Alteration of the mu opioid receptor: Ca²⁺ channel signaling pathway in a subset of rat sensory neurons following chronic femoral artery occlusion

The exercise pressor reflex, a crucial component of the cardiovascular response under physiological and pathophysiological states, is activated via metabolic and mechanical mediators that originate from contracting muscles and stimulate group III and IV afferents. We reported previously that stimulation of mu opioid receptors (MOR), expressed in both afferents, led to a significant attenuation of the reflex in rats whose femoral arteries had been occluded for 72 h. The present study examined the effect of arterial occlusion on the signaling components involved in the opioid-mediated modulation of Ca²⁺ channels in rat dorsal root ganglion neurons innervating the triceps surae muscles. We focused on neurons that were transfected with cDNA coding for enhanced green fluorescent protein whose expression is driven by the voltage-gated Na⁺ channel 1.8 (Na_V1.8) promoter region, a channel expressed primarily in nociceptive neurons. With the use of a small interference RNA approach, our results show that the pertussis toxin-sensitive Ga_i3 subunit couples MOR with Ca²⁺ channels. We observed a significant leftward shift of the MOR agonist [D-Ala2-N-Me-Phe4-Glycol5]-enkephalin concentration-response relationship in neurons isolated from rats with occluded arteries compared with those that were perfused freely. Femoral occlusion did not affect Ca²⁺ channel density or the fraction of the main Ca²⁺ channel subtype. Furthermore, Western blotting analysis indicated that the leftward shift did not result from either increased Ga_i3 or MOR expression. Finally, all neurons from both groups exhibited an inward current following exposure of the transient potential receptor vanilloid 1 (TRPV1) agonist, 8-methyl-N-vanillyl-6-nonenamide. These findings suggest that sensory neurons mediating the exercise pressor reflex express Na_V1.8 and TRPV1 channels, and femoral occlusion alters the MOR pharmacological profile.