[Xylazine Produced Analgesic Effect via Inhibits Hyperpolarization-activated Cyclic Nucleotide-gated Ion Channels...

To investigate the analgesic mechanism of xylazine by inhibiting the activation of hyperpolarized cyclic nucleotide-gated (HCN) ion channels.

HCN subchannel 1 (HCN1) knockout mice (HCN1^-/-) and HCN1 wild type mice (HCN1^+/+) were intraperitoneally injected with physiological saline and xylazine (10, 20, 30, and 40 mg/kg). Mechanical pain test and tail flick test were used to test the analgesic effect of xylazine by using the percentage of the maximal possible effect (%MPE); The control group and test groups of different concentrations of xylazine (12.5, 25, 50, and 100 μmol/L) were set up using HEK 293 cells transfected HCN1 plasmid and HCN subchannel 2 (HCN2) plasmid, respectively. The activated current of hyperpolarized HEK 293 cells expressing HCN1 and HCN2 ion channels and the inhibition rate of xylazine on hyperpolarization-activated currents were recorded using a whole cell patch clamp.

The results demonstrated that %MPE of the mechanical pain stimuli test and the thermal radiation stimuli test increased with the higher concentration of xylazine injected for both HCN1^+/+ mice and HCN1^-/-mice. When injecting xylazine by 30 mg/kg and 40 mg/kg, the %MPE of mechanical pain stimuli test for HCN1^-/- mice were %MPE= (62.06±14.72) % and %MPE= (69.92±16.09) %, respectively; and the percentages of tail flick tests were (52.50±1.97) % and %MPE= (64.74±6.34) %, respectively. But for HCN1^+/+ mice, the percentages of mechanical pain stimuli test were %MPE= (75.47±8.06) % and %MPE= (86.35±11.31) %; respectively, and the percentage of tail flick tests were %MPE= (57.83±4.82) % and (74.98±9.35) %. The analgesic effect results of the mechanical pain test and tail flick test of HCN1^+/+ mice were significantly different from HCN1^-/- mice ( P<0.05). Whole-cell patch clamp test results showed that xylazine had inhibitory effects on the currents of HCN1 and HCN2 ion channels, and the hyperpolarization-activated currents inhibition rate of HCN1 by xylazine (12.5-100 μmol/L) was between (24.62±23.62) %- (62.40±15.48) %; V of HCN1 was between (-79.58±1.56) mV- (-98.95±3.57) mV. The I inhibition rate of HCN2 by xylazine (12.5-100 μmol/L) was between (29.19±17.82) %- (80.02±6.64) %; with V of HCN2 between (-102.17±1.36) mV- (-117.48±2.38) mV.

Xylazine showed better analgesic effect on HCN1^+/+ mice than HCN1^-/- mice. Xylazine can produce analgesic effect by inhibiting HCN ion channel currents.