Objective  Vascular dementia (VD) is a common cognitive dysfunction associated with cerebrovascular disease. This study is aimed at investigating the therapeutic effect of anisodine hydromide (AH) on VD and the potential antioxidative stress mechanisms involved.

Methods  A VD model was established in Sprague-Dawley (SD) rats through permanent bilateral common carotid artery occlusion. The rats were divided into a sham group, a VD model group, and AH treatment groups receiving AH at low, medium, or high doses (n = 4). The neurological function of the rats in each group was evaluated using the Bederson scale, and limb coordination ability was assessed using the pole climbing test. Superoxide dismutase (SOD) and malondialdehyde (MDA) levels in the serum and brain were measured by enzyme-linked immunosorbent assay (ELISA) to assess the level of oxidative stress. In addition, apoptosis was assessed by TUNEL assay, and reactive oxygen species (ROS) levels in neuronal cells were determined using dichloro-dihydro-fluorescein diacetate (DCFH-DA) probe. The potential mechanism of action of AH on M receptors was investigated using M1-M5 inhibitors.

Results  Compared with the sham group, the nerve function and limb coordination of rats in the VD model group were significantly impaired (P < 0.01), and the SOD levels were significantly decreased in the serum (100.70 ± 18.95 U/mL vs. 44.22 ± 7.11 U/mL, P < 0.001) and the brain (131.77 ± 8.34 U/mg vs. 84.39 ± 4.10 U/mg, P < 0.01), MDA levels were significantly increased in the serum (12.03±1.01 nmol/mL vs. 17.74 ± 1.00 nmol/mL, P < 0.001) and the brain (4.41 ± 0.30 nmol/mg vs. 6.17 ± 0.70 nmol/mg, P < 0.05). AH treatment significantly improved the neurological function and limb coordination ability of VD rats. In comparison with the VD group, the high-dose AH treatment group, in particular, exhibited the most significant increase in SOD levels in the serum (44.22 ± 7.11 U/mL vs. 98.67 ± 0.86 U/mL, P < 0.001) and the brain (84.39 ± 4.10 U/mg vs. 162.83 ± 17.36 U/mg, P < 0.001), and the most significant decrease in MDA levels in the serum (17.74 ± 1.00 nmol/mL vs. 6.68 ± 0.06 nmol/mL, P < 0.001) and the brain (6.17 ± 0.70 nmol/mg vs. 3.96 ± 0.77 nmol/mg, P < 0.01). AH also reduced the number of TUNEL positive cells (P < 0.01) in a dose-dependent manner. The percentage of apoptotic cells was (36.10 ± 9.07)%, (9.60 ± 5.63)%, and (3.43 ± 0.92)%, respectively, for AH treatment at low, medium, and high concentrations, indicating that AH had an inhibitory effect on apoptosis. According to findings from the in vitro experiments, AH treatment reduced the MDA content (P < 0.01), increased the SOD activity (P < 0.01), and decreased the ROS levels of HT22 and NSC-34 cells in a dose-dependent manner. M2 receptor inhibitors could reduce the ROS level in oxidative stress injury, suggesting that AH, as an M receptor antagonist, might exert its effect by inhibiting the M2 receptor.

Conclusion AH modulates SOD and MDA levels and reduces oxidative stress injury, thereby improving neurological function and limb coordination and showing potential therapeutic effects in VD. The neuroprotective effects of AH may be related to its antioxidative stress and antiapoptotic mechanisms, and the M2 receptor may be a potential target of its actions. These findings provide an important theoretical basis for the development of new therapeutic strategies for VD.