Effects of Aged Garlic Extract on Cholinergic, Glutamatergic and GABAergic Systems with Regard to Cognitive Impairment in Aβ-Induced Rats

Abstract

Alzheimer's disease (AD) has been linked to the degeneration of central cholinergic and glutamatergic transmission, which correlates with progressive memory loss and the accumulation of amyloid-β (Aβ). It has been claimed that aged garlic extract (AGE) has a beneficial effect in preventing neurodegeneration in AD. Therefore, the objective of this study was to investigate the effects of AGE on Aβ-induced cognitive dysfunction with a biochemical basis in the cholinergic, glutamatergic, and GABAergic systems in rats. Adult male Wistar rats were orally administered three doses of AGE (125, 250, and 500 mg/kg) daily for 65 days. At day 56, they were injected with 1 μL of aggregated Aβ (1-42) into each lateral ventricle, bilaterally. After six days of Aβ injection, the rats' working and reference memory was tested using a radial arm maze. The rats were then euthanized to investigate any changes to the cholinergic neurons, vesicular glutamate transporter 1 and 2 proteins (VGLUT1 and VGLUT2), and glutamate decarboxylase (GAD) in the hippocampus. The results showed that AGE significantly improved the working memory and tended to improve the reference memory in cognitively-impaired rats. In addition, AGE significantly ameliorated the loss of cholinergic neurons and increased the VGLUT1 and GAD levels in the hippocampus of rat brains with Aβ-induced toxicity. In contrast, the VGLUT2 protein levels did not change in any of the treated groups. We concluded that AGE was able to attenuate the impairment of working memory via the modification of cholinergic neurons, VGLUT1, and GAD in the hippocampus of Aβ-induced rats.

Keywords: Alzheimer’s disease; aged garlic extract; amyloid-β; cholinergic neurons; glutamate decarboxylase; radial arm maze; vesicular glutamate transporters.

Figure 1

Schematic diagram of drug treatment and behavioural tests. Rats were injected with Aβ (1–42) into both sides of the lateral ventricle after 56 days of drug treatments. (RAM: radial arm maze test, Aβ: amyloid-β (1–42)).

Figure 2

The schematic diagram of the eight-arm radial maze. The animals were tested in the RAM with a 5 min delay.

Figure 3

Number of working memory errors and reference memory errors made while looking for the baited arm in the RAM on day 6 after Aβ injection. Data are presented as mean ± S.E.M. (n = 8), *, ** = significant differences from the vehicle control group at p < 0.05 and 0.01, respectively; ^#,^## = significant differences from the vehicle + Aβ group at p < 0.05 and 0.01, respectively.

Figure 4

The neuroprotective effect of AGE (aged garlic extract) on cholinergic neurons in the hippocampal region of Aβ-induced rats. (A–F) represent the photomicrographs of brain section showing the distribution of cholinergic neurons by double staining of Nissl stain with cresyl violet and immunohistochemistry stained with polyclonal ChAT (choline acetyltransferase) antibodies in the vehicle control group (A and B), vehicle + Aβ (C), AGE125 + Aβ (D), AGE250 + Aβ (E) and AGE 500 + Aβ (F). (G) represents the number of ChAT neurons. The cholinergic neurons are indicated with arrows. Data are presented as mean ± S.E.M. (n = 8), *** = significant differences from the vehicle control group at p < 0.001 and ^## = significant differences from the vehicle + Aβ group at p < 0.01.

Figure 5

Effects of AGE on amounts of VGLUT1 (A), VGLUT2 (B) and GAD 65 and 67 (C) in the rat hippocampus. Data are presented as mean ± S.E.M (n = 8), **, *** = significant differences from the vehicle control group at p < 0.01 and 0.001 respectively ^##, ^### = significant differences from the vehicle + Aβ group at p < 0.01 and 0.001, respectively. VGLUT1 and VGLUT2, vesicular glutamate transporter 1 and 2 proteins; GAD, glutamate decarboxylase.