The Effect of Neostigmine on Twitch Tension and Muscle Relaxant Concentration during Infusion of Mivacurium or Vecuronium

With approval from our local Institutional Review Board and after obtaining informed consent, we studied ten patients, ASA physical status 1 and 2, aged 18-46 yr, scheduled for elective surgery. Patients exceeding 130% of ideal body weight; those with renal, hepatic, neuromuscular, and/or electrolyte disorders; and those taking medication known to interfere with neuromuscular function were excluded.

After intravenous administration of 1-2 mg midazolam, anesthesia was induced with 3-4 micro gram/kg fentanyl and 2-3 mg/kg propofol. Tracheal intubation was performed without paralysis, [4]and ventilation was controlled to maintain normocapnia (end-tidal P^CO2of 30-35 mmHg; Datex Ultima, Helsinki). Anesthesia was maintained with 70% N²O and 1% end-tidal isoflurane. Electrocardiogram, Sp^O2, noninvasive blood pressure, and esophageal temperature were monitored continuously. Esophageal temperature was maintained between 35.5 degrees Celsius and 37.0 degrees Celsius.

After induction of anesthesia, the ulnar nerve was stimulated via subcutaneous needle electrodes at the wrist. Supramaximal stimuli of 0.2 ms duration were delivered in a train-of-four at 2 Hz every 12 s (Digistim II, Neuro Technology, Houston, TX). Preload was maintained at 200-400 g. The evoked twitch tension of the adductor pollicis muscle was measured using a calibrated force transducer (Myotrace, Houston, TX) and amplified (DC Bridge Signal Conditioner, Gould Electronics, Valley View, OH). Twitch tension was digitized (NB-M1O-16, National Instruments, Austin, TX), displayed (Lab View, National Instruments), and recorded on-line (Centris 650, Apple Computer, Hayward, CA). In addition, a strip chart recorded the evoked twitch tension (TA240, Gould Electronics). End-tidal isoflurane concentration was stable for > 20 min, and the first twitch response of each train (T1) was stable for > 10 min (the control twitch tension) before the muscle relaxant was administered.

For five subjects, mivacurium was infused at 1-3 micro gram *symbol* kg sup -1 *symbol* min sup -1. When twitch tension stabilized, the mivacurium infusion rate was adjusted, based on the Hill equation, targeting 90% twitch depression. [5]When twitch tension was stable at approximately 10% of the control value for > 10 min and the mivacurium infusion rate was unchanged for > 15 min, patients received atropine (25 micro gram/kg, not to exceed 1 mg) and 50 micro gram/kg neostigmine. The mivacurium infusion was continued unchanged throughout the remainder of the study.

After induction of anesthesia, 5 ml of venous blood was obtained to document normal values for plasma cholinesterase activity and dibucaine inhibition (SmithKline Beecham Clinical Laboratories, Van Nuys, CA). Venous blood samples (two 5-ml aliquots) were obtained before neostigmine (two samples separated by 10 min) and at 2, 4, 8, and 16 min after neostigmine. One aliquot was used to determine plasma cholinesterase activity, the other to determine mivacurium concentrations. To prevent mivacurium from degrading in vitro, phospholine iodide (1.25 mg in 100 micro liter of water) was added to these samples immediately; samples were iced within 1 min, and the plasma phase was separated and frozen within 1 h. Plasma cholinesterase activity was determined photometrically using acetylthiocholine as a substrate. Mivacurium concentrations were determined by high-pressure liquid chromatography using a modification of the technique described by Brown et al. [6]and a spectrofluorometric detector (RF-511PC, Shimadzu, Tokyo). The assay is sensitive to 5 ng/ml for each of the three stereoisomers and has a coefficient of variation less or equal to 16% at that concentration; the assay is not affected by the presence of neostigmine.

The remaining five subjects underwent the same protocol except that vecuronium was the muscle relaxant and plasma cholinesterase activity and dibucaine inhibition were not determined. Vecuronium concentrations were determined by gas-liquid chromatography, [7]sensitive to 10 ng/ml with a coefficient of variation < 15% at that concentration; the assay is not affected by the presence of neostigmine.

Peak twitch tension after antagonism and time to peak antagonism were determined. Antagonism was calculated as: Equation 1where baseline is the twitch tension immediately before neostigmine administration. For example, if twitch tension recovered from 11% of control to 53% of control, antagonism was (53% - 11%)/(100% - 11%) = 47%. Antagonism of mivacurium and vecuronium and time to peak effect were compared using Student's t test for unpaired data.

To document that concentrations of each of the mivacurium stereoisomers were at steady-state before neostigmine administration, the two plasma concentrations obtained before neostigmine were compared using Student's t test for paired data. Mivacurium concentrations and plasma cholinesterase activity values after neostigmine were compared to the control values using repeated-measures analysis of variance. Plasma vecuronium concentrations were analyzed in a manner similar to that for mivacurium. Values are reported as mean plus/minus SD. Statistical significance was accepted when P < 0.05.

With mivacurium, the infusion rate to maintain 90% twitch depression was 2.2 plus/minus 0.9 micro gram *symbol* kg sup -1 *symbol* min sup -1 (range 1.1-3.4 micro gram *symbol* kg sup -1 *symbol* min sup -1). Twitch tension immediately before neostigmine administration was 10.2 plus/minus 1.5%. Baseline values for plasma cholinesterase activity and dibucaine inhibition were normal for all subjects. After neostigmine, plasma cholinesterase activity decreased markedly (P < 0.05; Figure 1).

With vecuronium, the infusion rate to maintain 90% twitch depression was 0.65 plus/minus 0.17 micro gram *symbol* kg sup -1 *symbol* min sup -1 (range 0.53-0.94 micro gram *symbol* kg sup -1 *symbol* min sup -1). Twitch tension immediately before neostigmine administration was 10.2 plus/minus 0.6%.

Antagonism of mivacurium (39 plus/minus 11%) was less than that of vecuronium (54 plus/minus 9%, P < 0.05; Figure 2). Time to peak antagonism was more rapid with mivacurium (4.2 plus/minus 0.9 min; range 2.8-4.8 min) compared to vecuronium (7.0 plus/minus 2.2 min; range 3.4-9.4 min, P < 0.05).

Concentrations of each of the mivacurium stereoisomers did not vary before neostigmine administration. Concentrations of each of these stereoisomers increased after neostigmine (P < 0.05; Figure 3and Figure 4). Because of sampling difficulties, vecuronium concentrations were determined in only four patients. These values did not vary before or after neostigmine administration: 106 plus/minus 17 ng/ml before neostigmine, 107 plus/minus 18 ng/ml at 2 min, 102 plus/minus 20 ng/ml at 4 min, 106 plus/minus 17 ng/ml at 8 min, and 102 plus/minus 11 ng/ml at 16 min.

Administration of neostigmine during constant infusion of mivacurium decreased the activity of plasma cholinesterase, thereby increasing mivacurium's concentration. In addition, neostigmine antagonized mivacurium to a lesser extent than it antagonized vecuronium, presumably because of the increase in mivacurium concentrations after neostigmine. Although these findings can be predicted based on the known effects of neostigmine on plasma cholinesterase, they have not been demonstrated previously in vivo. In addition, our findings are consistent with Cook et al.'s observation that neostigmine decreased mivacurium's elimination in vitro.* Our results regarding antagonism of mivacurium by neostigmine must be considered cautiously; in clinical practice, mivacurium administration is discontinued before neostigmine is given. As a result, one would expect mivacurium's concentration to decrease rather than to increase (presumably transiently), as we observed. That neostigmine effectively antagonizes the neuromuscular effects of mivacurium has been demonstrated previously by Caldwell et al. [8]and Naguib et al. [9]In contrast, Kao et al. [1]demonstrated that, with 70 micro gram/kg neostigmine, recovery is slower than if no antagonist is given. Presumably, differences between studies can explain these disparate findings: Kao et al. gave neostigmine when twitch tension was 1-2% of control, whereas Caldwell et al. and Naguib et al. gave neostigmine when twitch tension recovered to 10% of control.

Our protocol was similar to that used to examine the dose-response relationship for antagonism of d-tubocurarine by edrophonium, [10]neostigmine, [11]and pyridostigmine. [11]In those studies, d-tubocurarine was infused to 90% steady-state block, after which single bolus doses of the antagonist were administered, and the infusion was continued unchanged. Under these experimental conditions, the neuromuscular effect of the antagonist was evaluated in the presence of a presumed constant plasma concentration of the muscle relaxant and unchanging anesthetic potentiation and P^CO2. Although d-tubocurarine concentrations were not measured in those experiments, its pharmacokinetic characteristics and metabolic pathways suggest that its concentrations remained constant from antagonist administration to peak effect. In contrast, neostigmine's effect on the activity of plasma cholinesterase suggested (as demonstrated in the current study) that neostigmine administration would increase plasma mivacurium concentrations.

The results of the current study provide insight into an issue that we raised in a similar study of mivacurium's antagonism by edrophonium. [3]In that study, 125-2,000 micro gram/kg edrophonium increased the concentration of mivacurium's two potent stereoisomers 48% and 79% (Figure 4) despite no change in the activity of plasma cholinesterase. We were unable to reconcile the lack of change in plasma cholinesterase activity with the rapid (peaking at 1-2 min), marked increase in plasma mivacurium concentrations and speculated that edrophonium might increase mivacurium concentrations by displacement from tissue rather than by altering metabolism. In the current study, neostigmine increased the concentration of mivacurium's two potent stereoisomers 20% and 35% at 2 min (Figure 4) despite nearly complete inhibition of plasma cholinesterase activity. This increase in mivacurium concentrations was smaller than that observed after edrophonium. This suggests that the increase in mivacurium concentrations with edrophonium did not result from edrophonium altering mivacurium's metabolism but rather from displacement of mivacurium from tissue. We are unable to explain why a similar displacement was not observed when edrophonium antagonized vecuronium.

In summary, we antagonized constant infusions of muscle relaxants with neostigmine, finding less effective antagonism of mivacurium compared to that of vecuronium. After neostigmine, plasma mivacurium (but not vecuronium) concentrations increased, at least partially explaining this difference. Our results may explain Kao et al.'s observation that neostigmine delayed recovery from profound mivacurium-induced neuromuscular blockade. In addition, our results suggest that the role of neostigmine in the antagonism of mivacurium may differ from its role in the antagonism of other nondepolarizing muscle relaxants and deserves further study.

The authors thank Ronald D. Miller, M.D., for assisting with study design. The spectrofluorometric detector for the mivacurium assay was purchased with an Educational Grant from Burroughs Wellcome, Inc.

* Cook DR, Chakravorti S, Brandom BW, Stiller RL: Effects of neostigmine, edrophonium and succinylcholine on the in vitro metabolism of mivacurium: Clinical correlates (abstract). ANESTHESIOLOGY 77:A948, 1992.