen in several midbrain regions, such as in the E-WN, following acute stress, we hypothesized that acute pain stress can result in the activation of the urocortinergic neurons in the E-WN. Fos immunoreactivity, the protein product of the immediate early gene c-fos, was used as a marker of cellular activity. Double-label immunohistochemical and double label immunofluorescence techniques were used in an acute pain stress model to reveal the colocalization of Fos-immunopositivity with urocortin-immunoreactivity (ir) within the E-WN. Our results showed that acute pain stress resulted in the activation of urocortin-ir neurons in the E-WN, peaking at 4 h after acute pain stress, based on the colocalization of Fos-ir with urocortin-ir, and the upregulation of urocortin mRNA transcripts in the E-WN. Based on these results, we suggest that the E-WN is a brain area that shows sustained activation by a painful stressor.