Journal: JCI Insight
Article Title: Developmental progression of respiratory dysfunction in a mouse model of Dravet syndrome
doi: 10.1172/jci.insight.184231
Figure Lengend Snippet: ( A ) Computer-assisted plot shows the location of RTN neurons from each genotype in the ventral parafacial region. Py, pyramidal tract; 7 N, facial motor nucleus. Numbers to the right of each section designate millimeters from bregma. ( B ) Double immunolabeling shows a Lucifer yellow–filled (LY-filled) CO 2 /H + -sensitive RTN neuron recorded in a slice from a Scn1a +/– mouse is Phox2b immunoreactive. DAPI was used to visualize the cell nucleus. Scale bar: 20 μm. We confirmed Phox2b immunoreactivity in RTN neurons from control ( n = 10) and Scn1a +/– ( n = 11) tissue. ( C ) Traces of firing rate and segments of holding current from RTN neurons in slices from control (black) and Scn1a +/– (red) mice show examples of spontaneous activity under control conditions (5% CO 2 , pH 7.3) and that neurons from both genotypes respond to 10% CO 2 with a washable and repeatable increase in activity (pH 7.0). ( D and E ) Summary data show that RTN neurons in Scn1a +/– tissue exhibit high baseline activity ( D ) but respond to 10% CO 2 by an amount similar to control neurons ( E ). ( F ) Traces of firing rate from RTN neurons in slices from control (black) and Scn1a +/– (red) mice show that exposure to 3% CO 2 strongly inhibits control neurons but causes only a modest inhibition of neurons in Scn1a +/– tissue. ( G ) Summary data ( n = 13/ genotype) show that RTN neurons in slices from Scn1a +/– mice are more excitable at 3% and 5% CO 2 . Means were compared using 2-way ANOVA followed by Šídák’s multiple-comparison test and slopes of neural activity between 3% and 10% CO 2 were compared using 1-way ANCOVA. *** P < 0.001 for differences between genotypes; #### P < 0.0001 for genotype differences from control.
Article Snippet: Slices were incubated overnight at room temperature with fresh blocking solution, goat anti-Phox2b antibody (1:150; R&D Systems, AF4940) and rabbit anti–Lucifer yellow antibody (1:450; Thermo Fisher Scientific, A-5750).
Techniques: Immunolabeling, Control, Activity Assay, Inhibition, Comparison
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