Effects of Ropivacaine on Voltage-dependent Sodium Channels in Cultured Rat Hippocampal Neurons<?xml:namespace prefix = o ns = "urn:schemas-microsoft-com:office:office" /> ABSTRACT Objective: Effects of ropivacaine on voltage-dependent sodium channels in cultured rat hippocampal neurons were investigated, and the relationship between adverse effects of local anesthetic on central nervous system and voltage-depentdent sodium channels were identified. Methods: Effects of ropivacaine on voltage-dependent sodium channels in cultured hippocampal neurons were observed using whole cell patch clamp technique. Results: As the holding potential was-90mv,1,10,50,100,1000,2000umol.L-1 ropivacaine reduced sodium peak currents by 7.35±2.29%,13.91±4.68%,32.67±5.11%,42.49±6.03%,78.59±7.98%,86.03±8.27%, (P<0.001), repectively. The IC50 was 157.66±22.49umol.L-1. Analysis of variance of single factor showed that the inhibitory effects of ropivacaine on sodium currents were significantly different in different holding potentials (P<0.001), i.e. the inhibitory effects on sodium currents were voltage-dependent. Conclusions: Ropivacaine had significant depressing effects on sodium current in a dose-dependent and voltage-dependent way. It is suggested that adverse effects of local anesthetic on central nervous system was related to the depression of sodium channels in hippocampal neurons. Key Words: Sodium channel, Voltage-dependent; Hippocampal neurons;Amides 罗哌卡因是一新型酰胺类局麻药,与布吡卡因相比,具有运动阻滞轻,心肌毒性作用低等优点[1]。然而,当其被误注血管或累积用药量超过正常范围,会出现中枢神经系统的中毒症状,初期表现为口唇麻木,耳鸣,视觉模糊,躁动不安,抽搐,甚至产生惊劂,如果不进行有效处理,最终可导致昏迷,发展为呼吸系统和心血管系统抑制。研究提示,中毒后期的抑制性症状可能与局麻药阻断钠通道有关[2]。钠通道的几种亚型广泛存在于中枢神经系统,它的生理作用在于产生快速去极化的动作电位,当处于激活状态下,钠通道开放,允许钠离子沿着电化学梯度流入神经细胞内部[3]。罗哌卡因用药过量导致中枢神经系统中毒症状是否与电压依赖性钠通道有关,至今尚不清楚,本研究拟通过测定罗哌卡因对大鼠海马神经元电压依赖性钠通道的作用,明确局麻药中毒的细胞机制。 材料和方法 1. 海马神经元的分散和培养 取新生Wistar大鼠,用75%乙醇消毒。在无菌条件下断头、取脑,解剖镜下分离出海马,用0.25%胰蛋白酶(37℃)消化30min,吹打分散细胞悬液,将细胞按1×105ml-1的密度接种在事先涂有多聚赖氨酸的35mm培养皿中,置于36℃10的二氧化碳培养箱中过夜,24h后更换培养基,将种植液换成2ml饲养液。以后每3天半量换液一次,培养细胞在8~12天期间用于膜片钳实验。为抑制非神经元过度增殖,在培养的第3天向培养基中加入适量阿糖胞苷(每皿中加阿糖胞苷储备液6μl,终浓度为3μg.ml-1)。
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