2.4.2 Recordings of capsaicin-induced TRPV1 currents in DRG neurons

TRPV1 currents were recorded by using an EPC10 amplifier (HEKA). DRG neurons were placed in an open recording bath filled with bathing solution contained 145 mM NaCl, 5 mM KCI, 2 mM CaCl₂, 1 mM MgCl₂, 10 mM HEPES, and 10 mM D-glucose, and pH was adjusted to 7.4. The osmolarity was adjusted to 315–325 mOsmol/L. Glass electrodes (recording electrodes, 1.65 ± 0.05 μm outer diameter) were pulled with a P97 puller (Sutter) and filled with an internal solution contained 140 mM KCl, 2 mM MgCl₂, 10 mM Na₂ATP, 2.5 mM CaCI₂, 10 mM EGTA, and 10 mM HEPES, and pH was adjusted to 7.2. The osmolarity was adjusted to 305–315 mOsmol/L. All testing was performed at approximately 22 °C.

The clamp voltage (Vh) was set to - 60 mV for all experiments, and capsaicin (CAP)-evoked currents were recorded in voltage clamp mode. Capsaicin-evoked whole-cell currents were filtered and the signal sampling frequency was 1 kHz. Membrane potentials were expressed as absolute values (millivolts, mV), and TRPV1 currents were expressed as absolute values (nanoamperes, nA) or multiples of changes compared with basal values. Capsaicin was dissolved into stock solution of 1 mmol/L by dehydrated alcohol, and then diluted into 1 μmol/L by the external solution. Capsazepine (CPZ), the competitive antagonist of the TRPV1 receptor, was prepared at a concentration of 10 μmol/L using the same method. All drugs were administered through a multi-channel rapid micro drug delivery system (Yibo Life Science Instrument).