Ginkgo biloba leaf extract improves the cognitive abilities of rats with D-galactose induced dementia

Abstract

Standardized Ginkgo biloba leaf extract has been used in clinical trials for its beneficial effects on brain functions, particularly in dementia. Substantial experimental evidences indicated that Ginkgo biloba leaf extract (EGB) protected neuronal cells from a variety of insults. We investigated the effect of EGB on cognitive ability and protein kinase B (PKB) activity in hippocampal neuronal cells of dementia model rats. Rats received an intraperitoneal injection of D-galactose to induce dementia. Forty-eight Spraque-Dawley rats were randomly divided into six groups, including the control group, D-galactose group (Gal), low-dose EGB group (EGB-L), mid-dose EGB group (EGB-M), high-dose EGB group (EGB-H) and treatment group. The EGB-L, EGB-M and EGB-H groups were administered with EGB and D-galactose simultaneously. Y-maze, cresyl violet staining, TUNEL assays and immunohistochemistry staining were performed to detect learning and memory abilities, morphological changes in the hippocampus, neuronal apoptosis and the expressing level of phospho-PKB, respectively. Rats in the Gal group showed decreased abilities of learning and memory, and hippocampal pyramidal cell layer was damaged, while EGB administration improved learning and memory abilities. The Gal group exhibited many stained, condensed nuclei and micronuclei, either isolated or within the cytoplasm of cells (39.5±1.4). Apoptotic cells decreased in the groups of EGB-L (35.9±0.9), EGB-M (16.8±1.0) and EGB-H (10.1±0.8), and there were statistical significances compared with the Gal group. Immunoreactivity of phospho-PKB was localized diffusely throughout the cytosol of cells in all groups, while the immunoreactivity of the Gal group was weak. EGB significantly attenuated learning and memory impairment in a dose-dependent manner, while it could decrease the nmber of TUNEL-positive cells, and increase the activity of PKB. Our results demonstrated that EGB attenuated memory impairment and cell apoptosis in galactose-induced dementia model rats by activating PKB.

Keywords: D-galactose; Ginkgo biloba extract; apoptosis; cognitive ability; dementia model; protein kinase B (PKB).

Fig. 1. Representative cresyl violet-stained coronal sections of the hippocampus from rats (×400).

Dementia in rats was induced with D-galactose. The rats were then treated with Ginkgo biloba leaf extract as detailed in Materials and Methods. A: Control group; B: D-galactose (Gal) group; C: EGB-L group; D: EGB-M group; E: EGB-H group; F: Therapy group. Normal arrangement and number of cells are seen in the hippocampus of the control group. The sections of the Gal group and treatment group show many cells with pyknotic nuclei and cytoplasm and they are deranged, especially for the Gal group. The sections from the groups of EGB-L and EGB-M demonstrate fewer cells with pyknotic nuclei and cytoplasm. Cells from the EGB-H group are identical to control, except for several deranged cells.

Fig. 2. Effects of Ginkgo biloba leaf extract on apoptosis of cells in the hippocampus of rats with D-galactose induced dementia (×400).

A: Control group; B: D-galactose (Gal) group; C: EGB-L group; D: EGB-M group; E: EGB-H group; F: Therapy group. Representative TUNEL-stained coronal sections (5 fields per section, n=8) of the hippocampus from rats. Apoptotic cells have classic condensation and fragmentation of their nuclei.

Fig. 3. Immunohistochemical analysis of phospho-AKt (ser473) in hippocampal tissues (×400).

A: Control group; B: D-galactose (Gal) group; C: EGB-L group; D: EGB-M group; E: EGB-H group; F: Therapy group. Representative sections from six groups showed that phospho-AKt (ser473) was localized in the cytosol of cells. The results are confirmed by immunostaining with at least four sections in every group. Immunoreactive intensity was the most intense in the control group and the weakest in the Gal group, while that of the EGB groups was more intense than the Gal group in a dose dependent manner.