Mechanism of heat treatment on exercise pressor reflex in hindlimb ischemia-reperfusion: Does the temperature gradient matter?

Purpose: To examine the effect of heat treatment (HT) with two temperature gradients in skeletal muscle: 1.5 and 3 °C, on the exercise pressor reflex (EPR) responses following limb ischemia-reperfusion (IR). Specifically, the involvement role of the P2Xs (receptors of ATP) pathway in the muscle afferent neurons was accessed. Methods: An experimental IR model was induced by 6 h of ischemia followed by 18 h of reperfusion in rats (IR rats). For HT groups, three HT sessions (muscle temperature increased by 1.5 or 3 °C) lasted for 30 mins each were applied. EPR responses were evoked by static muscle contraction (30s). Protein expression of P2X3 receptor in dorsal root ganglions (DRGs) was evaluated by western blot. In addition, a calcium imaging study was applied to detect calcium influx induced by activation of P2X3 in the isolated muscle DRG neurons of studied animal groups. Data are presented as mean ± standard deviation (M ± SD). Results: The mean arterial pressure (MAP) response to the static muscle contraction was significantly exaggerated in rats of IR 18 h (vs. sham, p < 0.01). The exaggerated BP response was attenuated with increasing Tm by 1.5 and 3 °C (vs. IR, p < 0.05). The expression of the P2X3 receptor was significantly enhanced in the DRGs of IR 18 h rats (vs. sham, p < 0.01). The upregulated P2X3 was suppressed in the DRGs of IR 18 h rats +HT of 1.5 °C and 3 °C (vs. IR, p < 0.05). In the isolated muscle afferent neurons, the Ca²⁺ entry induced by extracellular application of α,β-Me-ATP (30 μM) were significantly increased in IR rats (vs. sham, p < 0.001). Both HT protocols suppressed the enhanced IR-induced Ca²⁺ entry (vs. IR, p < 0.001). There was no difference between the BP responses in HT1.5 °C and 3 °C groups, nor P2X3 expression in muscle afferent DRG, P2X-mediated Ca²⁺ entry in isolated muscle afferent neurons (all p > 0.05). Conclusion: IR injury leads to upregulation of EPR responses, and HT attenuates this effect. The P2X3 signaling pathway was involved in the beneficial regulatory effect of HT on EPR in IR. The temperature gradient did not play a role in the extent of BP and muscle afferent P2X pathway activity attenuation in the present study.