Role of ZHX2 in regulating dorsal root ganglion μ-opioid receptor expression in mice with peripheral nerve injuryinduced pain hypersensitivity / 南方医科大学学报

OBJECTIVE@#To investigate the role of zinc-fingers and homeoboxes 2 (ZHX2) in regulating μ-opioid receptor expression in the dorsal root ganglion (DRG) in mice with peripheral nerve injury-induced pain hypersensitivity.@*METHODS@#Forty-eight male adult C57BL6J mice were randomized into 4 groups and subjected to chronic constriction injury (CCI) of the sciatic nerve or sham operation followed by microinjection of a specific small interfering RNA (siRNA) of ZHX2 or a negative control siRNA sequence (siNC) into the DRG. Seven days later, the mice were examined for changes in the hind paw withdrawal frequency (PWF), after which the DRG tissue was collected for detecting the expressions of μ-opioid receptor at the mRNA and protein levels using RT-qPCR and Western blotting. In another experiment, the DRG tissues were collected from 6 mice (21-day-old) for primary culture of the DRG neurons, which were transfected with ZHX2 siRNA or the siNC to observe the changes in the expressions of ZHX2 and μ-pioid receptor.@*RESULTS@#Microinjection of ZHX2 siRNA into the ipsilateral L3 and L4 DRGs significantly reversed CCI-induced μ-pioid receptor downregulation in the injured DRG and alleviated CCI-induced mechanical allodynia in the mice. In the cell experiment, ZHX2 knockdown obviously upregulated the mRNA and protein expressions of opioid receptor in the primary cultured DRG neurons.@*CONCLUSIONS@#ZHX2 knockdown in the DRG reverses CCI-induced down-regulation of μ opioid receptor to alleviate periphery nerve injury-induced pain hypersensitivity in mice.

Expression of µ-Opioid Receptor in CA1 Hippocampal Astrocytes

µ-opioid receptor (MOR) is a class of opioid receptors with a high affinity for enkephalins and beta-endorphin. In hippocampus, activation of MOR is known to enhance the neuronal excitability of pyramidal neurons, which has been mainly attributed to a disinhibition of pyramidal neurons via activating Gαi subunit to suppress the presynaptic release of GABA in hippocampal interneurons. In contrast, the potential role of MOR in hippocampal astrocytes, the most abundant cell type in the brain, has remained unexplored. Here, we determine the cellular and subcellular distribution of MOR in different cell types of the hippocampus by utilizing MOR-mCherry mice and two different antibodies against MOR. Consistent with previous findings, we demonstrate that MOR expression in the CA1 pyramidal layer is co-localized with axon terminals from GABAergic inhibitory neurons but not with soma of pyramidal neurons. More importantly, we demonstrate that MOR is highly expressed in CA1 hippocampal astrocytes. The ultrastructural analysis further demonstrates that the astrocytic MOR is localized in soma and processes, but not in microdomains near synapses. Lastly, we demonstrate that astrocytes in ventral tegmental area and nucleus accumbens also express MOR. Our results provide the unprecedented evidence for the presence of MOR in astrocytes, implicating potential roles of astrocytic MOR in addictive behaviors.

Effects of electroacupuncture on expression of μ-opioid receptor in nucleus ceruleus in rats with bone cancer pain-morphine tolerance / 中国针灸

<p><b>OBJECTIVE</b>To observe the effects of electroacupuncture (EA) on pain behavior in rats with bone cancer pain and morphine tolerance, and to explore partial action mechanism.</p><p><b>METHODS</b>Forty-two SD healthy female rats were randomly divided into a sham operation group (7 rats), a bone cancer pain group (8 rats), a morphine tolerance group (9 rats), an EA group (9 rats) and a sham EA group (9 rats). The rats in the sham operation group were treated with injection of phosphate buffer saline at medullary cavity of left-side tibia, and the rats in the remaining groups were injected with MRMT-1 breast cancer cells. After operation, no treatment was given to rats in the sham operation group and bone cancer pain group. 11 days after operation, rats in the morphine tole-rance group, EA group and sham EA group were treated with intraperitoneal injection of morphine hydrochloride, once every 12 hours, for 11 days to establish the model of bone cancer pain and morphine tolerance. One day after the establishment of this bone cancer pain model, the rats in the morphine tolerance group were injected with morphine, once every 12 hours (9:00 a.m. and 9:00 p.m.) for 7 days; the rats in the EA group and sham EA group were injected with morphine at 9:00 a.m., and treated with EA (2 Hz/100 Hz) and sham EA (only injected into the subcutaneous tissue) at bilateral "Zusanli" (ST 36) and "Kunlun" (BL 60), 30 min per treatment, once a day for 7 days. One day before cancer cell injection, 6 days, 8 days, 10 days after operation, after 30 min on 1 days, 5 days, 9 days, 11 days of morphine injection, and after 30 min on 1 days, 3 days, 5 days, 7 days of EA treatment, the paw withdrawal threshold (PWT) was measured in each group. On 11 day of morphine injection, HE staining was applied to observe the morphology and structure change of tibia in the sham operation group, bone cancer pain group and morphine tolerance group, random 2 rats in each group. On 7 days of EA treatment, fluorescent immunohistochemical method was applied to observe the expression of μ-opioid receptor positive cells in nucleus ceruleus in each group, random 4 rats in each one.</p><p><b>RESULTS</b>After 10 days of the cancer cells injection, the PWT of 28 rats of bone cancer pain model (8 rats in the bone cancer pain group, 8 rats in the morphine tolerance group, 6 rats in the EA group and 6 rats in the sham EA group) was significantly lower than that of 7 rats in the sham operation group (<0.01). After one day of morphine injection, the PWT of the morphine tolerance group, EA group and sham EA group was higher than that of the bone cancer pain group (all<0.01); on 11 d of morphine injection, the PWT of the morphine tolerance group, EA group and sham EA group was not significantly different from that of the bone cancer pain group (all>0.05). On 11 d of morphine injection, the tumor induced by cancer cells was observed in upper 1/3 tibia in the bone cancer pain group and morphine tolerance group, and the marrow cavity was filled with MRMT-1 cancer cells; no abnormal change was observed in the sham operation group. On 1 d, 3 d, 5 d and 7 d of EA treatment, the PWT of the cancer pain group, morphine tolerance group and sham EA group was lower than that of the EA group (all<0.01). On 7 d of EA treatment, the positive expression of MOR in nucleus ceruleus in the cancer pain group, morphine tolerance group, EA group and sham EA group was lower than that in the sham operation group (<0.01,<0.05), and that in the cancer pain group, morphine tolerance group and sham EA group was lower than that in the EA group (all<0.01).</p><p><b>CONCLUSIONS</b>EA can improve mechanical pain threshold in rats with bone cancer pain-morphine tolerance, and improve the abnormal pain, which is likely to be involved with improvement of the MOR positive cells expression in nucleus ceruleus by EA.</p>

Spicy Food Preference and Risk for Alcohol Dependence in Korean

OBJECTIVE: Previous studies have reported that both preference for spicy food and drinking behavior are associated with the activity of the opioid system in the central nervous system. The relationship between the preference for spicy food and the risk of alcohol dependence by comparing spicy food preference in alcohol-dependent patients vs. healthy controls was investigated. Also the association between the preference for spicy food and OPRM1 A118G was studied. METHODS: A total of 150 Korean male patients with alcohol dependence and 100 normal male control subjects were included in this study. Preference for spicy food was measured using the Food Preference Scale (FPS). DNA analysis was conducted to detect the A118G polymorphism. RESULTS: The mean FPS score was significantly higher in the alcohol-dependent patients (61.2±24.2) than in the normal control subjects (53.0±22.0). FPS scores differed significantly between alcohol-dependent patients and normal control subjects who had the G allele in OPRM1 A118G, but not between the two groups with the AA genotype. CONCLUSION: A strong preference for spicy food can be assumed to be a risk factor for alcohol dependence, particularly in those carrying the G allele in OPRM1 A118G.

Wheat peptides reduce oxidative stress and inhibit NO production through modulating μ-opioid receptor in a rat NSAID-induced stomach damage model / 中国天然药物

Non-steroidal anti-inflammatory drugs (NSAIDs) induce tissue damage and oxidative stress in animal models of stomach damage. In the present study, the protective effects of wheat peptides were evaluated in a NSAID-induced stomach damage model in rats. Different doses of wheat peptides or distilled water were administered daily by gavage for 30 days before the rat stomach damage model was established by administration of NSAIDs (aspirin and indomethacin) into the digestive tract twice. The treatment of wheat peptides decreased the NSAID-induced gastric epithelial cell degeneration and oxidative stress and NO levels in the rats. Wheat peptides significantly increased the superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities and decreased iNOS activity in stomach. The mRNA expression level of μ-opioid receptor was significantly decreased in wheat peptides-treated rats than that in in the control rats. The results suggest that NSAID drugs induced stomach damage in rats, wchih can be prevented by wheat peptides. The mechanisms for the protective effects were most likely through reducing NSAID-induced oxidative stress.

Effect on Distribution and Expression ofμ-opioid Receptor in Hippocampus of Premenstrual Syndrome Liver-qi Stagnation Rat Model by Shu-Yu Capsule / 世界科学技术-中医药现代化

This study was aimed to discuss the distribution and protein expression level ofμ-opioid receptor (MOR) in hippocampus of premenstrual syndrome (PMS) liver-qi stagnation rat model, in order to initially reveal the action mechanism of PMS liver-qi stagnation and intervention effect ofShu-Yu (SY) capsule. Chronic restraint stress method was used to copy PMS liver-qi stagnation rat model.SY capsule ofTiao-Gan prescription was given as intervention. Immunofluorescence (IF) and western blot (WB) technique were used to detect MOR in hippocampal CA1 and CA3 brain area of rats from each group. The results showed that compared with the normal group, the hippocampus MOR distribution arrangement was messy with increased protein concentration in the model group (P< 0.01). After drug intervention, the MOR protein level returned to normal level. It was concluded that the pathogenesis of PMS liver-qi stagnation may be associated with high expression of MOR in hippocampus CA1 and CA3 region of rats. SY capsule can effectively correct and restore it to nearly normal level. It may be one of the central mechanisms in SY capsule treatment of PMS liver-qi stagnation.

The coexistence expression of TRPV1 and μ-opoid receptor in spinal dorsal root ganglion projected to stomach and its clinical significance / 实用医学杂志

Objective To observe the coexistence expression of TRPV1 and μ-opoid receptorin spinal dorsal root ganglion (DRG) projected to stomach , and to investigate the relationship between TRPV1 andμ-opoid receptorand its clinical significance in rats with acute gastric mucosal lesion induced by water immersion and restraint stress. Methods FortyWistar rats were randomly assigned to 3 groups, including normal control group(group NC, n = 10), WIRS group (group WIRS, n = 20), and sufentanil pretreatment group (group SP, n = 10). A rat model of gsatric mucosal lesion was induced by WIRS. 6 hours after WIRS treatment, gastric tissues were excised and microscopically observed; ulcer index (GI) was noted. The coexistence expression of TRPV1 and μ-opioid receptor in DRG neurons was detected by immunofluorescence assay, and the levels of CGRP was measured by ELASA. Results As compared withgroup WIRS, the degree of gastric injury was obviously relieved in group SF. Coexistence of TRPV1 and μ-opioid receptor was detected in thoracic DRG neurons projected to stomach; the CGRP level was higher in group WIRS than in group NC. ConclusionsTRPV1 isinvolved in protection of acute gastric mucosal lesion. Activation of μ-opioid receptor can induce TRPV1 to release CGRP, resulting in protection of gastric mucosa.

Effects of Morphine on μOpioid Receptor-mediated ERK Phosphorylation / 医药导报

Objective To study the phosphorylation mode of extracellular regulated kinase( ERK)induced by acute and chronic morphine treatment on Chinese hamster ovary( CHO)cells expressed withμopioid receptors. Methods The time course of ERK phosphorylation 1 h and 36 h after morphine exposure as well as naloxone-precipitated withdrawal was detected by immunobloting. Results A transient enhancement of ERK phosphorylation was induced by 1 μmol · L-1 morphine with the peak effect at 5 min(P〈0. 01),and the effect was dose-dependent. No difference in ERK phosphorylation was found after 36h of treatment with 10 μmol · L-1 morphine compared with the control. However,5 or 10 min-naloxone precipitation induced remarkable decrease in ERK phosphorylation compared with the control(P〈0. 01). Conclusion Different changes of ERK phosphorylation were found under acute and chronic morphine treatment and naloxone precipitation,indicating a compensation of ERK related pathway induced byμ opioid receptors.

Effect of μ-opioid receptor agonist on acute lung injury induced by trauma-LPS two hits in rat model / 临床麻醉学杂志

Objective To explore the effect of μ-opioid receptor agonist on acute lung injury in-duced by trauma-LPS two hits in rat model.Methods Thirty-two aduld SD rats,were randomized in four groups(n=8):normal control group(group N),blank control group(group B),DAMGO group (group D)and DAMGO+CTOP group(group DC).A rat model was made by intraperitoneal injection of LPS at 6 hours after fracturing bilateral thighbone.Group N didn’t receive trauma and LPS,only anesthesia.The rats in group D received 200 μg/kg of DAMGO,group DC received 600 μg/kg of CTOP and 200 μg/kg of DAMGO,group B obtained the same amount of saline.6 hours after treat-ment,the arterial blood was collected for blood gas analysis,the lungs was harvested to observe lung tissue pathology change and dry-weight/wet-weight ratio,and the levels of MDA,TNF-α,IL-6,SOD activities in lung tissue were determined.Results The results of pathological observation showed that there was obvious inflammatory reaction in lung tissues after two-hits.Compared with group D, PaO2 ,pH and dry-weight/wet-weight ratio were significantly lower in group B and group DC(P <0.05),The score of Smith were significantly increased(P <0.05).The levels of MDA,TNF-α,IL-6 in lung tissue were significantly reduced in group D than those in group B and group DC(P <0.05), SOD activities in group D were significantly higher than those in group B and group DC(P <0.05 ). Conclusion μ-Opioid receptor DAMGO agonist has protective effect on acute lung injury induced by trauma-LPS two hits in rat model.

ENDOMORPHIN-1 IS MORE POTENT THAN ENDOMORPHIN-2 IN INHIBITION OF SYNAPTIC TRANSMISSION IN SUBSTANTIA GELATINOSA OF ADULT RAT SPINAL CORD / 神经解剖学杂志

Effects of endomorphin-1 (EM-1) and endomorphin-2 (EM-2) on synaptic transmission were investigated on neurons in substantia gelatinosa (SG) of the spinal dorsal horn by whole-cell voltage clamp recording. Both EM-1 (1 μmol/L) and EM-2 (1 μmol/L)remarkably reduced the frequency but not the amplitude of miniature excitatory postsynaptic currents (mEPSCs) and miniature inhibitory postsynaptic currents (mIPSCs). These effects were antagonized by 3-funaltrexamine ( β-FNA, 10 μmol/L), a selective μ-opioid receptor antagonist. Noticeably, EM-1 showed higher potency in decreasing the frequency of mEPSCs and mIPSCs than that of EM-2. These results indicate that EMs suppress both excitatory and inhibitory synaptic transmission by activating presynaptic μ-opioid receptors in the SG and EM-1, compared with EM-2, might be a more potent endogenous analgesic at the spinal cord level.

Inhibitory effects of κ-opioid receptor stimulation on ne-induced cardiac hypertrophy of the cultured myocardial cells / 中国药理学通报

AIM: To study the effects of κ-opioid receptor activation on the cardiac hypertrophy, cultured cardiomyocytes were to used study the inhibitory effects of U58,488H on cellular volume and norepinephrine(NE)-induced hypertrophy in the presence or absence of norbinaltorphimine(nor-BNI). METHODS: The protein content was assayed with Lowry's method. The cardiomyocytes' volumes was measured by an eyepiece graticule of inverted microscope. The contracting frequency of cultured myocytes was counted by the inverted microscope. RESULTS: A κ-opioid receptor agonist U50, 488H at (0.1 ∼ 10 μmol·L-1) inhibited the protein content of cultured myocardial cells in normal serum medium in a dose-dependent manner. The inhibitory effects of U50,488H were completely blocked by pretreatment with nor-BNI, a specific κ-opioid receptor antagonist at 1 μmol·L-1. U50,488H (0.1 ∼ 10 μmol·L-1) also inhibited NE-induced cellular hypertrophy in low serum medium, which also were abolished by nor-BNI (1 μmol· L-1). CONCLUSIONS: U50,488H inhibited NE-induced hypertrophy. The inhibitory effects of U50,488H are involved in mediating the action of NE-induced cardiac hypertrophy.