[Adenosine Al Receptor Mediated Neuroprotection of Shenmai Injection on Rat Cerebral Ischemia/Reperfusion Injury: an Experimental Study].

OBJECTIVE: To observe whether adenosine Al receptor (Al R) mediated neuroprotection of Shenmai Injection (SI) on rat cerebral ischemia/reperfusion (I/R) injury. METHODS: The focal cerebral I/R model was established by middle cerebral artery occlusion (MCAO). Totally 60 successfully modeled rats was divided into 5 groups according to randomized block principle, i.e., the model group, the SI group, the SI + AlR antagonist (1,3-dipropyl-8-cyclopentylxanthine, DPCPX) group, the AlR antagonist control group, and the dimethyl sulfoxide (DMSO) control group, 12 in each group. Besides, a sham-operation group was set up (n =12). SI at 15 mL/kg was peritoneally injected to mice in the SI group immediately after cerebral I/R. Equal volume of normal saline was injected to mice in the model group and the sham-operation group. DPCPX at 1 mg/mL was peritoneally injected to mice in the Al R antagonist control group 30 min before peritoneal injecting SI. DPCPX at 1 mg/kg and DMSO at 1 mL/kg were peritoneally injected to mice in the AlR antagonist control group and the DMSO control group 30 min immediately before cerebral I/R. Rats' neurobehavioral scores were assessed after 24 h reperfusion. The volume of cerebral infarction and Bcl-2 protein expression of cerebral infarction penumbra were also detected. Results Compared with the sham-operation group, neurobehavioral scores, the volume of cerebral infarction, and Bcl-2 protein expression increased (all P <0. 05). Compared with the model group, neurobehavioral scores and the volume of cerebral infarction obviously decreased, but Bcl-2 protein expression increased in the SI group (all P <0. 05). Compared with the SI group, neurobehavioral scores increased, the volume of cerebral infarction was obviously enlarged, and Bcl-2 protein expression was obviously reduced in the A1R antagonist control group (all P <0. 05). CONCLUSIONS: SI's neurobehavioral scores could be partially reversed in the Al R antagonist control group, the volume of cerebral infarction and Bcl-2 protein expression improved. AlR might possibly meditate neuroprotection of SI on MACO mire

[Local adenosine A1 receptors of baihui acupoint mediate cerebral ischemia tolerance induced by Electroacupuncture].

OBJECTIVE: To investigate whether local A1R of Baihui acupoint mediate cerebral ischemia tolerance induced by electro-acupuncture (EA). METHODS: Sixty SD rats were randomly divided into five groups, i.e., the sham-operation (S) group, the model group (M), the electroacupuncture (E) group, the CCPA group and the DMSO group. The focal cerebral ischemia/reperfusion model was established by middle cerebral artery occlusion (MCAO) in rats. Rats in the E group were received EA pretreatment baihui acupoint at 2 h before established MCAO. The rats in DMSO group and the CCPA group were injected with DMSO (20 µl) and CCPA (0.1 mmol/L) 20 µl into Baihui, respectively, at 2 h before established MCAO. After 24 h reperfusion, the rats' behavior, cerebral infarct volume, the cerebral Bcl-2 protein expression were assessed. RESULTS: Compared with M group, the rats' behavior was improved, the cerebral infarct volume was decreased and the Bcl-2 protein expression was up-regulated (P < 0.05) in the E group. Compared with M and DMSO group, the rats' behavior was improved, the cerebral infarct volume was decreased and the Bcl-2 protein expression was up-regulated (P < 0.05) in the CCPA group. There were no statistical differences between CCPA and E group. CONCLUSIONS: EA induced cerebral ischemia tolerance. Local A1R of Baihui acupoint possible mediate cerebral ischemia tolerance induced by Electroacupuncture.

[Effect of ginsenoside Rb1 on cerebral infarction volume and IL-1 beta in the brain tissue and sera of focal cerebral ischemia/reperfusion injury model rats].

OBJECTIVE: To investigate the effect of ginsenoside Rb1 on cerebral infarction volume as well as IL-1 beta in the brain tissue and sera of focal cerebral ischemia/reperfusion (I/R) injury model rats. METHODS: The I/R rat model was established by using thread according to Zea-Longa. SD rats were randomly divided into five groups, i.e., the sham-operation group, the model group, the low dose ginsenoside Rb1 (20 mg/kg) group, the medium dose ginsenoside Rb1 group (40 mg/kg), and the high dose ginsenoside Rb1 group (80 mg/kg), 12 in each group. Rats in the sham-operation group only received middle cerebral artery occlusion (MCAO) but without thread insertion. The MCAO model was prepared in the rest 4 groups, followed by MCAO2 h later. Ginsenoside Rb1 at each dose was peritoneally administrated to rats in corresponding groups immediately after cerebral ischemia. Equal volume of normal saline was administered to rats in the sham-operation group. Rats' cerebral infarction volume, integrals of neurologic defect degree, expression of IL-1 beta content in the brain tissue and sera were observed 24 h after 2-h cerebral I/R. RESULTS: In the model group, integrals of neurologic defect degree were improved (P < 0.01), IL-1 beta positive cells in the brain tissue increased and serum IL-1 beta content elevated (P < 0.05), when compared with the sham-operation group. In comparison of the model group, integrals of neurologic defect degree were lowered in the medium dose and high dose ginsenoside Rb1 groups (P < 0.05, P < 0.01). The cerebral infarction volume was all shrunken in each ginsenoside Rb1 group, IL-1 beta positive cells in the brain tissue decreased, and IL-1 beta content in serum reduced (P < 0.01, P < 0.05). Compared with the low dose ginsenoside Rb1 group, integrals of neurologic defect degree decreased, the cerebral infarction volume shrunken, and IL-1 beta content in serum reduced in the high dose ginsenoside Rb1 group (P < 0.01, P < 0.05). CONCLUSION: Ginsenoside Rb1 (20, 40, 80 mg/kg) might effectively release local cerebral ischemia by down-regulating the IL-1 beta expression.

[The role of adenosine deaminase in the electroacupuncture preconditioning induced rapid tolerance to focal cerebral ischemia].

OBJECTIVE: To observe the electroacupuncture (EA) pretreatment at Baihui (GV20) on the concentration of adenosine deaminase (ADA) and adenosine, and to evaluate its effects on the neurologic function score and the infarction volume after middle cerebral artery occlusion (MCAO) ischemia/reperfusion (I/R), thus exploring its mechanisms for relieving the ischemia/reperfusion injury. METHODS: Totally 54 male SD rats were randomly divided into 3 groups, the sham-EA group, the EA group, and the control group, 18 in each group. Rats in the control group were not intervened after anesthesia. Rats in the EA group were needled at Baihui (GV20) for 30 min. Rats in the sham-EA group received the same procedure as those performed in the EA group without electricity connected. The changes of adenosine and ADA contents were detected at 30, 60, and 120 min after EA respectively. The I/R model was established. Totally 48 male SD rats were randomly divided into 6 groups, i.e., the model group (Group A), the EA group (Group B), the EA +8-Cyclopentyl-1,3-dipropylxanthine (DPCPX) group (Group C), the EA + DMSO group (Group D), the Deoxycoformycin (Deo) group (Group E), and the normal saline group (Group F). Rats in Group B, C, and D received EA for 30 min before modeling. Rats in Group C and D were peritoneally injected with DPCPX (1 mg/kg) and DMSO (1 mL/kg) at 30 min before EA. The neurologic function score was evaluated and the infarct volumes were detected after 24-h reperfusion. RESULTS: Compared with the sham-EA group, there was no statistical difference in the contents of the adenosine or ADA in the control group at each time point (P > 0.05). Compared with the control group at the same time point, the content of ADA significantly decreased at 60 min in the EA group [(315.0 +/- 22.9 U/L), P < 0.05], and restored to the normal level at 120 min after EA. The content of adenosine increased in the EA group at 120 min [(20.4 +/- 2.2) ng/microL, P < 0.05]. Compared with the model group, the neurologic function score decreased (P < 0.05) and the infarct volumes were obviously reduced (P < 0.01) in Group B, D and E. There was no statistical difference in the neurologic function score or the infarct volumes in other groups, when compared with the model group (P > 0.05) CONCLUSION: EA at Baihui (GV20) showed protective effects on the cerebral I/R rats, which might be achieved through lowering the ADA concentration and elevating the adenosine content, and further activating adenosine A1 receptor.

Transcutaneous electrical acupoint stimulation in adult patients receiving gastrectomy/colorectal resection: A randomized controlled trial.

BACKGROUND: Acupuncture promotes the recovery of gastrointestinal function and provides analgesia after major abdominal surgery. The effects of transcutaneous electrical acupoint stimulation (TEAS) remain unclear. AIM: To explore the potential effects of TEAS on the recovery of gastrointestinal function after gastrectomy and colorectal resection. METHODS: Patients scheduled for gastrectomy or colorectal resection were randomized at a 2:3:3:2 ratio to receive: (1) TEAS at maximum tolerable current for 30 min immediately prior to anesthesia induction and for the entire duration of surgery, plus two 30-min daily sessions for 3 consecutive days after surgery (perioperative TEAS group); (2) Preoperative and intraoperative TEAS only; (3) Preoperative and postoperative TEAS only; or (4) Sham stimulation. The primary outcome was the time from the end of surgery to the first bowel sound. RESULTS: In total, 441 patients were randomized; 405 patients (58.4 ± 10.2 years of age; 247 males) received the planned surgery. The time to the first bowel sounds did not differ among the four groups (P = 0.90; log-rank test). On postoperative day 1, the rest pain scores differed significantly among the four groups (P = 0.04; Kruskal-Wallis test). Post hoc comparison using the Bonferroni test showed lower pain scores in the perioperative TEAS group (1.4 ± 1.2) than in the sham stimulation group (1.7 ± 1.1; P = 0.04). Surgical complications did not differ among the four groups. CONCLUSION: TEAS provided analgesic effects in adult patients undergoing major abdominal surgery, and it can be added to clinical practice as a means of accelerating postoperative rehabilitation of these patients.

[Effects of injecting adenosine A1 receptor agonist into baihui (GV20) on the cerebral cortex in ischemia/reperfusion injury model rats].

OBJECTIVE: To investigate the effects of injecting adenosine A1 receptor agonist (CCPA) into Baihui (GV20) on the cerebral cortex induced by the ischemia/reperfusion of middle cerebral artery occlusion (MCAO) in rats. METHODS: Twenty-four SD rats were randomly divided into four groups, i. e., the sham-operation group, the model group, the DMSO group, and the CCPA group. The MCAO model was established by thread embolism method. At the moment of ischemia/reperfusion, the rats in DMSO group and the CCPA group were injected with DMSO (20 microL) and CCPA (0.1 mmol) 20 microL into Baihui respectively. The rats' behavior, the histomorphology of ischemic penumbra in the cerebral cortex, the expressions of Bcl-2 protein, and the apoptosis rate of neurocytes were assessed. RESULTS: Compared with the model group and the DMSO group, the rats' behavior were markedly improved in the CCPA group (P<0.05). No obvious karyopyknosis and cytoplasm empty dye of neurons appeared. The Bcl-2 expressions in rats' cerebral cortex obviously increased (P<0.01). The apoptosis number of neurons obviously decreased (P<0.01). CONCLUSIONS: Injecting CCPA into Bahui improved the rats' behavior and histomorphology in the ischemic penumbra, elevated the expressions of Bcl-2 protein, and reduced the neurons apoptosis rate in the ischemic penumbra. It alleviated the cerebral ischemia-reperfusion injury. Therefore, it could be taken as a new treatment method.

Analgesia with 5' extracellular nucleotidase-mediated electroacupuncture for neuropathic pain.

BACKGROUND: Acupuncture is a treatment for neuropathic pain, but its mechanism remains unclear. Previous studies showed that analgesia was induced in rats with neuropathic pain when their spinal cord adenosine content increased after electroacupuncture (EA); however, the mechanism behind this electroacupuncture-induced increase has not been clarified. OBJECTIVE: This study aimed to determine the role that ecto-5'-nucleotidase plays in EA-induced analgesia for neuropathic pain. METHODS: We performed electroacupuncture at the Zusanli acupoint on the seventh day after establishing a rat model of neuropathic pain induced through chronic constriction injuries. We observed the mechanical withdrawal threshold and thermal pain threshold and detected the expression of ecto-5'-nucleotidase in the spinal cord using Western blot. Chronic constriction injury rat models were intraperitoneally injected with α,ß-methyleneadenosine 5'-diphosphate, an ecto-5'-nucleotidase inhibitor, 30 min before electroacupuncture. The adenosine content of the spinal cord was detected using high-performance liquid chromatography. Lastly, the adenosine A1 receptor agonist N6-cyclopentyladenosine was intrathecally injected into the lumbar swelling of the rats, and the mechanical withdrawal and thermal pain thresholds were reevaluated. RESULTS: Analgesia and increased ecto-5'-nucleotidase expression and adenosine content in the spinal cord were observed 1 h after electroacupuncture. α,ß-methyleneadenosine 5'-diphosphate was able to inhibit upregulation of adenosine content and electroacupuncture-induced analgesia. After administration of N6-cyclopentyladenosine, electroacupuncture-induced analgesia was restored. CONCLUSIONS: Our results suggest that electroacupuncture at Zusanli can produce analgesia in chronic constriction injury rat models, possibly via the increased ecto-5'-nucleotidase expression induced through electroacupuncture, thus leading to increased adenosine expression in the spinal cord.

Silencing miRNA-324-3p protects against cerebral ischemic injury via regulation of the GATA2/A1R axis.

Previous studies have suggested that miR-324-3p is related to the pathophysiology of cerebral ischemia, but the mechanism underlying this relationship is unclear. In this study, we found that miR-324-3p expression was decreased in patients with acute ischemic stroke and in in vitro and in vivo models of ischemic stroke. miR-324-3p agomir potentiated ischemic brain damage in rats subjected to middle cerebral artery occlusion, as indicated by increased infarct volumes and cell apoptosis rates and greater neurological deficits. In a PC12 cell oxygen-glucose deprivation/reoxygenation model, a miR-324-3p mimic decreased cell viability and expression of the anti-apoptotic protein BCL2 and increased expression of the pro-apoptotic protein BAX and rates of cell apoptosis, whereas treatment with a miR-324-3p inhibitor had the opposite effects. Silencing miR-324-3p increased adenosine A1 receptor (A1R) expression through regulation of GATA binding protein 2 (GATA2). These findings suggest that silencing miR-324-3p reduces ischemic brain damage via the GATA2/A1R axis.

[Melatonin receptor 2 mediated spinal dorsal horn astrocytes IL-17 inhibition contributes to electroacupuncture analgesia in neuropathic pain rats].

OBJECTIVE: To investigate the effect of electroacupuncture (EA) on pain behaviors and expression of spinal dorsal horn melatonin receptor 2 (MT2) and interleukin-17 (IL-17) in neuropathic pain rats, so as to explore its mechanism underlying pain relief. METHODS: The present study includes 3 parts. In the first part, eighteen male SD rats were randomly divided into 3 groups: sham operation, model and EA groups, with 6 rats in each group. The neuropathic pain model was established by chronic constriction injury (CCI) of the right sciatic nerve. On the 7th day following modeling, EA was applied to the right "Zusanli" (ST36) and "Sanyinjiao" (SP6) (1 mA,2 Hz/100 Hz) for 30 min. The mechanical pain threshold(MWT) and thermal pain thre-shold(TPT) of the affected limb were detected before modeling, 7 days following modeling and 60 min after EA. The expression of MT2 in spinal dorsal horn was detected by Western blot. The contents of melatonin （Mel） and IL-17 in the spinal dorsal horn were determined by ELISA. The expression of glial fibrillary acidic protein (GFAP) in the spinal dorsal horn was determined by Western blot and immunohistochemistry. In the second part, 30 rats were divided into 5 groups: sham operation, model, EA, MT2 antagonist (4-P-PDOT), and dimethyl sulfoxide (DMSO) groups, with 6 rats in each group. Rats of the 4-P-PDOT and DMSO groups were intrathecal injection with 10 µL MT2 antagonist 4-P-PDOT (100 µg) and equivalent DMSO 30 min before EA. The MWT and TPT of affected limb were detected. The GFAP expression and IL-17 content in the spinal dorsal horn was detected by Western blot， immunohistochemistry and ELISA, respectively. In the third part, 30 rats were randomly divided into 5 groups: sham operation, model, EA, recombinant IL-17, and normal saline groups, with 6 rats in each group. The recombinant IL-17 protein (100 ng, 10 µL) and the same amount of 0.9% sodium chloride solution were intrathecal injection into the rats of the recombinant IL-17 group and the normal saline group 30 min before the EA. The MWT and TPT of affected limb were measured. RESULTS: On the 7th day after modeling, the MWT of rats in the model group and the EA group were significantly higher, while TPT were lower than those before the modeling (P<0.05). At 60 min after EA, compared with the model group, the MWT and TPT of the EA group reversed significantly (P<0.05). The levels of GFAP and IL-17 were significantly increased, while the levels of Mel and MT2 were significantly decreased in the model group than in the sham operation group (P<0.05), and those were considerably reversed in the EA group than in the model group (P<0.05). Compared with the EA and DMSO groups, the MWT in the 4-P-PDOT group were significantly increased, while TPT were decreased (P<0.05), and the contents of GFAP and IL-17 were significantly increased (P<0.05). Compared to the EA and normal saline groups, MWT of the rats in the recombinant IL-17 group were significantly increased, while TPT decreased (P<0.05). CONCLUSION: EA of ST36 and SP6 can alleviate neuropathic pain in CCI rats, which is closely related to its effect in inhibiting the release of IL-17 from astrocytes mediated by MT2.

[Effect of Yes-associated protein in regulation of electroacupuncture pretreatment on cerebral ischemia reperfusion injury rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) pretreatment on inflammatory reaction, apoptosis and expression of Yes-associated protein (YAP) of ischemic penumbra of cerebral cortex in cerebral ischemia reperfusion injury rats, and to explore the possible mechanism of its neuroprotection effect. METHODS: A total of 84 SD rats were randomized into a sham operation group (12 rats), a model group (18 rats), an EA group (18 rats), an EA+YAP virus transfection group (18 rats) and an EA+virus control group (18 rats). Except for the sham operation group, thread embolization method was adopted to establish the middle cerebral artery occlusion (MCAO) model in rats of the other groups. EA was applied at "Baihui" (GV 20) and "Dazhui" (GV 14) for 30 min in the 3 EA intervention groups 2 h before model establishment, disperse-dense wave, 2 Hz/15 Hz in frequency and 1 mA in intensity. Adenovirus transfection technique was used to induce gene silencing of YAP in the EA+YAP virus transfection group, and adenovirus vectors was injected as negative control in the EA+virus control group 4 d before model establishment. Twenty-four hours after model establishment, neurological function score was evaluated, the relative cerebral infarction area was observed by TTC staining, the apoptosis in the ischemic penumbra of cerebral cortex was detected by TUNEL staining, the levels of inflammatory factors IL-1ß, IL-6 and TNF-α in the ischemic penumbra of cerebral cortex was detected by ELISA method, the expression of YAP was detected by Western blot and immunofluorescence. RESULTS: Compared with the sham operation group, the expression of YAP was increased in the model group (P<0.05); compared with the model group, the expression of YAP in the ischemic penumbra of cerebral cortex was increased in the EA group (P<0.05). Compared with the sham operation group, the neurological function score, the percentage of TUNEL positive cells and the levels of IL-1ß, IL-6 and TNF-α in the ischemic penumbra of cerebral cortex were increased in the model group (P<0.001, P<0.01); compared with the model group, the neurological function score, the relative cerebral infarction area, the percentage of TUNEL positive cells and the levels of IL-1ß, IL-6 and TNF-α in the ischemic penumbra of cerebral cortex were decreased in the EA group (P<0.05, P<0.01); compared with the EA group, the neurological function score, the relative cerebral infarction area, the percentage of TUNEL positive cells and the levels of IL-1ß, IL-6 and TNF-α in the ischemic penumbra of cerebral cortex were increased in the EA+YAP virus transfection group (P<0.01, P<0.05); compared with the EA+YAP virus transfection group, the neurological function score, the relative cerebral infarction area, the percentage of TUNEL positive cells and the levels of IL-1ß, IL-6 and TNF-α in the ischemic penumbra of cerebral cortex were decreased in the EA+virus control group (P<0.01, P<0.05). CONCLUSION: Electroacupuncture pretreatment can effectively improve the ischemia reperfusion injury, its mechanism may be related to up-regulating the expression of YAP in the ischemic penumbra of cerebral cortex and relieving the apoptosis and inflammatory reaction.

[Spinal transcription factor GATA4 mediated adenosine A1 receptor activation contributes to electroacupuncture analgesia in neuropathic pain rats].

OBJECTIVE: To investigate the effect of electroacupuncture (EA) on pain behaviors and expression of spinal transcription factor GATA-binding Protein 4 (GATA4) and adenosine A1 receptor in neuropathic pain rats, so as to explore its mechanism underlying pain relief. METHODS: The present study includes 2 parts. In the first part, 18 SD rats were randomly divided into control, adenovirus short-hairpin interference RNA for GATA4 (AV-shGATA4 RNA) and adenovirus empty vector (AV-control short-hairpin RNA, AV-shCTRL) groups, with 6 rats in each group. The expression of GATA4 protein in the lumbar spinal cord (L4-L6) was detected to evaluate the transfection efficiency of AV-shGATA4 RNA (silencing GATA4 expression). In the second part, thirty SD rats were randomly divided into 5 groups, namely sham operation, CCI model, EA, EA+AV-shGATA4 RNA, and EA+AV-shCTRL groups, with 6 rats in each group. The neuropathic pain model was established by chronic constriction injury (CCI) of the right sciatic nerve. On the 7th day following modeling, EA was applied to the right "Zusanli"(ST36) and "Taichong"(LR3) (1 mA，2 Hz /100 Hz) for 30 min. Rats of the EA+AV-shGATA4 RNA and EA+AV-shCTRL groups received intrathecal injection of AV-shGATA4 RNA and AV-shCTRL(1×1011 PFU/mL，10 µL)at the spinal L4-L6 segments, separately, 48 h before EA intervention. The mechanical pain threshold and thermal pain threshold of the affected limb were detected before molding, 7 days following molding and 60 min after EA. The expressions of adenosine A1 receptor and GATA4 protein in the spinal cord (L4-L6) were detected by Western blot. RESULTS: Outcomes of the first part showed that compared with the control group, no significant changes were found in the mechanical and thermal pain thresholds in both AV-shCTRL and AV-shGATA4 RNA groups and in the expression of spinal GATA4 protein of the AV-shCTRL group (P＞0.05). The expression of spinal GATA4 protein of the AV-shGATA4 RNA group was significantly lower than that of the AV-shCTRL group (P<0.05). In the second part of the study, before CCI modeling, there were no significant differences among the five groups in the mechanical and thermal pain thre-sholds (P> 0.05). On the 7th day following modeling, the mechanical and thermal pain thresholds were significantly lowered in compa-rison with their own pre-modeling of each group and with the sham operation group (P<0.05). At 60 min after EA and compared with the model group, the mechanical and thermal pain thresholds were significantly increased in both the EA and EA+AV-shCTRL groups (P<0.05) but not in the EA+AV-shGATA4 RNA group (P>0.05), suggesting a critical involvement of GATA4 in EA analgesia. The expression levels of adenosine A1 receptor and GATA4 protein were significantly increased in the model group than in the sham operation group (P<0.05), and considerably further up-regulated in both EA and EA+AV-shCTRL groups (P< 0.05), rather than in the EA+AV-shGATA4 RNA group (P>0.05), suggesting that the effects of EA in up-regulating the expression of A1 receptor and GATA4 were eliminated after silencing GATA4 protein. CONCLUSION: EA of ST36 and LR3 can relieve pain by increasing the expression of adenosine A1 receptor of the lumbar spinal cord in neuropathic pain rats, which is probably mediated by GATA4 protein.

[Study on efficacy specificity of electroacupuncture at "Zusanli "(ST36) and "Baihui" (GV20) in cerebral ischemia and inflammatory pain rats].

OBJECTIVE: To explore the efficacy difference between electroacupuncture (EA) at "Zusanli" (ST36) and "Baihui" (GV20) for inflammatory pain and cerebral ischemia-reperfusion injury (CIRI) in rats. METHODS: In 1st part of this study, 90 male SD rats were randomly divided into sham-operation, model (induced by occlusion of the middle cerebral artery and reperfusion), GV20 EA, ST36 EA，and sham EA groups (n=16 in each group). In the 2nd part of the study, 40 male SD rats were randomized into saline injection (control), inflammatory pain model (subcutaneous injection of complete Freund's adjuvant ［CFA］ into the right paw), ST36 EA, GV20 EA, and sham EA groups (n=8 in each group). In these two parts, EA (2 Hz/15 Hz, 1 mA) was applied to ST36 or GV20. The mechanical withdrawal threshold (MWT) and thermal withdrawal latency （TWL） were detected 2.5 h after administration of CFA by using Von Frey and plantar tester, respectively. The neurological deficit scores (NDS) were assessed by using Longa's method and the infarct size of the brain assessed after staining with 2% triphenyltetrazolium chloride (TTC). The expression of c-fos protein in the dorsal horns (DHs) of the spinal cord was detected by immunohistochemistry. RESULTS: (1) Twenty-four hours following CIRI, the NDS and infarct volume were significantly increased in the model group compared with the sham-operation group (P<0.01), and obviously decreased in the GV20 EA and ST36 EA groups relevant to the CIRI model group (P<0.05, P<0.01). There were no significant differences between the two EA groups in the NDS and infarct volume levels (P>0.05). (2) After administration of CFA, both the MPT and TPT were notably decreased in the inflammatory pain model group in contrast to the saline-injection group (P<0.01), but were considerably increased in both ST36 EA and GV20 EA groups (P<0.05), rather than in the sham EA group (P>0.05). The number of c-fos positive cells was significantly increased in the medial half of I-II and III-IV lamina of DHs in the L4-L6 segments of spinal cord in the inflammatory pain model group relevant to the saline-injection group (P<0.01,P<0.05), and was remarkably decreased in the lamina I-II (not in the deeper lamina) in both ST36 EA and GV20 EA groups (P<0.01), rather than in the sham EA group (P>0.05). No significant differences were found in the number of c-fos positive cells between the ST36 EA and GV20 EA groups (P>0.05). CONCLUSION: Our data do not support the specificity of functions at least between GV20 EA and ST36 EA in both CIRI and inflammatory pain model rats. This is the first study reporting the effect of EA at GV20 for relieving CFA-induced inflammatory pain.

Role of spinal adenosine A1 receptors in the analgesic effect of electroacupuncture in a rat model of neuropathic pain.

OBJECTIVE: The aim of this study was to determine the role of spinal adenosine A1 receptors (A1Rs) in the analgesic effects of electroacupuncture (EA) for neuropathic pain. METHODS: We performed EA for 30 minutes at the zusanli acupoint in the legs of rats with previously induced chronic constriction injuries and observed the mechanical and thermal pain thresholds 1 hour later. We also examined adenosine levels by high-performance liquid chromatography and A1R expression in the L4-6 spinal cord by western blot analysis. We then injected A1R short interfering RNA (AV-shA1RNA) into the L4-6 spinal cord to downregulate A1R expression and re-examined the mechanical and thermal pain thresholds. RESULTS: Adenosine levels and A1R expression in the L4-6 spinal cord were increased at 1 hour after EA. In addition, EA exhibited an analgesic effect that was reversed by AV-shA1RNA. CONCLUSIONS: Our results suggest that EA at the zusanli acupoint elicits an analgesic effect against neuropathic pain, mediated by A1Rs in the spinal cord.

[Involvement of miR-34a and p53 protein of cerebral cortex in electroacpuncture analgesia in mice with neuropathic pain].

OBJECTIVE: To explore the involvement of miR-34a in cerebral cortex mediated anti-hyperalgesic effect of electroacupuncture (EA) in mice with neuropathic pain induced by chronic constriction injury (CCI) of sciatic nerve, so as to reveal its mechanisms underlying improvement of neuropathic pain. METHODS: A total of 75 male C57BL/6 mice were equally randomized into 3 groups: sham, CCI model and CCI+EA (n=25 in each group). Mice of the sham group received simple separation of the right sciatic nerve without ligation. The CCI model was established by liagation of the right sciatic nerve. EA (2 Hz /15 Hz, 1 mA) was applied to bilateral "Zusanli" (ST36) and "Sanyinjiao" (SP9) for 30 min, once every other day. The mechanical and thermal pain threshold of the bilateral hind-paws was detected at the 3rd, 5th and 7th day after modeling, and the expression of miR-34a of bilateral cerebral cortex tissues and that of p53 protein of the left cerebral cortex were determined by using quantitive real time PCR and Western blot, respectively. RESULTS: The mechnical paw withdrawal frequency were significantly higher and the thermal paw withdrawal latencies (PWLs) were significantly shorter at the affected hind-limb (rather than at the healthy hind limb) on day 3, 5 and 7 in the CCI model group than those in the sham group (P<0.05), and considerably reversed at the affected hind-limb (rather than at the healthy hind limb) in the EA group than in the CCI model group (P<0.05), suggesting an analgesic effect of EA intervention. After modeling, the expression levels of miR-34a and p53 on day 3, 5 and 7 were significantly up-regulated in the left cerebral cortex tissue (rather than in the right cerebral cortex) of the CCI model group in comparison with the sham group (P<0.05). After EA intervention, the up-regulated expression levels of miR-34a and p53 in the left cerebral cortex tissue (rather than in the right cerebral cortex) were obviously suppressed in the EA group relevant to the CCI model group (P<0.05). CONCLUSION: EA stimulation of ST36 and SP9 can down-regulate the expression of miR-34a and p53 in the contra-lateral cerebral cortex tissue of the CCI mice, which may contribute to its anti-hyperalgesic effect.

[Effect of transcutaneous electrical acupoint stimulation on pulmonary function and oxidative stress response in lower extremity surgery patients using tourniquet].

OBJECTIVE: To investigate the effect of transcutaneous electrical acupoint stimulation (TEAS) on pulmonary function, superoxide dismutase (SOD) activity, and malondialdehyde (MDA) content in patients using tourniquet after lower extremity surgery. METHODS: A total of 40 patients who underwent lower extremity surgery were equally randomized into control group and TEAS group by using a random number table. All patients underwent lumbar epidural anesthesia combined with block anesthesia. The patients in the TEAS group were given TEAS at Zusanli (ST36) and Sanyinjiao (SP6) beginning from 30 min before surgery to the end of surgery, and those in the control group received TEAS at the same acupoints with minimum current intensity. Mean arterial pressure (MAP) and heart rate (HR) were recorded at each time point (T0: pre-surgery /TEAS, T1: 5 min after anesthesia, T2: 1 min before tourniquet-loosening, T3: 1 min after tourniquet-loosening, T4: 5 min after tourniquet-loosening, and T5: 6 h after tourniquet-loosening). Blood samples (4 mL) was collected from the radial artery before TEAS and 6 h after loosening tourniquet for analyzing blood gas parameters as partial pressure of caron dioxide(PCO2), arterial partial pressure of oxygen (PaO2), alveolar partial pressure of oxygen (PAO2), alveolar-arterial oxygen pressure difference (PA-aDO2) and respiratory index (RI) by using a blood gas analyzer, and plasma SOD activity and MDA content were assayed by using xanthine oxidase method and thiobarbituric acid colorimetry method, respectively. RESULTS: Intra-group comparison showed that compared with T0, a significant increase was found in PA-aDO2 and RI at T5 and a significant reduction in PaO2 and PaO2/ PAO2 (a/A) ratio in the control group (P<0.05), and the same changes in the TEAS group (P<0.05) except a/A ratio. Comparison between two groups showed that at T5, both PaO2 and a/A levels were significantly higher in the TEAS group than in the control group (P<0.05), and both PA-aDO2 and RI levels were obviously lower in the TEAS group than in the control group (P<0.05), suggesting an improvement of the pulmonary function after TEAS. At T5, plasma SOD activity was significantly decreased and plasma MDA content was remarkably increased in the control group relevant to T0 (P<0.05), SOD activity was significantly higher in the TEAS group than in the control group (P<0.05), and MDA content was evidently lower in the TEAS group than in the control group (P<0.05), suggesting a reduction of oxidative stress response after TEAS. CONCLUSION: TEAS at ST36 and SP6 can improve pulmonary function and attenuate oxidative stress response in patients using tourniquet after lower extremity surgery.

Electroacupuncture-induced neuroprotection against focal cerebral ischemia in the rat is mediated by adenosine A1 receptors.

The activation of adenosine A1 receptors is important for protecting against ischemic brain injury and pretreatment with electroacupuncture has been shown to mitigate ischemic brain insult. The aim of this study was to test whether the adenosine A1 receptor mediates electroacupuncture pretreatment-induced neuroprotection against ischemic brain injury. We first performed 30 minutes of electroacupuncture pretreatment at the Baihui acupoint (GV20), delivered with a current of 1 mA, a frequency of 2/15 Hz, and a depth of 1 mm. High-performance liquid chromatography found that adenosine triphosphate and adenosine levels peaked in the cerebral cortex at 15 minutes and 120 minutes after electroacupuncture pretreatment, respectively. We further examined the effect of 15 or 120 minutes electroacupuncture treatment on ischemic brain injury in a rat middle cerebral artery-occlusion model. We found that at 24 hours reperfusion,120 minutes after electroacupuncture pretreatment, but not for 15 minutes, significantly reduced behavioral deficits and infarct volumes. Last, we demonstrated that the protective effect gained by 120 minutes after electroacupuncture treatment before ischemic injury was abolished by pretreatment with the A1-receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (1 mg/kg, intraperitoneally). Our results suggest that pretreatment with electroacupuncture at the Baihui acupoint elicits protection against transient cerebral ischemia via action at adenosine A1 receptors.

Carnosic acid attenuates neuropathic pain in rat through the activation of spinal sirtuin1 and down-regulation of p66shc expression.

BACKGROUND: It has been reported that carnosic acid (CA) exhibits a range of biological activities including hepatoprotective, antioxidant and anti-inflammatory. However, the effect of carnosic acid in neuropathic pain remained elusive. METHODS: A neuropathic pain model of chronic constriction injury (CCI) was established in adult male Sprague-Dawley rats. Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were recorded, and western blot was performed to detect sirtuin1 and p66shc content. RESULTS: Intrathecal administration of carnosic acid attenuated mechanical allodynia and thermal hyperalgesia in rats following chronic constriction injury. Interestingly, carnosic acid analgesic effect was positively associated with spinal sirtuin1 activation; however, p66shc was inhibited by carnosic acid in the spinal cord. In additional, sirtuin1 inhibitor EX-527 reversed the anti-nociceptive effect of carnosic acid. CONCLUSIONS: Carnosic acid is effective in the treatment of the established CCI-induced pain. It may be possible that spinal sirtuin1 activition by carnosic acid attenuates neuropathic pain through a mechanism involving the down-regulation of p66shc expression.

[Effect of transcutaneous acupoint electrical stimulation combined with target controlled infusion of propofol on efficacy of general anesthesia for craniotomy].

OBJECTIVE: To observe the effect of transcutaneous acupoint electrical stimulation (TAES) combined with target controlled infusion of Propofol on the doses of Propofol and adjuvant drugs, and on the resuscitation time of general anesthesia for craniotomy patients. METHODS: Forty patients (aged 27 - 65 years), scheduled for craniotomy and signed the informed consent, were randomly and equally divided into TAES group and control group. Patients of the two groups received intravenous anesthesia mainly with target controlled infusion of Propofol. TAES (2 Hz/100 Hz, 1-12 mA) was applied to bilateral Yuyao (EX-HN 4) and Taiyang (EX-HN5) for 20 min first before surgery, and then to bilateral Hegu (LI 4), Quanliao (SI 18) and Fengchi (GB 20) during operation and till the end of the operation. The dosages of Propofol and adjuvant drugs, the duration of surgery and anesthesia, and the time of resuscitation and extubation were recorded. RESULTS: Compared with the control group, the dosages of Propofol and Nicardipine for craniotomy patients in the TAES group were significantly lower (P < 0.05), and the resuscitation time was obviously earlier and the tracheal catheter indwelling time markedly shorter in the TAES group (P < 0.05). CONCLUSION: TAES combined with target controlled infusion of Propofol can reduce the dosage of Propofol and Nicardipine, and shorten the resuscitation time and tracheal catheter indwelling time in craniotomy patients.