1Clinic of Anaesthesiology, University Hospital Eppendorf and Institute of Neural Signal Transduction, Centre for Molecular Neurobiology, University of Hamburg, Germany. 2Department of Anaesthesiology and Intensive Care Medicine, University of Bonn, Germany. 3Department of Anaesthesiology and Intensive Care Medicine, University of Bonn, Germany, and Departments of Anaesthesiology and Physiology, Weill Medical College of Cornell University, New York, NY, USA.*Corresponding author: Klinik für Anästhesiologie, Universitätsklinik Eppendorf, Martinistrasse 52, D-20246 Hamburg, Germany
Accepted for publication: January 28, 2002
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Abstract |
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Methods. Kv3 channels natively expressed in human SH-SY5Y cells were studied using a standard whole-cell patch-clamp protocol.
Results. Bupivacaine reversibly inhibited Kv3 channels in a concentration-dependent manner. The half-maximal inhibitory concentration (IC50) for conductance block was 57 µM and the Hill coefficient was close to unity. Bupivacaine accelerated macroscopic current decline by inducing inactivation-like behaviour. The midpoint of current activation was shifted to depolarized potentials in a concentration-dependent and reversible manner by a maximum of 26 mV. The IC50 was 47 µM and the Hill coefficient was 2.4. The free arterial plasma concentrations of bupivacaine that have been estimated to occur during convulsions in man would inhibit the Kv3 channels by at least 40% and would shift the midpoint of current activation by a minimum of 9 mV.
Conclusions. Both inhibition of potassium channels and a depolarizing shift of their activation midpoint would increase neuronal excitability. The effects of bupivacaine on human Kv3 channels are thus compatible with a contributory role of Kv channel alteration in bupivacaine-induced neuronal excitation.
Br J Anaesth 2002; 88: 8646
Keywords: anaesthetics local; complications, side-effects; complications, seizure
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Introduction |
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Information on the anaesthetic targets that may be involved in these neurotoxic effects is limited. Recent molecular evidence emphasizes the importance of human voltage-dependent potassium channels in the pathogenesis of epileptic disorders.3 Mutations in the genes encoding voltage-dependent potassium channels of the Kv and KCNQ channel families cause seizures in animal and man through a decrease in neuronal potassium currents.3 Local anaesthetic-induced potassium current suppression may thus contribute significantly to the excitatory side-effects of these anaesthetic agents.
The aim of this study was to characterize the effects of bupivacaine on human neuronal Kv3 channels. Kv3 channels are crucial for spike frequency adaptation in central neurones.4 Evidence from knockout mice demonstrates that suppression of Kv3 channels causes impaired motor skill, muscle contraction, abnormal EEG patterns and epileptic seizures.4 Inhibition of human Kv3 channels by bupivacaine may contribute to the neuronal excitation and epileptic seizures observed during accidental intravascular application of this drug.
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Methods and results |
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Bupivacaine reversibly inhibited human Kv3 potassium channels. The potassium current traces in Figure 1 demonstrate the inhibitory effect. Bupivacaine accelerated macroscopic current decline (Fig. 1) by inducing inactivation-like behaviour. In the presence of bupivacaine, the potassium currents declined more than 100150 times faster at potentials between +40 and +70 mV compared with potassium currents in the absence of the local anaesthetic. The time constants of this inactivation-like behaviour did not differ between test potentials of +40 and +70 mV. The time constants depended on concentration [4.4 (1.5) and 2.9 (1.2) ms at 10 and 100 µM respectively; P<0.05, n=810].
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Comment |
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Human Kv3 channels belong to a superfamily of voltage-dependent potassium channels consisting of six transmembrane domains.3 4 Mutations of several members of this potassium channel superfamily cause inherited forms of human epilepsy by suppression of neuronal potassium channel activity.3 Kv3 knockout mice exhibit abnormal neuronal behaviour ranging from impaired motor skill to abnormal EEG patterns and epileptic seizures.4 Estimates based on in vitro and animal experiments suggest that inhibition of voltage-dependent potassium channels by 2550% may cause epilepsy.3 As calculated from the Hill equation, the inhibition produced by the free plasma concentrations of bupivacaine that are estimated to cause seizure in man (35 µM)9 and dogs (32126 µM)10 would suppress the Kv3 channels by at least 40%. The observed inhibition of human Kv3 channels by bupivacaine would thus be sufficient to account for excitatory side-effects.
The depolarizing shift of the activation threshold by 1020 mV induced by bupivacaine would add to the possible increase in neuronal excitability induced by inhibition of the potassium channel conductance.8 Both a 50% channel block and a 15 mV depolarizing shift would alone induce spontaneous firing of the HodgkinHuxley squid axon model.8 In the presence of a 15 mV depolarizing shift in potassium channel activation, spontaneous firing of the axon would persist if 90% of the sodium channels were blocked.8 The combination of potassium channel inhibition and a depolarizing shift in the activation threshold, as observed in our study, would be compatible with neuronal excitation during bupivacaine intoxication. Establishing structural requirements for local anaesthetic action on human neuronal Kv channels may therefore help to elucidate the molecular determinants of the convulsive side-effects of this drug.
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Acknowledgements |
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References |
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2 Moore DC, Crawford RD, Scurlock JE. Severe hypoxia and acidosis following local anesthetic-induced convulsions. Anesthesiology 1980; 53: 25960[ISI][Medline]
3 Ashcroft FM. Voltage-gated K+ channels. In: Ashcroft FM, ed. Ion Channels and Disease. London: Academic Press, 2000; 97122
4
Rudy B, Chow A, Lau D, et al. Contributions of Kv3 channels to neuronal excitability. Ann N Y Acad Sci 1999; 868: 30443
5 Friederich P, Dilger JP, Pongs O, Urban BW. Kv3.1 expression in human neuroblastoma SH-SY5Y cells. Pflügers Arch 2000; 439: R427
6 Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 1981; 391: 85100[ISI][Medline]
7 Friederich P, Benzenberg D, Trellakis S, Urban BW. Interaction of volatile anesthetics with human Kv channels in relation to clinical concentrations. Anesthesiology 2001; 95: 9548[ISI][Medline]
8
Elliott AA, Elliott JR. Voltage-dependent inhibition of RCK1 K+ channels by phenol, p-cresol, and benzyl alcohol. Mol Pharmacol 1997; 51: 47583
9 Tucker GT. Pharmacokinetics of local anaesthetics. Br J Anaesth 1986; 58: 71731[ISI][Medline]
10 Feldman HS, Arthur GR, Covino BG. Comparative systemic toxicity of convulsant and supraconvulsant doses of intravenous ropivacaine, bupivacaine, and lidocaine in the conscious dog. Anesth Analg 1989; 69: 794801[Abstract]