Protocol for alleviating alcohol-induced conditioned place preference in mice using alternating bilateral sensory stimulation

Abstract

Alternating bilateral sensory stimulation (ABS) is a clinical physical therapy technique effective in treating post-traumatic stress disorder (PTSD). However, its utilization in treating conditions beyond PTSD remains limited. Here, we present a protocol to reduce ethanol-induced conditioned place preference (CPP) using 4 Hz ABS. We describe steps for the assembly of ABS apparatus, CPP training, and ABS treatment. This protocol has been validated in male mice and can be used to explore the mechanism behind the therapeutic effect of ABS. For complete details on the use and execution of this protocol, please refer to Lei et al.¹.

Keywords: Behavior; Health Sciences; Model Organisms; Neuroscience.

Graphical abstract

Figure 1

The apparatus for ethanol-induced CPP (A) Photographs of the three-chamber place conditioning place preference apparatus. (B) Image of the chambers described in the section of experimental setup preparation. (C) Diagram of the floor texture of the two chambers.

Figure 2

Equipment set-up for ABS treatment (A) Image of the ABS apparatus. (B) Diagram of our ABS apparatus with annotated dimensions. (C) Illustration of the structure of ABS device controlled by Arduino.

Figure 3

The experimental scheme for ethanol-induced CPP and ABS treatment (A) CON, conditioned side (paired with ethanol); UN, unconditioned side (paired with saline).

Figure 4

The behavioral results of ethanol-induced CPP after ABS treatment (A–E) Statistics of preference score during pre-training test and post-training test. (two-tailed paired t test, A: p = 0.9300; B: p < 0.0001; C: p < 0.0001; D: p = 0.1516; E: p = 0.0002). (F–J) Statistics of mean time spent in the conditioning chambers during pre-training test and post-training test. (two-tailed paired t test, F: p = 0.6723; G: p < 0.0001; H: p < 0.0001; I: p = 0.2371; J: p = 0.0004). (K and L) The comparative analysis of the preference score (K) and the mean time spent in the conditioning chambers (L) from the post-training test across all groups. (for K: one-way ANOVA, p < 0.0001, post hoc Dunnett’s test, CPP vs. Control: p = 0.0009, CPP vs. CPP+1 Hz ABS: p = 0.9977, CPP vs. CPP+4 Hz ABS: p = 0.0034, CPP vs. CPP+8 Hz ABS: p = 0.9294; for L: one-way ANOVA, p < 0.0001, post hoc Dunnett’s test, CPP vs. Control: p < 0.0001, CPP vs. CPP+1 Hz ABS: p = 0.9987, CPP vs. CPP+4 Hz ABS: p = 0.0014, CPP vs. CPP+8 Hz ABS: p = 0.9702). (M and N) The comparative analysis of the preference score (M) and the mean time spent in the conditioning chambers (N) from the post-training test across all groups. (for M: one-way ANOVA, p < 0.0001, post hoc Dunnett’s test, CPP vs. Control: p = 0.0011, CPP vs. CPP+1 Hz ABS: p = 0.8892, CPP vs. CPP+4 Hz ABS: p = 0.0329, CPP vs. CPP+8 Hz ABS: p = 0.9689; for N: one-way ANOVA, p < 0.0001, post hoc Dunnett’s test, CPP vs. Control: p = 0.0005, CPP vs. CPP+1 Hz ABS: p = 0.9502, CPP vs. CPP+4 Hz ABS: p = 0.0230, CPP vs. CPP+8 Hz ABS: p = 0.9688). (O and P) Locomotor activity during pre-training test. The total distance (O) and the average speed (P) displayed no significant difference in three groups. (one-way ANOVA with post hoc Tukey’s test, O: p = 0.1290; P: p = 0.1728). (Q and R) Locomotor activity during pre-training test. The total distance (Q) and the average speed (R) displayed no significant difference in three groups. (one-way ANOVA with post hoc Tukey’s test, Q: p = 0.8306; R: p = 0.6905) Data are showed as the mean ± SEM (n = 20 mice in Control group, EtOH group, EtOH+1 Hz ABS group, and EtOH+4 Hz ABS group; n = 21 mice in EtOH+8 Hz ABS group). Error bars are SEM. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ns for no significance. Data presented in (A-R) is reprinted with permission from Lei et al.