A Special Extract of Bacopa monnieri (CDRI-08) Restores Learning and Memory by Upregulating Expression of the NMDA Receptor Subunit GluN2B in the Brain of Scopolamine-Induced Amnesic Mice

Abstract

In the present communication, we have investigated effects of the CDRI-08, a well characterized extract of Bacopa monnieri, on expression of the GluN2B subunit of NMDAR in various brain regions of the scopolamine-induced amnesic mice. Our behavioral data reveal that scopolamine-treated amnesic mice exhibit significant decline in the spatial memory compared to the normal control mice. Our RT-PCR and immunoblotting data revealed that the scopolamine treatment resulted in a significant downregulation of the NMDAR GluN2B subunit expression in prefrontal cortex and hippocampus. Our enzyme assay data revealed that scopolamine caused a significant increase in the acetylcholinesterase activity in both the brain regions. Further, oral administration of the CDRI-08 to scopolamine-treated amnesic mice restored the spatial memory which was found to be associated with significant upregulation of the GluN2B subunit expression and decline in the acetylcholinesterase activity in prefrontal cortex as well as hippocampus towards their levels in the normal control mice. Our study provides the evidence for the mechanism underlying role of the Bacopa monnieri extract (CDRI-08) in restoring spatial memory in amnesic mice, which may have therapeutic implications.

Figure 1

Radial arm maze analysis of acquisition by mice during training period. Mouse of each experimental group was individually trained on the maze for searching food and time spent; reference memory errors, working memory errors, and reference-working memory errors were recorded. Graphs represent average value ± SEM of above parameters during acquisition trials. Data were analyzed by repetitive measures ANOVA followed by Dunnett's post hoc tests. ∗, P < 0.05 for mice groups in comparison to control within same day; #, P < 0.05 for mice groups on a particular day in comparison to that mice within groups on day 1. (a) Time taken to retrieve hidden food in radial arm maze. (b) and (c) Reference memory errors and working memory errors.

Figure 2

Radial arm maze analysis of spatial memory of mice of control and experimental groups: track record of movement mice in radial arms (a). Bar diagram showing the latency time for retrieving the hidden food (b). Mouse of each group was individually subjected to radial arm maze test and the time taken for retrieving food was recorded. Data represents mean ± SEM. C, vehicle-treated control; BME, Bacopa monnieri extract (CDRI-08) treated (200 mg/Kg/BW); SC, scopolamine-treated (2 mg/Kg BW); SC + BME, scopolamine-treated mice treated with CDRI-08. ∗ and #, P < 0.05 and ## and ∗∗, P < 0.01 were considered significant. ∗, comparison between control and other groups, and #, comparison between SC and other groups.

Figure 3

Radial arm maze tests for reference memory error (a), working memory error (b), and reference-working memory error (c). Mouse of each group was individually subjected to radial arm maze test for recording the errors. Data represents mean ± SEM. C, control; BME (Bacopa monnieri extract), CDRI-08-treated; SC, scopolamine-treated; SC + BME, scopolamine-treated mice treated with CDRI-08 as in Figure 2. ∗ and #, P < 0.05 and ## and ∗∗, P < 0.01 were considered significant. ∗, comparison between control and other groups. #, comparison between SC and remaining groups.

Figure 4

Acetylcholinesterase activity in prefrontal cortex (a) and hippocampus (b). Tissues obtained from 6-7 mice of each group were pooled and AChE activity was assayed. The AChE activity was expressed as unit/min/mg tissue. Data represents mean ± SEM. C, control; BME (Bacopa monnieri extract), CDRI-08-treated; SC, scopolamine-treated; SC + BME, scopolamine-treated mice treated with CDRI-08 as in Figure 2. ∗ and #, P < 0.05 and ## and ∗∗, P < 0.01 were considered significant. ∗, comparison between control and other groups. #, comparison between SC and remaining groups.

Figure 5

Western blot (a) and semiquantitative RT-PCR (b) analysis of GluN2B expression in prefrontal cortex. Prefrontal cortex from 6-7 mice of each group was pooled; lysates were prepared and detected for presence of GluN2B by ECL. X-ray film was scanned and the data was expressed as relative density value (RDV) by dividing the integrated density value of GluN2B by IDV of the β-actin. The data represents mean ± SEM. C, control; BME (Bacopa monnieri extract), CDRI-08-treated; SC, scopolamine-treated; SC + BME, scopolamine-treated mice treated with CDRI-08 as in Figure 2. ∗ and #, P < 0.05 and ## and ∗∗, P < 0.01 were considered significant. ∗, comparison between control and other groups. #, comparison between SC and the remaining groups.

Figure 6

Western blot (a) and semiquantitative RT-PCR (b) analysis of GluN2B expression in hippocampus. Hippocampus from 6-7 mice of each group was pooled; lysates were prepared and detected for presence of GluN2B by ECL. X-ray film was scanned and the data was expressed as relative density value (RDV) by dividing the integrated density value of GluN2B by IDV of the β-actin. The data represents mean ± SEM. C, control; BME (Bacopa monnieri extract), CDRI-08-treated; SC, scopolamine-treated; SC + BME, scopolamine-treated mice treated with CDRI-08 as in Figure 2. ∗ and #, P < 0.05 and ## and ∗∗, P < 0.01 were considered significant. ∗, comparison between control and other groups. #, comparison between SC and the remaining groups.