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. 2017 Jan 30;6:e19886. doi: 10.7554/eLife.19886

Figure 5. Bax deletion enhances EPSCs in adult born neurons.

(A) Whole cell recordings were made from immature GCs in control and BaxKOim slices at six weeks post-tamoxifen injection, using picrotoxin to isolate glutamatergic EPSCs. Simultaneous fEPSPs were recorded in the molecular layer as in Figure 1. (B) Immature GCs in control and BaxKOim tdT mice had a similar input resistance that was higher than mature GCs (n = 12, 12, 16, respectively; one-way ANOVA p=0.0004, *p<0.05,***p<0.0001 Bonferroni post hoc test). (C) Left, examples of fEPSPs (top) and EPSCs (bottom) recorded in immature GCs. Middle, an increase in synaptic transmission to immature BaxKO GCs was revealed by the EPSC plotted against fiber volley (two-way ANOVA, Fgenotype (1,143)=18.55 p<0.0001, n = 12 control tdT, 12 BaxKOim tdT; *p<0.05 with Bonferroni post-tests). Right, the EPSC/FV ratio for all stimulus intensities (control 1.24 ± 0.07, n = 80; BaxKOim1.59 ± 0.09, n = 75; unpaired t-test p=0.0029). (D) Schematic showing simultaneous recordings from adjacent tdT- (BaxWT) and tdT+. (BaxKO) GCs in slices from BaxKOim tdT mice at 16 weeks after tamoxifen. (E) Adult-generated BaxKO GCs had larger EPSCs than simultaneously recorded unlabeled mature GCs. EPSCs were normalized to the maximum amplitude of the unlabeled (BaxWT) GC in each slice (two-way ANOVA, Fgenotype (1,94)=11.59 p=0.001, n = 6 pairs), scale bars: 10 ms, 100 pA. Comparing raw EPSCs between pairs of unlabeled and tdT+ GCs across all stimulus intensities confirmed EPSCs were larger in tdT+ GCs (not shown, paired t-test, p<0.0013).

DOI: http://dx.doi.org/10.7554/eLife.19886.012

Figure 5.

Figure 5—figure supplement 1. No change in FV or fEPSP in slices from BaxKOim tdT mice.

Figure 5—figure supplement 1.

There was no difference in the FVs (left) or fEPSPs (middle) in BaxKOim mice with tdT-labeled immature neurons (two-way ANOVA p=0.535 and 0.345, stim intensity binned by 10 V). The fEPSP slope plotted against the FV also suggested no change in total synapse number (right, two-way ANOVA p=0.210, FVs binned by 100 μV). Data from 12 control slices and 12 BaxKOim slices. These results replicate those from Figure 1, showing that addition of tdT expression in immature GCs has no effect on the measures obtained in BaxKOim mice.
DOI: http://dx.doi.org/10.7554/eLife.19886.013
Figure 5—figure supplement 2. No change in PPR or sEPSCs in immature GCs from BaxKOim tdT mice.

Figure 5—figure supplement 2.

(A) A representative immature GC filled with biocytin during recording. Note the small dendritic tree compared to mature GCs (shown in Figure 1—figure supplement 2; Dieni et al., 2013, 2016). (B) Immature GCs in control and BaxKOim tdT cells had similar paired-pulse ratio (100 ms ISI). Unpaired t-test p=0.345; n = 12 control tdT and 11 BaxKOim tdT. (C) Example traces from spontaneous EPSC recordings (left). The frequency (middle) and amplitude (right) of spontaneous events were similar in control and BaxKOim tdT cells (unpaired t-tests, p=0.203 and 0.525 respectively, n = 9 control tdT and 10 BaxKOim tdT).
DOI: http://dx.doi.org/10.7554/eLife.19886.014
Figure 5—figure supplement 3. No differences in intrinsic properties between adult-born mature tdT+ (Bax -/-) and unlabeled mature GCs.

Figure 5—figure supplement 3.

There were no differences in the input resistance (unpaired t-test, p=0.51, control n = 21, BaxKOimn = 14), AP amplitude or AP frequency (unpaired t-tests, p=0.21 and 0.69 respectively, control n = 13, BaxKOimn = 11) between unlabeled mature GCs and tdT+ adult-born GCs in BaxKOim tdT mice. Note that this data is from16 weeks after TMX induced recombination, such that adult-born tdT+ GCs have mature intrinsic properties.
DOI: http://dx.doi.org/10.7554/eLife.19886.015
Figure 5—figure supplement 4. Global Bax levels are unaltered in BaxKOim hippocampus.

Figure 5—figure supplement 4.

Representative hippocampal western blot from control (Con), BaxKOim (KOim), and germ line Bax -/- hippocampal lysates for Bax protein and control β-tubulin. Band intensity was quantified and normalized to control protein. Unpaired t-test, p=0.28 (n = 3 mice, ± SEM).
DOI: http://dx.doi.org/10.7554/eLife.19886.016