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Objective To observe the therapeutic effect and mechanism of Modified Banxia Shumi Decoction on p-chlorophenylalanine(PCPA)-induced insomnia model rats.Methods Forty-eight male SD rats were randomly divided into six groups,i.e.,the normal group,the model group,the low-,medium-and high-dose groups of Chinese medicine and the Diazepam group,with 8 rats in each group.For 7 consecutive days before modeling,rats in the Chinese medicine low-,medium-and high-dose groups were treated with Modified Banxia Shumi Decoction for prophylactic treatment.Except for the normal group,PCPA-induced insomnia rat model was established in all groups.After modeling on day 1,each group continued to be administered the corresponding drug for 7 days.Body mass was monitored,open-field behavioral tests were performed,serum levels of orexin A(OXA)and orexin B(OXB)were detected by enzyme-linked immunosorbent assay(ELISA),the expression of hypothalamic orexin receptor 1(OX1R)was determined by immunohistochemistry,and hematoxylin-eosin(HE)staining was used to observe the pathologic changes in the hypothalamus of rats.Results(1)Before modeling,the growth trend of body mass of rats in each group was smooth,with no significant difference between groups;after modeling,except for the normal group,the growth rate of body mass of rats in each group slowed down or even declined;after 14 days of administration of Modified Banxia Shumi Decoction,the body mass of the Chinese medicine medium-dose group was significantly increased compared with that of the model group(P＜0.01).(2)Compared with the normal group,the model group showed an increase in the total distance of activity in the open field,the distance of activity in the central region and the number of times of entering the central region(P＜0.01),a significant increase in serum OXA and OXB contents(P＜0.01),a significant increase in the expression of hypothalamic OX1R(P＜0.01),and HE staining showed mild hyperplasia of the hypothalamic glial cells;compared with the model group,the total distance of activity in the open field,the distance of activity in the central region and the number of times entering the central region were reduced in the rats in the Chinese medicine medium-dose group and the Diazepam group(P＜0.01),the levels of serum OXA and OXB were significantly reduced(P＜0.01),the expression of hypothalamic OX1R was significantly reduced(P＜0.01),and the HE staining showed that a large number of neurons with perineurial interspace enlarged and the local glial cell hyperplasia.Conclusion Modified Banxia Shumi Decoction can improve insomnia and reduce anxiety in rats by down-regulating the levels of OXA and OXB in serum and the expression of OX1R in the hypothalamus.
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ObjectiveTo investigate the interventional effects of Shugan Jianpi Yangxin prescription on the expression of orexin-A （OXA）， orexin-1 receptor （OX1R）， and orexin-2 receptor （OX2R） in the mouse model of insomnia. MethodThe mouse model of insomnia was established by intraperitoneal injection of DL-4-chlorophenylalanine （PCPA）. Fifty BALB/c mice were randomized into a blank group， a model group， an eszopiclone （0.13 mg·kg-1） group， and low- and high-dose （8.4 and 33.6 g·kg-1， respectively） Shugan Jianpi Yangxin prescription groups and treated with the corresponding drugs for 14 consecutive days. The weight changes of mice were monitored， and Morris water maze and pentobarbital-induced sleep tests were conducted. Immunohistochemistry （IHC） was employed to examine the expression of OXA in the hypothalamus. Enzyme-linked immunosorbent assay was used to measure the levels of OXA and 5-hydroxytryptamine （5-HT） in the hypothalamus， serum， and spleen. Real-time fluorescence quantitative polymerase chain reaction was employed to determine the mRNA levels of OXA， OX1R， and OX2R in the hypothalamus. ResultCompared with the blank group， the model group had decreased body weight （P<0.01）， increased escape latency （P<0.01）， increased sleep latency （P<0.01）， shortened sleep duration （P<0.01）， elevated OXA level and lowered 5-HT level in the hypothalamus， serum， and spleen （P<0.05）， and up-regulated mRNA levels of OXA， OX1R， and OX2R in the hypothalamus （P<0.01）. Compared with the model group， the low- and high-dose groups of Shugan Jianpi Yangxin prescription showed increased body weight （P<0.05， P<0.01）， shortened escape latency （P<0.05）， shortened sleep latency and prolonged sleep duration （P<0.01）， and lowered OXA level and elevated 5-HT level in the hypothalamus， serum， and spleen （P<0.05， P<0.01）. Moreover， the two doses of Shugan Jianpi Yangxin prescription down-regulated the mRNA levels of OXA， OX1R， and OX2R in the hypothalamus （P<0.01）. ConclusionShugan Jianpi Yangxin prescription exerts sedative and hypnotic effects in mice by increasing the content of 5-HT in the brain and inhibiting the expression of OXA and its receptors in the hypothalamus.
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Objective:To evaluate the effect of orexin A on morphine-induced gastrointestinal dysfunction in mice.Methods:Forty SPF C57B/6 mice, aged 6-8 weeks, half male and half female, weighing 18-22 g, were divided into 5 groups ( n=8 each) using a random number table method: control group (group C), morphine group (group M) and morphine + different doses of orexin A groups (MOH, MOM and MOL groups). Normal saline 8 ml/kg was subcutaneously injected daily in group C, morphine 6 mg/kg was subcutaneously injected daily in the other four groups, and orexin A 75, 50 and 25 μg/kg were subcutaneously injected daily for 10 days at the same time in MOH, MOM and MOL groups.The fetal water content was calculated and averaged daily.After the last administration, the mice were gavaged with black nutrient paste, and the gastric emptying rate and small intestinal propulsion rate were detected 30 min later.Blood samples were collected from the orbit, and the concentration of serum gastrin (GAS) was detected by enzyme-linked immunosorbent assay.The mice were then sacrificed, and colon tissues were removed for determination of c-kit positive cell area (by immunohistochemistry) and expression of c-kit, substance P (SP) and neural nitric oxide synthase (nNOS) in colon tissues (by Western blot). Results:Compared with group C, the rate of fecal water content, gastric emptying rate, small intestinal propulsion rate and serum GAS concentration were significantly decreased, the area of c-kit positive cells was decreased, and the expression of c-kit and SP was down-regulated, and the expression of nNOS was up-regulated in group M ( P<0.05). Compared with group M, the small intestinal propulsive rate and serum GAS concentration were significantly increased, and the area of c-kit positive cells was increased, and the expression of c-kit was up-regulated in group MOH, the rate of fecal water content, gastric emptying rate, small intestinal propulsion rate and serum GAS concentration were significantly increased, the area of c-kit positive cells was increased, and the expression of c-kit and SP was up-regulated, and the expression of nNOS was down-regulated in group MOM, and the serum GAS concentration and c-kit positive cell area were significantly increased in group MOL ( P<0.05). Conclusions:Orexin A 50 μg/kg can effectively alleviate the gastrointestinal dysfunction induced by morphine in mice, and the mechanism may be related to promotion of GAS secretion, interstitial cells of Cajal growth and SP release and inhibition of NO release.
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Objective:to explore the mechanism of modified Tianwang Buxindan in improving abnormal glucose and lipid metabolism in mice with chronic sleep deprivation from the signal pathway of orexin A/ orexin receptor 1（OX1R）. Method:The 50 6-week-old male C57BL/6 mice were randomly divided into blank group ， model group ， estazolam group and Tianwang Buxindan low and high dose groups ，for ten mice of each group. Except the blank group， rats were deprived of sleep for 8 weeks by the method of multi-platform water environment. In the last 4 weeks， Tianwang Buxindan （8.5，17 g·kg-1）and estazolam solution（9.1 mg·kg-1）were given to the stomach， and the blank group and the model group were fed with pure water of the same volume. The food intake and body weight of mice were measured twice a week， on the 49th day， blood samples were collected from the tail vein for glucose tolerance test （GTT），on the 52nd day for insulin tolerance test（ITT）， was used to detect the expression of total cholesterol （TCH）， triglyceride（TG）and free fatty acid（FFA）in serum， and enzyme-linked immunosorbent assay（ELISA）was used to detect the expression of orexin A in serum and hypothalamus. Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR）and Western blot were used to detect the mRNA and protein expression of OX1R in hypothalamus. Result:After administration， the food intake of mice in each group was different， compared with the blank group， the body weight of model group was significantly reduced（P<0.05）， the glucose tolerance was significantly abnormal， and the TCH， TG， FFA values were significantly increased（P<0.01）. The expression of orexin A in serum and hypothalamus increased significantly（P<0.01）， and the mRNA and protein expression levels of OX1R in hypothalamus increased significantly（P<0.01）. Compared with the model group， the body weight of each group of Tianwang Buxindan was significantly increased（P<0.05）， with better glucose tolerance and insulin sensitivity， TCH， TG， FFA values were significantly reduced（P<0.05，P<0.01）， accompanied by serum and the expression of orexin A in the hypothalamus was significantly decreased（P<0.05，P<0.01）， the mRNA and protein expression levels of OX1R were significantly decreased（P<0.05，P<0.01）. Conclusion:Tianwang Buxindan can protect mice from abnormal glucose and lipid metabolism induced by chronic sleep deprivation， and its mechanism may be related to the down-regulation of orexin A/OX1R signal expression.
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Objective To analyze the correlation between serum orexin-A (OXA) and early postoperative cognitive function in elderly patients undergoing lumbar spinal surgery under general anesthesia. Methods A total of 76 elderly patients (age ≥65 years) underwent lumbar spine surgery under general anesthesia from December 2018 to December 2019 in the Affiliated Hospital of Inner Mongolia Medical University were collected. All the enrolled patients were evaluated by the same doctor with the Montreal cognitive assessment scale (MoCA) on one day before the surgery and one to three days after the surgery, and venous blood was extracted from the patient on the operation day and one day after the operation, and the serum levels of OXA and S100β were measured by ELISA. According to the results of cognitive function assessment, the patients were divided into postoperative cognitive dysfunction (POCD) group and non-postoperative cognitive dysfunction (NPOCD) group. The differences in serum OXA and S100β protein levels between the two groups and their correlation with MoCA scores were statistically analyzed. Results There was no statistically significant difference in the MoCA score, serum OXA level, and S100β protein level before surgery, and heart rate (HR), mean artery pressure (MAP), bispectral index (BIS) and pulse oxygen saturation (SpO2) levels before anesthesia induction (T0), at the start of surgery (T1), at 1h after the start of surgery (T2), at the withdrawal time (T3), and at 15 minutes after extubation (T4) between the two groups (P>0.05). At the first, second, and third day after operation, the MoCA scores of the POCD group were lower than those before the operation, and they were both lower than those of the NPOCD group, the difference was statistically significant (P<0.001). There was a statistically significant difference in serum OXA levels between the two groups after the operation (P<0.05). The postoperative S100β protein level was higher than that before the operation in the two groups, and the POCD group increased more significantly, the difference between the two groups after the operation was statistically significant (P<0.05). The postoperative OXA level was positively correlated with the MoCA score (r=0.545, 0.531, 0.779) and negatively correlated with the S100β protein level (r=-0.591, -0.362, -0.743) in the two groups, and the difference was statistically significant (P<0.05). Conclusions The level of serum OXA is positively correlated with early postoperative cognitive function in elderly patients undergoing lumbal spine surgery under general anesthesia, suggesting that OXA may be a potential target for reducing the risk of postoperative cognitive dysfunction in such patients, so as to provide new ideas for preventing and improving the postoperative cognitive dysfunction in elderly patients in the future.
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Objective To investigate the effects of orexin-A on firing activity of gastric distensionsensitive (GD) neurons in the basomedial amygdala (BMA) and food intake in diet-indaced obese rats.Methods Healthy male Wistar rats were selected,and the diet-induced obesity (DIO) rat model and dietinduced resistant (DR) rat model were established by high-fat diet.The effects of orexin-A and an opioid receptor antagonist naloxone on BMA GD neurons were observed by recording the extracellular potentials of single neurons.The effects of orexin-A and naloxone on the food intake of different rats were observed by using BMA catheterization.The mRNA expression and protein expression of orexin-1 receptor (OX-1R) and μ opioid receptor were detected by real-time PCR and Elisa,respectively.Results After microinjection of orexinA into the BMA,the firing frequency of GD-sensitive neurons in the normal rats was significantly increased (GD-E:(78.3±6.9)％,GD-Ⅰ:(55.5±4.7) ％,P＜0.01),and this effect was completely blocked by OX-1R receptor antagonist SB334867,and naloxone partially blocked the discharge-promoting effect of orexin-A;Compared with the normal rats,the firing frequency of GD-sensitive neurons in the DIO (GD-E:(91.6±7.1) ％,GD-Ⅰ:(67.9±8.1) ％) and DR(GD-E:(87.9±6.8) ％,GD-Ⅰ:(69.2±5.8) ％) rats was significantly increased after BMA injection of orexin-A (P＜0.05).After administration of orexin-A into the BMA,food intake of the normal rats,DIO rats and DR rats ((2.38±0.34) g,(3.75 ±0.32) g,(4.01 ±0.38) g,respectively) was significantly increased (P＜0.01),and the food intake of DR and DIO rats were significantly higher than that of normal rats (P＜0.05).After BMA was injected with naloxone,the food intake of rats was inhibited,and the food intake of the DIO rats was significantly lower than that of the DR rats (P＜0.05),food intake of the DR rats was significantly lower than that of the normal rats (P＜0.05).The results of real-time PCR showed that the mRNA levels of OX-1R in DIO and DR rats were(5.85±0.45)and (6.03±0.42)were higher than that of normal rats,and the difference was significant (P＜0.05);and mRNA levels of μ-opioid receptors in DIO and DR rats((4.51±0.42) and (8.31±0.41) times) were higher than those in normal rats (P＜0.05).The results of Elisa showed that the protein levels of OX-1R in DIO ((2.98±0.28) ng/μl)and DR rats ((3.05±0.31) ng/μl) were higher than those in normal rats ((1.53±0.31) ng/μl,P＜0.05).The content of μ-opioid receptor protein in DR rats ((4.21±0.35) ng/μl) was higher than that of DIO rats ((2.77±0.27) ng/μl),and higher than that of normal rats((1.48±0.32) ng/μ),the difference was significant (P＜0.05).Conclusion BMA orexin-A promotes the spontaneous discharge of GD-sensitive neurons and food intake in normal rats,DIO rats and DR rats,μ-opioid receptors may be involved in the regulation of this process.
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BACKGROUND: The trigeminal nucleus caudalis (Vc) is a primary central site for trigeminal transmitting. Noxious stimulation of the trigeminal nociceptors alters the central synaptic releases and neural expression of some inflammatory and trophic agents. Orexin-A and the orexin 1 receptor (OX1R) are expressed in pain pathways including trigeminal pain transmission. However, the the mechanism(s) underling orexin-A effects on trigeminal pain modulation have not been fully clarified. METHODS: Trigeminal pain was induced by subcutaneous injection of capsaicin in the upper lip in rats. The effect of trigeminal pain on cyclooxygenase-2 (COX-2) and brain-derived neurotrophic factor (BDNF) expression in the Vc of animals was determined by immunofluorescence. Subsequently, OX1R agonist (orexin-A) and antagonist (SB-334867-A) was administrated in the Vc to investigate the possible roles of the Vc OX1R on changes in COX-2 and BDNF levels following pain induction. RESULTS: The data indicated an increase in COX-2 and decrease in BDNF immuno-reactivity in the Vc of capsaicin, and capsaicin- pretreated with SB-334867-A (80 nM), groups of rat. However, the effect of capsaicin on COX-2 and BDNF expressions was reversed by a Vc microinjection of orexin-A (100 pM). CONCLUSIONS: Overall, the present data reveals that orexin-A can attenuate capsaicin-induced trigeminal pain through the modulation of pain effects on COX-2 and BDNF expressions in the Vc of rats.
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Animals , Rats , Brain-Derived Neurotrophic Factor , Capsaicin , Cyclooxygenase 2 , Facial Pain , Fluorescent Antibody Technique , Injections, Subcutaneous , Lip , Microinjections , Nociceptors , Orexin Receptor Antagonists , Orexins , Pain Measurement , Pain Perception , Trigeminal Caudal Nucleus , Trigeminal Neuralgia , Trigeminal NucleiABSTRACT
Objective To evaluate the role of hippocampal phosphatidylinositol 3-kinase∕serine-threonine kinase (PI3K∕Akt) signaling pathway in exogenous orexin A-induced improvement of isoflurane anesthesia-caused decline in memory function of mice. Methods A total of 100 pathogen-free healthy adult male C57BL∕6 mice, aged 8-12 weeks, weighing 20-25 g, in which the lateral ventricle was catheter-ized, were divided into 5 groups (n = 20 each) using a random number table: control group (group C), isoflurane group (group I), orexin A group (group OA), orexin A plus dimethyl sulfoxide group (group OA+D) and orexin A plus PI3K inhibitor LY294002 group (group OA+LY). Normal saline was administra-ted in group C and group I. Orexin A 1. 5 mmol∕L was given in group OA. Orexin A 1. 5 mmol∕L (dissolved in dimethyl sulfoxide) was given in group OA+D. Orexin A 1. 5 mmol∕L and LY29400210 mmol∕L were given in group OA+LY. Group C only inhaled pure oxygen for 2 h (oxygen flow rate 2 L∕min), all the rest groups inhaled 1. 4％ isoflurane for 2 h, and the corresponding drug 2 μl was injected into the lateral cere-bral ventricle according to the concentration mentioned above at 15 min before the end of anesthesia. Twelve mice were randomly selected from each group and trained for contextual fear conditioning test, and then fear memory retrieval was conducted at 24 h after training. The rest 8 mice in each group were sacrificed at 2 h after the end of anesthesia and their brains were removed for determination of the expression of PI3K, Akt and phosphor-Akt (p-Akt) protein by Western blot. Results Compared with group C, the freezing time was significantly shortened, the expression of PI3K, Akt and p-Akt was down-regulated, and p-Akt∕Akt ratio was decreased in group I (P<0. 05). Compared with group I, the freezing time was significantly pro-longed, the expression of PI3K, Akt and p-Akt was up-regulated, and p-Akt∕Akt ratio was increased in group OA (P<0. 05). There was no significant difference in each parameter mentioned above between group OA and group OA+D (P>0. 05). Compared with group OA+D, the freezing time was significantly short-ened, the expression of PI3K, Akt and p-Akt was down-regulated, and p-Akt∕Akt ratio was decreased in group OA+LY (P<0. 05). Conclusion Hippocampal PI3K∕Akt signaling pathway is involved in exoge-nous orexin A-induced improvement of isoflurane anesthesia-caused decline in memory function of mice.
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OBJECTIVE@#To compare the effect on post-stroke insomnia between the repetitive transcranial acupuncture stimulation (rTAS) and the conventional western medication in the patients and to explore the mechanism.@*METHODS@#Ninety patients of post-stroke insomnia were randomized into a rTAS group, a medication group and a placebo group, 30 cases in each one. In the rTAS group, patients were intervened by rTAS. The acupoints were Baihui (GV 20), Ningshen (Extra), emotion area, Wangu (GB 12), Taiyang (EX-HN 5), Neiguan (PC 6), Shenmen (HT 7), Sanyinjiao (SP 6), Zhaohai (KI 6), Zusanli (ST 36) and Taichong (LR 3). Fast twist with small amplitude was used at Baihui (GV 20) and emotion area for 2-3 min, 200-300 r/min, once 15 min. Electroacupuncture (EA) was applied at Baihui (GV 20) and Ningshen (Extra), bilateral Wangu (GB 12), Sanyinjiao (SP 6) and Zhaohai (KI 6) on the same side, 10 Hz, 0.5-1 mA. The treatment was given for 40 min in the rTAS group, once a day. Diazepam was prescribed orally in the medication group before sleep, 2.5 mg a day. Starch capsule was used in the placebo group before sleep, once a day. All the treatment was given for continuous 1 month. The level of serum orexin A was observed before and after treatment. The effects were compared. The recurrence rate was recorded 3 months after treatment.@*RESULTS@#The total effective rates in the rTAS group and the medication group were 86.7% (26/30) and 90.0% (27/30) repectively after treatment, which were better than 20.0% (6/30) in the placebo group (both 0.05). The total effective rates in the rTAS group and the medication group were 86.7% (26/30) and 86.7% (26/30) at follow-up repectively, which were better than 16.7% (5/30) in the placebo group (both <0.01).@*CONCLUSION@#The rTAS is safe and effective for post-stroke insomnia, which is similar to oral medication of diazepam. The decreasing serum orexin A may be one of the mechanisms.
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Humans , Acupuncture Therapy , Orexins , Sleep Initiation and Maintenance Disorders , Therapeutics , StrokeABSTRACT
Objective To investigate the expression of H1 protein in the prefrontal cortex of comatose rats which have suffered traumatic brain injury (TBI) after electrical stimulation of the median nerve (MNS).Methods Seventy-two Sprague-Dawley rats (weighing 250 to 300 g) were randomly divided into a stimulated group (MNS+ TBI),an antagonist group (MNS+TBI+OXR1 antagonist),a model group (TBI) and a control group,with 18 rats in each group.Traumatic brain injury was modeled in all of the rats except those of the control group.After the modeling,the stimulated group was given MNS,the antagonist group was provided with MNS and an OXR1 injection,and the model group was given MNS with a current intensity of 0.One hour after the experiment,the consciousness of each rat was evaluated using a double-blind method.Animals were sacrificed at 6,12 and 24 hours after the intervention and brain tissue was removed.H1 protein expression was examined using immunohistochemistry.Results One hour after the experiment,significant differences were observed in the consciousness of the 4 groups,with the 18 rats of the control group on consciousness level one.Thirteen rats in the stimulated group exhibited a righting reflex,compared with 9 in the antagonist group and 5 in the model group.Immunohistochemistry showed that H1 expression was strongest in the stimulated group,followed by the antagonist,control and model groups.The H1 expression was highest at 24 hours after the experiment,followed by that at 6 h and 12 h,but those differences were not statistically significant.Conclusion Median nerve electrical stimulation might modulate wakefulness after traumatic brain injury by promoting H1 expression via orexin-A in the prefrontal contex.
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Objective:To investigate the expression of 5-HT 2A receptor in the prefrontal cortex of traumatic brain injuryinduced coma rats after median nerve electrical stimulation.Method:A total of 72 SD rats (weighing 250-300g) were randomly divided into 4 groups:a stimulationgroup,an antagonist group,a sham-stimulation group and a control group.This traumatic brain injury model was established by a weight-drop head injury,and we evaluated the change of behavior through the six classical levels of consciousness.The animals were sacrificed and their brain tissues were removed at 6,12,and 24 hours after injury.5-HT 2A protein expression was examined by immunohistochemistry.Result:Thirteen rats exhibited righting reflex in the stimulation group.In the antagonist group,9 rats exhibited righting reflex.5 rats in the sham-stimulation group had the same response.The mean rank of consciousness degree were degree 9.50 in the control group,degree 52.75 in the sham-stimulation group,degree 37.61 in the stimulation group,degree 46.14 in the antagonist group.Comparison among groups presented an increasing consciousness degree:control group＜stimulation group＜antagonist group＜sham-stimulation group(P＜0.01).Resuits from immunohistochemistry showed that significant differences in the 5-HT 2A expression among the four groups (sham-stimulation＜control＜antagonist＜stimulation))(P＜0.05),and a within-group comparison showed that the 5-HT 2A expression level was as follows:6 hours＜24 hours ＜12 hours(P＜0.05).Conclusion:Median nerve electrical stimulation might modulate wakefulness by promoting the 5-HT 2A expression via orexin-A in the prefrontal cortex of rats with traumatic brain injury-induced coma.
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Objective To study the role and mechanism of orexin A in cell viability of Alzheimer's disease (AD) cell model PC12. Methods PC12 cells were treated with 0, 10, 20, 30, 40 and 50 μmol/L Aβ25-35 for 24 h, and then, cell viability was measured by MTT to confirm which concentration was the suitable one to establish the AD cell models. (1) AD cell models were treated with 0, 0.01, 0.1, 1 and 2μmol/L orexin A for 24 h, and then, 30μmol/L Aβ25-35 was added for 24 h; MTT assay was used to determine the cell viability to conform the suitable concentration of orexin A. (2) Inverted phase contrast microscope was employed to observe the morphology changes of PC12 cells from the control group, 30 μmol/L Aβ25-35 treatment group, and 0.01 μmol/L orexin A+30 μmol/L Aβ25-35 treatment group. (3) The PC12 cells were given pretreatment of orexin A receptor inhibitor SB408124 for 2 h, and cell viability was detected. Results (1) Aβ25-35 at concentration 30μmol/L was the suitable one to establish the AD cell models;after being pretreated with different concentrations of orexin A, the cell viability showed significant differences (F=27.120, P=0.000), and 0.01μmol/L orexin A was the suitable concentration. (2) Some of the cells from the 30μmol/L Aβ25-35 treatment group had breaking-off of protuberance and damaged soma;cells from 0.01μmol/L orexin A+30μmol/L Aβ25-35 treatment group had breaking-off of protuberance, and the degree of damaged soma was eased as compared with that in the 30μmol/L Aβ25-35 treatment group. (3) If the cell viability of the control group was 100％, cell viability of orexin A receptor inhibitor group was 109.10％±0.36％, which was significantly decreased as compared with that in the 0.01 μmol/L orexin A pretreated group (117.24％±2.72％, P<0.05). Conclusion Orexin A can improve the cell viability via combination of its specific receptor; orexin A and its specific receptor may be new targets for prevention and cure of AD.
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Objective To provide new evidences for understanding the mechanisms of promo-tive role of orexins in anesthetic emergence and the effect of microinjection of orexin-A/orexin-B into cerebral ventricle on the release of histamine.Methods Male SD rats were randomly divided into sa-line (control ), orexin-A and orexin-B groups. The microdialysis probe was inserted into hypothalamus under stereotaxic apparatus.The perfused fluid from the area of hypothalamic tube-romammillary nucleus was collected using in vivo microdialysis at 1 h,2 h and 3 h after 1 nmol or 5 nmol orexin-A or orexin-B microinjection into the cerebral ventricle (n =5 each).The concentrations of histamine at each time point in dialysates of perfused fluid were detected by high performance liquid chromatography (HPLC)to analyze its dynamic changes.After one week,each group was microin-jected with 10 nmol,15 nmol and 20 nmol orexin-A and orexin-B (n =5)into the cerebral ventricle respectively,dialysates was collect and histamine was detected at 1 h to analyze its dosage response. After one week,each group was microinjected 0.3 μl saline orexin-A and orexin-B (n =6)into the tu-beromammillary nucleus.Results Compared with the control group,microinjection of 1 nmol orexin-A significantly increased histamine release at 1 h,but the same dose of orexin-B had no such effect,5 nmol of orexin-A or orexin-B injections significantly facilitated histamine release at 2 h and 3 h (P <0.01).Microinjection of 10 nmol,15 nmol and 20 nmol orexin-A and orexin-B into ventricle caused an significant increase of histamine release at 1 h while the effect was the strongest in 20 nmol (P <0.05).Compared with the control group,microinjection of orexin-A significantly decreased time of the righting reflex (P <0.01),but the same dose of orexin-B had no such effect.Conclusion Micro-injection of both orexin-A or orexin-B into cerebral ventricle could promote the release of histamine, while the effect of orexin-A was stronger.Microinjection orexin-A into tuberomammillary nucleus sig-nificant facilitated recovery from isoflurane.
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Objective To investigate the wake-promoting action of median nerve electrical stimulation(MNES) in coma rats induced by traumatic brain injury(TBI) and its influence on the expression of α1-adrenergic receptor(α1 R) in the prefrontal contex (PFC).Methods Seventy-two healthy Sprague Dawley(SD) rats were randomly divided into the control group,sham-stimulated group(TBI),stimulated group (TBI+ MNES) and antagonist group(TBI+ OX1R antagonist +MNES).The control group had no any treatment.The TBI coma rat models were prepared in the other 3 groups.The sham stimulated group had no treatment.The antagonist group was injected with orexin receptor-l(OX1R) antagonist SB334867 into lateral ventricle,and both the antagonist group and stimulated group received MNES treatment.Then the behavior changes of rats in each group were observed and the α1 R expression level in PFC was detected by using the immunohistochemistry technique.Results Thirteen rats in the stimulated group and 8 rats in the antagonist group revived,while only 4 rats in the TBI group.The α1R levels from low to high were the blank control group,sham-stimulated group,antagonist group and stimulated group,showing the increasing trend,and the difference was statistically significant(P＜0.05).Conclusion MNES can improve the rat consciousness level after TBI coma,and its mechanism may be related with up-regulating the α1 R expression level in PFC area,moreover Orexin-A participates in this regulation process.
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Objective To explore the effect of vagus nerve stimualtion on wake-promoting and the expression of 5-hydroxytryptamine (5-HT) 2A receptor in the prefrontal contex of coma rats with traumatic brain injury. Methods 72 Sprague-Dawley rats were randomly divid-ed into control group, sham-stimulated group, stimulated group and antagonist group with 18 rats in each group. Traumatic brain injury mod-el was established by a weight-drop head injury. The antagonist group was injected with SB334867, and both the antagonist group and the stimulated group received vagus nerve stimulation. Their behaviors were recorded. And immunohistochemistry technique was used to detect the expression of 5-HT2A receptor in the prefrontal cortex. Results 12 rats in the stimulated group, 9 in the antagonist group and 4 in the sham-stimulated woke up. The expression of 5-HT2A receptor from low to high was ranged as the control group, the antagonist group, the sham-stimulated group and the stimulated group (χ2=11.464, P=0.009). Conclusion Vagus nerve stimulation could raise consciousness in co-ma rats after traumatic brain injury, which may be related to up-regulating the expression of 5-HT2A receptor.
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The cytoprotective enzyme heme oxygenase-1 (HO-1) influences endothelial cell survival, proliferation, inflammatory response, and angiogenesis in response to various angiogenic stimuli. In this study, we investigate the involvement of HO-1 in the angiogenic activity of orexin-A. We showed that orexin-A stimulates expression and activity of HO-1 in human umbilical vein endothelial cells (HUVECs). Furthermore, we showed that inhibition of HO-1 by tin (Sn) protoporphryin-IX (SnPP) reduced orexin-A-induced angiogenesis in vivo and ex vivo. Orexin-A-stimulated endothelial tube formation and chemotactic activity were also blocked in SnPP-treated vascular endothelial cells. Orexin-A treatment increased the expression of nuclear factor erythroid-derived 2 related factor 2 (Nrf2), and antioxidant response element (ARE) luciferase activity, leading to induction of HO-1. Collectively, these findings indicate that HO-1 plays a role as an important mediator of orexin-A-induced angiogenesis, and provide new possibilities for therapeutic approaches in pathophysiological conditions associated with angiogenesis.
Subject(s)
Antioxidant Response Elements , Endothelial Cells , Heme Oxygenase-1 , Human Umbilical Vein Endothelial Cells , Luciferases , Tin , OrexinsABSTRACT
Objective To investigate the correlation between orexin A and the serine proteinase inhibitor vaspin in obese rats with insulin resis?tance(IR)induced by high?fat diet and elaborate the possible action mechanism of orexin involvement in fat metabolism in IR pathological process. Methods A total of 75 4?week?old male Sprague?Dawley rats were randomly divided into the normal dietary group(NC group,n=20)and the high?fat dietary group(HF group,n=55)to establish the model of obese rats with IR,the euglycemic insulin clamp technique was used to determine re?lated indicators of insulin resistance and lipid metabolism. The rats were treated with orexin A(1×10-8?1×10-6 mol/L)by hypodermic injection. The serum levels of orexin A and vaspin in rats were detected with enzyme linked immunosorbent assay. Results After 6 weeks of high?fat diet,the se?rum glucose,insulin,TC,TG and LDL?C were increased significantly in HF group than in NC group,while GIR60~120 in obese rats was decreased significantly[(16.31 ± 1.54)vs(30.22 ± 2.76)mg/(kg · min),P<0.05]. The serum vaspin level was increased 177.08%in HF group compared with NC group(P<0.05). With hypodermic injection of orexin A(1×10-8 mol/L,1×10-7 mol/L and 1×10-6 mol/L),the levels of serum vaspin in?creased 25.00%,68.75%,and 120.83%in NC group and increased 7.52%,24.06%,and 40.60%in HF group. There was a positive correlation be?tween vaspin and orexin A,glucose,insulin,TC,TG,and LDL?C(r1=0.482,P1=0.02,r2=0.515,P2=0.02,r3=0.303,P3=0.04,r4=0.388,P4=0.03,r5=0.255,P5=0.04,r6=0.253,P6=0.04)and a negative correlation between vaspin and HDL?C(r=-0.226,P=0.04)in obese rats with IR. Conclusion High?fat diet can induce insulin resistance and obesity in rats,and orexin A is closely correlated to vaspin in obese rats with insu?lin resistance. Orexin A increases serum vaspin expression and thus involves in onset of insulin resistance in obese rats.
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Objective To investigate the interference effects of orexin A on cell proliferation of the insulin?secreting beta?cell line(INS?1 cells) through the orexin receptor 1(OX1R)and the AKT/PKB signaling pathway. Methods INS?1 cells were exposed to different concentrations of orexin A in vitro,and treated with OX1R antagonist(SB334867),PI3K antagonist(wortmannin),or AKT antagonist(PF?04691502). The INS?1 cell proliferation and apoptosis,insulin secretion,OX1R protein activity and AKT phosphorylation level were determined. Results Orexin A(10-10 to 10-6 mol/L)stimulated the proliferation and activation of INS?1 cells,prevented apoptpsis,and increased insulin secretion. Additionally,AKT phosphorylation was stimulated by orexin A(10-10 to 10-6 mol/L). The OX1R antagonist SB334867(10-6 mol/L),the PI3K antagonist wortmannin (10-8 mol/L)and the AKT antagonist PF?04691502(10-6 mol/L)weakened the effects of orexin A. Conclusion Orexin A activated the AKT sig?naling pathway through the mediation of orexin A?OX1R,and promoted cell proliferation in INS?1 cells.
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Obejective To explore the correlation of Orexin A and respiratory drive in chronic obstructive pulmo-nary disease (COPD) patients. Methods Pulmonary function was tested in 30 stable COPD patients and 20 normal healthy adults. Plasma orexin A level was measured with a radioimmunoassay kit. The correlation of Orexin A with BMI, age, BDI, FEV1, FVC, FEV1/FVC, MEP, MIP, P0.1 in COPD patients was analyzed. Results Plasma Orexin A levels in patients with COPD group(1.87±0.43)ng/L was higher than those in the control group(1.49±0.19)ng/L, P<0.01. Plasma Orexin A lev-els in patients with COPD correlated negatively with FEV1(r=-0.389,P < 0.05),and correlated positively with P0.1(r=0.728,P<0.01). Conclusion Plasma orexin A in COPD patients increased which may be caused by smoking and hyper-capnia. Orexin A may play a crucial role in regulating respiratory drive in COPD patients.
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Objective To explore the changes of plasma orexin-A level before and after operation in obstructive sleep apnea-hypopnea syndrome(OSAHS) children and its effect on their behavior performance.Methods 120 cases OSAHS children performed tonsillectomy and / or adenoidectomy and 30 cases normal children as control group.According to the AHI index,the OSAHS group was divided into mild group (5 times/h ≤ AHI ＜ 20times/h,n=13),moderate group (20/h ≤ AHI ＜40/h,n=76),and severe group (AHI ≥ 40 times / h,n=31).And at the same time,according to the sensory integration ratings,OSAHS group was divided into normal group(n =30),mildly abnormal group (n =47),moderately abnormal group (n =28),severely abnormal group (n =15).Before operation and 6 months after operation,plasma orexin-A levels and children's sensory integration were measured.Results Plasma orexin-A level of the OSAHS group ((0.41 ± 0.06) μg/ml) was significantly higher compared with the control group((0.31±0.04) μg/ml) (P＜0.01).In orexin-A level of different AHI groups before and after operation(mild group:(0.33±0.02) μg/ml vs (0.28± 0.03) μg/ml,moderate group:(0.39±0.04) μg/ml vs (0.29±0.03) μg/ml,severe group:(0.49±0.04) μg/ml vs (0.32± 0.02) μg/ml),there had significant differences (P＜0.01).In OSAHS children,AHI index had positive correlation with preoperative plasma orexin-A level (r=0.803,P＜0.01).There was a significant negative correlation between sensory integration scores and plasma orexinA level(r=-0.812,P＜0.01).Conclusions Plasma orexin-A level of OSAHS children is closely related to the severity of OSAHS and the changes of their behavioral ability.And it may become a diagnostic plasma marker of OSAHS children.