Functional and anatomical characterization of corticotropin-releasing factor receptor subtypes of the rat spinal cord involved in somatic pain relief

Abstract

Corticotropin-releasing factor (CRF) orchestrates our body’s response to stressful stimuli. Pain is often stressful and counterbalanced by activation of CRF receptors along the nociceptive pathway although the involvement of the CRF receptor subtypes 1 and/or 2 (CRF-R1 and CRF-R2, respectively) in CRF-induced analgesia remains controversial. This study examined CRF-R1 and CRF-R2 expression within the spinal cord of rats with Freund’s complete adjuvant-induced hindpaw inflammation using reverse transcriptase polymerase chain reaction, Western blot, radioligand binding, and immunofluorescence confocal analysis. Moreover, paw pressure algesiometry examined antinociceptive effects of intrathecal (i.t.) CRF and their possible antagonism through CRF-R1 and/or CRF-R2 selective antagonists as well as the opioid receptor antagonist naloxone. Our results demonstrated predominantly CRF-R2 mRNA, protein, binding sites and immunoreactivity in the dorsal horn of the rat spinal cord. Consistently, CRF as well as CRF-R2 agonist elicited potent, dose-dependent anti-nociceptive effects which were antagonized selectively by i.t. CRF-R2 (K41498) antagonist but not by CRF-R1 (NBI35965) antagonist. Moreover, the opioid antagonist naloxone dose-dependently reversed the i.t. CRF- as well as CRF-R2 agonist-elicited inhibition of somatic pain. Supporting these findings, double immunofluorescence confocal microscopy showed CRF-R2 on enkephalin (ENK)-containing inhibitory interneurons in close opposition of incoming mu-opioid receptor-immunoreactive nociceptive neurons. CRF-R2 was, however, not seen on pre- or on postsynaptic sensory neurons of the spinal cord. Taken together, these findings suggest that i.t. CRF or CRF-R2 agonist inhibit inflammatory somatic pain, occurring predominantly through CRF-R2 receptors located on spinal enkephalinergic inhibitory interneurons, which results in endogenous opioid-mediated pain inhibition.