Significant entry of tubocurarine into the brain of rats by adsorption to polysorbate 80-coated polybutylcyanoacrylate nanoparticles: an in situ brain perfusion study.

The possibility of using polysorbate 80-coated polybutylcyanoacrylate nanoparticles to deliver low molecular polar hydrophilic drugs to the CNS has been studied. Tubocurarine (a quaternary ammonium salt) does not penetrate the normal intact blood-brain barrier. However, the injection of this drug directly into the cerebral ventricles of the brain provokes the development of epileptiform seizures as assessed by electroencephalogram (EEG). An in situ perfused rat brain technique was used as an experimental technique together with a simultaneous recording of the EEG. Nanoparticles were prepared by butylcyanoacrylate polymerization in an acidic medium. Fifteen minutes after the introduction of tubocurarine-loaded polysorbate 80-coated nanoparticles into the perfusate, epileptiform spikes in the EEG appeared. Intraventricular injection of tubocurarine caused the appearance of the EEG seizures 5 min after administration. Neither tubocurarine solution nor tubocurarine-loaded nanoparticles without polysorbate 80 or a mixture of polysorbate 80 and tubocurarine were able to influence the EEG. Thus only the loading of tubocurarine onto the polysorbate 80-coated nanoparticles appears to enable the transport of this quaternary ammonium compound through the blood-brain barrier.

Mechanisms for the paradoxical resistance to d-tubocurarine during immobilization-induced muscle atrophy.

This study investigated whether immobilization-induced hyposensitivity to d-tubocurarine (dTC), up-regulation of acetylcholine receptors (AChRs) and changes in fiber size and motor endplate size persist indefinitely and whether they are causally related. Unilateral disuse of the tibialis muscle was produced in adult rats by pinning the knee and ankle joints at 90 degrees flexion. The contralateral unpinned and a separate group of sham-pinned legs served as controls. After 7, 14 or 28 days of disuse, the in vivo dose of dTC that produced 50% depression of nerve-evoked twitch (ED50) in the tibialis muscle increased 3.0-, 3. 2- and 2.1-fold (P < .05), and membrane AChRs increased 6.0- (P < . 05), 6.3- (P > .05) and 1.2-fold (P > .395) relative to control, respectively. Disuse caused muscle fiber atrophy (P < .01) but did not affect endplate size. Hence, the ratio of endplate size to fiber size increased. There was a transient increase in gene expression of all (including de novo expression of the gamma) subunits of the AChR, peaking at day 7 and returning to normal by day 28 of immobilization. The ED50 of dTC correlated directly with AChRs (R2 = 0.51; P < .0001) or the ratio of endplate size to fiber size (R2 = 0. 30; P < .001), and inversely with fiber size (R2 = 0.43, P < .0001). It is proposed that acting together, but not singly, the changes in AChRs, fiber size and relative endplate size contribute to the magnitude and time course of the resistance to dTC produced by chronic disuse.

Metocurine pharmacokinetics and pharmacodynamics in goats.

Non-depolarizing muscle relaxants can facilitate surgery and anaesthesia in numerous species. and volatile inhalational anaesthetics such as isoflurane potentiate their action. We studied the effect of isoflurane on the pharmacodynamics and pharmacokinetics of metocurine in six goats. Each was studied twice: once during barbiturate-opiate anaesthesia and once during isoflurane anaesthesia. The evoked response to sciatic nerve stimulation was measured using a force transducer attached to the hoof. Metocurine was infused until approximately 80-90% blockade. Plasma metocurine concentration was determined by high-performance liquid chromatography. Isoflurane increased the potency of metocurine significantly; IC50 (the concentration in the effect compartment at 50% paralysis) was 70 +/- 15 ng/mL during isoflurane anaesthesia and 129 +/- 42 ng/mL during barbiturate-opiate anaesthesia (P < 0.03). Volume of distribution (63 +/- 18 mL/kg), clearance (1.6 +/- 0.4 mL/min.kg) and elimination half-life (99 +/- 9 min) during barbiturate-opiate anaesthesia were not significantly different during isoflurane anaesthesia: 64 +/- 25 mL/kg, 1.5 +/- 0.7 mL/kg.min, 116 +/- 16 min respectively. We conclude that, relative to barbiturate-opiate anaesthesia, isoflurane potentiates metocurine in goats.

d-Tubocurarine accentuates the burn-induced upregulation of nicotinic acetylcholine receptors at the muscle membrane.

BACKGROUND: Increases in acetylcholine receptors (AChRs) at the muscle membrane, induced by burn injury, have been associated with a hyperkalemic response to succinylcholine and resistance to d-tubocurarine-like drugs. Muscle relaxants often are administered to burn-injured patients in the intensive care unit to facilitate mechanical ventilation. This study in rats tested whether continuous administration of d-tubocurarine in subparalytic doses exaggerates the upregulation of AChRs induced by burn trauma. Subparalytic doses were used to avoid the confounding effects of immobilization. METHODS: Three days after an approximate 50% body surface area burn or sham injury, the animals received an infusion of 3.03 +/- 0.05 micrograms/h of d-tubocurarine or equal volume of saline directly to the left gastrocnemius muscle via catheter connected to a subcutaneously implanted osmotic pump. After 7 days of d-tubocurarine or saline infusion, the AChRs were quantitated using 125I-alpha-bungarotoxin. The AChRs on the d-tubocurarine or saline-infused left gastrocnemius were compared to the contralateral gastrocnemius in the same group. The right or left gastrocnemius AChRs were compared to the ipsilateral muscles between groups. These intra- and intergroup comparisons allowed the delineation of the effects of catheter irritation, burns, or d-tubocurarine on AChRs. RESULTS: Daily examination of the withdrawal response to toe-pinch revealed no evidence of paralysis. Weight loss in the burn-injury animals receiving d-tubocurarine or saline was similar, confirming that the infusion of d-tubocurarine did not impair the mobility of the animals to move and feed. The plasma d-tubocurarine concentration after 7 days of infusion was 26.0 +/- 12 ng/ml (mean +/- SE). Regardless of burn or sham injury or of d-tubocurarine or saline infusion, the concentration of AChRs on the left was consistently greater than in the contralateral right gastrocnemius muscles within the same group, indicating that manipulation of the area alone can result in upregulation of AChRs. The AChRs in the right gastrocnemius of burn-injured animals were greater than those in the same muscle of sham-injured animals, regardless of saline (7.24 +/- 0.9 vs. 5.7 +/- 0.5 fmoles/mg protein, P = 0.06) or d-tubocurarine (7.3 +/- 0.4 vs. 5.7 +/- 0.5, P < 0.05) infusion to the burn-injury groups. AChRs in the left gastrocnemius of burn-injury animals receiving d-tubocurarine were significantly greater than those in burn- or sham-injury animals receiving saline (13.9 +/- 1.1 vs. 9.8 +/- 1.2 and 7.1 +/- 0.5 fmoles/mg protein, respectively, P < 0.05). CONCLUSIONS: Burn-induced upregulation of AChRs is accentuated by infusion of subparalytic doses of d-tubocurarine. Concomitant administration of d-tubocurarine to burn-injured patients may result in further exaggeration of the aberrant responses to neuromuscular relaxants.

A method for preferential delivery of volatile anesthetics to the in situ goat brain.

BACKGROUND: As part of studies aimed at better defining the effects of anesthetics at different anatomic sites, we have developed a model of preferentially delivering inhaled anesthetics to the in situ goat brain, using a bubble oxygenator and roller pump. We tested the hypotheses that (1) this model excludes the cerebral circulation from the body; (2) the concentration of halothane in the oxygenator exhaust correlates with the concentration of halothane in the oxygenator arterial blood. METHODS: After ligation of the occipital arteries in six halothane-anesthetized goats, we used a bubble oxygenator to perfuse the brain preferentially (exclusive of the body) via a carotid artery, draining cranial venous blood back into the oxygenator via the isolated jugular veins. (In goats, the vertebral arteries do not directly contribute to the cerebral circulation, and internal jugular veins and extracranial internal carotid arteries are absent). The extent of isolation was determined with radioactive microspheres injected into the left atrium during the following periods: (1) baseline; (2) during bypass when the blood pressure in the head equalled that in the body; (3) during bypass when the blood pressure in the body exceeded that in the head by approximately 30-35 mmHg; (4) when the bypass roller pump was stopped. We also measured the concentration of halothane in the arterial blood of the bypass unit. In three animals, systemic metocurine was administered during bypass to detect the presence of venous contamination. RESULTS: Baseline cerebral blood flow was 74 +/- 32 ml.100 g-1.min-1 (mean +/- SD). During bypass, cerebral blood flow originating from the systemic circulation was less than 1 ml.100 g-1.min-1, and isolation extended to the caudal medulla during periods 3 and 4, and to the first 1-cm segment of the spinal cord during period 2. The concentration of halothane in the oxygenator exhaust correlated reasonably well with the arterial halothane concentration (r = 0.82, P < 0.001). Systemic arterial metocurine concentrations peaked at 1 min (27 +/- 3.7 micrograms/ml) and decreased to 10.6 +/- 2.3 micrograms/ml at 10 min; head venous metocurine plasma concentrations gradually increased to 3.1 +/- 0.4 micrograms/ml at 10 min. CONCLUSIONS: This technique permits selective perfusion and delivery of inhaled anesthetics to the in situ goat brain, but is not adequate for selective delivery of fixed intravenous anesthetics.

Decreased sensitivity to metocurine during long-term phenytoin therapy may be attributable to protein binding and acetylcholine receptor changes.

Long-term phenytoin therapy induces resistance to the neuromuscular blocking effects of metocurine. The hypothesis that this is attributable to increased plasma protein binding of the drug (decreased free fraction) related to increased concentrations of alpha 1-acid glycoprotein (AAG) or attributable to the proliferation of acetylcholine receptors (AChR) at the muscle membrane was tested in the rat. After 14 days of phenytoin 40 mg.kg-1, administered intraperitoneally twice daily (n = 12), the neuromuscular pharmacodynamics were evaluated and compared with those of time-matched controls (n = 10). Protein binding was measured by equilibrium dialysis, AAG concentrations by radial immunodiffusion assay, and AChR by 125I-alpha-bungarotoxin binding. The effective dose for 50% inhibition of baseline twitch height (ED50) was significantly greater in the phenytoin group than in the control group (15.03 +/- 1.65 micrograms.kg-1 vs. 9.98 +/- 0.69 micrograms.kg-1, respectively). The concentrations of AAG increased gradually from 133.8 +/- 7.8 micrograms.ml-1 at day 0, to 343.1 +/- 58.0 micrograms.ml-1 at day 7, to 1,729.5 +/- 422.3 micrograms.ml-1 at day 14 in the phenytoin group. The induction of AAG concentrations in plasma was dependent on plasma phenytoin concentrations and was most prominent after 14 days of phenytoin (r = 0.77; P less than 0.01; n = 22). The free fraction of metocurine was significantly decreased in the phenytoin group compared to the control group (67.2 +/- 0.18% vs. 74.5 +/- 2.5%). There was a significant negative correlation between increased AAG concentrations and decreased free fraction (r = 0.65).(ABSTRACT TRUNCATED AT 250 WORDS)

Residual curarization in the neonate after caesarean section.

The transplacental transfer and the neonatal effects of atracurium 0.3 mg.kg-1 (ED95) were compared with those of d-tubocurarine at the usual clinical dose of 0.3 mg.kg-1 (ED90) in 46 patients undergoing elective Caesarean section. The atracurium group (25 patients) was similar to the d-tubocurarine group (21 patients) as far as age, parity and time intervals between precurarization, induction, skin incision, muscle relaxant administration, hysterotomy and birth. The transplacental transfer of atracurium was lower than that of d-tubocurarine, with a feto-maternal ratio of 9 +/- 3% for atracurium and 12 +/- 5% for d-tubocurarine (P less than 0.05). The transplacental transfer of laudanosine was low at 14 +/- 5%, with blood levels of 0.101 +/- 0.032 microM.L-1 in the umbilical vein. Newborns in the two groups were comparable in terms of Apgar scores at one, five and ten minutes, as well as for NACS scores (neurological and adaptive capacity scoring test) at two and 24 hours after birth. However, at 15 min after birth, only 55% of newborns in whom the mothers received atracurium had a normal NACS score (greater than or equal to 35/40) compared with 83% of newborns in whom the mothers received d-tubocurarine (P less than 0.05). Further analysis of the five variables related to active muscle tone revealed that the modal score for active extension of the neck of newborns from the atracurium group was lower than for newborns from the d-tubocurarine group (P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Determination of vecuronium in blood by HPLC with UV and electrochemical detection: a pilot study in man.

Vecuronium bromide is a non-depolarizing neuromuscular blocking agent with a rather low therapeutic level. Rapid, sensitive, and selective determination of vecuronium bromide in human blood or plasma was essential for pharmacokinetic study. We developed such a method using HPLC with electrochemical detection. Samples were first acidified, followed by a one-step liquid-liquid extraction. Tubocurarine, which has a structure similar to that of vecuronium, was used as the internal standard. The electrochemical detector, Ag/AgCl electrodes, was operated by setting the working electrodes, W1 and W2, at +0.65 V and +1.05 V, respectively. When vecuronium blood concentration was plotted versus peak area ratios (PAR) of vecuronium over tubocurarine, a linear relationship was observed over the range of 25 ng/mL to 500 ng/mL with a correlation coefficient greater than 0.997. Clinically possible sources of interference, such as atropine, apresoline, droperidol, fentanyl, labetalol, thiopentone, atracurium, and valium, were examined and none showed interference in the assay of vecuronium and tubocurarine. This method has been successfully applied in a preliminary study of the pharmacokinetics of vecuronium in a patient undergoing surgery. The low detection limit of this method in the patient was 3 ng/mL.

Exercise produces sensitivity to metocurine.

Chronic muscle disuse decreases the sensitivity of skeletal muscle to nondepolarizing relaxants, such as metocurine (MTC). In this study, the authors determined whether chronic conditioning would produce the opposite effect and increase the sensitivity of skeletal muscle to MTC. Five dogs were exercised by daily running over a period of 5 weeks. At the conclusion of this training period, a pharmacokinetic and pharmacodynamic study of the MTC dose-response relationship was performed. The same analysis was performed on four dogs housed in the same kennel who did not undergo conditioning. Neuromuscular blockade was measured and recorded bilaterally in both gastrocnemius muscles while the animal was anesthetized with nitrous oxide and pentobarbital, 30 ml.kg-1. Plasma concentrations of MTC were measured by radioimmunoassay. The MTC concentration estimated in the effect compartment which produced 50% paralysis was 0.114 +/- 0.008 micrograms.ml-1 (mean +/- SD) in exercised dogs and 0.189 +/- 0.038 micrograms.ml-1 in nonexercised dogs, which was significant at P less than 0.005). The MTC concentration versus response curves were parallel. This supports the authors' hypothesis that exercise increases sensitivity to the nondepolarizing muscle relaxant metocurine.

Resistance to metocurine-induced neuromuscular blockade in patients receiving phenytoin.

Recent reports have described resistance to pancuronium-induced neuromuscular blockade in patients chronically receiving anticonvulsants. This study examines the pharmacokinetics and pharmacodynamics of metocurine (MTC) in 12 patients undergoing craniotomy--six on chronic phenytoin therapy and six comparable controls. Each patient received MTC 0.2 mg/kg during the induction of general anesthesia. Quantification of plasma MTC concentration was performed by radioimmunoassay, while the response to MTC was evaluated by evoked compound electromyography (ECEMG). Patients in the phenytoin group were resistant to this dosage of MTC, as demonstrated by their response (83 +/- 16% compared with 98 +/- 2% depression of ECEMG in control patients, P less than 0.05) and by recovery index, defined as the time required for recovery from 25 to 75% of the control ECEMG (53 +/- 22 min compared with 125 +/- 54 min in control patients, P less than 0.01). Similarly, the total duration of neuromuscular blockade, measured to recovery to 90% of control ECEMG, was significantly shorter in the phenytoin group (122 +/- 25 min compared with 269 +/- 64 min in the control group, P less than 0.01). Plasma concentration-time curves were fit to biexponential equations for both groups. These were used to generate two-compartment models. Neither the model parameters nor the plasma concentrations of MTC at any time in the study were significantly different for the two groups. The pharmacodynamic analysis, however, showed that patients on phenytoin require a higher plasma concentration of MTC (0.415 +/- 0.095 microgram/ml compared with 0.249 +/- 0.066 microgram/ml in control patients at 50% ECEMG, P less than 0.01) to effect a given level of neuromuscular blockade.(ABSTRACT TRUNCATED AT 250 WORDS)

Decreased sensitivity to metocurine in patients with upper motoneuron disease.

Responses to the nondepolarizing muscle relaxant, metocurine, were studied in eight hemiplegic and eight unmatched patients with normal motor strength during the general anesthetic given for various neurosurgical operations. Metocurine, 0.3 mg/kg, was administered intravenously, and indirectly evoked thumb twitch tensions were measured on both sides in the hemiplegic patients, and on one side in the normal patients. Arterial blood samples were obtained as twitch tension was recovering, and serum metocurine concentrations were determined using a specific radioimmunoassay. Percentage of paralysis was plotted as a function of log [metocurine] and the data were compared by analysis of covariance. For the normal motor strength patients, r = 0.84; for the unaffected arm of the hemiplegic patients, r = 0.69; and for the affected arm of the hemiplegic patients, r = 0.86, all significant at P less than 0.001. The mean plasma metocurine concentrations at 20, 25, 50, 75, and 80% paralysis were significantly different for all groups (P less than 0.001). The regression lines, in turn, did not overlap and were significantly different, each from the other (P less than 0.005). We were, however, unable to detect any significant deviation from parallelism among the three regression lines. We also measured time to 50% return of single twitch height for each data group as follows (mean +/- SEM: for NMS patients, 242 +/- 73 min; for the unaffected arm of hemiplegic patients, 116 +/- 60 min; and for the affected arm of hemiplegic patients, 59 +/- 36 min. By ANOVA and the Bonferroni test, each value was different from the other at P less than or equal to 0.01.(ABSTRACT TRUNCATED AT 250 WORDS)

The pharmacokinetics of d-tubocurarine with surgery involving salvaged autologous blood.

The disposition of d-tubocurarine (dTc) was assessed when a bolus and infusion dosage regimen was used to obtain relaxation during major orthopedic surgery on the spine. Renal clearance of dTc was 0.63 +/- 0.23 ml X min-1 X kg-1 and was correlated with creatinine clearance. Total plasma clearance of 1.21 +/- 0.40 ml X min X -1 X kg-1 was lower than that found in many previous studies, and the predetermined continuous dTc infusion produced an apparent plateau in plasma concentrations of 1.8 +/- 0.3 micrograms X ml-1. Despite the operative blood loss, these concentrations were greater than anticipated and were associated with a more intense neuromuscular blockade than the infusion was designed to produce. Autologous blood transfusion was used to reduce the reliance on homologous donor blood, and the erythrocytes from the 2.2 +/- 1.2 1 of blood loss during the procedure were reinfused after intraoperative salvage, washing, and centrifugation. With 80 +/- 23 mg dTc administered, 1.4 +/- 0.8% was recovered from the fluid discarded after centrifugation. These results indicate that even massive intraoperative blood loss will not entail a significant reduction in the amount of dTc present in the body.

Canine gastrocnemius disuse atrophy: resistance to paralysis by dimethyl tubocurarine.

Ten dogs developed unilateral gastrocnemius disuse atrophy after unilateral hindlimb immobilization in a cast for 25 days. Dose-response curves to dimethyl tubocurarine (MTC) were determined during anesthesia with pentobarbital sodium-N2O. Bolus and continuous infusion increments of MTC every 30 min provided steady-state blood levels at each stage of paralysis. Both gastrocnemius tendons were sectioned and attached to transducers. Both sciatic nerves were stimulated every 30 min: 2 Hz for 2 s, a 15-s pause, 50 Hz for 2 s. Dose-response curves, computer calculated by nonlinear regression using a sigmoid maximal effect model of the Hill equation, were parallel for the data relating blocking of tetanus to dose of MTC. The 50% paralyzing dose (tetanus) for control vs. casted gastrocnemius muscle was 64 vs. 813 mg/kg; corresponding plasma concentrations were 0.12 vs. 2.0 micrograms/ml. Thus in vivo simultaneous tension measurements of both gastrocnemius muscles, one casted and one uncasted, demonstrated resistance to paralysis by MTC in muscle with disuse atrophy.

Dose-response of tubocurarine in patients with and without renal failure.

Different initial doses of tubocurarine (0.1, 0.2, 0.3 and 0.4 mg kg-1) were given to 20 patients with and 20 patients without renal failure (RF) during standard anaesthesia. Neuromuscular blockade was registered with a device based on electromyography. A fluorescent technique was used to measure plasma tubocurarine concentrations, for which blood samples were drawn during induction and at reversal of neuromuscular block. The dose-response curves for the induction period were slightly steeper in patients with RF. A linear correlation between tubocurarine concentration and twitch response was found in both groups, although the curve had shifted to the left in RF patients. The total amount of tubocurarine used for surgical relaxation was lower in the RF than in the non-RF patients.

Determinants of d-tubocurarine plasma protein binding in health and disease.

d-Tubocurarine protein binding was determined in plasma from normal surgical patients; patients with renal dysfunction; patients with alcoholic cirrhosis; and patients scheduled for coronary artery surgery and was compared with binding determined in solutions of isolated human serum albumin (HSA). The free fraction of d-tubocurarine was independent of plasma drug concentration throughout the range 0.1-3.0 micrograms/ml. The free fraction of d-tubocurarine in renal disease (0.44 +/- 0.05), liver disease (0.53 +/- 0.05), and cardiac disease (0.49 +/- 0.05) was not significantly different from that in normal patients (0.50 +/- 0.04). Furthermore, the free fraction of d-tubocurarine was unrelated to plasma concentrations of total protein, total globulin, albumin, total lipids, or alpha 1-acid glycoprotein in the four patient groups. The free fraction of d-tubocurarine (0.55 +/- 0.02, n = 7) in solutions of HSA (0.58 mM) was similar to that observed in plasma. Albumin, therefore, appears to be the major plasma binding protein for d-tubocurarine.

Metocurine requirements and plasma concentrations in burned paediatric patients.

A cumulative dose-response study for metocurine was performed on 11 burned and 11 non-burned children during thiopentone, nitrous oxide in oxygen and narcotic anaesthesia. Burned children required a significantly (P less than 0.001) greater dose (three-fold) of metocurine to achieve a given degree of neuromuscular blockade compared with non-burned children. For the same degree of twitch depression, burned children required twice the plasma concentration of metocurine. Despite the larger dose, the recovery time for the twitch was not significantly different in the two groups. It is speculated that there may be fundamental changes at the neuromuscular junction of burned children to account for their resistance to the effect of non-depolarizing neuromuscular blockers.