On the basis of sniffing time (social approach test), we observed that BALB/cJ were less sociable (p <0.05, Student’s t-test) at 30 day-of-age in comparison to the B6 mice (). There was no significant difference in the sniffing time between BALB/cJ and B6 mice at 50 and 70 day time point (p >0.05), which is consistent with a previously published study (Fairless et al., 2012
Bar diagram showing the mean and standard error in social sniffing time between BALB/cJ and B6 at different time points. Asterisk (*) represents significant differences with a p-value of ≤ 0.05.
2.1. Correlation between sociability and DTI
DTI data from most of the animals was without any substantial motion artifacts and in general, the signal-to-noise ratio (SNR) from the cortical region of the brain was over 100 (139.76±16.91) for the non-diffusion weighted (b0) images and it was over 75 (76.66±6.79) for the diffusion weighted (b786.73 s/mm2
) images. This resulted in excellent quality of the in-vivo DTI data as evident by the MD, FA and FA weighted color coded maps, shown in . A moderate but significant positive correlation was observed between MD values from the EC and social sniffing time of 30 day-old BALB/cJ mice (r =0.519; p =0.007). A moderate negative correlation was also observed between FA values from the Hippo (r =-0.523; p =0.006) and the FMC (r =-0.458; p =0.037) and sociability in these mice. None of the other areas demonstrated significant correlation between FA or MD and sociability of 30, 50 or 70 day old BALB/cJ mice (supplementary data Table S1
). Similarly, the B6 mice did not demonstrate any significant correlation between sociability and FA or MD values at any age (Supplementary data, Table S2
Fig. 2 Representative FA weighted color maps from the mid brain (A) and frontal (B) region of a 30-day-old BALB/cJ mouse. The region of interests (ROIs) on different gray and white matter regions are shown overlaid on the color maps. The color represents the (more ...)
2.2. Longitudinal changes in white matter regions in BALB/cJ mice
Significantly higher FA from the CC and EC was observed at 50 and 70 day in comparison to the 30 day time point (). However there were no significant differences between 50 and 70 day time points. When a linear mixed model was used to assess the differences in the growth pattern of WM areas, significant difference in the growth trajectory of the FA from the EC was observed in BALB/cJ mice in comparison to B6 mice (, ). The negative z score of the trajectory indicates that BALB/cJ mice exhibit a slower rate of change in FA values during this growth period (30-70 days) in the EC region in comparison to the B6 mice ().
Longitudinal and cross sectional changes at different time points in FA and MD (×10-3mm2/s) values in white matter regions in BALB/cJ and B6 mice
Longitudinal and cross sectional changes at different time points in FA and MD (10-3mm2/s) values in gray matter regions in BALB/cJ and B6 mice
Fig. 3 Growth trajectory of the FA values from external capsule in BALB/cJ and B6 mice. Higher FA value was observed in B6 mice at all-time points as compared to BALB/cJ mice. Black circles demonstrate the data points from B6 mice and open triangle represents (more ...)
2.3. Longitudinal changes in gray matter regions in BALB/cJ
The FA values from the Ctx and FMC at 30 day were significantly higher than the values at 50 and 70 day (). Significantly higher FA was also noted from the FMC at 50 day in comparison to the 70 day time points. When the differences in growth trajectories were assessed between BALB/cJ and B6 mice, significantly different trajectories of FA values from the Hippo, FMC and Th were observed (). Significantly positive developmental trajectories in Hippo and Th were noted in BALB/cJ mice, which demonstrate faster rate of change in FA values over the time in these regions in BALB/cJ as compared to B6 mice. On the other hand, a negative developmental trajectory in FMC demonstrated slower rate of change in FA values over time in BALB/cJ as compared to B6. These results indicate that different regions of the brain have different developmental trajectories in BALB/cJ mice.
Differences in the developmental trajectories of FA and MD (×10-3mm2/s) values in different gray and white matter regions between BABL/cJ and B6 using a multilevel model for trajectory analysis
2.4. Longitudinal changes in B6 mice at different time points
No significant differences in the WM regions were observed at any time point (). In the GM regions, the Amg region demonstrated lower FA at 30 days in comparison to the 50 day time point. The FA value was significantly higher at 50 day in comparison to both the 30 and 70 day time point. No significant difference in FA or MD values was observed from any other GM regions ().
2.5. Cross-sectional analysis of FA and MD values between BALB/cJ and B6 mice at different time points
Among the WM areas, significantly lower FA from the EC was observed in the BALB/cJ mice than B6 mice at 30 and 70 days ( and ). Additionally, the FA from the CC in BALB/cJ mice was higher than the B6 mice at 70 days. The MD values from the CC of BALB/cJ mice were higher than B6 mice at 30 and 50 days-of-age (). Among the GM areas, significantly lower FA from the Amg at 50 days was noted in BALB/cJ mice. However, the hippocampal FA values in BALB/cJ mice were significantly higher at 70 days. The hippocampal MD values in BALB/cJ mice at 50 days were also significantly higher than the B6 mice ().
Fig. 4 Box plots demonstrating the FA and MD (×10-3mm2/s) values from the external capsule (A and B) and frontal motor cortex (C and D) in BALB/cJ and B6 at 3 time points. Figure (A, C) show FA and (B, D) show MD values at 3 time points from BALB/cJ (more ...)
2.6. Histopathological Finding
Histological staining (H&E and Luxol fast blue) was performed from 2 BALB/cJ and 2 B6 mice at each time point. Due to the limited number of samples used, only a descriptive comparison between the staining pattern of BALB/cJ and B6 samples was performed. In comparison to BALB/cJ, the B6 brain samples demonstrated compact and darkly stained vacuolated nucleoli in EC and FMC on H&E stained sections (data not shown). When the LFB stained samples from 30-day-old brains were compared, the EC region of the brains in B6 mice appeared to have more compact and longer WM fibers than the BALB/cJ, in which the WM fibers appeared to be dispersed and comparatively smaller in size (white arrows, ). This difference was also observed from the samples taken from the 50 and 70 day-old animals ().
Fig. 5 Representative luxol-fast blue (LFB) stained sections from the external capsule region in B6 and BALB/cJ brain samples. Figure A-F represents the brain sections at 1.25x resolution. Figure G-L showing images from the rectangular box from external capsule (more ...)