Combined Changes in Chloride Regulation and Neuronal Excitability Enable Primary Afferent Depolarization to Elicit Spiking without Compromising its Inhibitory Effects
Fig 2
PAD-induced transient spiking in DRG neurons.
(A) Sample responses to virtual GABA conductance applied via dynamic clamp. Labels a-d on cartoon indicate testing conditions and are referred to in all subsequent panels. Most neurons, like the typical one illustrated here, spiked only for EGABA = -20 mV and after being made hyperexcitable by exposure to 2.5 mM 4-AP (point c). (B) Summary of the proportion of neurons responding with or without spikes to virtual PAD. Numbers inside each bar indicate the number of cells. A total of 29 neurons from naïve rats were tested before and after 4-AP and for each EGABA. A total of 10 neurons from nerve-injured rats were tested for each EGABA. The proportion of spiking/non-spiking cells was compared between conditions using Fisher’s exact tests (see Results). (C) Left panel summarizes the minimum ḡGABA required to elicit spiking in cells that spiked both before and after 4-AP for EGABA = -20 mV. Minimum ḡGABA was significantly reduced from 0.76 ± 0.19 to 0.20 ± 0.05 nS/pF (mean ± SEM) by 4-AP (p = 0.005, two-way repeated measures ANOVA and Tukey test). These values are lower than observed in simulations in Fig 1; therefore, we adjusted the neuron model to reproduce this higher sensitivity to ḡGABA. As illustrated in S1 Fig, this revised model shows the same relationship between EGABA and excitability (βw) as seen in Fig 1. Right panel shows minimum ḡGABA plotted against soma diameter. Soma diameter, which correlates with fiber type, did not significantly affect minimum ḡGABA or the effect of 4-AP (p = 0.61 and 0.29, respectively; two-way repeated measures ANOVA). (D) Summary of the minimum ḡGABA required to elicit spiking in cells that spiked for each EGABA value after 4-AP. The depolarizing shift in EGABA from -35 mV to -20 mV caused a significant reduction (p<0.022, paired t-test) from 0.30 ± 0.07 nS/pF to 0.11 ± 0.02 nS/pF. (E) Sample responses from a typical neuron tested with gGABA steps and ramps. The minimum ḡGABA required to elicit transient spiking when applied as a step was 40 nS (or 0.96 nS/pF after normalization by membrane capacitance) but a ramp with 2.5x greater peak amplitude failed to elicit spiking. EGABA = -20 mV.