The effect of win55, 212-2 on neuronal apoptosis in post-resuscitation in rats / 中华急诊医学杂志

Objective To investigate the pharmacological hypothermic effect of WIN55, 212-2 on neuronal apoptosis after cardiopulmonary resuscitation. Methods Cardiac Arrest ( CA ) was induced in Sprague-Dawley rats.Five minutes after onset of CA, cardiopulmonary resuscitation ( CPR) was carried out.At 30 minutes post-resuscitation, the animals were randomized into three groups (n=10 in each group): (1) WIN55, 212-2 hypothermia group [W group, WIN55, 212-2, 1 mg/(kg· h)].(2) Normothermia group (NT group, 5%DMSO);(3) WIN55, 212-2 with normothermia group (W+NT group, WIN55, 212-2, 1 mg/(kg· h).Animals in WIN55, 212-2 hypothermia group and WIN55, 212-2 with normothermia group were dealt with continuous intravenous infusion of WIN55, 212-2 [1 mg/(kg· h)] for 4 h, while rats in NT group were infused with equal volume of 5% DMSO instead.The survival time and neurological deficit score ( NDS) were observed.The CA models were established in three groups.After rats were sacrificed, the brains were harvested for detecting histopathological changes and apoptosis of neural cell at 24 h, 48 h and 72 h after ROSC respectively.Five animals of each group were chosen randomly ( random number ) .Results Body temperatures of rats in W group decreased from 37°C to 34°C in 4 hours.Accumulated survival rate in W group was higher than that in the other two groups ( P=0.02) .NDS was significantly improved in W group than that in the other two groups ( P<0.05) .Morphological change in W group was less serious than that in the other two groups.The number of neuron apoptosis in W group was smaller than that in the other two groups.Conclusions WIN55, 212-2 inducing pharmacologically hypothermia during post-resuscitation prolonged survival and improved cerebral function in rat cardiac arrest models.The beneficial effects of WIN55, 212-2 were associated with ameliorating the histopathological damage in brain and alleviating the neuron apoptosis.

Inhibition of 5-HT3 Receptors-activated Currents by Cannabinoids in Rat Trigeminal Ganglion Neurons / 华中科技大学学报（医学）（英德文版）

This study investigated the modulatory effect of synthetic cannbinoids WIN55,212-2 on 5-HT3 receptor-activated currents (I5-Hr3) in cultured rat trigeminal ganglion (TG) neurons using whole-cell patch clamp technique.The results showed that:(!) The majority of examined neurons (78.70％) were sensitive to 5-HT (3-300 μmol/L).5-HT induced inward currents in a concentrationdependent manner and the currents were blocked by ICS 205-930 (1 μmol/L),a selective antagonist of the 5-HT3 receptor; (2) Pre-application of WIN55,212-2 (0.01-1 μmol/L) significantly inhibited I5-HT3 reversibly in concentration-dependent and voltage-independent manners.The concentration-response curve of 5-HT3 receptor was shifted downward by WIN55,212-2 without any change of the threshold value.The EC50values of two curves were very close (17.5±4.5) μmol/L vs.(15.2±4.5) μmol/L and WIN55,212-2 decreased the maximal amplitude of I5-HI3 by (48.65±4.15)％; (3) Neither AM281,a selective CBI receptor antagonist,nor AM630,a selective CB2 receptor antagonist reversed the inhibition of I5-HT3by WIN55,212-2; (4) When WIN55,212-2 was given from 15 to 120 s before 5-HT application,inhibitory effect was gradually increased and the maximal inhibition took place at 90 s,and the inhibition remained at the same level after 90 s.We are led to concluded thatWIN55,212-2 inhibited I5-Hr3 significantly and neither CB1 receptor antagonist nor CB2 receptor antagonist could reverse the inhibition of I5-HT3 by WIN55,212-2.Moreover,WIN55,212-2 is not an open channel blocker (OCB) of 5-HT3 receptor.WIN55,212-2 significantly inhibited 5-HT-activated currents in a non-competitive manner.The inhibition of I5-HT3 by WIN55,212-2 is probably new one of peripheral analgesic mechanisms of WIN55,212-2,but the mechanism by which WIN55,212-2 inhibits I5-HT3 warrants further investigation.

Effects of cannabinoid on the expression of PKAC-β,c-fos and BDNF in cerebral cortex after ICH / 中国药理学通报

Aim To explore the effects of cannabinoid (WIN55,212-2) on mRNA and protein expressions of PKAC-β,c-fos and BDNF in cerebral cortex after intracerebral hemorrhage(ICH)in rats.Methods ICH model of rats were made by Ⅶ Collagenase,which were injected by the intraperitoneal route,thirty minutes after the operations.The rats were killed for brain tissue as specimens with 24 hours.The localizations of PKAC-β,c-fos and BDNF were assessed by immunohistochemistry;The expressions of PKAC-β,c-fos and BDNF mRNA were detected by RT-PCR;The expressions of PKAC-β and BDNF protein were revealed by Western blot.Results WIN55,212-2 increased not only the levels of BDNF mRNA and protein,but also c-fos mRNA in ipsilateral cerebral cortex.However,it decreased the levels of PKAC-β mRNA and protein.PKAC-β,c-fos,and BDNF proteins were expressed on membrane of neurons,nucleus of neurons or the cytoplasm of glial cells respectively.Conclusion WIN55,212-2 induces the expression of BDNF in the cerebral cortex,which provides a theoretical basis for the treatment of cerebrovascular disease.

Changes of PKAC-β, c-Fos and BDNF in cerebral cortex after intracerebral hemorrhage in rats treated with WIN55-212-2 / 中国病理生理杂志

AIM: To observe the effect of cannabinoid receptor (CB1R) agonist WIN55-212-2 on the expression of brain-derived neurotrophic factor (BDNF), c-Fos and protein kinase A beta-catalytic subunit (PKAC-β) in cerebrum cortex after intracerebral hemorrhage (ICH) in rats. METHODS: The intracerebral hemorrhage model of rat was made by the injection of collagenase Ⅶ, and WIN55-212-2 was intraperitoneally (ip) injected 30 min later. The rats were killed for sampling the brain tissues as specimens 24 h after ICH. The methods of immunohistochemical analysis and Western blotting were adopted to detect the expression of PKAC-β and BDNF. The mRNA expression of PKAC-β, c-Fos and BDNF was determined by RT-PCR. RESULTS: WIN55-212-2 obviously improved some nervous deficit symptoms and increased the expression of BDNF at mRNA and protein levels with upregulating the mRNA expression of c-Fos and downregulating the expression of PKA at mRNA and protein levels in the ipsilateral cerebral cortex. The proteins of PKAC-β, c-Fos and BDNF were expressed on the membrane or nucleus of the neuron or in the cytoplasm of glial cells, respectively. CONCLUSION: The expression of BDNF is induced not only by upregulation of c-Fos, but also by downregulation of PKA in WIN55-212-2 treated rats.

The Cannabinoid Agonist WIN55,212-2 Suppresses Opioid-induced Pruritus in Mice

BACKGROUND: Cannabinoid receptor agonists can reverse opioidinduced nausea and vomiting in animals, but have not yet been tested against opioid-induced pruritus. This study tests the hypothesis that a cannabinoid receptor agonist will prevent opioidinduced pruritus and evaluates if the use of a cannabinoid receptor agonist will increase the analgesic efficacy of opioids. METHODS: Various doses of fentanyl were injected subcutaneously in mice to obtain a dose-response curve with the use of a writhing test. To observe the analgesic potentiation of the cannabinoid agonist WIN55,212-2 in the writhing test, mice were pretreated with various concentrations of WIN55,212-2 (0.25, 0.5, 1.0, 2.0 mg/kg) 10 min prior to the injection of an ED50 dose of fentanyl, as determined from the dose-response curve. To observe the antipruritogenic effect of WIN55,212-2 in a scratching test, mice were pretreated with WIN55,212-2 (0.25, 0.5 mg/kg) 20 min prior to fentanyl injection. A CB1 receptor selective antagonist, AM251 (3 mg/kg), was used to confirm the cannabinoid receptor selectivity. RESULTS: The ED50 of fentanyl in the writhing test was 0.018 mg/kg (range, 0.011?0.025 mg/kg). A dose of 1 mg/kg WIN55,212-2 increased the analgesic efficacy of fentanyl significantly (P < 0.001), but doses of 0.25 mg/kg and 0.5 mg/kg did not increase the analgesic efficacy. A dose of 0.25 mg/kg WIN55,212-5 reduced the scratching response of fentanyl significantly (P < 0.001) and this action was a cannabinoid receptor selective response. CONCLUSIONS: These results demonstrate that 0.25 mg/kg WIN55,212-2 can prevent opioid-induced pruritus. The antipruritogenic activity of WIN55,212-2 occurs at CB1 receptors even if the analgesic efficacy of fentanyl cannot be increased.

Effect of WIN55,212-2 on the Expression of PGE_2 in Cultured Bovine Trabecular Meshwork Cells / 中国中医药信息杂志

Objective To study the effect of WIN55,212-2 on the expression of PGE2 in cultured bovine trabecular meshwork cells(TMCs) and discuss its mechanism of reducing the intraocular pressure.Methods TMCs were obtained through the cultured tissue,bovine TMCs were prmiarily cultured and subcultured.The third generation bovine TMCs were incubated with different dosages of WIN55,212-2 and the supernatant was collected.The concentration of PGE2 in the medium was measured by ELISA method.PGE2mRNA in TMCs was analyzed by RT-PCR.Result PGE2 in the supernatant and PGE2mRNA in TMCs increased depending on the dosages of WIN55,212-2.Conclusion Certain concentration of WIN55,212-2 may promote PGE2 secretion of TMCs,so as to reduce the intraocular pressure.

Effects of WIN 55,212-2 on IK Current in Cultured Trigeminal Ganglion Neurons of Rat / 华中科技大学学报（医学）（英德文版）

To investigate the effects of WIN 55,212-2 on IK in cultured rat trigeminal ganglion (TG) neurons, whole-cell patch clamp techniques were used to record the IK before and after WIN 55,212-2 perfusion at different concentrations. 30 μmol/L WIN 55,212-2 markedly (35.7 %±7.3%, P＜0. 01, n=8) inhibited IK currents, and the currents were partially recovered after washing.30μmol/L WIN 55,212-2 also induced a significant depolarizing shift in conductance-voltage parameters (control: V0.5 =10.43 ± 4.25 mV, k= 16.27±3.86; WIN 55,212-2: V0. 5 =24.71±3.91mV, k =16.69±2.75; n = 8, P＜0.01 for V0. 5). 0.01μmol/L WIN 55,212-2 slightly (27.0 %± 7.9 %, P＜0.05, n=7) increased IK currents, but had no significant change in conductance voltage parameters (control: V0.5 =10. 74±5. 27 mV, k=17. 33±2. 96; WIN 55,212-2: V0.5 =11.06±2.05 mV, k=19. 69±6. 60; n=7, P＞0.05 for V0.5 and k). These results suggested that WIN 55,212-2 has dual action, which might be through different receptors.

Effects of cannabinoid on the expression of PKAC-?,c-fos and BDNF in cerebral cortex after ICH / 中国药理学通报

Aim To explore the effects of cannabinoid (WIN55,212-2) on mRNA and protein expressions of PKAC-?,c-fos and BDNF in cerebral cortex after intracerebral hemorrhage(ICH)in rats.Methods ICH model of rats were made by Ⅶ Collagenase,which were injected by the intraperitoneal route,thirty minutes after the operations.The rats were killed for brain tissue as specimens with 24 hours.The localizations of PKAC-?,c-fos and BDNF were assessed by immunohistochemistry;The expressions of PKAC-?,c-fos and BDNF mRNA were detected by RT-PCR;The expressions of PKAC-? and BDNF protein were revealed by Western blot.Results WIN55,212-2 increased not only the levels of BDNF mRNA and protein,but also c-fos mRNA in ipsilateral cerebral cortex.However,it decreased the levels of PKAC-? mRNA and protein.PKAC-?,c-fos,and BDNF proteins were expressed on membrane of neurons,nucleus of neurons or the cytoplasm of glial cells respectively.Conclusion WIN55,212-2 induces the expression of BDNF in the cerebral cortex,which provides a theoretical basis for the treatment of cerebrovascular disease.

Neuroprotective effect of repeated WIN 55,212-2 preconditioning on focal cerebral ischemia-reperfusion injury in rats / 解放军医学杂志

Objective To explore the neuroprotective effect of repeated preconditioning with cannabinoid receptor agonist WIN 55,212-2 on focal cerebral ischemia-reperfusion injury in rats.Methods Focal cerebral ischemia was induced by middle cerebral artery occlusion(MCAO)for 120min.Fifty male SD rats were randomly assigned to five groups(10 each):rats in control group and dimethyl sulphoxide group(DMSO group)were intraperitoneally administered 0.3ml normal saline and 0.3ml DMSO once a day for 5 days.Rats in WIN 55,212-2 preconditioning groups(including WIN1,WIN3 and WIN5 group)received intraperitoneal injection of 1mg/kg WIN 55,212-2(dissolved with 0.3ml DMSO)once a day for 1d,3d and 5d,respectively.All animals underwent MCAO operation 24h after last pretreatment to reproduce temporal(120min)focal cerebral ischemia model.The neurological function score(NFS)was evaluated at 24,48 and 72h after reperfusion.Brain infarct was identified with 2% 2,3,5-triphenyltetrazolium chloride(TTC)staining 72h after reperfusion,and the brain infarct volume was expressed as percentage of normal cerebral hemisphere volume.Results The NFSs of rats in WIN 55,212-2 preconditioning groups(WIN1,WIN3 and WIN5 group)were significantly higher,and the infarct volumes were significantly smaller than that in control group and DMSO group at 24h,48h and 72h after reperfusion(P0.05).Conclusion The neuroprotective effect of repeated preconditioning with cannabinoid receptor agonist WIN 55,212-2 may be enhanced by increased time of pretreatment.