Figure 2.

Knockdown of Pfn2 in the IPN is sufficient to increase anxiety. (A) Representative midbrain coronal section schematic and fluorescence microscopy images of WT mouse injected with either lenti-pGIPZ-Pfn2-shRNA-tGFP (left image) or lenti-pGIPZ-Scramble-tGFP (right image) in the IPN. Nuclei were labeled with DAPI (blue). (B) Knockdown of Pfn2 expression was measured by RT-qPCR using RNA from laser-captured IPN tissue infected with lenti-pGIPZ-Pfn2-shRNA-tGFP compared to lenti-pGIPZ-scramble-tGFP control. mRNA expression was normalized to β-2-microglobulin. Statistical analysis using an unpaired, two-tailed t-test compared lenti-Pfn2 shRNA mice to lenti-scramble controls (t₁₁ = 2.272, P < 0.05). ^(*)P < 0.05; n = 6–7. (C) Average time spent in the open arms and total number of arm entries in the elevated plus maze of mice injected with either lenti-Pfn2 shRNA (stippled bars) or lenti-scramble shRNA control (open bars) in the IPN. Statistical analysis used an unpaired two-tailed t-test to compare time spent in open arms (t₃₃ = 3.583, P = 0.0011) and total number of arm entries (t₃₃ = 0.7355, P = 0.4672) of lenti-Pfn2 shRNA mice to lenti-scramble controls. ^(**)P < 0.01 n = 16–19. Box plot edges are 25^th and 75^th percentile, central line is the median and whiskers are max and min. (D) Representative midbrain coronal section schematic and fluorescence microscopy image of WT mouse injected with lenti-pGIPZ-Pfn2-shRNA-tGFP in the VTA. Nuclei were labeled with DAPI (blue). (E) Average time spent in the open arms and average total number of arm entries in the elevated plus maze of mice injected with either lenti-Pfn2 shRNA (stippled bars) or lenti-scramble shRNA control (open bars) in the VTA. Unpaired, two-tailed t-test was used for statistical analysis of time spent in open arms (t₁₆ = 1.09, P = 0.2918) and total arm entries (t₁₆ = 0.4153, P = 0.6835). n = 9. Box plot edges are 25^th and 75^th percentile, central line is the median and whiskers are max and min.