Depression stands out as a leading cause of heightened self-harm and suicide rates, significantly compromising the well-being and overall health of women in the perimenopausal stage[25]. The exploration of the pathogenesis and therapeutic drugs of PMD are urgently needed. While there are various hypotheses concerning the pathogenesis of PMD, there is a medical consensus that ovarian dysfunction is a crucial factor contributing to the development of PMD[26]. Hence, while estrogen has emerged as a primary research direction for PMD, the controversy surrounding its impact on female depression and the significant side effects of estrogen replacement therapy have impeded the progress of estrogen-focused investigations into the pathogenesis of PMD[27]. ALLO, as a metabolite of progesterone and a positive allosteric modulator of GABAAR, plays a crucial role in female neurocognitive disorders such as postpartum depression and premenstrual dysphoric disorder. Scholars in the field of PMD research have gradually recognized the significance of ALLO in the context of PMD[28].
Compared to the Sham group, the OVX + CUMS group exhibited a significant decrease in serum levels of E2, 5-HT, and NE, accompanied by depressive-like behavior. The observed changes are consistent with the clinical presentation of PMD patients, indicating the successful establishment of PMD rats in this study. The significant decrease in ALLO and GABA levels in the serum of OVX + CUMS group aligns with previous clinical research findings[14, 29–31]. As a positive allosteric modulator of GABAAR, ALLO is likely to be involved in the pathogenesis of PMD by regulating the expression of GABAAR subunits. Therefore, we employed Q-PCR to assess the mRNA expression of Gabra4, Gabrb2, and Gabrd in brain regions. The study revealed a significant upregulation of Gabra4 and Gabrb2 mRNA expression in the prefrontal cortex, hippocampus, and hypothalamus of the OVX + CUMS group, along with a significant increase in Gabrd mRNA expression in the hypothalamus and amygdala. Many laboratories have demonstrated that ALLO withdrawal leads to a transient increase in the expression of GABAAR α4 subunit in various brain regions of the hippocampus in female mice, as well as in an in vitro neuronal system[32, 33]. The above evidence supports the findings of this study. GABAR β2 subunit is associated with neuropsychiatric disorders including bipolar disorder, epilepsy, autism spectrum disorder, Alzheimer's disease, depression, and premenstrual dysphoric disorder[34]. Compared to wild-type mice, Gabrb2 knock-out mice exhibit lower levels of depression[35, 36]. In comparison to normal young mice, perimenopausal mice exhibit a reduced mRNA expression of GABAAR β2 in the prefrontal cortex[37]. Furthermore, there is no significant difference in prefrontal cortex GABAAR β2 mRNA expression between perimenopausal mice and those subjected to chronic unpredictable stress[37]. Clinical investigations have observed a notable reduction in mRNA transcription levels of the GABAAR β2 subunit in the prefrontal cortex among individuals suffering from depression and mood disorders[38]. Due to GABAARβ2 expression being subject to epigenetic regulation, and given that epigenetic regulation varies with development, genotype, and disease states, the expression of GABAARβ2 in different mood disorders is not consistent[39, 40]. Many studies identifie a close association between the GABAAR δ subunit and depressive emotions. In comparison to the Sham group, the OVX + CUMS group showed no significant difference in Gabrd mRNA expression in the prefrontal cortex and hippocampus, while there was a significant increase in the hypothalamus and amygdala brain regions. Zhang[24] found that the expression of the δ subunit showed no significant difference between liver-qi stagnation syndrome rats with premenstrual dysphoric disorder and normal rats, moreover, overexpression of the δ subunit in normal rats did not induce depressive behavior. Currently, there is no research reporting the relationship between Gabrd and PMD, and most literature suggests that the decrease in δ subunit expression is associated with depressive emotions[37]. Therefore, we speculate that the GABAAR δ subunit may not be involved in the occurrence and development of PMD.The significant increase in mRNA expression of the δ subunit in the hypothalamus and amygdala in this study may be related to the decrease in ALLO. A study found that compared to the pregnancy period, postpartum mice showed an increasing trend in the expression of GABAAR δ subunit[41].
The existing evidence preliminarily indicates that ALLO and the GABAergic system are associated with the pathogenesis of PMD. To further validate the hypothesis that the decrease in ALLO levels mediates changes in the mRNA expression of GABAAR subunits, leading to the onset of PMD, this study administered ALLO through intraperitoneal injection to increase ALLO levels in PMD rats. And than we observed depressive-related indicators, ALLO levels in serum, and changes in GABAAR subunit mRNA expression in the brain regions of PMD rats. The study findings indicate that intraperitoneal injection of ALLO increased the serum ALLO levels in PMD rats. This improvement in ALLO levels was associated with amelioration of depressive-like behavior in rats and an increase in serum levels of E2, 5-HT, and NE, suggesting that ALLO has a therapeutic effect on PMD. Following ALLO intervention, a significant decrease in Gabra4 mRNA expression was observed in the prefrontal cortex, and a significant increase in Gabra4 mRNA expression was noted in the hippocampus of PMD rats. Furthermore, there were significant reductions in Gabrb2 mRNA expression in the prefrontal cortex and the hypothalamus, while significant increases in Gabrd mRNA expression were observed in the prefrontal cortex and hippocampus. Many studies have also found that ALLO can enhance the expression of α4 and δ subunits in the hippocampus of female rats[42–44].Based on relevant literature and the results of previous studies, we believe that ALLO-mediated expression of Gabra4, Gabrb2 in the prefrontal cortex, and Gabrb2 in the hypothalamus is one of the pathological mechanisms of PMD. At the same time, the α4 and δ subunits in the prefrontal cortex and hippocampus may be potential targets for the therapeutic effects of ALLO on PMD. Interestingly, we found a significant increase in hippocampal Gabra4 mRNA expression in PMD rats, and even after ALLO intervention, Gabra4 mRNA expression remained significantly elevated. Therefore, further research is needed to explore the underlying reasons for this observation.
Rutin, a monomer derived from traditional Chinese herbal medicine, exhibits antidepressant effects with minimal toxic side effects[45]. This study revealed that rutin treatment significantly ameliorated depressive-like behavior in PMD rats, elevated serum levels of 5-HT and ALLO, and concurrently reduced the mRNA expression of Gabra4 and Gabrb2 in the prefrontal cortex. The optimal therapeutic effect for PMD was observed in the low-dose group of rutin (8.65 mg/kg). Based on preliminary research, we propose that rutin exerts its therapeutic effects on PMD by upregulating ALLO levels in the serum, subsequently mediating the downregulation the mRNA expression of Gabra4 and Gabrb2 in the prefrontal cortex.
This study, for the first time, investigated the mRNA expression of ALLO-mediated GABAAR α4, β2, and δ subunits in multiple brain regions of PMD rats. By pinpointing the brain regions and subunits regulated by ALLO, it has provided numerous clues and insights for subsequent research into the pathogenesis of PMD.Simultaneously, it was discovered that rutin possesses therapeutic effects on PMD, offering a novel option for clinical treatment of PMD. However, this study has several limitations. For instance, it only investigated the mRNA expression of ALLO-mediated GABAAR α4, β2, and δ subunits in emotion-related brain regions. Subsequent research should continue to explore the protein expression of these subunits mediated by ALLO. The overall functional expression of GABAARs is crucial in the pathogenesis of PMD. Various GABAARs contain α4, β2, and δ subunits, with the α4β2δGABAR receptor having the closest association with depression and ALLO. Therefore, future studies should emphasize the impact of ALLO on the function of α4β2δGABAR.