Objective To investigate the role of microglial activation in spinal cord in a rat model of persistent postoperative pain evoked by skin/muscle incision and retraction (SMIR) .Methods Seventy-two male SD rats weighing 200-250 g in which intrathecal (IT) catheter was successfully inserted were randomly divided into 3 groups ( n = 24 each) : group sham operation; group SMIR and group SMIR + FT minocycline (a specific microglia inhibitor) . The rat model of persistent postoperative pain evoked by SMIR was established according to the method described by Flatters. Pain behavior was assessed by paw mechanical withdrawal threshold ( MWT) to von Frey filament stimulation at 1 day before (T0,baseline) and 3, 7, 12, 22 and 32 days after operation (T1-5,) . Four animals were sacrificed at each time point in each group for detection of the expression of Iba-1 (a specific marker of microglia) in the spinal dorsal horn by immunofluorescence and the microglia was counted. Results MWT was significantly decreasedat T1-4, while the expression of Iba-1 and microglia counts in the spinal dorsal horn were significantly increased at T1, 2 by SMIR in group Ⅱ. IT minocycline significantly attenuated the hyperalgesia induced by SMIR at T1-4 and decreased Iba-1 expression and microglia counts at T1,2 in group Ⅲ. Conclusion Microglial activation in the spinal cord plays an important role in the development and maintenance of SMIR-evoked persistent postoperative pain in rats.

Objective To investigate the role of gamma-aminobutyric acid transporter-1 (GAT-1) in the spinal cord in a rat model of bone cancer pain.Methods Eighty female SD rats weighing 150-180 g were randomly divided into 5 groups (n =16 each): sham operation group(group Ⅰ ),bone cancer pain group(group Ⅲ ),sham operation+ NO-711 group(group Ⅲ ),Ⅳ group BCP + NO-711 group(group Ⅳ ) and BCP + vehicle group (group Ⅴ ).Bone cancer pain was induced by inoculating Walker-256 mammary gland carcinoma cells into medullary cavity of tibia.NO-711 (20 μg,10 μl) was administered intrathecally once a day for 3 consecutive days from the 14th day after operation.Mechanical withdrawl threshold (MWT) of mechanical stimulus was determined the day before operation and at days 3,5,7,10,14 and 16 after operation.The animals were sacrificed on the 16th day after operation,and then the spinal cords were removed for determination of the expression of GAT-1 and double immunostaining of GAT-1 and glial fibrillary acidic protein (GFAP,astrocyte marker).Results MWT were significantly decreased in groups Ⅱ,Ⅳ and Ⅴ as compared with groups.Ⅰ and Ⅲ.The expression of GAT-1 significantly up-regulated in groups Ⅱ,Ⅴ as compared with groups Ⅰ and Ⅲ.NO-711 significantly increased MWT,while decreased the expression of GAT-1 in group Ⅳ compared with groups Ⅱ and Ⅴ.The expression of GAT-1 up-regulation appeared colocalizes with in astrocytes activation in spinal dorsal horn.Conclusion The up-regulation of expression of GAT-1 in spinal cord is involued in the development and maintenance of bone cancer pain,which may be related to the astrocytes activation.

BACKGROUND: The disorder of energy metabolism and cerebral edema after cerebral ischemia-reperfusion are the important factors to inducecerebral ischemia-reperfusion injury. The Chinese herb, breviscapine, whose effective component is scutellarin, can prevent the activation of protein kinase C evoked by ischemia-reperfusion, relieve calcic overload and reduce the volume of ischemia infarcted focus volume, and then alleviate cerebral ischemia-reperfusion injury. But what are the influences of breviscapine on energy metabolism and cerebral edema after cerebral ischemia-reperfusion?OBJECTIVE: To observe the effects of breviscapine parenteral solution on energy metabolism and cerebral edema after cerebral ischemia-reperfusion in gerbils.DESIGN: A randomized control trial.SETTING: Affiliated Hospital of Jining Medical College, Jiangsu Provincial Key Laboratory of Anesthesiology, Affiliated Hospital of Xuzhou Medical College.MATERIALS: The experiment was carried out in the Jiangsu Provincial Key Laboratory of Anesthesiology Between February and August 1999. Seventy-two male gerbils were used.METHODS: The gerbils were randomly divided into sham-operated group (n=8), normothermia control group (n=32) and breviscapine group (n=32).According to the reperfused time, the normothermia control group and breviscapine group were divided into 4 subgroups with 8 gerbils in each subgroup: 0, 10, 30 and 60-minute reperfusion groups. The gerbils in the normothermia control group and breviscapine group were made into models of forebrain ischemia reperfusion, treated with ischemia for 10 minutes. In the sham-operated group, only bilateral common carotid arteries were freed but not occluded. In the breviscapine group, the gerbils were given intraperitoneal injection of breviscapine psrenteral solution (90 mg/kg) at 15 minutes before ischemia. The gerbils in the sham-operated group and normothermia control group were treated with saline of the same volume. The brain water was determined by drying electrothermostat. The contents of adenosine triphosphate (ATP), adenosine diphosphate (ADP) and adenosine phosphate (AMP) in hippocampus were determined with high performance liquid chromatography and ultraviolet detector.MAIN OUTCOME MEASURES: ① ATP, ADP and AMP contents in hippocampus; ② Water contents in cerebral cortex.RESULTS: All the 72 gerbils were involved in the analysis of results without deletion. ① ATP, ADP and AMP contents in hippocampus: At 0, 10, 30 and 60 minutes after reperfusion, ATP and adenine nucleotide pool contents in hippocampal tissue in the normothermia control group were obviously decreased, the ATP contents were 68%, 56%, 49% and 50% of those in the sham-operated group respectively (P ＜ 0.01), and adenine nucleotide pool contents were 62%, 50%, 51% and 52% of those in the sham-operated group respectively (P ＜ 0.01). The ATP contents at each time point in the breviscapine group were 84%, 69%, 64% and 63% of those in the sham-operated group respectively, and the adenine nucleotide pool contents were 86%, 72%, 68% and 69% of those in the sham-operated group respectively, which were all obviously higher than those in the normothermia control group (P ＜ 0.05). ②Water contents in cerebral cortex: The water content after cerebral ischemia-reperfusion in the normothermia control group was obviously higher than that in the sham-operated group, and gradually aggravated with the prolongation of reperfusion. The water content in the breviscapine group was obviously higher than that in the sham-operated group, but markedly lower than that in the normothermia control group (P ＜ 0.05).CONCLUSION: Breviscapine can play a role in protecting brain through inhibiting disorder of energy metabolism and relieving cerebral edema.

Objective To investigate the glial activation in the spinal cord in a rat model of persistent postoperative pain. Methods Forty-eight adult male SD rats weighing 200-250 g were randomly divided into 2 groups ( n = 24 each): group Ⅰ sham operation (group S) and group Ⅱ persistent postoperative pain. Persistent postoperative pain was evoked by skin/muscle incision and retraction (SMIR) as described by Flatters. Pawwithdrawal threshold to yon Frey hair stimulation was measured before operation (baseline) and at 1, 3, 12, 22and 32 d after establishment of the model. Four animals were sacrificed at each time point and lumbar segment of the spinal cord was removed for determination of expression of glial fibrillary acidic protein (GFAP) in the astrocytes by immunofluorescence histo-chemistry assay. Results The mechanical threshold started to decrease at 1 d after establishment of the model, and peaked at 12 d after establishment of the mode. Immunofluorescence histochemistry assay demonstrated that GFAP expression in the dorsal horn was significantly increased at 3 d after estabhshment of the model and reached the peak at 12 d and was maintained at the high level until 22 d after establishment of the model. Conclusion Glial activation is involved in the mechanism of persistent postoperative pain evoked by SMIR.

Objective To investigate the effect of intrathecal γ-aminobutyric acid transporter-1 ( GAT-1 )small interfering RNA (siRNA) on neuropathic pain in rats. Methods Male SD rats weighing 200-250 g were studied. The experiment was performed in 3 parts. Part Ⅰ Twenty rats were randomly divided into 5 groups ( n =4 each): GAT-1 siRNA-1 group, GAT-1 siRNA-2 group, GAT-1 siRNA-3 group, negative control siRNA group and DEPC treatment group. Two days after ligation of sciatic nerve, intrathecal siRNA 2 μg or equal volume of DF-PC was injected once a day for 3 consecutive days. The rats were killed and the lumbar segment of the spinal cord was removed at 2nd day after the last intrathecal injection for determination of the expression of GAT-1 in the spinal dorsal horn by Western Blot. Part Ⅱ Thirty rats were randomly divided into 3 groups ( n = 10 each): GAT-1 siRNA-3 + lipo2000 group, GAT-1 siRNA-3 mismatch siRNA + lipo2000 group, and DEPC treatment + lipo2000group. Paw-withdrawl threshold (PWT) to thermal and mechanical stimulation was measured before ligation of sciatic nerve, 3 days after ligation of sciatic nerve and at 1, 3, 5, 7 and 10 days after consecutive administration for 3 days. Part Ⅲ Eighty-four rats were randomly divided into 3 groups as described in Part Ⅱ ( n = 28 each). Four rats were killed at each time point and the lumbar segment of the spinal cord was removed for determination of the expression of GAT-1 in the spinal dorsal horn by Western blot. Results PWT to thermal and mechanical stimulation was significantly inreased and the GAT-1 expression was down-regulated after the injection of GAT-1 siRNA.Conclusion Intrathecal GAT-1 siRNA can reduce the neuropathic pain by inhibiton of up-regulation of the GAT-1 expression in the spinal dorsal horn in rats.

Objective To test the hypothesis that isoflurane-preconditioning (ISO-P) provides added protection of myocardium with hypothermia against ischemia-reperfusion injury through KATP channels. Methods Thirty-two SD rats of both sexes weighing 230-270 g were studied. The animals were anesthetized with intraperitoneal ketamine 100 mg?kg-1 and heparinized. Chest was opened and heart was immediately removed and perfused in a Langendorff apparatus with oxygenated (95% O2, 5% CO2) Krebs-Hensleit buffer (KHB) for 50 min,followed by 150 min of hypothermic-ischemia (perfusion was suspended and heart was immersed in 17℃ KHB) and 60 min of normothermic reperfusion (heart was again reperfused with 37.7℃ KHB and immersed in 37.7℃ KHB) (I/R). The animals were randomly divided into 4 groups of 8 animals:(l) control group; (2) ISO-P group; (3) ISO-P + GLB group and (4) ISO-P + 5-HD group. The control group received no pretreatment before I/R. In ISO-P group the heart was perfused with KHB gassed with 1% isoflurane for 15 min followed by 15 min wash-out before I/R. In group 3 and 4 the heart was perfused with KHB containing glibenclamide 20?mol?L-1 (group 3) or 5-hydroxydecanoate 150?mol ?L-1 (group 4) in addition to 1% isoflurane before I/R. Cardiac function was measured with a fluid filled latex balloon inserted in left ventricle (LVSP, LVDP, LVEDP, dp/dtmax, dp/dtmin). During hypothermic ischemia left ventricle pressure (LVP),S-time (the interval between the beginning of ischemia to the time point when LVP increased by 1 mm Hg from the baseline pressure) and C-time (the interval between the beginning of ischemia to the time point when LVP was lmmHg below the plateau pressure) were measured. Other criteria included coronary flow (CF),infarct size and creatine kinase.Results (l) S-time and C-time were negatively correlated with infarct size.(2)In ISO-P group (group 2) S-time and C-time were significantly longer during ischemia; LVDP and LVDP x HR were significantly higher during normothermic reperfusion and infarct size was significantly smaller than those in control group (groupl) (P

Objective To investigate the effects of sodium hydroxybutyrate (?-OH) on neuronal apoptosis in neonatal rat cortex induced by hypoxic-ischemic (HI) insult. Methods One-hundred and fifty 7-day old newborn SD rats were randomly assigned to one of S groups ( n = 30 each): sham operation group (A); HI + normal saline (NS) group (B) and three HI + ?-OH groups (C, D, E) . HI was induced by ligation of left common carotid artery followed by 2 h inhalation of hypoxic air (8% O2 92% N2). In sham operation group (A) left common carotid artery was exposed but not ligated and no hypoxic air was inhaled after operation. In group B (HI + NS) NS 0.2 ml?10 g-1 was injected intraperitoneally (IP) t.i.d immediately after HI until the animals were killed; whereas in HI +?-OH groups ?-OH 50 (C) or 100 (D) or 200 (E) mg?kg-1 was injected IP instead of NS. Six animals were killed at 1 h, 3 h, 1 d, 3 d and 7 d in each group and brains were removed. The number of apoptotic neurons in the left cortex was detected using TUNEL staining technique. Results The number of apoptotic neurons at 1 h-7 d after HI was significantly greater in group B, C, D and E than in group A ( sham operation) . The expression of apoptosis positive neurons reached the peak at 1 day after HI. The number of apoptotic neurons at 3 h-3 d was significantly greater in group E (?-OH 200 mg?kg-1) than in group C and D (?-OH 50 and 100 mg?kg-1) . There was no significant difference in the number of apoptotic neurons between group B (HI + NS) and E (HI +?-OH 200 mg?kg-1). Conclusion Sodium hydroxybutyrate 50 and 100 mg?kg-1 can attenuate neuronal apoptosis in neonatal rat cortex induced by hypoxic-ischemic insult.