Sleep-improving mechanisms of Jiu Wei Bu Xue Oral Liquid on regulating Glu/GABA balance in insomnia rats based on network pharmacology and experimental verification / 药学学报

This study aimed to investigate the mechanism of Jiu Wei Bu Xue Oral Liquid on insomnia rats combining the methods of network pharmacology, molecular docking and experimental verification. UPLC-Q-TOF-MS/MS method and TCMIP, TCMSP databases were used to collect the ingredients and targets of Jiu Wei Bu Xue Oral Liquid. Protein-protein interactions and network analysis were performed to screen the key network targets and putative active ingredients of Jiu Wei Bu Xue Oral Liquid in treatment of insomnia, and then following by biological function and KEGG pathway analysis. Then binding ability for key network targets and putative active ingredients were predicted with molecular docking. The prediction targets were validated in para-chlorophenylalanine (PCPA) induced insomnia rats with administration of Jiu Wei Bu Xue Oral Liquid (2, 4, 8 mL·kg-1) for 7 days. Pentobarbital sodium induced sleeping test were performed to evaluate the synergistic sleep-aiding effect of Jiu Wei Bu Xue Oral Liquid. Then glutamic acid (Glu), γ-aminobutyrate (GABA) content and glutamate decarboxylase 1 (GAD67) activity in hypothalamus or hippocampus were evaluated, and the expressions of GAD67, γ-aminobutyric acid receptor subunit α1 (GABRA1) and γ-aminobutyric acid receptor subunit β2 (GABRB2) in hippocampus were detected by qRT-PCR and Western blot methods. Animal experiments were approved by the Institutional Committee on Animal Care of Guangxi Institute of Chinese Medicine & Pharmaceutical Science (the number of permission: 2022060802). Results showed that 16 key network targets and 16 putative active ingredients were obtained by analyzing the herbs-ingredients-targets network of Jiu Wei Bu Xue Oral Liquid in treatment of insomnia. Network pharmacology and molecular docking all indicated these active ingredients, for example atractylenolide Ⅲ, showed better binding ability with GABRA1 and GABRB2. Animal study indicated that, compared to PCPA-induced insomnia model, Jiu Wei Bu Xue Oral Liquid remarkably shortened the sleeping latency and increased the sleeping duration, increased GAD67 activity and the production of GABA in hippocampus of insomnia rats, as well as the expressions of GAD67, GABRA1 and GABRB2, while decreased Glu content in hypothalamus, leading to decreasing of Glu/GABA ratio and recovery of Glu-GABA balance. These results indicated that Jiu Wei Bu Xue Oral Liquid improved insomnia symptoms and helped maintain the Glu-GABA balance within hypothalamus and hippocampus, and reduced the excitatory neurotoxicity within brain. The mechanism may due to the elevation of GAD67 expression and enzyme activity, and the enhancement of type-A GABA receptor (GABAAR)-mediated neurons inhibition.

Phytohormone abscisic acid boosts pentobarbital-induced sleep through activation of GABA-A, PPARß and PPARγ receptor signaling / O fito-hormônio ácido abscísico estimula o sono induzido por fenobarbital por meio de ativação da sinalização de receptores GABA-A, PPARß e PPARγ

ABSTRACT Background: Sleep disorders induce anxiety and forgetfulness and change habits. The chemical hypnotic drugs currently used have serious side effects and, therefore, people are drawn towards using natural compounds such as plant-based healing agents. Abscisic acid (ABA) is produced in a variety of mammalian tissues and it is involved in many neurophysiological functions. Objective: To investigate the possible effect of ABA on pentobarbital-induced sleep and its possible signaling through GABA-A and PPAR (γ and β) receptors, in male Wistar rats. Methods: The possible effect of ABA (5 and 10 µg/rat, intracerebroventricularly) on sleep onset latency time and duration was evaluated in a V-maze model of sleep. Pentobarbital sodium (40 mg/kg, intraperitoneally) was injected to induce sleep 30 min after administration of ABA. PPARβ (GSK0660, 80 nM/rat), PPARγ (GW9662, 3 nM/rat) or GABA-A receptor (bicuculline, 6 µg/rat) antagonists were given 15 min before ABA injection. Diazepam (2 mg/kg, intraperitoneally) was used as a positive control group. Results: ABA at 5 µg significantly boosted the pentobarbital-induced subhypnotic effects and promoted induction of sleep onset in a manner comparable to diazepam treatment. Furthermore, pretreatment with bicuculline significantly abolished the ABA effects on sleep parameters, while the amplifying effects of ABA on the induction of sleep onset was not significantly affected by PPARβ or PPARγ antagonists. The sleep prolonging effect of ABA was significantly prevented by both PPAR antagonists. Conclusions: The data showed that ABA boosts pentobarbital-induced sleep and that GABA-A, PPARβ and PPARγ receptors are, at least in part, involved in ABA signaling.

RESUMO Introdução: Os distúrbios do sono induzem a ansiedade e esquecimento e mudam hábitos. Os medicamentos hipnóticos químicos utilizados atualmente têm efeitos colaterais graves e, portanto, as pessoas são atraídas para o uso de compostos naturais, como agentes de cura à base de plantas. O ácido abscísico (ABA) é produzido em uma variedade de tecidos de mamíferos e está envolvido em muitas funções neurofisiológicas. Objetivo: Investigar o possível efeito do ABA no sono induzido por pentobarbital e sua possível sinalização por meio dos receptores GABA-A e PPAR (γ e β), em ratos Wistar machos. Métodos: O possível efeito do ABA (5 e 10 µg/rato, intracerebroventricularmente) no tempo de latência e duração do início do sono foi avaliado em um modelo de labirinto em V de sono. Pentobarbital sódico (40 mg/kg, intraperitonealmente) foi injetado para induzir o sono 30 minutos após a administração de ABA. PPARβ (GSK0660, 80 nM/rato), PPARγ (GW9662, 3 nM/rato) ou antagonistas do receptor GABA-A (bicuculina, 6 µg/rato) foram administrados 15 minutos antes da injeção de ABA. Diazepam (2 mg/kg, intraperitonealmente) foi utilizado como grupo de controle positivo. Resultados: ABA a 5 µg aumentou significativamente os efeitos sub-hipnóticos induzidos por pentobarbital e promoveu a indução do início do sono de forma comparável ao tratamento com diazepam. Além disso, o pré-tratamento com bicuculina aboliu significativamente os efeitos do ABA nos parâmetros do sono, ao passo que os efeitos amplificadores do ABA na indução do início do sono não foram significativamente afetados pelos antagonistas do PPARβ ou PPARγ. O efeito de prolongamento do sono do ABA foi significativamente prevenido por ambos os antagonistas do PPAR. Conclusões: Os dados mostraram que o ABA estimula o sono induzido por pentobarbital e que os receptores GABA-A, PPARβ e PPARγ estão, pelo menos em parte, envolvidos na sinalização ABA.

Antinociceptive activity of Thyme (Thymus vulgaris L. ) and interactions with neurotropics and analgesics

The plant world represents an important source of potential therapeutic agents, but concomitant administration of herbal and conventional medications may result in interactions with subsequent beneficial or adverse effects. This study was designed to examine the analgesic effect of thyme tincture and thyme syrup, two commonly used thyme formulations, and their interactions with codeine, paracetamol, pentobarbital and diazepam in mice. The identification and quantification of thymol and carvacrol were carried out by GC/MS and GC/FID. The analgesic activity was studied using a hot plate method. Effects of thyme syrup on diazepam-induced motor coordination impairment in rotarod test and on pentobarbital-induced sleeping time were also determined. Thymol (175.3 µg/mL and 9.73 µg/mL) and carvacrol (10.54 µg/mL and 0.55 µg/mL) concentrations were measured in tincture and syrup, respectively. Thyme syrup and tincture exhibited effective analgesic activity in the hot plate pain model. Pretreatment with thyme formulations reduced analgesic activity of codeine, and potentiated the analgesic activity of paracetamol. Co-administration of thyme formulations has led to potentiation of diazepam and pentobarbital depressive central nervous system effects. Thyme formulations interacted with tested conventional drugs, probably through interference with their metabolic pathways and succeeding altered concentrations and pharmacological effects.

Optimization of extraction technology of Guizhi Zhumian Capsule by orthogonal design and pharmacodynamics test / 中草药

Objective: To optimize the extraction technology of Guizhi Zhumian Capsules (GZC). Methods: Based on the analysis of the single factor experiment Results:, the orthogonal test method was adopted to study the three factors including the amount of water added, extraction time, and extraction frequency by taking the dry extract yield of the medicinal materials and transfer rate of geniposide as indicators to optimize the water extraction process of GZC. To optimize the alcohol precipitation process of GSC, these factors including the concentration of the medicinal materials, alcohol content, and time of alcohol precipitation were investigated. The extracts before and after alcohol precipitation were compared by improving sleep pharmacodynamics. Results: The best water extraction technology of GZC was decocted three times with 10 times of water, 0.5 h each time. The optimal alcohol precipitation process was to concentrate the filtrate of water extraction to 1 mL, which was equivalent to 1 g of the original medicinal materials, with 80% alcohol content and 12 h alcohol precipitation time. Compared with the negative control group, the water extract group of GZC could prolong the sleep time of pentobarbital sodium mice and increase the sleep rate of mice under the lower dose of pentobarbital sodium valve (P 0.05). Conclusion: The pharmaceutical efficacy of the extract before and after alcohol precipitation was different. In order not to affect the drug effect, the water extraction process was finally selected as the best extraction process.

Effectiveness of three different anesthetic techniques in intraventricular catheterization and its effect on survival rate of the rats / 中国比较医学杂志

Objective To investigate the effectiveness of three different anesthetic techniques in intraventricular catheterization and its effect on the survival rate of rats. Methods Thirty Wistar rats were equally allocated into 3 groups:chloral hydrate group,pentobarbital sodium group and isoflurane group. Intraventricular catheterization was performed in the rats after anesthesia with i. p. injection of chloral hydrate and pentobarbital sodium, and isoflurane inhalation, respectively. Levels of blood glucose were detected before and at 15 and 30 minutes and 1, 3, 7, 14, 28 days after anesthesia. Body mass and 24-hour food intake were recorded before and at 1, 3, 7 days after anesthesia. The onset time and effective time of anesthesia, operation time and the survival rates on 30 days of the rats were compared and analyzed. Results The onset time and effective time of anesthesia, and the operation time in the isoflurane group were shorter than that in the chloral hydrate group, while these parameters in this group were shorter than that in the pentobarbital sodium group. Blood glucose in the chloral hydrate group was apparently increased during the surgical operation, while the body mass, 24-hour food intake and blood glucose were decreasing since one day after operation, and all the rats in this group died during the 30-day observation, mainly, due to enteroplegia. Blood glucose in the pentobarbital sodium group was mildly increased after anesthesia, while the body mass, 24-hour food intake and blood glucose were mildly decreased at one day after operation and recovered within one week. In this group, 3 rats died of respiratory distress due to overdose anesthesia and one rat died during the 30 day-observation. The blood glucose in the isoflurane group was mildly increased after operation, while the 24-hour food intake and blood glucose did not markedly changed, the body mass was stably increased, and no rat died during the 30-day-observation. Conclusions Intraperitoneal injection of chloral hydrate is not suitable for intraventricular catheterization in rats. Intraperitoneal injection of pentobarbital sodium can be only carefully applied for intraventricular catheterization under poorly-limited conditions. Isoflurane inhalation anesthesia is recommended for intraventricular catheterization in rats.

Effect of different anesthetics and blood sampling methods on blood routine test in SD rats and Kunming mice / 中国职业医学

OBJECTIVE: To compare the effects of different anesthetics and blood sampling methods on blood routine test results in experimental animals. METHODS: A total of 42 specific pathogen free( SPF) male Sprague Dawley( SD) rats and 59 SPF male Kunming( KM) mice were randomly divided into 4 groups( control group,ether group,chloral hydrate group and pentobarbital sodium group). Ether group animals were treated with ether inhalation anesthesia; animals in chloral hydrate group and pentobarbital sodium group were injected intraperitoneally with chloral hydrate or pentobarbital sodium. The control group received no anesthesia treatment. Blood samples were collected by different ways: orbital venous plexus,abdominal aorta or eyeball enucleation. White blood cell( WBC) count,red blood cell( RBC) count,platelet(PLT) count,hemoglobin(Hb) level and hematocrit(HCT) in blood samples were analyzed. RESULTS: The RBC count,Hb level and HCT of SD rats in pentobarbital sodium group were significantly lower than those in control group( P <0. 05). The HCT of SD rats in ether group was lower than that in control group( P < 0. 05). The WBC count of orbital venous plexus of KM mice was lower than that taken by eyeball enucleation in control group( P < 0. 05),but the WBC count of orbital venous plexus was higher than that taken by eyeball enucleation in chloral hydrate group( P < 0. 05). The RBC count,Hb level,HCT of KM mice in pentobarbital sodium group were significantly lower than those in control group(P < 0. 05). CONCLUSION: The anesthetic can affect the blood routine test results of experimental animals. Different blood sampling methods have effects on blood routine test results of KM mice.

Effects of isoflurane and pentobarbital on the establishment of subarachnoid block model in rats / 西安交通大学学报(医学版)

Objective To compare the effects of isoflurane and pentobarbital on the establishment of subarachnoid block model in rats.Methods 60 SD rats aged 4 months were randomly divided into Group A (n =30) and Group B (n=30).Rats in Group A received intraperitoncal injection of 10 g/L pentobarbital sodium solution 30 mg/kg and 1/4 of the initial dosage was added according to the operation effect.The induction and maintenance of anesthesia were achieved by isoflurane inhalation in Group B during operation.We recorded the time of anesthesia induction,quality of anesthesia,time of anesthesia,time of operation,and recovery time.The heart rate,respiration frequency,temperature,and saturation of blood oxygen were recorded during operation.We compared death from anesthesia and success of modeling in the two groups.Results There was no significant difference between the groups with regard to age,weight,body temperature or saturation of blood oxygen (P＞ 0.05).Compared to Group B,heart rate decreased 1-60 minutes after anesthesia and respiration frequency decreased 5 minutes after anesthesia in Group A (P＜0.05).The time of anesthesia induction,time of anesthesia,time of operation,and recovery time were shorter in Group B (P＜0.05).The quality of anesthesia was better in Group B (P＜0.05).The success rate of modeling was higher but mortality rate of anesthesia was lower in Group B than in Group A (P＜0.05).Conclusion Compared with intraperitoneal injection of pentobarbital sodium,isoflurane inhalation can provide a better anesthetic effect during the operation to establish a rat model of subarachnoid block.

Ethanol Extract of Perillae Herba Enhances Pentobarbital-Induced Sleep and Non-Rapid Eye Movement (NREM) Sleep through GABA(A)-ergic Systems

Perillae Herba has been traditionally used for the sedation in the oriental countries. Therefore, this study was conducted to determine whether Perillae Herba ethanol extract (PHEE) enhances pentobarbital-induced sleeping behaviors in animals. In addition, the possible mechanisms are demonstrated. PHEE (12.5, 25 and 50 mg/kg. p.o.) reduced the locomotor activity in mice. PHEE reduced sleep latency and augmented the total sleep time in pentobarbital (42 mg/kg, i.p.)-induced sleep in mice. Furthermore, the number of sleeping mice treated with sub-hypnotic pentobarbital (28 mg/kg, i.p.) increased. PHEE (50 mg/kg. p.o.) decreased the sleep/wake cycles and wakefulness, and increased total sleeping time and NREM sleep in electroencephalogram (EEG) of rats. In addition, PHEE (0.1, 1.0 and 10 µg/ml) increased the intracellular Cl⁻ level through the GABA receptors in the hypothalamus of rats. Moreover, the protein of glutamate decarboxylase (GAD) was overexpressed by PFEE. It was found that PHEE enhanced pentobarbital-induced sleeping behaviors through GABA(A)-ergic transmissions.

Rosmarinic Acid Potentiates Pentobarbital-Induced Sleep Behaviors and Non-Rapid Eye Movement (NREM) Sleep through the Activation of GABA(A)-ergic Systems

It has been known that RA, one of major constituents of Perilla frutescens which has been used as a traditional folk remedy for sedation in oriental countries, shows the anxiolytic-like and sedative effects. This study was performed to know whether RA may enhance pentobarbital-induced sleep through γ-aminobutyric acid (GABA)(A)-ergic systems in rodents. RA (0.5, 1.0 and 2.0 mg/kg, p.o.) reduced the locomotor activity in mice. RA decreased sleep latency and increased the total sleep time in pentobarbital (42 mg/kg, i.p.)-induced sleeping mice. RA also increased sleeping time and number of falling sleep mice after treatment with sub-hypnotic pentobarbital (28 mg/kg, i.p.). In electroencephalogram (EEG) recording, RA (2.0 mg/kg) not only decreased the counts of sleep/wake cycles and REM sleep, but also increased the total and NREM sleep in rats. The power density of NREM sleep showed the increase in δ-waves and the decrease in α-waves. On the other hand, RA (0.1, 1.0 and 10 μg/ml) increased intracellular Cl− influx in the primary cultured hypothalamic cells of rats. RA (p.o.) increased the protein expression of glutamic acid decarboxylase (GAD(65/67) ) and GABA(A) receptors subunits except β1 subunit. In conclusion, RA augmented pentobarbital-induced sleeping behaviors through GABA(A)-ergic transmission. Thus, it is suggested that RA may be useful for the treatment of insomnia.

Potentiation of decursinol angelate on pentobarbital-induced sleeping behaviors via the activation of GABA(A)-ergic systems in rodents

Angelicae Gigantis Radix (AGR, Angelica gigas) has been used for a long time as a traditional folk medicine in Korea and oriental countries. Decursinol angelate (DCA) is structurally isomeric decursin, one of the major components of AGR. This study was performed to confirm whether DCA augments pentobarbital-induced sleeping behaviors via the activation of GABA(A)-ergic systems in animals. Oral administration of DCA (10, 25 and 50 mg/kg) markedly suppressed spontaneous locomotor activity. DCA also prolonged sleeping time, and decreased the sleep latency by pentobarbital (42 mg/kg), in a dose-dependent manner, similar to muscimol, both at the hypnotic (42 mg/kg) and sub-hypnotic (28 mg/kg) dosages. Especially, DCA increased the number of sleeping animals in the sub-hypnotic dosage. DCA (50 mg/kg, p.o.) itself modulated sleep architectures; DCA reduced the counts of sleep/wake cycles. At the same time, DCA increased total sleep time, but not non-rapid eye movement (NREM) and rapid eye movement (REM) sleep. In the molecular experiments. DCA (0.001, 0.01 and 0.1 µg/ml) increased intracellular Cl- influx level in hypothalamic primary cultured neuronal cells of rats. In addition, DCA increased the protein expression of glutamic acid decarboxylase (GAD(65/67)) and GABA(A) receptors subtypes. Taken together, these results suggest that DCA potentiates pentobarbital-induced sleeping behaviors through the activation of GABA(A)-ergic systems, and can be useful in the treatment of insomnia.

Sinomenine, an Alkaloid Derived from Sinomenium acutum Potentiates Pentobarbital-Induced Sleep Behaviors and Non-Rapid Eye Movement (NREM) Sleep in Rodents

Sinomenium acutum has been long used in the preparations of traditional medicine in Japan, China and Korea for the treatment of various disorders including rheumatism, fever, pulmonary diseases and mood disorders. Recently, it was reported that Sinomenium acutum, has sedative and anxiolytic effects mediated by GABA-ergic systems. These experiments were performed to investigate whether sinomenine (SIN), an alkaloid derived from Sinomenium acutum enhances pentobarbital-induced sleep via γ-aminobutyric acid (GABA)-ergic systems, and modulates sleep architecture in mice. Oral administration of SIN (40 mg/kg) markedly reduced spontaneous locomotor activity, similar to diazepam (a benzodiazepine agonist) in mice. SIN shortened sleep latency, and increased total sleep time in a dose-dependent manner when co-administrated with pentobarbital (42 mg/kg, i.p.). SIN also increased the number of sleeping mice and total sleep time by concomitant administration with the sub-hypnotic dosage of pentobarbital (28 mg/kg, i.p.). SIN reduced the number of sleep-wake cycles, and increased total sleep time and non-rapid eye movement (NREM) sleep. In addition, SIN also increased chloride influx in the primary cultured hypothalamic neuronal cells. Furthermore, protein overexpression of glutamic acid decarboxylase (GAD(65/67)) and GABA(A) receptor subunits by western blot were found, being activated by SIN. In conclusion, SIN augments pentobarbital-induced sleeping behaviors through GABA(A)-ergic systems, and increased NREM sleep. It could be a candidate for the treatment of insomnia.

Neuropharmacological effects of the ethanolic extract of Sida acuta

ABSTRACT Sida acuta Burm. f., Malvaceae, is regarded as astringent, tonic and useful in treating urinary diseases and blood disorders, bile, liver and as treatment for nervous diseases. Different methods were developed: sodium pentobarbital-induced sleeping time, anxiolytic activity, test for muscle-effects, pentylenetetrazole (PTZ)-induced seizures, effect on normal body temperature. All experiments were performed in an isolated room with 12/12 h light/dark cycles at 22 ± 1 ºC. The effects described in this work for Sida acuta are according to what is known in traditional medicine, where is used as sedative agent. At the higher doses used in this work (500 and 1000 mg/kg), the Sida acuta extract reduced the latency time (T1) and increased the sleeping time (T2) induced by pentobarbital, indicating a sedative and hypnotic effect of the plant's extract. The extract of Sida acuta shows an increase in open arm exploration (anxiolytic activity). Results obtained in the rota-rod test showed that only the elevated dose (750 mg/kg) of Sida acuta extract, acutely administered, promotes significant changes, at 60 and 120 min post-administration, in the time of permanence in the rod. The ethanolic extract from the leaves and stems of Sida acuta, causes effects on the central nervous system in experimental animals.

Rhynchophylline, One of Major Constituents of Uncariae Ramulus et Uncus Enhances Pentobarbital-induced Sleep Behaviors and Rapid Eye Movement Sleep in Rodents

Rhynchophylline (RP) is a major tetracyclic oxindole alkaloid of Uncariae Ramulus et Uncus which has been used to treat hypertension, seizures, pain and anxiety in the oriental countries. A recent report revealed that RP attenuated ischemia-induced neuronal damage and kainite-induced convulsions in animals. This study was performed to investigate whether RP enhances pentobarbital-induced sleep behaviors and modulates sleep architecture in mice. Locomotor activity was significantly inhibited by RP at 0.25 and 0.5 mg/kg, similar to 2 mg/kg diazepam (a benzodiazepine agonist) in mice. RP shortened sleep latency and increased total sleep time in a dose-dependent manner when administrated with pentobarbital (42 mg/kg, i.p.). RP also increased the number of sleeping mice and total sleep time by concomitant administration with the sub-hypnotic dosage of pentobarbital (28mg/kg, i.p.). On the other hand, RP (0.25mg/kg, p.o.) itself significantly inhibited sleep-wake cycles, prolonged total sleep time, and rapid eye movement in rats. In addition, RP also increased chloride influx in the primary cultured hypothalamic neuronal cells. In addition, we found that glutamic acid decarboxylase (GAD(65/67)) was activated by RP. In conclusion, RP augments pentobarbital-induced sleeping behaviors, and can be a candidate for treating insomnia.

Quinpirole Increases Melatonin-Augmented Pentobarbital Sleep via Cortical ERK, p38 MAPK, and PKC in Mice

Sleep, which is an essential part of human life, is modulated by neurotransmitter systems, including gamma-aminobutyric acid (GABA) and dopamine signaling. However, the mechanisms that initiate and maintain sleep remain obscure. In this study, we investigated the relationship between melatonin (MT) and dopamine D2-like receptor signaling in pentobarbital-induced sleep and the intracellular mechanisms of sleep maintenance in the cerebral cortex. In mice, pentobarbital-induced sleep was augmented by intraperitoneal administration of 30 mg/kg MT. To investigate the relationship between MT and D2-like receptors, we administered quinpirole, a D2-like receptor agonist, to MT- and pentobarbital-treated mice. Quinpirole (1 mg/kg, i.p.) increased the duration of MT-augmented sleep in mice. In addition, locomotor activity analysis showed that neither MT nor quinpirole produced sedative effects when administered alone. In order to understand the mechanisms underlying quinpirole-augmented sleep, we measured protein levels of mitogen-activated protein kinases (MAPKs) and cortical protein kinases related to MT signaling. Treatment with quinpirole or MT activated extracellular-signal-regulated kinase 1 and 2 (ERK1/2), p38 MAPK, and protein kinase C (PKC) in the cerebral cortex, while protein kinase A (PKA) activation was not altered significantly. Taken together, our results show that quinpirole increases the duration of MT-augmented sleep through ERK1/2, p38 MAPK, and PKC signaling. These findings suggest that modulation of D2-like receptors might enhance the effect of MT on sleep.

4-Hydroxybenzaldehyde, One of Constituents from Gastrodiae Rhizoma Augments Pentobarbital-induced Sleeping Behaviors and Non-rapid Eye Movement (NREM) Sleep in Rodents

In the previous experiments, we reported that ethanol extract of Gastrodiae Rhizoma, the dried tuber of Gastrodia ElataBlume (Orchidaceae) increased pentobarbital-induced sleeping behaviors. These experiments were undertaken to know whether 4-hydroxybenzaldehyde (4-HBD), is one of the major compounds of Gastrodiae Rhizoma increases pentobarbital-induced sleeping behaviors and changes sleep architectures via activating GABA(A)-ergic systems in rodents. 4-HBD decreased locomotor activity in mice. 4-HBD increased total sleep time, and decreased of sleep onset by pentobarbital (28 mg/kg and 40 mg/kg). 4-HBD showed synergistic effects with muscimol (a GABA(A) receptor agonist), shortening sleep onset and enhancing sleep time on pentobarbital-induced sleeping behaviors. On the other hand, 4-HBD (200 mg/kg, p.o.) itself significantly inhibited the counts of sleep-wake cycles, and prolonged total sleep time and non-rapid eye movement (NREM) in rats. Moreover, 4-HBD increased intracellular Cl- levels in the primary cultured cerebellar cells. The protein levels of glutamic acid decarboxylase (GAD) and GABA(A) receptors subunits were over-expressed by 4-HBD. Consequently, these results demonstrate that 4-HBD increased NREM sleep as well as sleeping behaviors via the activation of GABA(A)-ergic systems in rodents.

Sedative-Hypnotic and Receptor Binding Studies of Fermented Marine Organisms

This study was performed to investigate the sedative-hypnotic activity of gamma-aminobutyric acid (GABA)-enriched fermented marine organisms (FMO), including sea tangle (FST) and oyster (FO) by Lactobacillus brevis BJ20 (L. brevis BJ20). FST and FO were tested for their binding activity of the GABA(A)-benzodiazepine and 5-HT(2C) receptors, which are well-known molecular targets for sleep aids. We also measured the sleep latency and sleep duration during pentobarbital-induced sleep in mice after oral administration of FST and FO. In GABA(A) and 5-HT(2C) receptor binding assays, FST displayed an effective concentration-dependent binding affinity to GABA(A) receptor, similar to the binding affinity to 5-HT(2C) receptor. FO exhibited higher affinity to 5-HT(2C) receptor, compared with the GABA(A) receptor. The oral administration of FST and FO produced a dose-dependent decrease in sleep latency and increase in sleep duration in pentobarbital-induced hypnosis. The data demonstrate that FST and FO possess sedative-hypnotic activity possibly by modulating GABA(A) and 5-HT(2C) receptors. We propose that FST and FO might be effective agents for treatment of insomnia.

Effects of mild hypothermia induced by pentobarbital sodium on hematology in male BALB/C mice / 中国比较医学杂志

Objective To observe the effects of mild hypothermia induced by pentobarbital sodium on hematology in male BALB/C mice.Method Healthy male BALB/C mice were divided randomly into two groups:control group ( C) and mild hypothermia group(M).The body temperature of the mild hypothermia group was maintained between 28℃ to 30℃( anal temperature ) for 4 hours induced by pentobarbital sodium injected intraperitoneally , then recover unaffected . Anal temperature, coagulation, electrolytes, and blood cell indexes were examined in 2, 24, 72 hours after treated by mild hypothermia;Control group was given equal volume of saline volume at constant temperature .Results The body temperature and coagulation in mild hypothermia group showed no significant difference compared with the control group ( P﹥0.05),but the concentration of K +and Na +in mild hypothermia group were higher than control group (P﹤0.01), the number of WBC in mild hypothermia group was lower than control group ( P﹤0.01或P﹤0.05 ) , and the RBC、HGB、MCH、MCHC in mild hypothermia group were lower than control group transiently (P﹤0.01或P﹤0.05).Conclusion Mild hypothermia induced by pentobarbital sodium affects some of hematological values in mice considerably .

Study of Rhizoma Coptidis alkaloids on promoting sleep in mice / 中国药理学通报

Aim To study the hypnotic effect of five alkaloids extracted from Rhizoma Coptidis (berberine , coptisine,palmatine,epiberberine,jatrorrhizine )in mice,and preliminarily explore its underlying mecha-nism.Methods The experiments of locomotor activity and hypnosis induced by suprathreshold and subthresh-old doses of pentobarbital sodium were used to evaluate the effect of drugs on sleep behavior in mice.Then, HPLC-FLD was used to detect the contents of NE,DA and 5-HT on PCPA mice model.Results Compared with control group,berberine and coptisine notably in-hibited spontaneous activity in behavioral experiments (P <0.05),and increased the sleeping percentage of mice under subthreshold dose of pentobarbital sodium. Berberine and coptisine shortened the period of sleep latency,and prolonged the sustained period of sleeping at suprathreshold dose in mice (P <0.05 or P <0.01 ).Other alkaloids had no significant differences in sleep latency and period of sleep observed in this current experiment.Compared with PCPA mice model group,berberine and coptisine remarkably increased the contents of NE and 5-HT (P <0.01 ),but they had no effects on DA.Conclusions Berberine and coptisine may play a sedative and hypnotic role in PC-PA mice by increasing contents of 5-HT and NE in hy-pothalamus,and the sedative and hypnotic effects of berberine are stronger than those of coptisine.Other alkaloids have no effects on sleeping in mice.

Pachymic Acid Enhances Pentobarbital-Induced Sleeping Behaviors via GABA(A)-ergic Systems in Mice

This study was investigated to know whether pachymic acid (PA), one of the predominant triterpenoids in Poria cocos (Hoelen) has the sedative-hypnotic effects, and underlying mechanisms are mediated via gamma-aminobutyric acid (GABA)-ergic systems. Oral administration of PA markedly suppressed locomotion activity in mice. This compound also prolonged sleeping time, and reduced sleep latency showing synergic effects with muscimol (0.2 mg/kg) in shortening sleep onset and enhancing sleep time induced by pentobarbital, both at the hypnotic (40 mg/kg) and sub-hypnotic (28 mg/kg) doses. Additionally, PA elevated intracellular chloride levels in hypothalamic primary cultured neuronal cells of rats. Moreover, Western blotting quantitative results showed that PA increased the amount of protein level expression of GAD65/67 over a broader range of doses. PA increased alpha- and beta-subunits protein levels, but decreased gamma-subunit protein levels in GABA(A) receptors. The present experiment provides evidence for the hypnotic effects as PA enhanced pentobarbital-induced sleeping behaviors via GABA(A)-ergic mechanisms in rodents. Taken together, it is proposed that PA may be useful for the treatment of sleep disturbed subjects with insomnia.

Effect of different anesthesia methods on erythrocyte immune function in mice

OBJECTIVE@#To explore effect of different anesthesia methods and different anesthetics on erythrocyte immune function in mice.@*METHODS@#The mice were anesthetized by isoflurane and ether inhalation, and also under intraperitoneal anesthesia with sodium pentobarbital and chloral hydrate. Blood was collected from the ventro-cardinal vein. Automatic blood cell analyzer was used for routine blood examination, and the canthine oxidase method was used to measure the superoxide dismutase (SOD) activity. Lipid peroxidation product malondialdehyde (MDA) was measured with TBA, and glutathione peroxidase (GSH-Px) was measured with DTNB, and then the effect of different anesthesia methods and different anesthetics on erythrocyte immune function in mice was observed.@*RESULTS@#Hct level of chloral hydrate intraperitoneal injection group was significantly higher than the other three groups (P<0.05). And the MDA levels in the pentobarbital sodium group were significantly higher than the other three groups (P<0.05). SOD and GSH-Px of the chloral hydrate and sodium pentobarbital intraperitoneal injection group were significantly lower than the other two groups; RBC-C 3bRR and RBC-ICR of the chloral hydrate and sodium pentobarbital intraperitoneal injection group were significantly lower than the other two groups.@*CONCLUSIONS@#Different drugs can induce changes in immune function of mice at different levels. Isoflurane and ether have less damage to animal body, while chloral hydrate and sodium pentobarbital intraperitoneal injection have a certain inhibitory effect on the animal body respiratory system and can cause greater damage to the body. Therefore, the reasonable selection and control of anesthetics are very important in order to avoid the experimental errors caused by anesthesia.