Spinal Nerve Axotomy: Effects on I In Vivo and HCNs in DRG Neurons.

In vitro experiments performed on dissociated dorsal root ganglion (DRG) neurons suggest the involvement of the hyperpolarization-activated cation current (I) in enhancing neuronal excitability, potentially contributing to neuropathic pain. However, the more confirmative in vivo information about how nerve injury interacts with I is lacking. In this study, I was recorded in vivo using the dynamic single-electrode voltage clamp (dSEVC) technique on L5 DRG neurons of normal rats and those seven days after spinal nerve axotomy (SNA). Compared to normal rats, SNA unexpectedly inhibited the activity of I channels on A-fiber DRG neurons: (a) the I current magnitude, density, and conductance were consistently diminished; and (b) the I activation velocity was slowed and the voltage for I activation was hyperpolarized. The half-activation voltage (V) exhibited a negative shift, and the time constant for I activation was prolonged across all test potentials, indicating the reduced availability of I after SNA. To further investigate the mechanisms of SNA on I, the underlying HCN channels and the correlated mRNA were quantified and compared. The mRNA expression level of 1-4 was uniformly enhanced after SNA, which might have contributed to the increased cytoplasmic HCN1 intensity observed in both medium- and large-sized DRG neurons. This finding contradicted the functional reduction of I after SNA. Surprisingly, the HCN labeling pattern was altered after SNA: the labeling area of HCN1 and HCN2 at the membranous ring region of the axotomized large neurons became significantly thinner or absent. We concluded that the diminished ring immunoreactivity for HCN1 and HCN2 correlated with a reduced availability of I channels, elucidating the observed decrease in I in axotomized A-fiber neurons.