Escitalopram Oxalate Improved Insomnia by Targeting SLC6A4 to Regulate the Hypothalamic-Pituitary-Adrenal Axis

The hypothalamus-pituitary-target gland axis is thought to be linked with insomnia, yet there has been a lack of further systematic studies to prove this. This study included 30 patients with primary insomnia (PI), 30 patients with depression-comorbid insomnia (DCI), and 30 healthy controls for exploring the alterations in the hypothalamus-pituitary-adrenal/thyroid axes’ hormones and gonadotropin-releasing hormone (GnRH). The Pittsburgh Sleep Quality Index was used to evaluate sleep quality in all subjects. The serum concentrations of corticotrophin-releasing hormone (CRH), thyrotrophin-releasing hormone (TRH), GnRH, adrenocorticotropic hormone (ACTH), thyroid stimulating hormone (TSH), cortisol, total triiodothyronine (TT3), and total thyroxine (TT4) in the morning (between 0730 h and 0800 h) were detected. Compared to the controls, all hormonal levels were elevated in the insomniacs, except ACTH and TSH in the PI group. Compared to the DCI patients, the PI patients had higher levels of CRH, cortisol, TT3, and TT4 but lower levels of TRH, GnRH, and ACTH. Spearman’s correlation analysis indicated that CRH, TRH, GnRH, TSH, cortisol, TT4, and TT3 were positively correlated with the severity of insomnia. The linear regression analysis showed that only CRH, GnRH, cortisol, and TT3 were affected by the PSQI scores among all subjects, and only CRH was included in the regression model by the “stepwise” method in the insomnia patients. Our results indicated that PI patients may have over-activity of the hypothalamus-pituitary-adrenal/thyroid axes and an elevated level of GnRH in the morning.

This study aims to investigate the mechanism through which Xiangshao Granules treat anxiety and depression using metabolomics and gut microbiota techniques, combined with animal experiments. Sixty female ICR mice were selected for the experiment and randomly divided into six groups: a control group, a model group, a low-dose Xiangshao Granules group, a medium-dose Xiangshao Granules group, a high-dose Xiangshao Granules group, and an estradiol(positive drug) group. Except for the control group, rats in other groups were induced for anxiety and depression model by ovariectomy(OVX) combined with chronic unpredictable mild stress(CUMS). After successful modeling, the mice received oral administration of Xiangshao Granules or estradiol for three weeks. Anxiety and depression behaviors in mice were evaluated using light-dark box tests, open field tests, and elevated plus-maze tests. The levels of substances closely related to anxiety and depression, such as serotonin(5-HT) and estrogen(E_2), were quantified in plasma and hippocampal tissue using enzyme-linked immunosorbent assay(ELISA). Metabolomics and 16S rDNA amplicon sequencing techniques were employed to analyze the regulatory effects of Xiangshao Granules on plasma metabolites and metabolic pathways in anxiety and depression mice, as well as their impact on the distribution of gut microbiota. Finally, the correlation between plasma metabolites and differential gut microbiota was constructed using the Spearman correlation coefficient method. Behavioral experimental results indicated that, compared to the control group, the model group exhibited significantly decreased dwell time in the light box, reduced total distance in the open field, and diminished dwell time in the open arm. In contrast, high dose of Xiangshao Granules were found to increase the dwell time in the light box and total distance in the open field. ELISA results indicated that the levels of 5-HT, gamma-aminobutyric acid(GABA), E_2 were significantly decreased, luteinizing hormone(LH), adrenocorticotropic hormone(ACTH), and corticosterone(CORT) were significantly elevated in the anxiety and depression mice, and treatment with middle, high dose of Xiangshao Granules reversed the levels of these substances. Additionally, in the anxiety and depression mouse model, the levels of follicle-stimulating hormone(FSH) were significantly increased, whereas middle, high dose of Xiangshao Granules decreased FSH levels. Metabolomics analysis revealed that Xiangshao Granules significantly changed the metabolic profile of the anxiety and depression mice, affecting central carbon metabolism, amino acid biosynthesis, and ABC transporter pathways. The results from 16S rDNA amplicon sequencing showed that Xiangshao Granules improved the relative abundance of genera such as Bacteroidia, Bacilli, Lactobacillales, and Lactobacillus. Spearman correlation analysis indicated a close association between specific differential gut microbiota and plasma differential metabolites. This study suggests that Xiangshao Granules significantly ameliorate anxiety and depression symptoms in mice by altering the levels of substances associated with these conditions, including 5-HT, GABA, E_2, LH, and ACTH. The metabolomics and gut microbiota data suggest that the therapeutic mechanism may be closely related to the regulation of amino acid biosynthesis, central carbon metabolism, and the alteration of key microbial community compositions.

e16604 Background: Ectopic ACTH and/or CRH-secreting pheochromocytomas are rare catecholamine-secreting tumors originating from adrenal medulla, which could also secrete ectopic adrenocorticotropic hormone (ACTH) and corticotrophin-releasing hormone (CRH), mimicking Cushing syndrome and complicating the diagnosis clinically. The extremely low incidence limits the comprehensive understanding and management for specific individuals accompanied with this kind of tumor. Furthermore, mechanisms of tumorigenesis and ectopic ACTH secretion in pheochromocytomas are unknown. Studying this rare type of tumor will help tailor therapies to the specifically molecular and metabolic characteristics of pheochromocytomas. Methods: Fresh tumor and peritumoral specimens were collected from three ACTH+&CRH+ pheochromocytoma patients and three ACTH-&CRH- pheochromocytoma patients, and then were digested and sent to construct single-cell library as well as sequencing. Subsequent analysis and immunohistochemical staining were conducted to explore the difference of ACTH+&CRH+ pheochromocytomas from the ACTH-&CRH- ones and normal tissues. Results: We constructed the linear trajectory of 2 types of pheochromocytomas. The cluster of intermediate cells that differentiated into mature ACTH+&CRH+ pheochromocytomas showing high expression of HES1, and the immunohistochemical staining revealed that HES1 was higher in ACTH+&CRH+ ones. Besides, Higher expression of POMC and PNMT was observed in ACTH+&CRH+ pheochromocytomas compared with ACTH-&CRH- ones, being in accordance with the clinical information that three ACTH+&CRH+ pheochromocytoma patients had relatively higher catecholamine levels. T cells in ACTH+&CRH+ pheochromocytomas were activated by signals, like HLA-B, promoting their cytotoxic functions, while T cells in ACTH-&CRH- ones were subject to inhibitory signals, especially from M2 macrophages. What's more, a population of endothelial cells marked by SELE was identified in ACTH+&CRH+ pheochromocytomas, showing strong interaction with CD8+ T cells and facilitating lymphocyte infiltration, particularly the GZMK⁺IL7R⁺ T cells. While ACTH-&CRH- pheochromocytomas were enriched in glycolysis and HIF signaling pathways, and characterized by lacking T cell infiltration, with macrophages being the dominant immune cells. Conclusions: HES1 plays a significant role in the fate transition of pheochromocytomas and may drive ACTH secretion. In addition, ACTH+&CRH+ pheochromocytomas are considerated as relatively immunologically“hot”tumors but poor in angiogenesis. In contrast, ACTH-&CRH- pheochromocytomas lack immune infiltration while exhibiting extensive angiogenesis. Immunotherapies may be effective for the former, and the latter may benefit from anti-angiogenic therapies.

The present study aimed at further investigating trait aspects of sleep-related cognitive arousal and general cognitive arousal and their association with both objective and subjective sleep parameters in primary insomnia patients. A clinical sample of 182 primary insomnia patients and 54 healthy controls was investigated using 2 nights of polysomnography, subjective sleep variables, and a questionnaire on sleep-related and general cognitive arousal. Compared to healthy controls, primary insomnia patients showed both more sleep-related and general cognitive arousal. Furthermore, sleep-related cognitive arousal was closely associated with measures of sleep-onset and sleep-maintenance problems, while general cognitive arousal was not. Cognitive-behavioral treatment for insomnia might benefit from dedicating more effort to psychological interventions that are able to reduce sleep-related cognitive arousal.

Guhan Yangsheng Jing mitigates hippocampal neuronal pyroptotic injury and manifies learning and memory capabilities in sleep deprived mice via the NLRP3/Caspase1/GSDMD signaling pathway

Post-traumatic stress disorder (PTSD) is a long-term mental disorder. Zhi zhu xiang (ZZX) which is the root and rhizome of Valeriana jatamansi Jones has been used for anti-anxiety and sedation. Because of PTSD occurring in combination with anxiety and depression, this study aimed at exploring anti-PTSD activity of ZZX and possible mechanisms. Effects 95% ethanol extract of ZZX on behavior of PTSD mice induced by single prolonged stress & foot-shock were evaluated by open field test (OFT), elevated plus maze (EPM), force swimming test (FST) and fear conditioning response (FCR). Potentially active components, targets and pathways of ZZX involved in PTSD were analyzed by network pharmacology and molecular docking. Enzyme-linked immunosorbent assay was used to detect levels of neurotransmitters, including 5-hydroxytryptamine (5-HT), norepinephrine (NE), dopamine (DA), and γ-aminobutyric acid (GABA) and glutamic acid (Glu) in the hippocampus, as well as the levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT) and corticotropin releasing hormone (CRH) in the hypothalamic pituitary adrenal (HPA) axis of serum. Levels of cannabinoid receptor1 (CB1), diacylglycerol lipase α (DAGLα), monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) mRNA in the hippocampus were detected by real-time PCR. The results showed that ZZX (80 mg·kg-1, ZZX-M) increased PTSD mice' movement distance, residence time and frequency of exploring in the central area of OFT and the open arm of EPM, and reduced the immobility time of mice in the FST and the freezing time of mice in the FCR. Results of network pharmacology and molecular docking showed that 47 potentially active compounds of ZZX exerted an anti-PTSD effect by acting on key targets, including CNR1(same as CB1), MAOA, NR3C1, MAPK14, MAPK8, HTR2C, DRD2 through multiple signaling pathways such as serotonergic, dopaminergic, glutamatergic and retrograde endocannabinoid signaling. The experimental results showed that ZZX-M restored levels of 5-HT, NE, DA, GABA, Glu in the hippocampus and ACTH, CORT, CRH in serum. Meanwhile, ZZX-M up-regulated levels of CB1 and DAGLα genes, but down-regulated levels of MAGL and FAAH genes in the hippocampus. This study demonstrates that ZZX alleviates PTSD-like behavior in mice by restoring neurotransmitters and HPA axis and regulating endocannabinoid related genes.

Post-traumatic stress disorder (PTSD) is a long-term mental disorder. Zhi zhu xiang (ZZX) which is the root and rhizome of Valeriana jatamansi Jones has been used for anti-anxiety and sedation. Because of PTSD occurring in combination with anxiety and depression, this study aimed at exploring anti-PTSD activity of ZZX and possible mechanisms. Effects 95% ethanol extract of ZZX on behavior of PTSD mice induced by single prolonged stress & foot-shock were evaluated by open field test (OFT), elevated plus maze (EPM), force swimming test (FST) and fear conditioning response (FCR). Potentially active components, targets and pathways of ZZX involved in PTSD were analyzed by network pharmacology and molecular docking. Enzyme-linked immunosorbent assay was used to detect levels of neurotransmitters, including 5-hydroxytryptamine (5-HT), norepinephrine (NE), dopamine (DA), and γ-aminobutyric acid (GABA) and glutamic acid (Glu) in the hippocampus, as well as the levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT) and corticotropin releasing hormone (CRH) in the hypothalamic pituitary adrenal (HPA) axis of serum. Levels of cannabinoid receptor1 (CB1), diacylglycerol lipase α (DAGLα), monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) mRNA in the hippocampus were detected by real-time PCR. The results showed that ZZX (80 mg·kg-1, ZZX-M) increased PTSD mice' movement distance, residence time and frequency of exploring in the central area of OFT and the open arm of EPM, and reduced the immobility time of mice in the FST and the freezing time of mice in the FCR. Results of network pharmacology and molecular docking showed that 47 potentially active compounds of ZZX exerted an anti-PTSD effect by acting on key targets, including CNR1(same as CB1), MAOA, NR3C1, MAPK14, MAPK8, HTR2C, DRD2 through multiple signaling pathways such as serotonergic, dopaminergic, glutamatergic and retrograde endocannabinoid signaling. The experimental results showed that ZZX-M restored levels of 5-HT, NE, DA, GABA, Glu in the hippocampus and ACTH, CORT, CRH in serum. Meanwhile, ZZX-M up-regulated levels of CB1 and DAGLα genes, but down-regulated levels of MAGL and FAAH genes in the hippocampus. This study demonstrates that ZZX alleviates PTSD-like behavior in mice by restoring neurotransmitters and HPA axis and regulating endocannabinoid related genes.

Post-traumatic stress disorder (PTSD) is a long-term mental disorder. Zhi zhu xiang (ZZX) which is the root and rhizome of Valeriana jatamansi Jones has been used for anti-anxiety and sedation. Because of PTSD occurring in combination with anxiety and depression, this study aimed at exploring anti-PTSD activity of ZZX and possible mechanisms. Effects 95% ethanol extract of ZZX on behavior of PTSD mice induced by single prolonged stress & foot-shock were evaluated by open field test (OFT), elevated plus maze (EPM), force swimming test (FST) and fear conditioning response (FCR). Potentially active components, targets and pathways of ZZX involved in PTSD were analyzed by network pharmacology and molecular docking. Enzyme-linked immunosorbent assay was used to detect levels of neurotransmitters, including 5-hydroxytryptamine (5-HT), norepinephrine (NE), dopamine (DA), and γ-aminobutyric acid (GABA) and glutamic acid (Glu) in the hippocampus, as well as the levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT) and corticotropin releasing hormone (CRH) in the hypothalamic pituitary adrenal (HPA) axis of serum. Levels of cannabinoid receptor1 (CB1), diacylglycerol lipase α (DAGLα), monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) mRNA in the hippocampus were detected by real-time PCR. The results showed that ZZX (80 mg·kg-1, ZZX-M) increased PTSD mice' movement distance, residence time and frequency of exploring in the central area of OFT and the open arm of EPM, and reduced the immobility time of mice in the FST and the freezing time of mice in the FCR. Results of network pharmacology and molecular docking showed that 47 potentially active compounds of ZZX exerted an anti-PTSD effect by acting on key targets, including CNR1(same as CB1), MAOA, NR3C1, MAPK14, MAPK8, HTR2C, DRD2 through multiple signaling pathways such as serotonergic, dopaminergic, glutamatergic and retrograde endocannabinoid signaling. The experimental results showed that ZZX-M restored levels of 5-HT, NE, DA, GABA, Glu in the hippocampus and ACTH, CORT, CRH in serum. Meanwhile, ZZX-M up-regulated levels of CB1 and DAGLα genes, but down-regulated levels of MAGL and FAAH genes in the hippocampus. This study demonstrates that ZZX alleviates PTSD-like behavior in mice by restoring neurotransmitters and HPA axis and regulating endocannabinoid related genes.

Post-traumatic stress disorder (PTSD) is a long-term mental disorder. Zhi zhu xiang (ZZX) which is the root and rhizome of Valeriana jatamansi Jones has been used for anti-anxiety and sedation. Because of PTSD occurring in combination with anxiety and depression, this study aimed at exploring anti-PTSD activity of ZZX and possible mechanisms. Effects 95% ethanol extract of ZZX on behavior of PTSD mice induced by single prolonged stress & foot-shock were evaluated by open field test (OFT), elevated plus maze (EPM), force swimming test (FST) and fear conditioning response (FCR). Potentially active components, targets and pathways of ZZX involved in PTSD were analyzed by network pharmacology and molecular docking. Enzyme-linked immunosorbent assay was used to detect levels of neurotransmitters, including 5-hydroxytryptamine (5-HT), norepinephrine (NE), dopamine (DA), and γ-aminobutyric acid (GABA) and glutamic acid (Glu) in the hippocampus, as well as the levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT) and corticotropin releasing hormone (CRH) in the hypothalamic pituitary adrenal (HPA) axis of serum. Levels of cannabinoid receptor1 (CB1), diacylglycerol lipase α (DAGLα), monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) mRNA in the hippocampus were detected by real-time PCR. The results showed that ZZX (80 mg·kg-1, ZZX-M) increased PTSD mice' movement distance, residence time and frequency of exploring in the central area of OFT and the open arm of EPM, and reduced the immobility time of mice in the FST and the freezing time of mice in the FCR. Results of network pharmacology and molecular docking showed that 47 potentially active compounds of ZZX exerted an anti-PTSD effect by acting on key targets, including CNR1(same as CB1), MAOA, NR3C1, MAPK14, MAPK8, HTR2C, DRD2 through multiple signaling pathways such as serotonergic, dopaminergic, glutamatergic and retrograde endocannabinoid signaling. The experimental results showed that ZZX-M restored levels of 5-HT, NE, DA, GABA, Glu in the hippocampus and ACTH, CORT, CRH in serum. Meanwhile, ZZX-M up-regulated levels of CB1 and DAGLα genes, but down-regulated levels of MAGL and FAAH genes in the hippocampus. This study demonstrates that ZZX alleviates PTSD-like behavior in mice by restoring neurotransmitters and HPA axis and regulating endocannabinoid related genes.

To observe the clinical effect of Tiaoshen Guben holistic therapy of acupuncture and moxibustion (holistic treatment with acupuncture and moxibustion by adjusting the mind and consolidating the root) on comorbidity of depression and insomnia. Twenty-four patients with comorbidity of depression and insomnia were included and treated with Tiaoshen Guben holistic therapy of acupuncture and moxibustion. Acupuncture was applied to Baihui (GV20), Guanyuan (CV4), bilateral Neiguan (PC6), etc. The refined moxibustion therapy was delivered at Zhongwan (CV12), Qihai (CV6), bilateral Yongquan (KI1), etc. Subcutaneous embedding therapy with thumb-tack needle was adopted at bilateral Xinshu (BL15), bilateral Pishu (BL20), etc. The intervention was operated once every other day, 3 treatments a week, and for 6 consecutive weeks. Before and after treatment completion, and in 1 month after treatment, Pittsburgh sleep quality index (PSQI) and Hamilton's depression scale (HAMD-17) were adopted to assess sleep quality and depression symptoms in the patients, respectively. Before and after treatment completion, using functional magnetic resonance imaging (fMRI), the functional connectivity (FC) of locus coeruleus (LC) in brain regions was evaluated; and the levels of serum norepinephrine (NE), cortisol (CORT), adrenocorticotropic hormone (ACTH) and corticotropin releasing hormone (CRH) were detected. Compared with the scores before treatment, PSQI and HAMD-17 scores after treatment completion and in 1 month after treatment were reduced (P<0.01); and strengthened FC was revealed between the right LC and the pars opercularis of the left inferior frontal gyrus, as well as the lateral occipital lobe region. After treatment completion, serum NE was elevated (P<0.01), the levels of CORT, ACTH and CRH were reduced (P<0.01). Before and after treatment completion, the difference in FC between the right LC and the pars opercularis of the left inferior frontal gyrus was negatively correlated with the differences in PSQI score (r = -0.484, P = 0.016) and HAMD-17 score (r = -0.233, P = 0.027). Tiaoshen Guben holistic therapy of acupuncture and moxibustion can effectively alleviate depression symptoms and improve sleep quality in the patients with comorbidity of depression and insomnia, which is obtained probably through reducing the levels of serum CORT, ACTH and CRH, increasing serum NE, strengthening the FC of the right LC with the pars opercularis of the left inferior frontal gyrus and the lateral occipital lobe region.

The corticotropin releasing factor (CRF) type 1 receptor (CRF1) is a class B family G protein-coupled receptor that regulates the hypothalamic-pituitary-adrenal stress axis. Astressin is an amino-terminal truncated analog of CRF that retains high affinity binding to the extracellular domain of the receptor and is believed to act as a neutral competitive antagonist of receptor activation. Here we show that despite being unable to activate the CRF1 receptor, astressin binding results in the internalization of the receptor. Furthermore, entirely different pathways of internalization of CRF1 receptors are utilized following CRF and astressin binding. CRF causes the receptor to be phosphorylated, recruit beta-arrestin2, and to be internalized rapidly, likely through clathrin-coated pits. Astressin, however, fails to induce receptor phosphorylation or beta-arrestin2 recruitment, and internalization is slow and occurs through a pathway that is insensitive to inhibitors of clathrin-coated pits and caveolae. The fate of the internalized receptors also differs because only CRF-induced internalization results in receptor down-regulation. Furthermore, we present evidence that for astressin to induce internalization it must interact with both the extracellular amino terminus and the juxtamembrane domain of the receptor. Astressin binds with 6-fold higher affinity to full-length CRF1 receptors than to a chimeric protein containing only the extracellular domain attached to the transmembrane domain of the activin IIB receptor, yet two 12-residue analogs of astressin have similar affinities for both proteins but are unable to induce receptor internalization. These data demonstrate that agonists and antagonists for CRF1 receptors promote distinct conformations, which are then differentially regulated.

The aim of the present study was to examine the effects of corticotropin-releasing factor (CRF) in conscious dogs and to determine whether the stimulation of adrenocorticotropic hormone (ACTH) release by angiotensin II (ANG II) results from potentiation of the action of CRF. In addition, the possible role of CRF in the stimulation of vasopressin released by ANG II was investigated. The following experiments were performed: 1) intravenous saline infusion; 2) ANG II (10 ng.kg-1.min-1) alone; 3) vasopressin (1 ng.kg-1.min-1) alone; 4) CRF (0.001, 0.01, or 0.1 microgram/kg iv) bolus; 5) vasopressin (1 ng.kg-1.min-1) and CRF (0.1 microgram/kg) together; 6) CRF (0.001, 0.01, or 0.1 microgram/kg) and ANG II (10 ng.kg-1.min-1) together; 7) ANG II (10 ng.kg-1.min-1) followed 15 min later with CRF (0.001, 0.01, or 0.1 microgram/kg). Each dose of CRF was tested on a different day. Infusion of ANG II alone stimulated the release of ACTH, cortisol, and vasopressin. Administration of CRF produced dose-dependent increases in plasma ACTH and cortisol concentrations, and the highest dose of CRF increased plasma vasopressin concentration. CRF given together with ANG II did not potentiate the stimulation of ACTH release by CRF. Vasopressin at the dose tested did not stimulate ACTH release but potentiated the ACTH response to CRF. ANG II stimulated vasopressin release but did not potentiate the AVP response to CRF. These results show that, in conscious dogs, ANG II and CRF each increase plasma ACTH concentration and that the ACTH response to CRF is potentiated by vasopressin but not by ANG II.(ABSTRACT TRUNCATED AT 250 WORDS)

To observe the effect of electroacupuncture (EA) at "Hegu" (LI4) and "Taichong" (LR3) on DNA methylation of the solute carrier family 6 member 4 (SLC6A4) gene promoter region in the hippocampus of depressed rats, and to explore the potential antidepressant mechanism of EA. Thirty male Sprague-Dawley rats were randomly divided into a blank group, a model group, a medication group, a 5-Azacytidine (5-AZA) group, and an EA group, 6 rats in each group. Depression models were established in the model group, the medication group, the 5-AZA group, and the EA group using chronic unpredictable mild stress (CUMS) combined with solitary housing. The medication group was treated with intragastric administration of fluoxetine hydrochloride capsules; the 5-AZA group was treated with intraperitoneal injection of 5-AZA; the EA group was treated with EA at bilateral "Hegu" (LI4) and "Taichong" (LR3), with disperse-dense wave, frequency of 2 Hz/100 Hz, and intensity of 1-1.2 mA, 20 min each session. All the treatment was given in three groups once daily for 21 consecutive days. Behavioral changes were evaluated by sucrose preference test, open field test, and novelty-suppressed feeding test. Serum levels of serotonin (5-HT), dopamine (DA), and norepinephrine (NE) were measured by ELISA. The expression of SLC6A4 and 5-HT1AR protein and mRNA in hippocampus was detected by Western blot and real-time quantitative PCR, respectively. DNA methylation status of the SLC6A4 promoter region in hippocampal tissue was analyzed by bisulfite sequencing PCR (BSP). Compared with the blank group, the model group showed decreased sucrose preference, reduced total locomotor distance, and prolonged latency to feeding (P<0.05), decreased serum 5-HT, DA, and NE levels (P<0.05), downregulated hippocampal SLC6A4 and 5-HT1AR protein and mRNA expression (P<0.05), and increased CpG site methylation rate of the SLC6A4 promoter region (P<0.05). Compared with the model group, the medication group, the 5-AZA group, and the EA group exhibited increased sucrose preference, increased total locomotor distance, shortened latency to feeding (P<0.05), elevated serum 5-HT, DA, and NE levels (P<0.05), upregulated hippocampal SLC6A4 and 5-HT1AR protein and mRNA expression (P<0.05), and reduced CpG site methylation rate of the SLC6A4 promoter (P<0.05). Compared with the medication group and the 5-AZA group, the EA group showed higher sucrose preference, greater total locomotor distance, shorter latency to feeding (P<0.05), and increased serum DA and NE levels (P<0.05). EA could improve depressive behaviors in depressed rat models. The underlying mechanism may involve inhibition of SLC6A4 hypermethylation in the hippocampus on the serotonergic system, upregulation of SLC6A4 and 5-HT1AR protein and mRNA expression, and elevation of monoamine neurotransmitters such as 5-HT.

In response to stress corticotropin releasing hormone (CRH) is secreted from the hypothalamus and travels to the anterior pituitary where it stimulates the release of proteins derived from the precursor protein, proopiomelanocortin (POMC), including adrenocorticotropic hormone (ACTH) and β-endorphin. β-endorphin interacts with opioid receptors and relieves stress while ACTH travels through the blood stream to the adrenal cortex where it stimulates the release of cortisol. Cortisols actions include releasing glucose from stored glycogen and stimulating breakdown of fat from adipose tissue, induction of gluconeogenesis and modulation of the immune system. As a negative feedback control, cortisol inhibits CRH release from the hypothalamus and ACTH from the anterior pituitary. During pregnancy the placenta synthesizes and secretes CRH into the systemic circulation causing a dramatic increase in the circulating levels of CRH throughout pregnancy that peaks at delivery and unlike the situation in the hypothalamus, cortisol has a positive effect on placental CRH production. While the CRH binding protein (CRH-BP) may attenuate the action of placental CRH on the pregnant womans pituitary, there is evidence that the CRH-BP does not completely cut off access of placental CRH to the pituitary. For example, CRH-BP levels decrease in late pregnancy lowering the ability to decrease CRH activity. In late pregnancy when circulating CRH levels rise dramatically and superimpose over falling levels of the CRH-BP, the anterior pituitary may desensitize to placental CRH so that ACTH and cortisol levels do not become pathologically elevated. In non-pregnant individuals, disturbances in the hypothalamic CRH - pituitary ACTH axis are associated with mood disturbances and in pregnant women who have elevated CRH levels during pregnancy an association with prenatal depression has been shown. More work needs to be done to investigate the possible roles of CRH in pregnancy related depression. Besides effects on the pituitary, placental CRH may exert physiological regulation on the uterus. Higher than normal levels of CRH in mid-pregnancy are associated with preterm pregnancy suggesting a role for CRH in the timing of parturition. In preterm pregnancy when the uterus has to be kept quiescent, CRH receptors are coupled to various cell signaling systems including the cyclic AMP system that inhibits myosin light chain phosphorylation in smooth muscle thereby dampening uterine contraction. As the time for labour approaches, CRH receptors may alter coupling to cellular signaling systems that phosphorylate myosin light chain and initiate rhythmic contraction of the uterus. The possibility of using modulators of CRH action such as CRH receptor agonists to correct pathological conditions in the expectant mother provides a powerful motivation for future studies to fully elucidate the roles of CRH in pregnancy. Keywords: Corticotrophin releasing hormone (CRH), CRH binding protein (CRH-BP), CRH receptors, placenta, uterus, pregnancy, depression

Objective: Probiotics have been reported to exert beneficial effects on sleep through the gut-brain axis. Therefore, this randomized, double-blind, placebo-controlled trial assessed the effects of Lacticaseibacillus paracasei 207-27 supplementation on sleep quality and its safety and potential mechanisms. Method and study design: Healthy adults under mild stress aged 18-35 years consumed low or high doses of L. paracasei 207-27 or a placebo for 28 days. Fecal samples, blood samples, and questionnaires were collected at the baseline and the end of the intervention. Sleep quality was measured using wearable devices and Pittsburgh sleep quality index (PSQI) questionnaire. Serum inflammatory markers, corticotropin-releasing hormone, adrenocorticotropic hormone (ACTH), cortisol (COR), γ-aminobutyric acid, and 5-hydroxytryptamine levels were detected using enzyme-linked immunosorbent assay. The gut microbiota was analyzed using 16S rRNA sequencing and bioinformatics. Short-chain fatty acids levels were detected using gas chromatography-mass spectrometry. Results: Both the low-dose and high-dose groups exhibited significant improvements in wearable device- measured sleep duration compared to the placebo group. The global scores of PSQI in three groups significantly decreased after intervention without statistical difference between groups. At the phylum level, the low-dose group exhibited a higher relative abundance of Bacteroidota and a lower Firmicutes-to-Bacteroidetes (F/B) ratio. At the genus level, two treatment groups had higher relative abundance of Bacteroides and Megamonas, alongside lower levels of Escherichia-Shigella. Furthermore, the low-dose group exhibited significant increases in acetic acid, propionic acid, butyric acid, and valeric acid levels, while two treatment groups exhibited a significant decrease in COR levels. Correlation analysis revealed that the increased levels of acetic acid and butyric acid in the low-dose group may be associated with decreased ACTH. Conclusion: L. paracasei 207-27 administration in healthy adults resulted in improvements in gut microbiota community and sleep duration. The mechanisms might involve modulation of the gut microbiota structure to regulate the function of the gut-brain axis, including increases in SCFA levels and decreases in hypothalamic-pituitary-adrenal axis activity. The Chinese clinical trial registry number is ChiCTR2300069453 (https://www.chictr.org.cn/showproj.html?proj=191193, registered 16 May 2023 - retrospectively registered).