Fig. 3: Hypercapnia fails to stimulate active expiration in anesthetized Tcf4^tr/+ mice.

Diaphragm and abdominal EMG activity was measured in isoflurane (1.5%) anesthetized Tcf4^+/+ and Tcf4^tr/+ (50 days old) during exposure to graded increases in CO₂. A Traces of raw and integrated (∫) diaphragm and abdominal EMG activity show that Tcf4^+/+ mice treated with saline (30 µL; I.P.) respond to 5 and 7% CO₂ with proportional increases in Dia_EMG and Abd_EMG activity. Conversely, saline (30 µL; I.P.) treated Tcf4^tr/+ mice show a diminished Dia_EMG response to CO₂ and completely lacked Abd_EMG activity, even at 7% CO₂. Systemic (I.P.) administration of PF-04531083 (40 mg/kg) increased CO₂-dependent Dia_EMG but not Abd_EMG activity in Tcf4^tr/+ mice. B, C Summary data show effects of CO₂ on Dia_EMG amplitude (B F_2,10 = 40.74, p < 0.0001, two-way ANOVA) and frequency (C F_2,10 = 27.96, p < 0.0001, two-way ANOVA) in Tcf4^+/+ and Tcf4^tr/+ mice (n = 6 biologically independent animals/genotype, data are presented as mean values ± SEM). D, E summary data show effects of CO₂ on Abd_EMG amplitude (D F_2,10 = 145.0, p < 0.0001, two-way ANOVA) and frequency (E F_2,10 = 655.4, p < 0.0001, two-way ANOVA) in Tcf4^+/+ and Tcf4^tr/+ mice (n = 6 biologically independent animals/genotype, data are presented as mean values ± SEM). These results are consistent with anatomical evidence that Phox2b+ neurons in the lateral parafacial region are severely depleted in Tcf4^tr/+ mice, and the possibility that these cells are a key determinant of expiratory activity (i.e., function as expiratory pF_L neurons). Asterisk (*) indicate the different from 0% CO₂ within condition; #, different between genotypes/conditions. One symbol = p < 0.05, two symbols = p < 0.01, three symbols = p < 0.001, four symbols = p < 0.0001 (two-way RM-ANOVA with Tukey’s multiple comparison test).