Fig. 2

The extent of hippocampal injury and cognitive impairment is titrated by the concentration of the initial viral inoculum. Hippocampal pathology was assessed by H&E histology at 7 dpi (A-J) and 45 dpi (K-N) in mice inoculated with 200,000 PFU (A, E, I, K), 50,000 PFU (B, F, L), 12,500 PFU (C, G, J, M), or 3125 PFU (D, H, N) of TMEV. Extensive destruction of the CA1 pyramidal layer is observed at 7 dpi in mice inoculated with the highest concentration of TMEV (A, E, I, O). CA1 destruction is more variable but still readily apparent at 7 dpi in mice inoculated with 50,000 PFU of TMEV (B, F, O) and in some mice the extent of CA1 damage observed at 45 dpi (L) is as extensive as that observed in mice inoculated with 200,000 PFU of TMEV (K). Mice inoculated with 12,500 PFU of TMEV exhibit preservation of CA1 neurons at 7 dpi, with many mice exhibiting nearly complete sparing of CA1 (C, G, O) and only small, discrete areas of injury at high magnification (J). This preservation was verified at 45 dpi in the 12,500 PFU cohort (M). The absence of injury observed in this group occurs despite the presence of infiltrate apparent in the H&E stained sections (C, G, J). Finally, mice inoculated with 3125 PFU of TMEV exhibit essentially no evidence of CA1 injury at 7 dpi (D, H, O) or 45 dpi (N) and show only weak inflammatory infiltration. (O) Quantitation of hippocampal injury at 7 dpi reveals profound preservation in mice inoculated with 12,500 PFU of TMEV. F_(3,22) = 37.2861, P < 0.0001 by one-way ANOVA; Shapiro–Wilk P = 0.0020; Dunn's method pairwise analysis: 200,000 PFU vs 12,500 PFU: P = 0.0021; 200,000 PFU vs 3125 PFU: P = 0.0005; 12,500 PFU vs 3125: P = 1.000; d_s = 2.1; pooled results from 2 separate experiments. Mice were inoculated with 200,000 PFU or 12,500 PFU of TMEV and cognitive performance was assessed starting at 45 dpi using the Barnes maze (P–Q). P Latency to acquisition of the escape hole (s) was measured over 4 testing days in infected mice and compared to performance in uninfected mice inoculated with vehicle. Mice inoculated with 200,000 PFU do not learn to navigate to the escape while mice inoculated with 12,500 PFU perform as well as control mice. Latency: F_(11,56) = 16.6172, P < 0.0001 by two-way ANOVA (inoculum x day); Shapiro–Wilk P < 0.0001; Tukey HSD pairwise analysis: @ day 1: 200,000 PFU vs vehicle: P = 1.000; 12,500 PFU vs vehicle: P = 0.9999; 200,000 PFU vs 12,500 PFU: P = 0.9971; @ day 4: 200,000 PFU vs vehicle: P = 0.0109; 12,500 PFU vs vehicle: P = 1.000; 200,000 PFU vs 12,500 PFU: P = 0.0143; d_s = 0.55. Each symbol represents an individual animal. Q Analysis of error rate also indicates normal spatial memory acquisition in the mice infected with 12,500 PFU but not in mice inoculated with 200,000 PFU. Error rate: F_(11,56) = 18.9760, P < 0.0001 by two-way ANOVA (inoculum x day); Shapiro–Wilk P = 0.0011; Tukey HSD pairwise analysis: @ day 1: 200,000 PFU vs vehicle: P = 0.9190; 12,500 PFU vs vehicle: P = 0.9997; 200,000 PFU vs 12,500 PFU: P = 1.000; @ day 4: 200,000 PFU vs vehicle: P < 0.0001; 12,500 PFU vs vehicle: P = 0.9530; 200,000 PFU vs 12,500 PFU: P = 0.0004; d_s = 0.62. Scale bars: D = 1 mm (A–D); H = 100 μm (E–H); J = 100 μm (I, J); N = 100 μm (K–N). ** indicates P < 0.01 in panel (O). Arrowheads in E–J highlight representative examples of inflammatory infiltrate