Oral Administration of Efavirenz Dysregulates the Tph2 Gene in Brain Serotonergic Areas and Alters Weight and Mood in Mice

Abstract

Most HIV-antiretroviral drugs have adverse effects. Efavirenz (EFV) is an example of a drug with neuropsychiatric effects, such as anxiety, depression, and suicidal thoughts, in people living with HIV (PLWH). The mechanisms by which EFV causes neuropsychiatric alterations in PLWH are complex, multifactorial, and not fully understood, although several studies in animals have reported changes in brain energy metabolism, alterations in monoamine turnover, GABA, and glutamate levels, and changes in 5-HT receptors. In this report, we studied the effects of EFV on the serotonergic system in healthy mice, specifically, whether EFV results in alterations in the levels of the tryptophan hydroxylase 2 (Tph2) gene in the brain. EFV (10 mg/kg) and distilled water (1.5 µL/kg) (control group) were orally administered to the mice for 36 days. At the end of the treatment, Tph2 expression levels in mouse brains were measured, and mood was evaluated by three trials: the forced swim test, elevated plus maze, and open field test. Our results revealed dysregulation of Tph2 expression in the brainstem, amygdala, and hypothalamus in the EFV group, and 5-HT levels increased in the amygdala in the EFV group. In the behavioral tests, mice given EFV exhibited a passive avoidance response in the forced swim test and anxiety-like behavior in the elevated plus maze, and they lost weight. Herein, for the first time, we showed that EFV triggered dysregulation of the Tph2 gene in the three serotonergic areas studied; and 5-HT levels increased in the amygdala using the ELISA method. However, further studies will be necessary to clarify the increase of 5-HT in the amygdala as well as understand the paradoxical decrease in body weight with the simultaneous increase in food consumption. It will also be necessary to measure 5-HT by other techniques different from ELISA, such as HPLC.

Keywords: 5-HT; anxiety; depression; efavirenz; tryptophan hydroxylase 2.

Figure 1

Effects of EFV on Tph2 expression (A) and 5-HT levels (B) in the brainstem, amygdala, and hypothalamus in mice. Diminished Tph2 mRNA expression following EFV administration compared to that in the control group is shown (A). Each dot represents three pooled samples of tissues from three different animals. In contrast, 5-HT levels were increased after EFV administration only within the amygdala (B). Each dot represents two pooled samples of tissues from three different animals. Unpaired t test with Welch’s correction. * p < 0.05; *** p < 0.001; **** p < 0.0001.

Figure 2

Behavioral tests. Three behavioral paradigms were used in this study to assess depression-like behavior (force swim test) or anxiety (open field test and elevated plus-maze test) in mice following 36 days of oral EFV administration (10 mg/kg). (A) A significant increase in immobility time was observed in the EFV group compared to the control group. Unpaired t test with Welch’s correction (F_3,3 = 6.409; p < 0.05). (B) In the open field test, no difference was detected between the EFV group and the control group (Welch’s t test; F_4,5 = 2.950; p > 0.05). (C) In the elevated plus maze test, a significantly lower percentage of time spent in close arms and in the center was observed in mice treated with EFV, and the percentage of entries increased (Welch’s t test; p < 0.01). The percentage of time spent on the EPM test was calculated as the time spent on the arms in seconds divided by the total duration spent on the EPM test, which was 300 s × 100%. Each dot represents an animal. The values are the means ± SEMs. Welch’s t test. * p < 0.05; ** p < 0.01.

Figure 3

Effects of EFV on body weight (A) and food consumption (B). Twenty-four mice per group were fed and housed in groups of six with water available ad libitum in each home cage (n = 48, 8 cages). Every day, the mice received EFV (10 mg/kg) via the oral route or distilled water (1.5 µL/kg) via the oral route to determine whether EFV alters body weight or feeding. The body weight of each mouse and the food consumption of each cage were measured daily. This experiment continued until 36 days after EFV administration. A significant decrease in body weight was observed in the EFV group, beginning with respect to the final weight (F_35,74 = 4.828; p < 0.0001). With respect to food intake, an increase in consumption was observed in the EFV group compared with the control group (F_{21, 21} = 6.498; p < 0.0001). The distribution of the values of the studied parameters was tested for normality using the Shapiro–Wilk test. Body weight was analyzed with two-way repeated-measures ANOVA followed by the Tukey post hoc test. Food intake was analyzed by an unpaired t test. The values are the means ± SEMs. **** p < 0.0001.