Anesthesiology 2009; 110:1244 –52 Copyright © 2009, the American Society of Anesthesiologists, Inc. Lippincott Williams & Wilkins, Inc. Pharmacological Characteristics of the Inhibition of Nondepolarizing Neuromuscular Blocking Agents at Human Adult Muscle Nicotinic Acetylcholine Receptor Malin Jonsson Fagerlund, M.D., Ph.D.,* Michael Dabrowski, Ph.D.,† Lars I. Eriksson, M.D., Ph.D.‡ Background: Nondepolarizing neuromuscular blocking agents (NMBAs) are classic competitive-inhibitors at the muscle nicotinic acetylcholine receptor (nAChR). Although the fetal subtype mus- cle nAChR has been extensively studied at a molecular level, less is known about the interaction between nondepolarizing NMBAs and the human adult muscle nAChR. The aim of this study was to investigate the effect of clinically used nondepolarizing NMBAs at human adult muscle nAChRs and the mechanisms behind the inhibition. Methods: Human subunits for the adult  1  1  muscle nAChR were cloned and expressed into Xenopus oocytes and thereafter studied with two-electrode voltage clamp. The effect of the clini- cally used nondepolarizing NMBAs, including atracurium, cis-atra- curium, mivacurium, pancuronium, rocuronium, vecuronium, and d-tubocurarine, on acetylcholine-induced and dimethylphe- nylpiperazinium-induced currents were investigated. Results: All nondepolarizing NMBAs tested inhibited acetylcho- line- and dimethylphenylpiperazinium-induced currents in hu- man adult  1  1  muscle nAChRs, and no receptor activation was seen. Interestingly, acetylcholine desensitized the human adult  1  1  muscle type receptor and attenuated the inhibition caused by nondepolarizing NMBAs, as evident by lack of increase in IC 50 values for the nondepolarizing NMBAs with increased concentra- tions of acetylcholine. In contrast, dimethylphenylpiperazinium- induced currents were competitively inhibited by the nondepolar- izing NMBAs. Conclusions: This study demonstrates that nondepolarizing NMBAs inhibit human adult muscle nAChRs expressed in Xenopus oocytes by mixed mechanisms. When using the nondesensitizing agonist dimethylphenylpiperazinium, inhibition by the NMBA is competitive, whereas activation with high concentrations of acetyl- choline in combination with NMBA induces a noncompetitive inhi- bition, which the authors speculate can involve receptor desensitiza- tion similar to that observed in the neuromuscular junction. NONDEPOLARIZING neuromuscular blocking agents (NMBAs) are extensively used in routine practice of anes- thesia and intensive care medicine to provide muscle re- laxation. It is well known that nondepolarizing NMBAs block the transmission in the neuromuscular junction by inhibition of nicotinic acetylcholine receptors (nAChRs), both presynaptically and postsynaptically. 1,2 The nAChR is the prototype of the ligand-gated ion channel superfamily and is composed by five subunits arranged around a central cation pore. 3 The nicotinic receptors are further subdivided into muscle and neuro- nal subtypes, the muscle subtypes being composed of two 1 subunit: one  1 , , and / subunit, and the neuronal subtypes are  and  heteromers or  ho- momers. 4 In the neuromuscular junction, neuronal  3  2 nAChRs are presynaptical autoreceptors, whereas the adult muscle  1  1  and/or the fetal  1  1  subtype are located postsynaptically. Finally, during denervation/im- mobilization the neuronal  7 nAChR has been demon- strated postsynaptically. 5,6 The fetal and adult subtypes of nAChRs have individual biophysical and pharmacolog- ical properties 7,8 ; historically, however, the fetal muscle nAChR has been more extensively studied, and less is therefore known about the adult muscle subtype. Al- though affinity and potency data for nondepolarizing NMBAs at human adult muscle nAChRs are sparse, 1 rodent data on adult muscle nAChRs expressed in Xeno- pus oocytes has been published previously with some- what divergent result. 9 –11 Garland et al. found vecuro- nium to be more potent at mouse adult nAChRs compared to d-tubocurarine and pancuronium, whereas Paul et al. demonstrated that pancuronium was the most potent NMBA of the three. 10,11 In addition, a recent study found d-tubocurarine to have the highest affinity of several clinically used NMBAs at the mouse adult nAChR. 12 Notably, previous studies have shown that nondepolarizing NMBAs can act as both agonists, com- petitive and noncompetitive antagonists at rodent mus- cle nAChRs, 11–14 but we lack information on the mech- anisms behind the human receptor inhibition. Although there is a large sequence homology between human and rodent nAChRs, differences in single amino acid se- quences can cause relatively large functional affinity and kinetic differences. 8,12 To our knowledge, only one study has described the effect of clinically used nonde- polarizing NMBAs at the human adult muscle nAChR, and no further characterization has been done. 1 Therefore, the aim of this study was to investigate the inhibition at the human adult muscle nAChR produced by clinically used nondepolarizing NMBAs. * Resident, Department of Anesthesiology and Intensive Care Medicine, Karo- linska University Hospital and Karolinska Institutet, Stockholm, Sweden; † Asso- ciate Principle Scientist, Molecular Pharmacology, AstraZeneca R&D, So ¨derta ¨lje, Sweden; ‡ Professor, Department of Anesthesiology and Intensive Care Medi- cine, Karolinska University Hospital and Karolinska Institutet, Stockholm, Sweden. Received from the Department of Anesthesiology and Intensive Care Medi- cine, Karolinska University Hospital, Stockholm and Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden. Submitted for publication July 3, 2008. Accepted for publication January 15, 2009. Supported by grants from the Swedish Research Council (project number 10401), the Stockholm County Council, the Swedish Society of Medicine, Karolinska Institu- tet funds (all Stockholm, Sweden). Organon is a part of Schering-Plough, Rose- land, New Jersey. Presented in part as a poster discussion at the Annual Meeting of the American Society of Anesthesiologists, Chicago, Illinois, October 14 –18, 2006. Address correspondence to Dr. Jonsson Fagerlund: Department of Anesthesiology and Intensive Care Medicine, and Karolinska Institutet, Karolinska University Hospi- tal, Stockholm, SE-171 76 Stockholm, Sweden. malin.jonsson.fagerlund@ki.se. 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