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. 2020 May 18;9:e55342. doi: 10.7554/eLife.55342

Figure 6. Manipulation of α2-GABAA Receptors in NAc Mimicked Fear Deficits in Chat-Mecp2-/y Mice.

(A) Schematic of AAV/RNAi viral construct. Confocal image of injected NAc slice demonstrating colocalization of GFP expression with ChAT antibody, co-stained with DAPI. (B, C) mIPSCs of AAV/RNAi-infected NAc neurons from Chat-Mecp2-/y mice were rescued. (B) Examples of mIPSC traces. (C) Cumulative distribution of mIPSC amplitude of NAc neurons infected by AAV-Sham or AAV/RNAi. P-values were calculated by one-way analysis of variance (ANOVA) with Bonferroni’s multiple comparisons test. F (2, 39)=5.816, p=0.0062. n = 14 neurons from four mice for each group. (D) Representative images of NAc slices from Chat-Mecp2-/y mice injected with AAV/GFP or AAV/RNAi. (E) Statistically integrated immunofluorescence data of α2-GABAA receptors normalized to AAV/GFP levels. P-values were calculated by two-tailed unpaired t-test. t = 2.985, df = 39, p=0.0049. n = 27 neurons from three mice (AAV/RNAi), 14 neurons from three mice (AAV/Sham). (F) Left: Schematic of cannula implantation. Right: Infusion sites of drugs, stained with DAPI. (G, H) Fear deficit was measured by retrieval process in fear conditioning test 20 min after drug application. P-values were calculated by two-tailed unpaired t-test. t = 2.542, df = 23, p=0.0182 for (E). t = 3.609, df = 23, p=0.0015 for (F). n = 14 mice for ACSF, n = 11 mice for L-838,417. Data are means ± SEM. *p<0.05, **p<0.01.

Figure 6—source data 1. Statistical reporting of Figure 6.

Figure 6.

Figure 6—figure supplement 1. Knockdown of α2-GABAA Receptors in Cholinergic Interneurons Did Not Affect mIPSC Frequency in Cholinergic Interneurons, Related to Figure 6.

Figure 6—figure supplement 1.

Cumulative distribution of mIPSC amplitude in NAc neurons infected by AAV-Sham or AAV/RNAi. P-values were calculated by one-way analysis of variance (ANOVA) with Bonferroni’s multiple comparisons test. F (2, 39)=1.066, p=0.3541. n = 14 neurons from four mice for each group. NS means no significance.
Figure 6—figure supplement 2. Retrograde tracing from NAc cholinergic neurons.

Figure 6—figure supplement 2.

(A) Sequential injections of AAV-FLEX-TVA-GFP, AAV-FLEX-Rabies-glycoprotein and G-deleted EnA-rabies-mCh into NAc of a ChatCre mouse labeled starter cells (yellow). (B, C) Transsynaptic retrograde labeling neurons in the BLA (B) and (vHPC). BLA: basolateral amygdala. CeA: central nucleus of the amygdala. vHIP: ventral hippocampus.