Fig. 6: CRHR1 activation decreases TRN cell bursting.
a Left, current-clamp recordings from a TRN neuron showing low-threshold bursts followed by tonic firing in response to depolarizing steps, before (Bsl) and after CRH application (1 µM). Right, bath-applied CRH significantly reduced bursting across a range of current steps (n = 9). b CRH decreases bursting in a concentration-dependent manner (average of 50–125 pA steps: 0 nM, n = 12; 10 nM, n = 10; 200 nM, n = 11; 500 nM, n = 10; 1 µM, n = 9). c Photostimulation of CRH afferents (TG, 20 Hz, 5-ms pulses, 22 s) mimicked the effect of exogenous CRH, with no effect in wild-type mice (WT, n = 10; TG, n = 11). d CRHR1 antagonist NBI35965 (NBI, 3 µM) abolished the effects of CRH (CRH, n = 10; NBI + CRH, n = 8; Opto-CRH, n = 11; NBI + Opto-CRH, n = 15). e Left, example biphasic currents in a TRN neuron before and after CRH. CaV3-channel- and SK2-channel-mediated contributions to total charge are color-coded. Right, CRH only affected the negative component (p = 0.076), otherwise stable in control recordings (Ctr, n = 7; CRH, n = 8). f Left, example tail currents mediated by SK2 channels at the end of depolarizations to +20 mV. Right, percentage change of charge and decay time, indicating no significant impact of CRH on SK2-mediated currents (n = 7, p > 0.05). g Left, isolated CaV3 currents (magnified portion in the inset). Right, CRH (500 nM, n = 9) induced a decrease in CaV3 currents that was prevented by NBI35965 (3 µM, n = 6). h Repetitive depolarizations revealed a progressive decrease in CaV3 current peaks following CRH application (n = 5). Data are mean ± SEM. Voltage-clamp traces are the average of 12–15 recordings. Statistical tests: a, c, e, g repeated-measures two-way ANOVA, b one-way ANOVA, and h repeated-measures one-way ANOVA with Holm–Šídák’s test; d two-tailed Mann–Whitney test, e two-tailed paired t-test, and f, g two-tailed Wilcoxon test. #p < 0.1, *p < 0.05, **p < 0.01, ***p < 0.001. See Supplementary Table 1 and Source data file for details.
