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Objective To evaluate the effects of different concentrations of chloroprocaine on KCNQ2/Q3 channel currents in HEK2936 cells.Methods Human embryonic kidney (HEK293) cells served as an expression system.KCNQ2 and KCNQ3 cDNAs and green fluorescent protein were transfected into HEK293 cells by using lipofectamine.The KCNQ2/Q3 currents were recorded by using the whole-cell patch-clamp technique.Part Ⅰ The transfected HEK293 cells were randomly divided into 4 groups (n =11 each):control group,and 1,10 and 100 mmol/L chloroprocaine groups.The KCNQ2/Q3 channel currents produced by different concentrations of chloroprocaine were recorded under different holding potentials (-40,0 and 40 mV) and the action time was 1 min.Part Ⅱ The transfected HEK293 cells were randomly divided into 2 groups (n =5 each):control group and 10 mmol/L chloroprocaine.The KCNQ2/Q3 channel currents were recorded under different holding potentials (-80-30 mV)and the action time was 1 min.Different test potentials were normalized and fitted to Boltzmann function,and KCNQ2/Q3 channel Ⅰ-Ⅴ curve was then obtained.The activation and deactivation currents were both fitted to a single exponential function and the time constants for current activation and for current deactivation were calculated.Results Part Ⅰ When the holding potential was 40,0 and-40 mV,the suppression rate of KCNQ2/Q3 channel currents in HEK293 cells was higher in 1,10 and 100 mmol/L chloroprocaine groups than in control group (P ＜0.05 or 0.01).Part Ⅱ Compared with control group,the time constant for the current activation at 0 mV of holding potential was prolonged,the time constant for the current deactivation was shortened when the holding potential was-80 mV,and the half-activation voltage of KCNQ2/Q3 channels was increased,the activation curve shifted to the depolarized potentials,and KCNQ2/Q3 channel Ⅰ-Ⅴ curve slope was decreased in 10 mmol/L chloroprocaine group (P ＜ 0.05).Conclusion Chloroprocaine concentration-dependently suppresses KCNQ2/Q3 channel currents in HEK2936 cells.The KCNQ2/Q3 channel is closed in advance due to KCNQ2/Q3 channel opening delay induced by chloroprocaine thus decreasing the activity of KCNQ2/Q3 channels.
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Objective To investigate the effect of KCNQ2/3 channel opener retigabine on the median effective dose (ED50) of bupivacaine and chloroprocaine for induction of convulsion in mice and the relationship between KCNQ2/3 channels and the neurotoxicity of local anesthetics.Methods Pathogen-free female Kunming mice,weighing 20-30 g,were used in the study.The experiment was performed in two parts.Part Ⅰ Sixty mice were randomly divided into 2 groups (n =30 each):control group (group C) and retigabine group (group R).The C and R groups were further divided into 3 subgroups with different doses of chlorprocaine (C + L1,C + L2 and C+ L3 groups,and R+ L1,R+ L2 and R+ L3 groups,n =10 each).In groups C and R,0.9％ normal saline 0.005 ml/g and retigabine 20 mg/kg were injected intraperitoneally,respectively,and chlorprocaine was injected intraperitoneally 20 min later.The doses of chlorprocaine were 150.0,172.5 and 198.4 mg/kg in C + L1,C + L2 and C + L3 groups,respectively,and 198.4,228.2 and 262.4 mg/kg in R+ L1,R+ 12 and R+ L3 groups,respectively.Part Ⅱ Eighty mice were randomly divided into 2 groups (n =40 each):control group (group C) and retigabine group (group R).The C and R groups were further divided into 4 subgroups with different doses of bupivacaine (C + B1,C + B2,C + B3 and C + B4 groups,and R + B1,R + B2,R + B3 and R + B4 groups,n =10 each).In groups C and R,0.9％ normal saline 0.005 ml/g and retigabine 20 mg/kg were injected intraperitoneally,respectively,and bupivacaine was injected intraperitoneally 20 min later.The doses of bupivacaine were 37.8,43.5,50.0 and 57.5 mg/kg in C + B1,C + B2,C + B3 and C + B4 groups,respectively,and 50.0,57.5,66.1 and 76.0 mg/kg in R + B1,R + B2,R + B3 and R + B4 groups,respectively.The ED50 and 95％ confidence interval (CI) of bupivacaine and chloroprocaine for induction of convulsion were calculated by Probit analysis.Results The ED50(95％ CI) of chloroprocaine was 165.3 (155.8-175.0) mg/kg,and the ED50(95％CI) of bupivacaine was 41.1 (36.7-44.5) mg/kg in C group.The ED50 (95％ CI) of chloroprocaine was 212.4 (200.2-224.3) mg/kg,and the ED5o (95％ CI)of bupivacaine was 51.5 (945.1-56.0)mg/kg in R group.Compared with group C,the ED50 of bupivacaine and chloroprocaine for induction of convulsion was significantly increased in group R (P ＜ 0.01).Conclusion KCNQ2/3 channel opener retigabine can significantly increase the ED50 of bupivacaine and chloroprocaine for induction of convulsion and reduce convulsion induced by bupivacaine and chloroprocaine in mice,indicating that the neurotoxicity of local anesthetics is related to inhibition of KCNQ2/3 channels.
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Objective To compare the effects of different doses of dexmedetomidine on the median effective concentration (EC50) of ropivacaine for epidural block and investigate the appropriate dose.Methods One hundred and twenty ASA Ⅰ patients of both sexes,aged 20-55 yr,weighing 50-75 kg,scheduled for knee arthroscopic operation,were randomly divided into 4 groups (n =30 each):no dexmedetomidine group (group D0),0.25 μg/kg dexmedetomidine group (group D0.25),0.50μg/kg dexmedetomidine group (group D0.50),and 1.00μg/kg dexmedetomidine group (group D1.00).In group D0,ropivacaine 20 ml was injected into epidural space.The ropivacaine-dexmedetomidine mixtures containing 0.25,0.50 and 1.00 μg/kg dexmedetomidine were injected into epidural space in groups D0.25,D1.00 and D0.50 respectively.The volume of mixtures was 20 nl in groups D0.25,D1.00 and D0.50.The initial concentration of ropivacaine was set at 0.40 ％,0.40 ％,0.28 ％ and 0.20 ％ in groups D0,D0.25,D0.50 and D1.00 respectively and then the EC50 was determined by up-and-down technique.The concentration of ropivacaine was increased/decreased by 0.02％ in the next patient.The analgesic effect was assessed using VAS score.VAS score =0 was considered as effective analgesia.The EC50 and 95％ confidence interval (CI) of ropivacaine were calculated using probit method.Adverse effects were recorded.Results The EC50 and 95 ％ CI of ropivacaine was 0.38％ (0.35-0.41)％,0.34％ (0.31-0.36)％,0.22％ (0.20-0.24)％ and 0.14％ (0.12-0.15) ％ in groups D0,D0.25,D0.50 and D1.00 respectively.The EC50 of ropivacaine was decreased gradually in groups D0,D0.25,D0.50 and D1.00 ( P ＜ 0.05).Compared with group D0,the incidonce of hypotension and bradycardia was significantly increased in group D1.00 ( P ＜ 0.05),while no significant change was found in the incidence of adverse effects in groups D0.25 and D0.50 (P ＞ 0.05).Conclusion The appropriate dose of dexmedetomidine for epidural analgesia is 0.50 μg/kg.