Fig. 8: Cholesterol supplementation has dual effects on HCN channel gating and attenuates neuronal excitability in SNI pain models.

a Representative G-V relationship of HCN channel activation of naive small DRG neurons with and without WSC treatment. b–d Summary of the effects of WSC treatment on HCN gating parameters of naive DRG neurons: V_1/2, slope factor, and free energy for channel activation, as in panel (a), mean ± s.e.m., n = 7 cells, *p = 0.04 using two-sided student’s t-test. e WSC treatment increased the CTxB-based FLIM-FRET efficiency in ipsilateral small DRG neurons, mean ± s.e.m., n = 12 cells without WSC and n = 6 with WSC, **p = 7e-4 using two-sided student’s t-test. f Summary of the effects of WSC on action potential firing in ipsilateral small DRG neurons. Data shown are mean ± s.e.m., n = 9 for contralateral side neurons, n = 12 for ipsilateral side neurons (*p = 0.02), n = 8 for ipsilateral neurons with WSC, **p = 0.002 comparing with and without WSC for ipsilateral side neurons, one-way ANOVA (no adjustment). g Summary of the number of action potentials of small DRG neurons versus the amplitude of current injection (same sample size as in panel (f), 0 pA current injection), mean ± s.e.m. With the +20 pA current injection, *p = 0.04 comparing with and without WSC for ipsilateral neurons. No statistical significance was detected in other amounts of current injection. h HCN currents elicited by hyperpolarizing voltages in WSC-treated ipsilateral small DRG neurons. i–k Summary of gating parameters of HCN currents with (n = 9 cells) and without (n = 8 cells) WSC treatment for ipsilateral small neurons in the SNI model: V_1/2 (p = 0.45), slope factor (**p = 0.007), and free energy (*p = 0.04) for channel activation, mean ± s.e.m., two-sided student’s t-test. l Summary data of HCN deactivation τ, with (n = 9) and without (n = 7) WSC treatment for ipsilateral small DRG neurons, mean ± s.e.m., *p = 0.02 using two-sided student’s t-test. m Schematic illustrating cholesterol’s dual effects on HCN channel gating: via OMD expansion or direct binding. HCN3-based structure (PDB: 8INZ) and membrane lipids are shown in ordered (right) vs. disordered (left) states. Created in BioRender. Dai, G. (2024) BioRender.com/f24v785.