Contribution of nerve growth factor to upregulation of P2X3 expression in DRG neurons of rats with femoral artery occlusion

Abstract

Femoral artery occlusion augments the sympathetic nerve and pressor responses to muscle contraction and muscle metabolites injected into the arterial blood supply of the hindlimb muscles in rats. The underlying mechanism by which these reflex responses are enhanced after muscle vascular insufficiency is unclear. Purinergic P2X3 receptor has been reported to contribute to the metabolic component of the exercise pressor reflex. Thus the purpose of this study was to examine if chronic femoral occlusion would alter the expression of P2X3 in dorsal root ganglion (DRG) neurons of rats. Also, P2X3-mediated sympathetic responsiveness was examined after femoral occlusion. In addition, the role played by nerve growth factor (NGF) in regulating the expression and response of P2X3 was examined. Western blot analysis showed that 24 h of femoral ligation increased the levels of P2X3 (optical density: 0.93 ± 0.07 in control and 1.37 ± 0.10 after occlusion; P < 0.05 vs. control). The fluorescence immunohistochemistry further demonstrated that the occlusion elevated P2X3 expression in DRG neurons (percentage of P2X3-positive cells: 33 ± 3% in control and 51 ± 3% in occlusion; P < 0.05 vs. control). Furthermore, the results showed that responses of renal sympathetic nerve activity and blood pressure to stimulation of P2X were greater in occluded rats than responses in control rats by injection of α,β-methylene ATP into the arterial blood supply of the hindlimb muscle. Finally, infusion of NGF in the hindlimb muscles of healthy rats increased P2X3 (optical density: 0.98 ± 0.12 in control and 1.37 ± 0.16 with NGF; P < 0.05 vs. control). The pressor response to injection of α,β-methylene ATP was increased in the rats with NGF infusion. Likewise, blocking NGF attenuated exaggeration of the reflex response induced by α,β-methylene ATP in occluded rats. The findings of this study suggest that the levels of P2X3 in primary afferent neurons are upregulated as the blood supply to the hindlimb is deficient under ischemic conditions, leading to augmentation of the muscle reflex. NGF is closely related to increases in P2X3 receptor expression and response.