Involvement of Opioid Peptides in the Analgesic Effect of Spinal Cord Stimulation in a Rat Model of Neuropathic Pain / 神经科学通报·英文版

Spinal cord stimulation (SCS)-induced analgesia was characterized, and its underlying mechanisms were examined in a spared nerve injury model of neuropathic pain in rats. The analgesic effect of SCS with moderate mechanical hypersensitivity was increased with increasing stimulation intensity between the 20% and 80% motor thresholds. Various frequencies (2, 15, 50, 100, 10000 Hz, and 2/100 Hz dense-dispersed) of SCS were similarly effective. SCS-induced analgesia was maintained without tolerance within 24 h of continuous stimulation. SCS at 2 Hz significantly increased methionine enkephalin content in the cerebrospinal fluid. The analgesic effect of 2 Hz was abolished by μ or κ opioid receptor antagonist. The effect of 100 Hz was prevented by a κ antagonist, and that of 10 kHz was blocked by any of the μ, δ, or κ receptor antagonists, suggesting that the analgesic effect of SCS at different frequencies is mediated by different endorphins and opioid receptors.

Clinical trials of acupuncture treatment of post-stroke shoulder pain / 针刺研究

OBJECTIVE: To observe the clinical therapeutic effect of "Tiaoshen Zhitong" (mental regulating and pain relieving) needling and its influence on serological indicators in the treatment of post-stroke shoulder pain, so as to provide new therapeutic thoughts and approach for post-stroke shoulder pain. METHODS: A total of 80 inpatients with post-stroke shoulder pain were randomly divided into a control group (routine needling, 39 cases) and an observation group ("Tiaoshen Zhitong" needling, 41 cases) according to the random number table. Patients of the two groups accepted basic medication treatment including anticoagulants, hypotensive drugs, hypoglycermic drugs, lipid-lowering drugs, etc. In addition, patients of the control group were also treated by routine acupuncture stimulation (uniform reinforcing-reducing stimulation) of Jianyu (LI15), Jianqian (EX-UE12), Jianhou (Extra), Jianliao (TE14), Waiguan (TE5) and Hegu (LI4) on the affected side, and those of the observation group also treated by "Tiaoshen Zhitong" needling of Ear-Shenmen (MA-TF1), bilateral Neiguan (PC6, lifting-thrusting-reducing method), Shuigou (GV26, lifting-thrusting-reducing method), and Jianyu (LI15), Jianliao(TE14), Jianzhen (SI9) and Yanglingquan (GB34, the latter 4 points were stimulated with uniform reinforcing-reducing method) on the affected side. The treatment was given once every day, 6 days a week for 4 weeks. The pain severity was assessed by using visual analogue scale (VAS), the upper limb function evaluated by using Fugl-Meyer assessment (FMA) scale, the shoulder-joint function evaluated by using Constant-Murley score (CMS) questionnaire, and the daily living ability assessed by using Barthel index (BI) scale. The enzyme linked immunosorbent assay (ELISA) was used to determine the contents of serum beta-endorphin (β-EP), enkephalin (ENK) and dynorphin (Dyn). The clinical therapeutic effect was evaluated by using Nimodipine scale method. RESULTS: Of the 39 and 41 cases in the control and observation groups, 7(17.95%) and 12(29.27%) were basically cured, 12(30.77%) and 13(31.71%) experienced marked improvement, 8(20.51%) and 11(26.83%) were effective, 12(30.77%) and 5 (12.19%) failed, with the total effective rate being 69.23% and 87.80%, respectively. The effective rate of the observation group was significantly higher than that of the control group (P<0.05). After the treatment, the VAS score was obviously reduced (P<0.01), and the scores of FMA scale, CMS questionnaire and BI scale, and contents of serum β-EP, ENK and Dyn were all increased obviously in the two groups compared with their own pre-treatment (P<0.01). The therapeutic effect of "Tiaoshen Zhitong" needling was significantly superior to that of the routine needling in lowering VAS, and in raising scores of FMA scale, CMS questionnaire and BI scale and in up-regulating serum β-EP, ENK and Dyn levels (P<0.01). CONCLUSION: "Tiaoshen Zhitong" needling is effective in reducing post-stroke shoulder pain and improving the motor function of the upper limb and shoulder-joint as well as the quality of daily life in stroke patients with shoulder pain. Its analgesic effect is probably related to the increase of the levels of serum β-EP, ENK and Dyn.

Investigation of Protonated and Lithiated Leucine-Enkephalin by Hydrogen/Deuterium Exchange and Theoretical Calculations / 分析化学

Differences of protonated and lithiated leucine-enkephalin(LE) were investigated by hydrogen deuterium exchange-mass spectrometry(HDX-MS) combined with quantum chemistry calculation. The results revealed that the protonated ions possessed very high product yield with all hydrogen atoms being exchanged, while the reaction of lithiated LE stopped after exchanging five hydrogen atoms in the same experimental conditions. The different HDX behaviours probably indicated their conformational differences. To further clarify the experimental results, the most stable conformations of protonated and lithiated leucine-enkephalin were calculated by density functional theory. It was found that terminal amino group was the most thermodynamically stable protonation site,while Li+in coordination of four carbonyl oxygen atoms formed the most favourable lithiated LE. The reaction field reduction of lithium LE was probably due to the less acidity of hydrogen atoms and the increasing rigid conformation change induced by lithium ion.

Effects of methionine enkephalin on immune enhancement by reducing myeloid derived suppressor cells and reprogramming liver metabolism in colon cancer mice / 中国药理学与毒理学杂志

OBJECTIVE To investigate enhanced immune function of methionine encephalin (MENK) and its anti-tumor mechanism in CT26 colon cancer mouse model. METHODS 3×106 CT26 cells were implanted subcutaneously in BALB/c mice. Four days after, MENK was peritoneally administrated at the concentration of 20 mg·kg-1 for 14 d. The percentage of MDSCs in bone marrow, spleen, blood, tumor and liver were detected by flow cytometry. Non- esterified fatty acid (NEFA), triglycerides (TG) and total cholesterol (T-CHO) in liver homogenate were tested by a NEFA test kit, a TG test kit and a T- CHO test kit respectively. qRT- PCR and Western blot were used to measure mRNA and protein levels of inflammation-, glycometabolsim- and lipometabolsim-associated indexes in liver. RESULTS MENK decreased percentages of MDSCs in bone marrow, spleen, blood and tumor in colon cancer mice. MENK-treated mice displayed elevated ratio of CD4+T and CD8+T cells in spleen as well as increased T and B lymphocytes proliferation. Meanwhile, MENK also ameliorated liver damage reflected by lower levels of GPT and GOT in serum and reduced risks of cancer- associated index including inflammation, high lipid and high glucose. Furthermore, MENK lowered down the levels of NEFA, TG and T- CHO in liver homogenate. MENK treatment decreased expression of p- STAT3, increased expression of p-AKT, IRS1 and Glut4 at protein level as well as reduced lipogenesis-associated genes and elevated glycolysis-associated genes in liver of tumor bearing mice. Also, abated expression of genes associated with MDSCs generation (M-CSF, GM-CSF, IL-6, IL-1β) and migration (S100A9, KC) was observed within shrunken subcutaneous tumor by MENK intervention. CONCLUSION MENK has the ability to strength immune function against colon cancer by reducing MDSCs and improving liver metabolism.

Computational Approach for Protein Structure Prediction / 대한의료정보학회지

OBJECTIVES: To predict the structure of protein, which dictates the function it performs, a newly designed algorithm is developed which blends the concept of self-organization and the genetic algorithm. METHODS: Among many other approaches, genetic algorithm is found to be a promising cooperative computational method to solve protein structure prediction in a reasonable time. To automate the right choice of parameter values the influence of self-organization is adopted to design a new genetic operator to optimize the process of prediction. Torsion angles, the local structural parameters which define the backbone of protein are considered to encode the chromosome that enhances the quality of the confirmation. Newly designed self-configured genetic operators are used to develop self-organizing genetic algorithm to facilitate the accurate structure prediction. RESULTS: Peptides are used to gauge the validity of the proposed algorithm. As a result, the structure predicted shows clear improvements in the root mean square deviation on overlapping the native indicates the overall performance of the algorithm. In addition, the Ramachandran plot results implies that the conformations of phi-psi angles in the predicted structure are better as compared to native and also free from steric hindrances. CONCLUSIONS: The proposed algorithm is promising which contributes to the prediction of a native-like structure by eliminating the time constraint and effort demand. In addition, the energy of the predicted structure is minimized to a greater extent, which proves the stability of protein.

Signal transduction mechanism of macrophages polarization induced by methionine enkephalin / 中国药学杂志

OBJECTIVE: To investigate the signal transduction mechanism of macrophages polarization induced by methionine enkephalin (MENK), which can promote tumoricidal responses in vitro. METHODS: The phenotype of macrophages were assessed by the quantitative analysis of key surface molecules on macrophages with flow cytometry (CD64, CD206). The expressions of NF-kB/STAT6 signal transduction were analyzed with Western-blotting. RESULTS: MENK(10-12 mol · L-1) could significantly decrease the expression of CD206 (P < 0.01), and at the same time, it could increase the expression of CD64 significantly (P < 0.01). MENK could increase the expression of NF-κB compared with MENK-untreated group. Furthermore MENK could inhibit the activation of STAT6 which is mediated by IL4 (P < 0.05). CONCLUSION: MENK could inhibit the activation of STAT6 signaling pathway mediated by IL4. Meanwhile, MENK could increase the expression of NF-κB and be conducive to the combination of NF-kB sites with M1-type cytokines, consequently MENK could induced the conversion of macrophages from M2 to M1 phenotype effectively.

The Neuroprotective Effect of delta-opioid Receptor Stimulation with D-Ala2, D-Leu5 Enkephalin Against Ischemic Neuronal Injury

PURPOSE: Oxygen is indispensable for survival and aerobic metabolism in all mammalian cells. Inadequate oxygen triggers a multifaceted cellular response negatively impacting important physiological functions which are observed in clinical diseases such as stroke, drowning, cardiac arrest, hazardous gas poisoning, myocardial infarction and vascular dementia. In this study, we investigated the neuroprotective effect of a synthetic delta-opioid agonist, [D-Ala2, D-Leu5] enkephalin (DADLE), and its role in ischemic neuronal injury. METHODS: This experiment was conducted in vitro using a primary culture of rat cortical neurons. Ischemia induction was performed using a hypoxic chamber. To test the degree of neuronal viability, as protected by delta-opioid stimulation with DADLE under ischemia, we used three independent approaches including a lactate dehydrogenase assay, MTT assay, and an immunofluorescent staining assay for viable cells. In addition, the gene expressions of caspase-3 and heat shock protein 70 were analyzed using real-time PCR. RESULTS: Incubation of the cortical neurons with DADLE protected them from ischemia-induced cytotoxicity, as observed by all three independent viability assays. Also, we found that its neuroprotective effect might be related with suppression of the caspase-3 gene. CONCLUSION: The results of this study suggested that DADLE exhibits a neuroprotective effect against ischemia-induced neuronal cell death.

The Neuroprotective Effect of delta-opioid Receptor Stimulation with D-Ala2, D-Leu5 Enkephalin Against Ischemic Neuronal Injury

PURPOSE: Oxygen is indispensable for survival and aerobic metabolism in all mammalian cells. Inadequate oxygen triggers a multifaceted cellular response negatively impacting important physiological functions which are observed in clinical diseases such as stroke, drowning, cardiac arrest, hazardous gas poisoning, myocardial infarction and vascular dementia. In this study, we investigated the neuroprotective effect of a synthetic delta-opioid agonist, [D-Ala2, D-Leu5] enkephalin (DADLE), and its role in ischemic neuronal injury. METHODS: This experiment was conducted in vitro using a primary culture of rat cortical neurons. Ischemia induction was performed using a hypoxic chamber. To test the degree of neuronal viability, as protected by delta-opioid stimulation with DADLE under ischemia, we used three independent approaches including a lactate dehydrogenase assay, MTT assay, and an immunofluorescent staining assay for viable cells. In addition, the gene expressions of caspase-3 and heat shock protein 70 were analyzed using real-time PCR. RESULTS: Incubation of the cortical neurons with DADLE protected them from ischemia-induced cytotoxicity, as observed by all three independent viability assays. Also, we found that its neuroprotective effect might be related with suppression of the caspase-3 gene. CONCLUSION: The results of this study suggested that DADLE exhibits a neuroprotective effect against ischemia-induced neuronal cell death.

Effects of δ-optoid receptor agonist on hemodynamics and oxygen metabolism in rats with sepsis / 中华麻醉学杂志

Objective To investigate the effects of δ-opioid receptor agonist DADLE (D-Ala2-D-Leu5-enkephalin) on the hemodynamics and oxygen metabolism in rats with sepsis. Methods Eighty healthy male SD rats were randomly divided into 4 groups ( n = 20 each) : sham operation group (group S), sepsis group (group SEP) ,DADLE, group and DADLE2, group. Sepsis was induced by cecum ligation and puncture (CLP) in SEP, DADLE,and DADLE2 groups. In DADLE1 and DADLE2 groups, 0.5 mg/ml DADLE 10 mg/kg was injected intraperitoneally (IP) 0.5 h before CLP and immediately after CLP respectively. Mean arterial pressure (MAP), left ventricular systolic pressure (LVSP) and ± dp/dtmax were recorded at 0, 2, 4 and 6 h after CLP (T1-4). Blood samples from left common carotid artery and right external jugular vein were collected at T4 for blood gas analysis. The cardiac index (CI), O2 delivery (DO2), O2 consumption (VO2) and O2 extraction rate (ERO2) were calculated.Results Compared with group S, MAP and LVSP were significantly increased at T2, while decreased at T3,4, and ± dp/dtmax was significantly increased in group SEP, MAP was significantly increased at T2, while decreased at T3,4, LVSP was significantly increased at T2,3, while decreased at T4 , and ± dp/dtmax was significantly increased in DADLE, and DADLE2 groups, and CI, DO2 and VO2 were significantly decreased and ERO2 was increased in SEP, DADLE, and DADLE2 groups (P＜0.05). Compared with group SEP, MAP, LVSP and ± dp/dtmax at T3,4 and CI, DO2 and VO2 were significantly increased, while ERO2 was significantly decreased in DADLE1 and DADLE2 groups (P ＜ 0.05). There was no significant difference in the parameters mentioned above between DADLE1 and DADLE2 groups ( P ＞ 0.05). Conclusion δ-opioid receptor agonist DADLE can obviously improve the hemodynamics and oxygen metabolism in septic rats.

Effects of a delta-opioid Agonist on the Brainstem Vestibular Nuclear Neuronal Activity of Rats

This study was undertaken to investigate the effects of [D-Ala2, D-Leu5]-enkephalin (DADLE) on the spontaneous activity of medial vestibular nuclear neurons of the rat. Sprague-Dawley rats, aged 14 to 16 days, were anesthetized with ether and decapitated. After enzymatic digestion, the brain stem portion of medial vestibular nuclear neuron was obtained by micropunching. The dissociated neurons were transferred to a recording chamber mounted on an inverted microscope, and spontaneous action potentials were recorded by standard patch-clamp techniques. The spontaneous action potentials were increased by DADLE in 12 cells and decreased in 3 cells. The spike frequency and resting membrane potential of these cells were increased by DADLE. The depth of afterhyperpolarization was not affected by DADLE. The potassium currents were decreased in 20 cells and increased in 5 cells. These results suggest that DADLE increases the neuronal activity of the medial vestibular nuclear neurons by altering resting membrane potential.

The Effects of Dopamin D-1 Agonist (SKF-38393) on Expression of Substance-P & Enkephalin mRNA in the Striatum of Hemiparkinsonian Rat Model

BACKGROUND: The selective D1 dopamine agonist has been known to play a stimulatory role in substance-P synthesis in the striatum, but its effect on the enkephalin mRNA expression has not been well known in the striatum of unilateral 6-hydroxydopamine(OHDA)-lesioned rat. So we investigated the effect of selective D1 dopaminergic agonist on substance-P, enkephalin mRNA expression in the striatum with different time interval. METHODS: The lesioned rats were divided into two groups (treated and non-treated). Each group was subdivided according to time course after lesioning (2nd, 4th and 8th week). Dopamine 1 receptor agonist, SKF-38393 (5 mg/kg/ip) and the same volume of saline was injected to treated and nontreated group respectively. The levels of enkephalin and substance-P mRNA were determined by in situ-hydridization and the expression of mRNA levels were compared between the groups. RESULTS: The expression of striatal enkephalin mRNA was increased on lesioned side in all groups. Especially, SKF-38393 enhanced the striatal expression of enkephalin mRNA after lesioning 2nd week. After lesioning 4th week and 8th week, the effect of SKF-38393 was not significant. The striatal expression of substance-P mRNAs was significantly decreased on the lesioned side, especially at the 2nd weeks. This decrement of substance-P mRNA was reversed by SKF-38393 at the 2nd week. CONCLUSIONS: These data show that the selective D1 agonist SKF-38393 have an agonistic effect on direct pathway but antagonistic effect on indirect pathway in early course of Parkinsonian rat model.

The Effects of Dopamin D-1 Agonist (SKF-38393) on Expression of Substance-P & Enkephalin mRNA in the Striatum of Hemiparkinsonian Rat Model

BACKGROUND: The selective D1 dopamine agonist has been known to play a stimulatory role in substance-P synthesis in the striatum, but its effect on the enkephalin mRNA expression has not been well known in the striatum of unilateral 6-hydroxydopamine(OHDA)-lesioned rat. So we investigated the effect of selective D1 dopaminergic agonist on substance-P, enkephalin mRNA expression in the striatum with different time interval. METHODS: The lesioned rats were divided into two groups (treated and non-treated). Each group was subdivided according to time course after lesioning (2nd, 4th and 8th week). Dopamine 1 receptor agonist, SKF-38393 (5 mg/kg/ip) and the same volume of saline was injected to treated and nontreated group respectively. The levels of enkephalin and substance-P mRNA were determined by in situ-hydridization and the expression of mRNA levels were compared between the groups. RESULTS: The expression of striatal enkephalin mRNA was increased on lesioned side in all groups. Especially, SKF-38393 enhanced the striatal expression of enkephalin mRNA after lesioning 2nd week. After lesioning 4th week and 8th week, the effect of SKF-38393 was not significant. The striatal expression of substance-P mRNAs was significantly decreased on the lesioned side, especially at the 2nd weeks. This decrement of substance-P mRNA was reversed by SKF-38393 at the 2nd week. CONCLUSIONS: These data show that the selective D1 agonist SKF-38393 have an agonistic effect on direct pathway but antagonistic effect on indirect pathway in early course of Parkinsonian rat model.

Pain-reducing Effect by Transplants of Isolated Xenogeneic Chromaffin Cells in Mouse

OBJECTIVE: Adrenal medullary chromaffin cells are known to release analgesic substances such as opioides and catecholamines. Transplantation of them is a novel method that challenges current approaches in treating chronic pain. The transplantation of xenogeneic chromaffin cells into the central nervous system(CNS) supply antinociception in animals. In this study, we investigated the analgesic effects of rat adrenal medullary chromaffin cells transplanted into the CNS of the mouse. To study the antinociceptive efficacy of transplanted chromaffin cells, the survival of rat adrenal medullary chromaffin cells transplanted into the CNS of mouse was determined. METHODS: The adrenal medullary chromaffin cells isolated from rat were transplanted into the striatum of mouse. These cells were confirmed of the release of Met-enkephalin and Leu-enkephalin by HPLC, and immunoblots for tyrosine hydroxylase(TH). Two weeks after transplantation, we performed immunohistochemistry for TH to determine the survival of implanted cells and assessed pain sensitivity at the same time. RESULTS: The isolated rat adrenal medullary chromaffin cells were positive for anti-TH antibody and released Met-enkephalin and Leu-enkephalin more than rat endothelial cells. Transplanted rat chromaffin cells were stained with anti-TH antibody in striatum of mouse after 2 weeks. Pain sensitivity was reduced on the chromaffin cell-transplanted mouse compared to endothelial cell-transplanted mouse by the hot plate test. CONCLUSION: These results suggest that the rat chromaffin cells were suitably transplanted into the CNS of mouse. This approach could be used as a therapy for reducing of chronic pain induced by cancer or neuronal injury.

Clinical study on opioid polypeptides involved in epidemic encephalitis-B pathogenesis / 中国实用内科杂志

Objectives In order to study the endogenous opioid polypeptides involved in pathogenesis of epidemic encephalitis-B and the clinical therapeutic efficacy.Methods ①Leucine-enkephalin,?-endophin and Dynorphin levels in plasma and and CSF of patients with epidemic encephalitis-B during critical stage and convalescent stage were measured by radio-immunoassay.②Naloxone therapeutic efficacy in patients with epidemic encephalitis-B were investigated.Results Opioid polypeptides levels In plasma and CSF were significantly higher in critical stage and dropped to normal levels in convalescent stage.③We demonstrated that endogenous opioid polypeptides special antagnoist agent-Naloxone was a very important therapy agent for epidemic encephalitis-B patients.Therapy group efficacy was significantly better than control group.Conclusions These results demonstrate that endogenous opioid polypeptides involves in the physiopathologic changes of epidemic encephalitis-B.

Regulation on phenotypic and functional maturation of dendritic cell by met-enkephalin / 中国免疫学杂志

Objective:To explore the mechanism of immunomodulatory effects of methionine-enkephalin(MENK)on dendritic cells(DC).Methods:We used scanning electronic microscope for DC morpholopy,assay for acid phosphatas activity,flow cytometry(FCM) and ELISA to study the effects of DC by MENK.Results:MENK(10-12mol/L) could increase the expression of MHC classⅡ,CD86 and CD40 molecules on DC surface(P

EFFECT OF METHIONINE ENKEPHALIN ON MIGRATION OF MACROPHAGES FROM MICE WITH IMPAIRED LIVER / 药物分析学报

Objective To observe the effects of methionine enkephalin (M-Enk) on migration of macrophages from mice with impaired liver and its immunomodulatory mechanisms. MethodsLiver of mice was impaired by feeding CCl4 and macrophage migration inhibitory factor (MMIF) was produced by Con A-stimulated spleen lympho- cytes. Inhibition of macrophage migration was measured in reaction system by adding M-Enk. Results Migration of macrophages in both liver-impaired and control group were suppressed by MMIF, but the suppression might be re- versed by adding 1 μmol/L M-Enk (P＜0. 05). M-Enk could significantly inhibit in vitro both of the combination of MMIF with macrophages and production of MMIF from lymphocytes (P＜0. 01). Macrophages from liver-imparied group showed a higher sensitivity compared to the control group (P＜0. 05). ConclusionThe study suggests that opi- oid peptieds play an important role in the modulation of the immune response under stress as liver impairment.

The Gene Expression of Dynorphin, Enkephalin, c-Fos in the Neuropathic Rat / 대한마취과학회지

BACKGROUND: We studied the time course of gene expression of dynorphin, enkephalin, c-fos, and the changes of allodynia, and the effect of chemical sympathectomy on the gene expression and allodynia in neuropathic rat. METHODS: In two groups of rat (Sprague-Dawley), the left L5 and L6 spinal nerves were tight ligated. In gene expression group (N=25), behavioral tests for mechanical allodynia and cold allodynia were perfomed for the next two weeks. After the test of allodynia, the expression of dynorphin, enkephalin, c-fos were assessed by Northern blot hybridization. In chemical sympathectomy group (N=16), after chemical sympathectomy (guanethidine 70 mg/kg intraperitoneally, from postoperative 7 days to 9 days), the changes of allodynia and the gene expression of enkephalin, c-fos were tested. RESULTS: Mechanical allodynia and cold allodynia was developed on the postoperative 3, 5, 7, 14 days. Preprodynorphin mRNA expression was reached peak level at the postoperative 8 hrs, sustained increase by the postoperative 3 days, but preproenkephalin mRNA expression increased slightly after operation. c-Fos mRNA expression was increased immediately at the postoperative 30 min, 1 hr, returned to normal level thereafter, and increased again on the postoperative 3, 5, 7 days that neuropathic pain was developed. Mechanical allodynia and cold allodynia were decreased by chemical sympathectomy. The increased c-fos mRNA expression and pain at postoperative 7 days was reduced by chemical sympathectomy. CONCLUSION: These results suggest that the transient gene expression of dynorphin and c-fos after tight ligation of L5 and L6 spinal nerves induces the development neuropathic pain, and late c-fos expression is related to neuropathic pain.

Effects of clonidine on releases of enkephalin and substance P from periaqueductal gray / 中华麻醉学杂志

Objective:The stimulative effects of clonidine on release of enkephalin and substance P from periaqueductal gray was studied to analyse the analgesic neurotransmitters related to alpha 2 adrenoceptor activation. Method: Twenty SD rats were allocated in pairs to receiving intraperitoneal clonidine 40?g in clonidine group or normal saline in control group. Immunohistochemical technique was used to measure the contents of enkephalin and substance P in slice of rat midbrain. Result:The immunoreaction of enkephalin from slice of rat midbrain was decreased significantly in clonidine group compared with control group(P

The Effect of Unilateral Dopaminergic Deafferentation on Kainic acid Induced Seizures in the Rat

There has been quite a number of research regarding the role of dopamine in epilepsy and it has been still very controversial, In this study, the effect of dopaminergic deafferentation on kainic acid induced seizures was evaluated with behavioral study and molecular biologic method. We produced unilateral dopaminergic deafferentation via injection of 6-hydroxydopamine at the location of right substantia nigra in the Spague- Dawley rats (250-300gm) using the stereotaxic technique under pentobarbital anesthesia. Four weeks after this procedure, kainic acid (10m9/kg) was injected into the peritoneal cavity for induction of seizures. Observations of seizure pattern and mRNA expression of c-fos, dynorphin and enkephalin were obtained in the lesion group and were compared with those in the non-lesion group in terms of behavior characteristics and in situ hybridization histochemistry. In behavior study, the results demonstrated that more severe seizure patterns (rearing, falling, tonic-clonic seizure and status epilepticus) and more fast seizure evolution time(from onset to stage V) in the lesion group than those in the non-lesion group, which may indicated statistically significant (16.21+/-12.84 min. vs 35.88+/-16.55 min., p(0.05) The results of in situ hybridization revealed that the expression of c-fos, dynorphin and enkephalin mRNAs in certain areas of brain was higher in the lesion group than that in non-lesion group; c-fos in cerebral cortex, dynorphin in dentate gyrus, enkephalin in the entorhinal cortex and amygdala nuclei. Although the lesion was unilateral, the mRNA expression patterns appeared symmetrical shape. The characteristic behavior and molecular biologic study results are suggested that the dopamine may plays a important role in control of kainic acid induced seizures.

Expression and location of fos-protein and enkephalin in central nervous system following enflurane anesthesia / 中华麻醉学杂志

Objective To investigate the expression and co-existence of fos-protein and enkephalin in CNS following enflurane anesthesia. Methods Twelve adult SD rats of both sexes weighing 195-223g were divided randomly into two equal groups: control group and enflurane group: The animals in enflurane group breathed 2% enflurane for 2h. The animals in control group underwent the same experimental steps except enflurane inhalation. Before experiment the animals were kept in a quiet place for 24h and strong light was avoided. After enflurane inhalation, chest was opened and 100 ml of normal saline was infused via left ventricle to wash Out blood from whole body, then followed by infusion of 4% polymerized formaldehyde 0.1mol/L PB 500 ml for fixation of tissue. 90 mm later the whole brain and spinal cord were harvested for determination of fos-protein and enkephalin expression and their location using double-labelled immunohistochemical technique. Results The control group showed more ELI(enkephalin like immunoactivity) neurons, less FLI(fos like immuneactivity) neurons and FLI/ELI(fos and enkephalin like immunoactivity) neurons were very rare. The enflurane group showed more FLI, ELI and FLI/ELI neurons. They were mainly distributed in frontal-cortex, lateral septal nucleus, basolateral amygdaloid nucleus, hippocampus CA1, paraventricular nucleus, ventral posterolateral nucleus, habenular nucleus, ventromedial hypothalamus nucleus, supraoptic nucleus, substantia nigra nucleus, interpeduncular nucleus, periaqueductal gray and dorsal horn. In enflurane group the number of FLI and ELI neurons in these nuclei was significantly higher(P