Figure 4.

Early aging alters presynaptic markers in the LPP terminal zone. A, Section through hippocampus shows the two zones in the DG outer molecular layer (OML) used for analysis of synaptic proteins. B, Deconvolved immunofluorescence images show the localization of p-FAK (red) and vesicular protein synaptophysin (SYN; green) in rat. Arrows point to double-labeled puncta. Scale bar, 10 µm. C, FDT analysis of the density of p-FAK immunoreactivity (ir) colocalized with synaptophysin. Measurements were made for >100,000 individually reconstructed, double-labeled (2×) synapses per slice. Values for each slice were plotted as the percentage of 2× synapses (y-axis) fitting into an ascending series of labeling-density bins (x-axis). The resultant curves for individual slices were then averaged for a group and plotted as mean ± SEM values. There was a small but significant left shift in the curve for the 8-month-old group (N = 12) relative to that for the 3-month-old group (N = 15). Scatter plot summarizes the percentage of 2× synapses with high-density p-FAK-ir (right of vertical dotted line; *p < 0.05). D, Same analysis as in C but for the density of synaptophysin-ir: there was a pronounced rightward skew in the plot for 8-month-old rats (toward greater synaptic densities) versus the 3-month-old rats, and a large increase in the proportion of terminals with dense synaptophysin-ir (scatter plot; ***p < 0.001). E, Deconvolved images show dual immunofluorescence for TrkB (red) and synaptophysin (green) in mouse. Arrows point to 2× puncta. Scale bars: 4 µm; inset, 1 µm. F, Analysis of the density of TrkB-ir in LPP terminals in mice: there was a pronounced left shift in the density frequency distribution for the 8-month-old group relative to the 3-month-old group (N = 6/group), resulting in a large reduction in the proportion of terminals with dense TrkB-ir (scatter plot; ***p < 0.001).