Fig. 1: Hypofunction of T-Ca²⁺ channels and decreased CaV3.3 expression in TRN neurons of adult Gclm-KO mice, resulting in alteration of their bursting profile.

A Smaller proportion of TRN neurons generating burst firing at resting membrane potential (RMP) in KO as compared to WT mice (WT n = 13; KO n = 12; Fisher exact test, p = 0.02). B Weaker density of T-Ca²⁺ currents activated at RMP in TRN neurons of KO (n = 11) as compared to WT (n = 10) mice (p = 0.034, one-tailed t-test). C Representative recording of single bursting in a TRN neuron. D Threshold for the initial membrane potential required to induce single bursting upon depolarization. E Representative recording of repetitive bursting in a TRN neuron. F Threshold for the initial membrane potential required to induce repetitive bursting upon depolarization. Note that KO (n = 6) TRN neurons require a more hyperpolarized membrane potential, particularly for exhibiting repetitive bursts, as compared to WT mice (n = 8). G, H Top: Representative recordings of T-Ca²⁺ and SK currents induced by a short constant depolarization step (from −110 to −40 mV), with their amplitudes increasing with greater hyperpolarizing initial membrane potential (going from −30 mV to −110 mV). Bottom: Density of T-Ca²⁺ (G) and SK currents (H) activated from each of the initial membrane potentials. Compared to WT mice, TRN neurons in KO display overall smaller T-Ca²⁺ (F = 31.92 DFn = 1 DFd = 289; p < 0.0001; WT n = 9; KO n = 10) and SK current densities (F = 36.34 DFn = 1 DFd = 289; p < 0.0001; WT n = 9; KO n = 10). I Correlation between T-Ca²⁺ and SK current densities in TRN neurons of both genotypes (Spearman test, r = −0.99 for WT; r = −1 for KO; p < 0.0001 for both). J Micrographs showing immunofluorescent labeling for parvalbumin (PV, red) and CaV3.2 (green) in the TRN of adult WT and KO mice. K No significant difference of CaV3.2 labeled profiles (number of CaV3.2-IR voxels) in TRN of WT and KO mice (n = 5 for both genotypes). L Regional CaV3.2 quantification within the anterior, medial, and posterior sections of the TRN. M Micrographs showing immunofluorescent labeling for PV (red) and CaV3.3 (green) in the TRN of adult WT and KO mice. N Reduced CaV3.3 labeled profiles (number of CaV3.3-IR voxels) in TRN of KO as compared to WT mice (n = 4 for both genotypes). O Reduced expression of CaV3.3 mostly in the anterior and medial sections of the TRN in KO as compared to WT mice. Unpaired t-tests or two-way ANOVAs with Bonferroni post-tests; *p < 0.05 **p < 0.01 ***p < 0.001. Data are presented as means ± s.e.m.