The effects of ICAM5 overexpression on the expression levels of F-actin, G-actin, and cytoskeleton-related proteins CaMKII and Arc in the hippocampal cells of alcohol-dependent mice were analyzed using Western blot.
- Under fluorescence microscopy, a distinct red fluorescence was observed in the hippocampal region of the mice, which primarily co-localized with MAP2, indicating successful expression of AAV-ICAM5 in the neurons of the mouse hippocampal region.
- Behavioral tests: (1) Two-bottle choice drinking test: When the ethanol concentration was 3%, 6%, 9%, and 12%, there were no significant differences in ethanol consumption or preference between the EtOH group and the ICAM5 + EtOH group. When the ethanol concentration increased to 15%, during the fourth cycle of drinking, the ethanol consumption in the ICAM5 + EtOH group was significantly reduced compared to the EtOH group (P < 0.001, n = 11), and the preference for ethanol also significantly decreased (P < 0.05, n = 11). During the experiment, there were no significant differences in total liquid consumption and body weight among the four groups of mice. (2) Saccharin and quinine preference tests: In the saccharin two-bottle choice test, there were no statistical differences in the consumption and preference of different concentrations of saccharin (0.04%, 0.08%) among the four groups of mice. In the quinine two-bottle choice test, as the concentration of quinine increased (60 μM), the consumption and preference for quinine decreased in all four groups of mice, but there were no significant differences between the groups. (3) Reintroduction of 15% ethanol after 4 weeks of withdrawal: Compared to the EtOH group, the ethanol consumption and preference in the ICAM5 + EtOH group were significantly reduced (P < 0.05, n ≥ 13). (4) Novel object recognition (NOR) test: Compared to the Water group, the discrimination index of the EtOH group was significantly lower (P < 0.001, n ≥ 13). However, after overexpression of ICAM5 in the hippocampal region, the discrimination index of the mice significantly increased (P < 0.05, n ≥ 13). (5) Beam balance test (BBT): Compared to the EtOH group, the time taken to cross the balance beam was significantly shortened (P < 0.01, n ≥ 9), and the number of foot slips was significantly reduced (P < 0.001, n ≥ 9) after overexpression of ICAM5 in the hippocampal region.
