Acid-sensing ion channel subtype 3 function and immunolabelling increases in skeletal muscle sensory neurons following femoral artery occlusion

Sympathetic nerve activity and arterial blood pressure responses to static hindlimb muscle contractions are greater in rats with femoral arteries that were previously ligated (24-72 h earlier) than in control rats. Studies further demonstrate that acid-sensing ion channel subtype 3 (ASIC3) in thin-fibre muscle afferents contributes to the amplified reflex muscle responses observed in occluded rats, probably due to enhanced ASIC3 expression in muscle sensory neurons. The purpose of this study was to characterize acid-induced current with activation of ASIC3 in dorsal root ganglion (DRG) neurons of control rats and rats with 24 h of femoral occlusion using whole-cell patch clamp methods. Also, immunohistochemistry was employed to examine existence of ASIC3 expression in DRG neurons of thin-fibre afferents. DRG neurons from 4-to 6-week-old rats were labelled by injecting the fluorescence tracer DiI into the hindlimb muscles 4-5 days prior to the recording experiments. The results of this study show that similar to 90% of current responses evoked by pH 6.7 in DRG neurons innervating the hindlimb muscles are ASIC3-like. The peak current amplitude to pH 6.7 is significantly attenuated with application of rAPETx2, a specific ASIC3 antagonist. In addition, ASIC3-like current responses to pH 6.7 are observed in small, medium and large DRG neurons, and size distribution of DRG neurons is similar in control and occluded animals. However, the peak current amplitude of DRG neuron response induced by ASIC3 stimulation is larger in occluded rats than that in control rats. Moreover, the percentage of DRG neurons with ASIC3-like currents is greater after arterial occlusion comparedwith control. Furthermore, results fromdouble immunofluorescence experiments show that femoral artery occlusion mainly augments ASIC3 expression within DRG neurons projecting C-fibre afferents. Taken together, these data suggest that (1) the majority of current responses to pH 6.7 are ASIC3-like in DRG neurons with nerve endings in the hindlimb muscles, (2) a greater acid-induced current responding to pH 6.7 develops when hindlimb arterial blood supply is deficient under ischaemic conditions, and (3) increased ASIC3 expression is largely observed in thin C-fibres of DRG neurons after hindlimb ischaemia.