CLINICS 2011;66(4):663-671 Figure 1 - Diagram of an amygdaloid neuron and the scheme of dendritic quantification. DBP – Dendritic branching points; DI – Dendritic intersections; CC – Concentric circles; S – Soma; and 1 to 5 – Concentric zones. Quantification of dendritic branching points and dendritic intersections The concentric circle method of Sholl (1956) was used for dendritic quantification.40 Concentric circles on a transpar- ent sheet with a radial distance of 20 mm between circles were used for dendritic quantification (i.e., dendritic branching points and intersections). The sheet was placed on a neuron tracing so that the center of the cell body of the neuron coincided with the center of the concentric circles. The number of branching points between the two concentric circles, i.e., within each successive 20-mm concentric zone (ring), was counted. The dendritic intersection was the point where a dendrite intersected a given concentric circle (Fig. 1). The dendritic intersections at each concentric circle were counted. Both branching points and intersections were counted up to a radial distance of 100 mm from the center of the somata. The mean number of dendritic branching points in each concentric zone and the number of dendritic intersections at each concentric circle were calculated. The number of dendritic processes that arose from the somata of amygdaloid neurons was also counted. Statistical analysis The data were analyzed using an analysis of variance (ANOVA) followed by Bonferroni’s test (post hoc) using GraphPad Prism, version 2.01. RESULTS The rats treated with all doses of BM showed an improvement in spatial learning performance and enhanced memory retention compared with normal control rats.34 Amygdaloid neuronal dendritic analysis in rats treated with all doses of BM (20, 40 and 80 mg/kg) for 2 weeks did not reveal altered dendritic arborization. However, treatment with BM 40 and 80 mg/kg for 4 weeks (Figs. 2 and 3; Tables 1 and 2) and with BM extract 20, 40 and 80 mg/kg for 6 weeks resulted in a significant increase in dendritic length (dendritic intersections) and the number of branching points 665 Effect of Bacopa monniera on amygdaloid neurons Vollala VR et al. (Figs. 4 and 5; Tables 3 and 4). In addition, treatment for 6 weeks with higher doses of BM extract (40 and 80 mg/kg) increased the number of dendritic processes that arose from the somata (Table 3). There was no difference in dendritic length or branching pattern between the control and gum acacia-treated rats, which suggested that the daily handling of the rats (i.e., handling stress and vehicle) itself did not alter the dendritic pattern. Because there were no significant differences in dendritic length and branching, only the comparisons between the control and experimental groups were subsequently detailed, and all of the figures ignore the vehicle control group data. Four-week treatment group Dendritic intersections (Table 1). There were no significant changes in dendritic intersections at any of the concentric circles in the BM 20-mg/kg group when compared with the normal control group. Both the BM 40- and 80-mg/kg groups showed significant increases in dendritic intersections at 20 mm, 40 mm, 60 mm and 80 mm concentric circles. Dendritic branching points in different concentric zones (Table 2). No significant changes were observed in the dendritic branching points in any of the concentric zones in the BM 20-mg/kg group. However, both the BM 40- and 80- mg/kg groups showed a significant increase in dendritic branching points in concentric zone 20 to 40 mm, concentric zone 40 to 60 mm, and concentric zone 60 to 80 mm. Total number of branching points (Table 2). There was no significant change in the total number of branching points in the BM 20-mg/kg group. However, the total number of branching points was increased significantly in the BM 40- and 80-mg/kg groups. Number of dendritic processes (Table 1). There were no significant differences in the number of dendritic processes that arose from the somata in any of the groups that were treated with BM (20, 40 and 80 mg/kg). Six-week treatment group Dendritic intersections (Table 3). All of the groups that were treated with BM showed significant increases in dendritic intersections in the 40-mm and 60-mm concentric circles. In addition, the BM 40- and 80-mg/kg groups showed significantly increased numbers of dendritic intersections in the 20-mm, 80-mm and 100-mm concentric circles. Dendritic branching points in different concentric zones (Table 4). All of the groups that were treated with BM showed significant increases in dendritic branching points in the 20- to 40-mm and 40- to 60-mm concentric zones. In addition, the BM 40- and 80-mg/kg groups showed significantly increased numbers of dendritic branching points in the 0- to 20-mm, 60- to 80-mm and 80- to 100-mm concentric zones. Total number of branching points (Table 4). The total number of branching points was increased significantly in all three groups that were treated with BM (20, 40 and 80 mg/kg) compared with the normal control group. Number of dendritic processes (Table 3). There were no significant differences in the number of dendritic processes that arose from the somata in the group treated with BM 20 mg/kg. However, the BM 40- and 80-mg/kg groups showed significantly higher numbers of dendritic processes that arose from the somata.