A forced swim-based rat model of premenstrual depression: effects of hormonal changes and drug intervention

Abstract

Premenstrual dysphoric disorder (PMDD), a form of premenstrual syndrome (PMS), is a severe health disturbance that affects a patient's emotions; it is caused by periodic psychological symptoms, and its pathogenesis remains unclear. As depression-like symptoms are found in a majority of clinical cases, a reliable animal model of premenstrual depression is indispensable to understand the pathogenesis. Herein, we describe a novel rat model of premenstrual depression, based on the forced swimming test, with a regular estrous cycle. The results showed that in the estrous cycle, the depression-like behavior of rats occurred in the non-receptive phase and disappeared in the receptive phase. Following ovariectomy, the depression-like symptoms disappeared and returned after a hormone priming regimen. Moreover, fluoxetine, an anti-depressant, could reverse the behavioral symptoms in these model rats with normal estrous cycle. Further, the model rats showed significant changes in the serum levels of estrogen and progesterone, hippocampal levels of allopregnanolone, 5-hydroxytryptamine, norepinephrine, and γ-aminobutyric acid (GABA), and in the expression of GABA_A receptor 4α subunit, all of which were reversed to physiological levels by fluoxetine. Overall, we established a reliable and standardized rat model of premenstrual depression, which may facilitate the elucidation of PMS/PMDD pathogenesis and development of related therapies.

Keywords: Gabra4; animal model standardization; estrus cycle; forced swimming test; premenstrual dysphoric disorder.

Figure 1

Schedule of the experimental design including the two experimental parts. (A) Schedule of experimental part 1. (B) Schedule of experimental part 2. D1, Diestrus 1 phase; D2, Diestrus 2 phase; P/E, Proestrus/Estrus phase; M, Metestrus phase; FST, Forced swimming test; i.g., intragastrically.

Figure 2

Results of the forced swimming test after model establishment and screening. (A) Results of immobility duration. (B) Results of immobility number. (C) Results of immobility latency. N1, the first test in the nonreceptive phase; N2, the second test in the non-receptive phase; R, the test in the receptive phase. *p<0.05, **p<0.01,***p<0.001, ****p<0.0001 (n=8 in the control group and n=16 in the model group; two-way ANOVA followed by post-hoc Sidak’s multiple comparisons test).

Figure 3

Results of the forced swimming test after ovariectomy. (A) Results of immobility duration. (B) Results of immobility number. (C) Results of immobility latency. N, the test in the non-receptive phase; R, the test in the receptive phase (n=8 in the control group and n=16 in the model group; two-way ANOVA followed by post-hoc Sidak’s multiple comparisons test).

Figure 4

Results of the forced swimming test after the hormone priming regimen. (A) Results of immobility duration. (B) Results of immobility number. (C) Results of immobility latency. N, the test in the non-receptive phase; R, the test in the receptive phase. ****p<0.0001 (n=8 in the control group and n=16 in the model group; two-way ANOVA followed by post-hoc Sidak’s multiple comparisons test).

Figure 5

Results of the forced swimming test after fluoxetine treatment of the hormone-primed rats. (A) Results of immobility duration. (B) Results of immobility number. (C) Results of immobility latency. N, the test in the non-receptive phase; R, the test in the receptive phase. **p<0.01 compared to the control group, ***p<0.001 compared to the control group, ****p<0.0001 compared to the control group, ###p<0.001 compared to the model group, ####p<0.0001 compared to the model group (n=8 in each group; two-way ANOVA followed by post-hoc Sidak’s multiple comparisons test).

Figure 6

Results of the forced swimming test after fluoxetine treatment in rats with a normal estrous cycle. (A) Results of immobility duration. (B) Results of immobility number. (C) Results of immobility latency. N, the test in the non-receptive phase; R, the test in the receptive phase. *p<0.05 compared to control group, **p<0.01 compared to the control group, #p<0.05 compared to the model group, ###p<0.001 compared to the model group (n=8 in each group; two-way ANOVA followed by post-hoc Sidak’s multiple comparisons test).

Figure 7

Estrogen (E2) and progesterone (P) levels in serum. (A) E2 levels in serum. (B) P levels in serum. FLX, fluoxetine. **p<0.01 compared to the control group, ##p<0.01 compared to the model group (n=8 in each group; one-way ANOVA followed by post-hoc Tukey’s multiple comparisons test).

Figure 8

5-hydroxytryptamine (5-HT), norepinephrine (NE), γ-aminobutyric acid (GABA), and allopregnanolone (ALLO) levels in the hippocampus. (A) 5-HT levels in the hippocampus. (B) NE levels in the hippocampus. (C) GABA levels in the hippocampus. (D) ALLO levels in the hippocampus. FLX, fluoxetine. **p<0.01 compared to the control group, ***p<0.001 compared to the control group, ##p<0.01 compared to the model group, ###p<0.001 compared to the model group (n=8 in each group; one-way ANOVA followed by post-hoc Tukey’s multiple comparisons test).

Figure 9

The mRNA and protein expression levels of GABRA4 in the hippocampus. (A) The mRNA expression levels of GABRA4 in the hippocampus. (B) The protein expression levels of GABRA4 in the hippocampus. FLX, fluoxetine. *p<0.05 compared to the control group, #p<0.05 compared to the control group, ##p<0.01 compared to the model group (n=8 in each group; one-way ANOVA followed by post-hoc Tukey’s multiple comparisons test).