An integrated liver, hippocampus and serum metabolomics based on UPLC-Q-TOF-MS revealed the therapeutical mechanism of Ziziphi Spinosae Semen in p-chlorophenylalanine-induced insomnia rats.

Ziziphi Spinosae Semen (ZSS), a well-known herbal medicine for treating insomnia, is popular in not only China but also in Europe, India and Iran. However, its underlying mechanisms remain unclear. In this work, taking the targeted organs of insomnia, the liver and hippocampus, as the objects, a combination metabolomics based on ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was established to illustrate the abnormality of metabolic characteristics of the liver, hippocampus and serum of p-chlorophenylalanine (PCPA)-induced insomnia rats and to demonstrate the mechanism of ZSS in treating insomnia. The results showed that ZSS could restore the brain cell morphology, decrease the degree of hepatocyte necrosis and regulate the disturbance of neurotransmitters and hormones in insomnia rats. In terms of metabolomics, a total of 33 liver metabolites, 25 hippocampal metabolites and 18 serum metabolites were finally selected as the potential biomarkers and an important pathway of phenylalanine, tyrosine and tryptophan biosynthesis was common in three tissues in PCPA rats. Meanwhile, ZSS significantly reversed the levels of 23 liver metabolites, 15 hippocampal metabolites and 5 serum metabolites. The present study demonstrates the actions of ZSS in treating insomnia by enhancing both cerebral and hepatic functions.

Effects of aqueous extracts and volatile oils prepared from Huaxiang Anshen decoction on p-chlorophenylalanine-induced insomnia mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Insomnia occurs frequently in modern society, and its common symptoms include difficulty in falling asleep and decreased sleep quality and time, memory, and attention. With the advantages of having few side-effects and reduced drug-dependence, a compound traditional Chinese medicine (TCM) prescription called Huaxiang Anshen Decoction (HAD) has been widely used in clinical practice in China mainly for primary insomnia treatment. Although the effects of volatile oils from TCM herbs have been increasingly reported, volatile oils in HAD are conventionally neglected because of its preparation process and clinical usage. Therefore, exploring the anti-insomnia effects of volatile oils from HAD is of great importance. AIM OF THE STUDY: The sedative and hypnotic effects of the conventional aqueous extracts, the volatile oils from HAD, and their combinations were investigated. METHODS: The main components in HAD volatile oils (HAD-Oils), were analyzed through gas chromatography-mass spectrometry (GC-MS). The HAD volatile oil inclusion complex (HAD-OIC) was prepared with ß-cyclodextrin, and characterized. P-chlorophenylalanine (PCPA) was used to induce insomnia mice model and the test groups of HAD aqueous extract (HAD-AE), HAD-OIC and their combination (AE-OIC). An open field test was used in evaluating the mice's activities, and the levels of 5-hydroxytryptamine (5-HT) in mice sera, glutamate (Glu) in the hypothalamus, and γ-aminobutyric acid (γ-GABA) and dopamine (DA) in the brain tissues were assayed by enzyme-linked immunosorbent assay (ELISA). RESULTS: A total 74 components in HAD-Oil were determined by GC/MS, and cyperenone (20.46%) and α-cyperone (10.39%) had the highest relative content. The characterization results of the physical phase showed that volatile oils were successfully encapsulated by ß-cyclodextrin and HAD-OIC was produced. The average encapsulation rates of cyperenone and α-cyperone were 79.93% and 71.96%, respectively. The results of pharmacology study showed that all the test groups increased the body weight and decreased voluntary activity when compared with the model group (P < 0.05). The HAD-AE, HAD-OIC, and AE-OIC groups increased the levels of 5-HT in the sera and DA and Glu/γ-GABA in the brains, and AE-OIC groups showed better performance than the other test groups. CONCLUSIONS: HAD-Oil exerts sedative and hypnotic effects, which are increased when it is used with HAD-AEs. This result provides a favorable experimental evidence that volatile oils should be retained for the further development of HAD.

Efficacy of electroacupuncture stimulating Shenmen (HT7), Baihui (GV20), Sanyinjiao (SP6) on spatial learning and memory deficits in rats with insomnia induced by para-chlorophenylalanine: a single acupoint combined acupoints.

OBJECTIVE: To investiage the effect of electroacupuncture (EA) at a single acupoint of Shenmen (HT7), Baihui (GV20), Sanyinjiao (SP6) and at combined acupoints of Shenmen (HT7) and Baihui (GV20) and Sanyinjiao (SP6) on the PKA/CREB and BDNF/TrkB signaling, as well as neuroapoptosis and neurogenesis in hippocampus and elucidate the underlying mechanism of single and combined acupoints on ameliorating spatial learning and memory deficits in a rat model of primary insomnia. METHODS: Primary insomnia was modeled by intraperitoneal injection of para-chlorophenylalanine (PCPA) once daily for 2 d. EA was applied at Shenmen (HT7), Baihui (GV20), Sanyinjiao (SP6), or Shenmen (HT7) + Baihui (GV20) + Sanyinjiao (SP6) (combined) for 30 min daily for 4 d. Spatial learning and memory function was evaluated by the Morris water maze (MWM) test. Protein expressions of hippocampal cAMP-dependent protein kinase (PKA)-Cß, phosphorylated cAMP-responsive element-binding protein (p-CREB), brain-derived neurotrophic factor (BDNF), and tyrosine kinase receptor B (TrkB) were evaluated by Western blotting. Neuronal apoptosis in the hippocampus was detected with the transferase-mediated dUTP-X nick end labeling assay. Endogenous neurogenesis was examined with bromodeoxyuridine staining. The MWM test and hippocampal p-CREB, BDNF, and TrkB protein levels in the combined acupoints group were evaluated after the administration of a PKA-selective inhibitor (H89). RESULTS: Spatial learning and memory were significantly impaired in rats with insomnia. The spatial learning deficits were ameliorated in the Shenmen (HT7), Baihui (GV20), Sanyinjiao (SP6), and combined groups; this improvement was significantly greater in the combined group than the single acupoint groups. The spatial memory impairment was improved in the combined, Baihui (GV20), and Shenmen (HT7) groups, but not the Sanyinjiao (SP6) group. The expressions of PKA-Cß, p-CREB, BDNF, and TrkB were decreased in rats with insomnia. All these proteins were significantly upregulated in the combined group. PKA/p-CREB protein levels were elevated in the Baihui (GV20) and Shenmen (HT7) groups, whereas BDNF/TrkB expression was upregulated in the Sanyinjiao (SP6) group. The staining results showed significant attenuation of hippocampal cell apoptosis and increased numbers of proliferating cells in the combined group, whereas the single acupoint groups only showed decreased numbers of apoptotic cells. In the combined group, the PKA inhibitor reversed the improvement of spatial memory and upregulation of p-CREB expression caused by EA, but did not affect its activation of BDNF/TrkB signaling. CONCLUSIONS: EA at the single acupoints Baihui (GV20), Shenmen (HT7), or Sanyinjiao (SP6) had an ameliorating effect on the spatial learning and memory deficits induced by insomnia. EA at combined acupoints exerted a synergistic effect on the improvements in spatial learning and memory impairment in rats with insomnia by upregulating the hippocampal PKA/CREB and BDNF/TrkB signaling, facilitating neurogenesis, and inhibiting neuronal apoptosis. These findings indicate that EA at combined acupoints [(Baihui (GV20), Shenmen (HT7), and Sanyinjiao (SP6)] achieves a more pronounced regulation of hippocampal neuroplasticity than EA at single acupoints, which may partly explain the underlying mechanisms by which EA at combined acupoints exerts a better ameliorative effect on the cognitive dysfunction caused by insomnia.

Electroacupuncture of the cymba concha alleviates p-chlorophenylalanine-induced insomnia in mice.

OBJECTIVE: Insomnia is the most common sleep disorder and is often comorbid with mental and physical diseases. The present study was designed to investigate the hypnotic effect of electroacupuncture (EA) of the cymba concha to stimulate the auricular branch of the vagus nerve (ABVN). METHODS: Mice were intraperitoneally injected with p-chlorophenylalanine (PCPA, 300 mg/kg·d) for 2 days to induce insomnia and subsequently received EA or manual acupuncture (MA) of the cymba concha for 30 min once daily for 5 consecutive days, or no treatment. The phenobarbital-induced sleep test was used to analyze the hypnotic effects and the open field test was used to analyze the locomotor activities and anxiolytic effects of EA/MA of the cymba concha. In addition, the levels of gamma-aminobutyric acid (GABA) and glutamate (Glu) in the hypothalamus and peripheral blood were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: PCPA injection significantly decreased sleep duration, increased sleep latency and induced anxiety-like behaviors in mice. In PCPA-insulted mice, EA of the cymba concha improved the sleep disturbance by significantly prolonging sleep duration, while no change in sleep latency was observed. Moreover, EA of the cymba concha improved PCPA-induced anxiety-like behaviors without decreasing locomotor activities in the open field test. EA of the cymba concha increased the level of GABA in the hypothalamus and peripheral blood, while Glu concentrations remained unchanged. CONCLUSION: These findings indicate that EA of the region innervated by the ABVN upregulates GABA levels in the hypothalamus and ameliorates the symptoms of insomnia and anxiety, suggesting that EA of the cymba concha might have potential value as an intervention for insomnia.

A comprehensive study on the relieving effect of Lilium brownii on the intestinal flora and metabolic disorder in p-chlorphenylalanine induced insomnia rats.

CONTEXT: The bulb of Lilium brownii F. E. Brown (Liliaceae) (LB) is a common Chinese medicine to relieve insomnia. OBJECTIVE: To investigate the molecular mechanism of LB relieving insomnia. MATERIALS AND METHODS: Insomnia model was induced by intraperitoneally injection p-chlorophenylalanine (PCPA) in Wistar rats. Rats were divided into three groups: Control, PCPA (400 mg/kg, i.p. 2 days), LB (598.64 mg/kg, oral 7 days). The levels of 5-hydroxytryptamine (5-HT), norepinephrine (NE), melatonin (MT), and the expression of GABAA, 5-HT1A and MT receptors, as well as pathological changes in hypothalamus, were evaluated. 16S rDNA sequencing and UPLC-MS/MS were used to reveal the change of the intestinal flora and metabolic profile. RESULTS: The adverse changes in the abundance and diversity of intestinal flora and faecal metabolic phenotype altered by PCPA in rats were reversed after LB treatment, accompanied by the up-regulated levels of 5-HT as 8.14 ng/mL, MT as 16.16 pg/mL, 5-HT1A R and GABAA R, down-regulated level of NE as 0.47 ng/mL, and the improvement of pathological phenomena of cells in the hypothalamus. And the arachidonic acid metabolism and tryptophan metabolism pathway most significantly altered by PCPA were markedly regulated by LB. Besides, it was also found that LB reduced the levels of kynurenic acid related to psychiatric disorders and trimethylamine-N-oxide associated with cardiovascular disease. CONCLUSION: The mechanism of LB relieving insomnia involves regulating flora and metabolites to resemble the control group. As a medicinal and edible herb, LB could be considered for development as a health-care food to relieve increasing insomniacs in the future.

Armillaria mellea fermentation liquor ameliorates p-chlorophenylalanine-induced insomnia associated with the modulation of serotonergic system and gut microbiota in rats.

In China, Armillaria mellea (Vahl) P. Kumm. has been used as a folk medicine to treat insomnia for several hundred years. However, the underlying mechanisms involved are currently unknown. In this study, the anti-insomnia efficacy of A. mellea fermentation liquor (AFL) was evaluated in p-chlorophenylalanine-induced insomnia rats by measuring the serotonergic systems and gut microbiota. Our results demonstrate that all doses of AFL significantly reduced locomotor activity and alleviated decreasing weights in insomnia rats. Further, AFL exhibited better sedative effects by reducing sleep latency and increasing sleep duration in pentobarbital-treated rats. AFL treatment also elevated serum glutathione peroxidase and superoxide dismutase levels, while reducing serum interleukin-6, tumor necrosis factor-α, and interleukin-1ß levels. Furthermore, AFL alleviated insomnia by enhancing 5-hydroxytryptamine content and the expression 5-HT1A and 5-HT2A receptor in the hippocampus. Meanwhile, AFL treatment normalized the composition of gut microbiota in insomnia-model rats, while increasing relative abundance of Lachnospiraceae, Ruminococcaceae, and Saccharimonadaceae restores the gut microbial ecosystem altered in insomnia rats. The experiments show that A. mellea alleviated insomnia by modulating serotonergic system and gut microbiota. PRACTICAL APPLICATIONS: Insomnia has become a serious health issue of global concern. As a well-known traditional Chinese medicine, Armillaria mellea has been clinically employed in the treatment of insomnia for centuries in Asia with significant efficacy. In the present study, we firstly reported A. mellea fermentation liquor potentially relieved insomnia rats by alteration of gut microbiota and serotonergic systems and could guide future clinical studies. As a popular edible and medicinal mushroom, A. mellea also can be potentially used in the development and production of novel food products in the future.

Comprehensive 16S rDNA Sequencing and LC-MS/MS-Based Metabolomics to Investigate Intestinal Flora and Metabolic Profiles of the Serum, Hypothalamus and Hippocampus in p­Chlorophenylalanine-Induced Insomnia Rats Treated with Lilium brownie.

Gut microbiota homeostasis in the organism and insomnia have been reported to influence each other. In the study, a method of 16S rRNA gene sequencing combined with ultra-high performance liquid chromatography-mass/mass spectrometry was adopted to evaluate the effects of Lilium brownie (LB) on intestinal flora and metabolic profiles of serum, hypothalamus and hippocampus in insomnia rat induced by p­chlorophenylalanine (PCPA). It was observed that the imbalance in the diversity and abundance of gut microbiota induced by PCPA was restored after LB intervention. Among these, the Porphyromonadaceae, Lactobacillus and Escherichia were significantly adjusted at the genus level by PCPA and LB, respectively. It was also found that the most of metabolic phenotypes in serum, hypothalamus and hippocampus perturbed by PCPA were regulated towards normal after LB intervention, especially 5-hydroxy-L-tryptophan of the hypothalamus involving in 5-HT metabolism. Moreover, the arachidonic acid metabolism in serum, hypothalamus and hippocampus, and the serotonergic synapse in hypothalamus and hippocampus were the most fundamentally and significantly affected pathways after LB intervention. The results of correlation analysis showed that several floras including Pseudoruegeria have an outstanding contribution to the change of differential metabolites. In brief, the results confirm that gut microbiota is significantly returned to normal and may interact with the corresponding metabolites to relieve insomnia under LB intervention.

The effect of Bailemian on neurotransmitters and gut microbiota in p-chlorophenylalanine induced insomnia mice.

Bailemian (BLM) is reportedly used for the treatment of insomnia as a traditional Chinese medicine in China for many years. However, the anti-insomnia mechanisms of BLM are still unknown. The present study aims to investigate the anti-insomnia activity of BLM by evaluating its influence on the relevant neurotransmitters and gut microbiota in p-chlorophenylalanine (PCPA) induced insomnia mice. The results indicated that the level of GABA, 5-HT, DA, and NE is significantly decreased in the PCPA-induced insomnia model group compared with the control group, while the level of Glu is higher than the control group. Treatment with BLM could ameliorate the symptoms of insomnia and significantly modulate the levels of the neurotransmitters mentioned above in brain and colonic faeces. Furthermore, the structure and composition of gut microbiota were changed after the administration of BLM and can increase the percentage of beneficial bacterial species in gut microbiota. These results indicated that Bailemian could ameliorate the symptoms of insomnia, and its effects may be through modification of the neurotransmitters levels and gut microbiota composition.

iTRAQ-based proteomics analysis on insomnia rats treated with Mongolian medical warm acupuncture.

OBJECTIVE: To explore the proteomic changes in the hypothalamus of rats treated with Mongolian medical warm acupuncture for insomnia therapy based proteomics. METHOD: We used an iTRAQ-based quantitative proteomic approach to identify proteins that potential molecular mechanisms involved in the treatment of insomnia by Mongolian medical warm acupuncture. RESULT: In total, 7477 proteins were identified, of which 36 proteins showed increased levels and 45 proteins showed decreased levels in insomnia model group (M) compared with healthy control group (C), 72 proteins showed increased levels and 44 proteins showed decreased levels from the warm acupuncture treated insomnia group (W) compared with healthy controls (C), 28 proteins showed increased levels and 17 proteins showed decreased levels from the warm acupuncture-treated insomnia group (W) compared with insomnia model group (M). Compared with healthy control groups, warm acupuncture-treated insomnia group showed obvious recovered. Bioinformatics analysis indicated that up-regulation of neuroactive ligand-receptor interaction and oxytocin signaling was the most significantly elevated regulate process of Mongolian medical warm acupuncture treatment for insomnia. Proteins showed that increased/decreased expression in the warm acupuncture-treated insomnia group included Prolargin (PRELP), NMDA receptor synaptonuclear-signaling and neuronal migration factor (NSMF), Transmembrane protein 41B (TMEM41B) and Microtubule-associated protein 1B (MAP1B) to adjust insomnia. CONCLUSION: A combination of findings in the present study suggest that warm acupuncture treatment is efficacious in improving sleep by regulating the protein expression process in an experimental rat model and may be of potential benefit in treating insomnia patients with the added advantage with no adverse effects.

Shuangxia decoction alleviates p-chlorophenylalanine induced insomnia through the modification of serotonergic and immune system.

As a Traditional Chinese Medicine (TCM), Shuangxia Decoction (SXD) has been used to treat insomnia in oriental countries for more than thousands of years and it presents remarkable clinical effects. However, its active pharmacological fraction and the mechanism of sedative-hypnotic effects have not been explored. In this paper, we investigated active pharmacological fraction and revealed the detailed mechanisms underlying the sedative-hypnotic effects of SXD. It showed that SXD water extract compared to ethanol extract possessed better sedative effects on locomotion activity in normal mice and increased sleep duration in subhypnotic dose of sodium pentobarbital-treated mice. SXD alleviated p-chlorophenylalanine (PCPA) -induced insomnia by increasing the content of 5-HT in cortex [F (4, 55) = 12.67], decreasing the content of dopamine (DA) and norepinephrine (NE). Furthermore, SXD enhanced the expression of 5-HT1A and 5-HT2A receptors in hypothalamic and reduced serum levels of IL-1,TNF-α [F (5, 36) = 15.58]. In conclusion, these results indicated that SXD produced beneficial sedative and hypnotic bioactivities mediated by regulating the serotonergic and immune system.

Determination of five neurotransmitters in the rat brain for the study of the hypnotic effects of Ziziphi Spinosae Semen aqueous extract on insomnia rat model by UPLC-MS/MS.

Ziziphi Spinosae Semen (ZSS) has been used for treatment of insomnia in China for centuries. To reveal the influence of insomnia on the levels of the neurotransmitters including serotonin (5-HT), glutamic acid (Glu), γ-aminobutyric acid (GABA), noradrenaline (NE) and dopamine (DA), and to study the role of ZSS aqueous extract in the treatment of insomnia, an UPLC-ESI- MS/MS method was developed and validated for simultaneous determination of five neurotransmitters in the rat brain. The brain samples were pretreated by one-step direct protein precipitation with acetonitrile. The analytes were detected in positive mode with multiple reaction monitoring (MRM) and the procedure was completed in less than 10 min. The method showed a good linearity (R2 > 0.9967) with the other validation parameters were within acceptance range. The results indicated that the concentration of 5-HT, GABA and DA is significantly lower (P < 0.01) in para-chlorophenylalanine (PCPA)-induced insomnia rat model group, while Glu and NE significantly higher than those in control group (P < 0.01). Treatment with ZSS aqueous extract (4 or 8 g·kg-1·d-1 for seven days) could ameliorate the symptoms of insomnia by significantly changing the levels of the neurotransmitter parameters mentioned above. The data obtained in this study demonstrate that ZSS aqueous extract could ameliorate the symptoms of insomnia by modulating the levels of monoamines and amino acid neurotransmitters in the brain.

Anticonvulsive activity of duloxetine: A new choice for the epileptic patients with depression.

The aim of this study was to investigate the antiepileptic effects of duloxetine in the maximal electroshock test and convulsions induced by four compounds: Pentylenetetrazole, 3-mercaptopropionic acid, thiosemicarbazide, and bicuculline. Duloxetine exhibited moderate anticonvulsive activity with an ED50 (median effective dose) of 48.21 mg/kg in the maximal electroshock test in mice. The anticonvulsive action of duloxetine was also confirmed in chemical-induced seizure tests, where this drug decreased tonic convulsions. Single administration of duloxetine (6.25-25 mg/kg) significantly increased the anticonvulsant effects of valproate, carbamazepine, and oxcarbazepine in the maximal electroshock test. Furthermore, pretreatment with thiosemicarbazide (an inhibitor of GABA synthesis enzyme) significantly increased the ED50 of duloxetine, suggesting the GABAergic system may contribute to the anticonvulsive action of duloxetine. These results support the use of duloxetine in the treatment of coexisting depression and epilepsy.

An extract of Synedrella nodiflora (L) Gaertn exhibits antidepressant properties through monoaminergic mechanisms.

Synedrella nodiflora (SNE) has been used traditionally for many neurological conditions and some of these neuroactive effects have been scientifically substantiated. The usefulness of SNE in depression has however not been investigated despite the availability of data in other disease models indicating it may be useful. The present study therefore examined the effect of SNE in acute murine models of depression and the possible mechanisms mediating its activities in these models. Preliminary qualitative phytochemical and high performance liquid chromatography (HPLC) screening were conducted on SNE. The behavioural effects of SNE (100, 300 and 1000 mg/kg) pre-treated mice were examined in the forced swimming (FST) and tail suspension (TST) tests. Behavioural events such as mobility (swimming, climbing, curling and climbing), and immobility, were scored. The possible involvement of monoamines in the effects of SNE was assessed in the TST by pre-treating mice with α-methyldopa, reserpine and para-chlorophenylalanine (pCPA) in separate experiments. Flavonoids, tannins, saponins, alkaloids, cardiac glycosides, coumarins, triterpenes, sterols, anthraquinones and phenolic compounds were present in SNE. HPLC analysis revealed the presence of two major constituents observed at retention times 42.56 and 46.51 min, with percentage composition of 45.72% and 36.88% respectively. SNE significantly reduced immobility scores in both FST and TST, suggesting antidepressant effects. The antidepressant properties of SNE were reversed by the pre-treatment of α-methyldopa, reserpine and pCPA, suggesting a possible involvement of monoamines (noradrenaline and serotonin) in its mechanism(s) of actions. SNE exhibits antidepressant effects, possibly mediated through an interplay of enhancement of noradrenergic and serotoninergic mechanisms.

Evidence for the Involvement of Monoaminergic Pathways in the Antidepressant-Like Activity of Cymbopogon citratus in Mice.

Objectives Depression is a complex neuropsychiatric disorder, which affects the quality of life of the sufferers and treatment approach is associated with serious adverse effects and sometimes therapeutic failures. Cymbopogon citratus leaf (CC) has been reported to exert anti-depressant effect but its mechanism of action is yet to be elucidated hence, the need for this study. Methods The anti-depressant-like effect of Cymbopogon citratus aqueous leaf was evaluated using forced swim test (FST), tail suspension test (TST) and yohimbine-induced lethality test (YLT) in aggregated mice. Interaction studies involving p-chlorophenylalanine (pCPA), an inhibitor of serotonin biosynthesis and yohimbine, α2-adrenergic receptor antagonist were carried out to evaluate the role of monoaminergic system in the anti-depressant-like effect of CC. The effect of CC on spontaneous motor activity (SMA) was also assessed using activity cage. ResultsCymbopogon citratus (25 and 50 mg/kg, p.o.) demonstrated antidepressant-like activity devoid of significant stimulation of the SMA in mice. However, the antidepressant-like property of CC was significantly (p<0.05) attenuated by pretreatment with yohimbine suggesting involvement of noradrenergic pathway in the action of the extract. Also, pCPA reversed the anti-immobility effect of CC, indicating the role of serotonergic system in the mediation of its antidepressant activity. Moreover, CC (25 and 50 mg/kg) potentiated the lethal effect of yohimbine in aggregated mice, which further suggest the involvement of monoaminergic systems in its action. Conclusions The results of the study showed that C. citratus might be interacting with serotonergic and noradrenergic pathways to mediate its anti-depressant-like effect in mice.

Serotonergic and dopaminergic systems are implicated in antidepressant-like effects of chotosan, a Kampo formula, in mice.

We previously demonstrated that chotosan (CTS), a traditional herbal formula called Kampo medicine, improves diabetes-induced cognitive deficits. In the present study, we investigated the antidepressant-like effects of CTS in mice. The administration of CTS (1.0 g/kg, for 3 days) decreased the immobility time in the forced-swim test, and this decrease was prevented by the prior administration of sulpiride (an antagonist of D2/3 receptors) and WAY100635 (an antagonist of 5-HT1A receptors). None of the treatments tested altered the locomotor activity of mice. These results suggest that CTS exerts antidepressant-like effects through changes in the serotonergic and dopaminergic systems.

cis-3-Hexenol and trans-2-hexenal mixture prevents development of PTSD-like phenotype in rats.

Several green leaf volatiles have anxiolytic/antidepressant properties and attenuate adrenocortical stress response in rodents. However, it remains unknown whether a mixture of cis-3-hexenol and trans-2-hexenal so-called 'green odor (GO)' affects fear-associated post-traumatic stress disorder (PTSD)-like behavior. In the present study, fear memory of the initial conditioning stimulus was stably maintained by weekly presentation of conditioned tone. Examination of open field behavior, acoustic startle response, prepulse inhibition, and immobility in the forced swim test for 2 weeks after initial conditioning revealed that conditioned rats sustained anxiety, enhanced startle response, hypervigilance, depression-like behavior, and hypocortisolism, which is consistent with PTSD symptoms. Daily, not acute, GO presentation facilitated fear extinction and reduced PTSD-like behavioral and endocrinal responses. To further investigate the mechanism of effect of GO, we examined the effect of paroxetine (a selective serotonin reuptake inhibitor), p-chlorophenylalanine (PCPA, an irreversible serotonin synthesis inhibitor), alone or in combination of GO on PTSD-like phenotype. The alleviative effects of GO were masked by simultaneous paroxetine administration. PCPA-induced serotonin depletion abolished the effects of GO. Our results suggest that daily GO presentation facilitates fear extinction and prevents development of PTSD-like symptoms.

Inhalation of Cedrus atlantica essential oil alleviates pain behavior through activation of descending pain modulation pathways in a mouse model of postoperative pain.

ETHNOPHARMACOLOGICAL RELEVANCE: Cedrus atlantica essential oil (CaEO) presents analgesic and anti-inflammatory sedative properties. However, it remains unknown whether CaEO alleviates acute postoperative pain. MATERIALS AND METHODS: Here, we investigated the effect of CaEO on postoperative pain and its mechanisms related to the descending pain control in Swiss males mice induced by a plantar incision surgery (PIS) in the hindpaw. RESULTS: Inhalation of CaEO (5', 30' or 60') markedly reduced mechanical hypersensitivity. This effect was prevented by pre-treatment with naloxone or p-chlorophenylalanine methyl ester (PCPA, 100mg/kg, i.p.)-induced depletion of serotonin. In addition, p-alpha-methyl-para-tyrosin (AMPT, 100mg/kg, i.p.)-induced depletion of norepinephrine, intraperitoneal injection of the α2-adrenergic receptor antagonist yohimbine (0.15 mg/kg, i.p.) or haloperidol (1mg/kg, i.p.) an antagonist of dopaminergic (D1 and D2) receptors prevented the effect of CaEO on hypersensitivity. CONCLUSIONS: These findings suggest that CaEO alleviates postoperative pain by activating the descending pain modulation pathways on the opioidergic, serotonergic, noradrenergic (α2-adrenergic) and dopaminergic (dopamine D1 and D2 receptors) systems.

In silico Analysis and Experimental Validation of Lignan Extracts from Kadsura longipedunculata for Potential 5-HT1AR Agonists.

OBJECTIVES: Kadsura longipedunculata (KL) has been widely used for the treatment of insomnia in traditional Chinese medicine. The aim of this study was to explore the mechanism of the sedative and hypnotic effects of KL. MATERIALS AND METHODS: The content of KL was evaluated by HPLC-TOF-MS, and a potential target was found and used to construct its 3D structure to screen for potential ligands among the compounds in KL by using bioinformatics analysis, including similarity ensemble approach (SEA) docking, homology modeling, molecular docking and ligand-based pharmacophore. The PCPA-induced insomnia rat model was then applied to confirm the potential targets related to the sedative effects of KL by performing the forced swimming test (FST), the tail suspension test (TST) and the measurement of target-related proteins using western blotting and immunofluorescence. RESULTS: Bioinformatics analysis showed that most of lignan compounds in KL were optimal ligands for the 5-HT1A receptor (5-HT1AR), and they were found to be potential targets related to sedative effects; the main lignan content of KL extracts was characterized by HPLC-TOF-MS, with 7 proposed lignans detected. Administration of KL could significantly reduce FST and TST immobility time in the PCPA-induced 5HT-depleted insomnia rat model. The expressions of proteins related to the 5-HT1AR pathway were regulated by extracts of KL in a concentration-dependent manner, indicating that extracts of KL had 5-HT1AR agonist-like effects. CONCLUSION: In silico analysis and experimental validation together demonstrated that lignan extracts from KL can target 5-HT1AR in insomniac rats, which could shed light on its use as a potential 5-HT1AR agonist drug.

Investigation of the GPR39 zinc receptor following inhibition of monoaminergic neurotransmission and potentialization of glutamatergic neurotransmission.

Zinc can regulate neural function in the brain via the GPR39 receptor. In the present study we investigated whether inhibition of serotonin, noradrenaline and dopamine synthesis and potentialization of glutamate, via administration of p-chlorophenylalanine (pCPA), α-methyl-p-tyrosine (αMT) and N-methyl-D-aspartatic acid (NMDA), respectively, would cause changes in GPR39 levels. Western blot analysis showed GPR39 up-regulation following 3-day administration of αMT and NMDA in the frontal cortex, and GPR39 down-regulation following 10-day administration of pCPA, αMT, and NMDA in the hippocampus of CD-1 mice. There were no changes in serum zinc levels. Additionally, we investigated tryptophan, tyrosine and glutamate concentrations in the hippocampus and frontal cortex of GPR39 knockout (GPR39 KO) mice. Liquid chromatography-mass spectrometry (LC-MS) showed a significant decrease in tryptophan and tyrosine, but not in glutamate concentrations in the hippocampus of GPR39 KO mice. There were no changes in the frontal cortex between GPR39 KO and wild type. These results indicate a possible role of the GPR39 receptor in monoaminergic and glutamatergic neurotransmission, which plays an important role in the pathophysiology of depression.

The Involvement of Serotonin in the Hypoglycemic Effects Produced by Administration of the Aqueous Extract of Xylaria nigripes with Steroid-Induced Insulin-Resistant Rats.

Xylaria nigripes (XN) is a medicinal fungus with a high-economic value. The aim of this study was to explore the hypoglycemic effects and mechanisms of the XN aqueous extract in steroid-induced insulin-resistant (SIIR) rats. Significant hypoglycemic effects were observed 60 min after administration of XN aqueous extract. In normal Wistar, hypoglycemic effects were 21% (the plasma glucose level decreased from 128.6 ± 12.5 to 100.9 ± 10.7 mg/dL). In SIIR, hypoglycemic effects were 26% (the plasma glucose level decreased from 177.6 ± 12.5 to 133.3 ± 29.7 mg/dL) rats refer to their baseline. The signaling proteins for insulin-receptor substrate-1 and glucose transporter-4 increased 0.51-fold and 1.12-fold, respectively, as determined by Western blotting; the increase in the proteins was 13% and 9%, respectively, as determined by immunohistochemistry. The serotonin antagonist, α-p-chlorophenylalanine, effectively blocked the hypoglycemic effects and increased the signaling protein levels. After XN administration, none of the animals showed significant changes in plasma-free fatty acids in 60 min. In summary, the XN extract may have hypoglycemic effects in normal Wistar and SIIR rats that may have a serotonin-related hypoglycemic effect and enhance insulin sensitivity in the SIIR rats.