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Objective: To observe the effect of electroacupuncture (EA) on urodynamics of neurogenic bladder and pituitary adenylate cyclase activating peptide(PACAP)/cyclic adenosine monophosphate (cAMP)/protein kinase A(PKA) signaling pathway in detrusor tissue of rats after suprasacral spinal cord injury (SCI), so as to explore its possible mechanism in improving detrusor hyperreflexia bladder function after shock stage of suprasacral SCI., Methods: Female SD rats were randomly divided into control, sham operation, model and EA groups, with 12 rats in each group. T10 spinal cord transection (SCT) was performed by surgery. Rats in the EA group were given EA (10 Hz/50 Hz, 20 min) at "Ciliao" (BL32), "Zhongji" (CV3) and "Sanyinjiao" (SP6) once daily for 7 days. After the intervention, urodynamics testing was detected in each group. HE staining was used to observe the morphological changes of bladder detrusor. The protein and mRNA expression of PACAP38 in detrusor was detected by immunohistochemistry, Western blot and real-time quantitive PCR, respectively. The contents of cAMP and PKA were determined by ELISA., Results: Compared with the control and sham operation groups, the maximum bladder capacity and bladder com-pliance, and the protein and mRNA expression of PACAP38, and the contents of cAMP and PKA of the model group were significantly decreased ( P <0.01, P <0.05), while the base pressure and leakage point pressure of bladder were significantly increased ( P <0.01). After EA intervention, the above indexes were all reversed in the EA group relevant to those of the model group ( P <0.01, P <0.05)., Conclusion: EA at BL32, CV3 and SP6 can improve the bladder function in rats with bladder detrusor hyperreflexia after SCI, which may be related to its effect in activating the PACAP/cAMP/PKA signaling pathway in detrusor tissue.

Objective: To investigate the effect of protein kinase A (PKA) activation on aggregation funetion of platelets in vitro., Methods: The peripheral blood of healthy adults were collected, and the washed platelets were gained from collected peripheral blood. The washed platelets were treated with PKA activator Forskolin, then the platelet aggregation was induced by using Ristocetin, Thrombin, Collagen and ADP respectively, the platelet aggregation level was detected by the platelet aggregator., Results: Compared with the controls, 5 μmol/L forskolin significantly inhibited ADP and collagen-induced platelet aggregation (P＜0.001), and showed mild inhibiting effect on Thrombin-induced platelet aggregation (P＜0.05). 2.5-10 μmol/L forskolin significantly inhibited ADP and Collagen -induced platelet aggregation (P＜0.001); but not showed significantly inhibitory effects on Ristocetin-induced platelet aggregation (P＞0.05)., Conclusion: PKA activation inhibits agonists-induced platelet aggregation.

Objective To observe the effect of low-frequency pulsed electromagnetic fields(PEMFs) on bone formation in rat osteoblasts(ROBs) and explore the mechanism of action of the cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA)/cyclic adenosine effect binding protein(CREB) signaling pathway.Methods The skulls of newborn Wistar rats were harvested,and the ROBs were obtained by multiple enzymatic digestion methods for subculture. After treatment with 50 Hz 0.6 mT PEMFs for 3,6,and 9 days,the alkaline phosphatase(ALP) concentration in ROBs was detected;after 0,15,30,60,90,and 120 min,the expression of bone formation-related factor(RUNX2),the protein expression of osteogenesis-associated transcription factor(OSX),the cAMP concentration,as well as the protein expressions of p-PKA,p-CREB,and CREB were detected. The p-CREB nuclear translocation was observed. After interference with IFT88 by RNA interference,the expressions of RUNX2,OSX,p-PKA,and p-CREB protein in ROBs were detected.Results After treatment with PEMFs for 3,6,and 9 days,the ALP activity values in ROBs were 24.356±4.911,37.688±2.151,and 39.922±5.486,respectively,which were significantly higher than 18.531±2.401(P=0.0121),33.675±4.366(P=0.0324),and 36.574±1.339(P=0.0134) in the control groups. RUNX2 and OSX activities in ROBs were significantly higher than untreated group after PEMFs treatment for 30(P=0.0042 and P=0.0058),60(P=0.0097 and P=0.0079),and 90 min(P=0.0083 and P=0.0098). After PEMFs treatment for 30(P=0.0012) and 60 min(P=0.0035),the cAMP concentrations in ROBs were significantly higher than that in untreated group. After PEMFs treatment for 15(P=0.0018),30(P=0.0087),90(P=0.0250),and 120 min(P=0.0350),the p-PKA levels in ROBs were significantly higher than that in the untreated group. After PEMFs treatment for 15(P=0.0075),30(P=0.0017),60(P=0.0074),and 90 min(P=0.0096),the level of p-CREB in the ROBs was significantly higher than in the untreated group. After PEMFs treatment of ROBs for 15 min,CREB phosphorylated and accumulated in the nuclei. PKA and p-PKA were co-localized with primary cilia and stained,and it was found that p-PKA was localized on the primary cilia. After the primary cilia was removed by RNA interference,the protein expression levels of p-PKA(F=78.602,P=0.0270),p-CREB(F=76.082,P=0.0089),RUNX2(F=41.064,P=0.0230) and OSX(F=57.524,P=0.0310) were significantly lower than those of the non-interfered group.Conclusion PEMFs promote bone formation in ROBs by activating the primary cilia-associated cAMP/PKA/CREB signaling pathway.

Objective: We have demonstrated that electroacupuncture (EA) of "Weizhong" (BL 40) and "Huantiao" (GB 30) could evidently relieve mechanical hyperalgesia in spared nerve injury (SNI) rats. The present study was designed to observe the effect of EA on levels of cAMP, and protein kinase A (PKA) and cAMP response element binding protein (CREB) in the lumbar spinal cord of the same pain model rats, so as to explore its mechanisms underlying improvement of neuropathic pain., Methods: One hundred male SD rats were randomly divided into 5 groups: sham operation, model, EA, AP-5 and L-NAME groups ( n ＝20 in each group). The sham operation group underwent a simple separation of the sciatic nerve but without ligation and abscission. The neuropathic pain model was established by abscission of the right tibial and common peroneal nerve. EA (2 Hz, 1－3 mA) was applied to right BL 40 and GB 30 for 30 min, once a day for 7 days, starting from day 11 after surgery. For rats of the AP-5 and L-NAME groups, AP-5 (a competitive antagonist for NMDA receptor, 0.7 mg•kg －1 •d －1 ) and L-NAME (a non-selective antagonist for nitric oxide synthase, NOS, 60 mg•kg －1 •d －1 ) were respectively administrated by intraperitoneal injection, once daily for 7 days. The mechanical pain threshold was measured, the cAMP content of spinal cord (L 4－L 6) was determined by radioimmunoassay, and the expression of PKA, p-PKA and CREB proteins of spinal cord (L 4－L 6) was detected by Western blot., Results: The content of cAMP and the expression levels of PKA, p-PKA and CREB were significantly higher in the model group than in the sham operation group ( P <0.01), and considerably lower in the EA group than in the model group ( P <0.01, P <0.05). Compared to the model group, the expression levels of PKA and p-PKA were significantly decreased in the AP-5 and L-NAME groups ( P <0.01, P <0.05). Compared to the EA group, the content of cAMP and the expression levels of CREB were significantly higher ( P <0.05), and the expression of p-PKA was significantly lower in the AP-5 and L-NAME groups ( P <0.05)．., Conclusion: EA intervention-induced down-regulation of cAMP, PKA and CREB levels in the lumbar spinal cord may contribute to its analgesic effect in neuropathic pain rats, suggesting an involvement of reduction of cAMP/PKA/CREB signaling of spinal cord in EA analgesia.

Objective: To explore the effect of PKA gene on acute T lymphocyte leukemia cells in children and its mechanism., Methods: Jurkat and Sup-T1 cells were divided into 2 group: control group (Jurkat and Sup-T1 cells treated with non-specific siRNA) and transfected group (Jurkat and Sup-T1 cells transfected with PKA siRNA). The effects of down-regulating the expression of PKA gene on the viability, proliferotion, migration and cell cycle distribution of Jurkat and Sup-T1 cells in 2 groups were analyzed by CCK-8 assay, transwell experiment, cell colony-formation test and flow cytometry; the cyclin-related protein levels after transfection with PKA siRNA were detected by Western blot., Results: It was revealed that the expression of PKA in Jurkat and Sup-T1 cells decreased to different degree after siRNA transfection(P<0.05). CCK-8 assay showed that the proliferation of Jurkat and Sup-T1 cells in the transfected group was slower than that in the control group(P<0.05). Transwell experiment showed that the migration and invasion ability of Jurkat cells in the transfection group was weaker than that in the control group (P<0.05). The cell colong formation number of Jurkat and Sup-T1 cells in the transfection group decreased significantly (P<0.05). The cell level of Jurkat and Sup-T1 cells in G 0 /G 1 phase increased after tansfection (P<0.05). Western blot assay revealed that the expression levels of CDK2, CyclinD1 and p-Rb in the Jurkat and Sup-T1 cells of the transfection group were suppressed (P<0.05)., Conclusion: The down-regulating PKA gene expression can decrease the proliferation and migration of tumor cells, and also can restrict the cell proliferation through related cell cycle proteins.

Objective: To study the effect of electro-acupuncture (EA) at Neiguan (PC6) and Lieque (LU7) on the expression of protein kinases in cardiomyocytes of myocardial ischemia (MI) rats., Methods: Healthy male SD rats were randomly divided into the control group, the model group, the Neiguan point group, the Lieque point group, and the non-meridian non-acupoint group, 10 in each group by random digit table. The MI rat model was established by injecting isoprenaline hydrochloride (85 mg/kg). EA at Neiguan (PC6), Lieque (LU7), and non-meridian non-acupoint were respectively performed. Changes of the expression of protein kinases [such as protein kinase A (PKA), protein kinase C (PKC), protein kinase G (PKG)] in rat cardiomyocytes were observed using Western blot., Results: Compared with the control group, expression levels of PKA, PKC, and PKG increased obviously in the model group (P < 0.01). Compared with the model group, expression levels of PKA, PKC, and PKG decreased in the Neiguan point group and the Lieque point group (P < 0.01, P < 0.05). Expression levels of PKA decreased in the non-meridian non-acupoint group (P < 0.01). Compared with the Neiguan point group, expression levels of PKA, PKC, and PKG increased in the non-meridian non-acupoint group and the Lieque point group (P < 0.01, P < 0.05). Compared with the Lieque point group, expression levels of PKA, PKC, and PKG increased in the non-meridian non-acupoint group (P < 0.01, P < 0.05)., Conclusion: EA at Neiguan (PC6) and Lieque (LU7) could decrease protein expression levels of PKA, PKC and PKG in rat myocardial cells, and the effect of acupuncture at Neiguan (PC6) was better than that obtained by EA at Lieque (LU7).

Objective: To explore the potential biochemical mechanisms of neuromuscular junction transmission (NMJT) dysfunction induced by organophosphorus insecticides., Methods: Ten rats were dosed with phoxim (1,144 mg/kg) and 5 of them developed myasthenia. The NMJT function was evaluated by the mean consecutive differences (MCD) measured by stimulation single fiber electromyography (SSFEMG) with the frequency of stimuli at 20 Hz. The activity of Ca(2+)-ATPase, Na(+), K(+)-ATPase, cAMP dependent protein kinase (PKA), Ca(2+)/phospholipid dependent protein kinase (PKC), and tyrosine protein kinase (TPK) were determined., Results: In comparison with the control and non-myasthenic rats, the results in myasthenic rats showed that: (1) the MCDs increased; (2) the activities of Ca(2+)-ATPase, Na(+), K(+)-ATPase and PKA decreased and were negatively correlated with MCD; (3) the activities of PKC and TPK increased, and were positively correlated with MCD; (4) the phosphorylation of serine residuals in sarcolemma was weaker and the phosphorylation of tyrosine residuals was stronger., Conclusions: The NMJT dysfunction is likely associated with the decrease in Ca(2+)-ATPase and Na(+), K(+)-ATPase activity. The acceleration of desensitization and prolongation of resensitization of nicotinic acetylcholine receptors occur following the increase in tyrosine phosphorylation and the decrease in serine phosphorylation induced by OPs poisoning.