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Summary Adrenomedullary chromaffin cells are catecholamine (CA)-producing cells originating from trunk neural crest (NC) via sympathoadrenal progenitors (SAPs). We generated NC and SAPs from human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) in vitro via BMP2/FGF2 exposure, ascertained by qPCR and immunoexpression of SOX10, ASCL1, TFAP2α, and PHOX2B, and by fluorescence-activated cell sorting selection for p75NTR and GD2, and confirmed their trunk-like HOX gene expression. We showed that continuing BMP4 and curtailing FGF2 in vitro, augmented with corticosteroid mimetic, induced these cells to upregulate the chromaffin cell-specific marker PNMT and other CA synthesis and storage markers, and we demonstrated noradrenaline and adrenaline by Faglu and high-performance liquid chromatography. We showed these human cells' SAP-like property of migration and differentiation into cells expressing chromaffin cell markers by implanting them into avian embryos in vivo and in chorio-allantoic membrane grafts. These cells have the potential for investigating differentiation of human chromaffin cells and for modeling diseases involving this cell type., Highlights • Human ESCs and iPSCs differentiated to chromaffin cells via sympathoadrenal stage • Chromaffin-like cells are identified by PNMT expression • Adrenaline and noradrenaline production is revealed by FAGLU and by HPLC • Selection of GD2+ sympathoadrenal cells improves chromaffin cell differentiation, In this article, Newgreen and colleagues show that human pluripotent cells can be induced to form chromaffin-like cells, which produce adrenaline and noradrenaline in vitro. The method uses the dual-inhibitor method to induce neural crest progenitor and sympathoadrenal-like cells then extended BMP4 and corticosteroid exposure for the chromaffin-like stage, with curtailed FGF2 exposure to limit sympathetic neuron induction.

Background Central core disease and malignant hyperthermia are human disorders of skeletal muscle resulting from aberrant Ca2+ handling. Most malignant hyperthermia and central core disease cases are associated with amino acid changes in the type 1 ryanodine receptor (RyR1), the skeletal muscle Ca2+-release channel. Malignant hyperthermia exhibits a gain-of-function phenotype, and central core disease results from loss of channel function. For a variant to be classified as pathogenic, functional studies must demonstrate a correlation with the pathophysiology of malignant hyperthermia or central core disease. Objective We assessed the pathogenicity of four C-terminal variants of the ryanodine receptor using functional analysis. The variants were identified in families affected by either malignant hyperthermia or central core disease. Methods Four variants were introduced separately into human cDNA encoding the skeletal muscle ryanodine receptor. Following transient expression in HEK-293T cells, functional studies were carried out using calcium release assays in response to an agonist. Two previously characterized variants and wild-type skeletal muscle ryanodine receptor were used as controls. Results The p.Met4640Ile variant associated with central core disease showed no difference in calcium release compared to wild-type. The p.Val4849Ile variant associated with malignant hyperthermia was more sensitive to agonist than wild-type but did not reach statistical significance and two variants (p.Phe4857Ser and p.Asp4918Asn) associated with central core disease were completely inactive. Conclusions The p.Val4849Ile variant should be considered a risk factor for malignant hyperthermia, while the p.Phe4857Ser and p.Asp4918Asn variants should be classified as pathogenic for central core disease.

Summary Background Infant spinal anesthesia with levobupivacaine has been promoted as a technique to reduce both the risk of postoperative apnea and exposure to volatile anesthesia. There is, however, no pharmacokinetic data to support the currently recommended doses. Aims Our aim was to determine whether infant levobupivacaine spinal anesthesia is associated with plasma concentrations consistent with a low risk of local anesthetic systemic toxicity. Methods This was an open-label pharmacokinetic safety and tolerability study of levobupivacaine spinal anesthesia in infants

Significance In human muscle with ryanodine receptor (RyR) variants, there is expected to be a change in the basal RyRs Ca 2+ leak that may alter local Ca 2+ dynamics. It is not currently possible to assess these basal Ca 2+ dynamics in human muscle. By trapping a Ca 2+ -sensitive fluorescent dye in the transverse tubular (t)-system of freshly biopsied human skeletal muscle fibers we could detect tiny local RyR-dependent Ca 2+ movements. We show that malignant hyperthermia (MH) causative RyR variant human muscle displays a chronically different RyR Ca 2+ leak which reconfigures the Ca 2+ -handling properties by the t-system. This approach can be used to assess human muscle for function and pathogenicity of other RyR variants of uncertain significance, MH susceptibility, and assessment of drugs targeted to the Ca 2+ -handling proteins.

Glioma cells release glutamate through expression of system xc-, which exchanges intracellular glutamate for extracellular cysteine. Lack of the excitatory amino acid transporter 2 (EAAT2) expression maintains high extracellular glutamate levels in the glioma microenvironment, causing excitotoxicity to surrounding parenchyma. Not only does this contribute to the survival and proliferation of glioma cells, but is involved in the pathophysiology of tumour-associated epilepsy (TAE). We investigated the role of the peroxisome proliferator activated receptor gamma (PPARγ) agonist pioglitazone in modulating EAAT2 expression in glioma cells. We found that EAAT2 expression was increased in a dose dependent manner in both U87MG and U251MG glioma cells. Extracellular glutamate levels were reduced with the addition of pioglitazone, where statistical significance was reached in both U87MG and U251MG cells at a concentration of ≥ 30 μM pioglitazone (p < 0.05). The PPARγ antagonist GW9662 inhibited the effect of pioglitazone on extracellular glutamate levels, indicating PPARγ dependence. In addition, pioglitazone significantly reduced cell viability of U87MG and U251MG cells at ≥ 30 μM and 100 μM (p < 0.05) respectively. GW9662 also significantly reduced viability of U87MG and U251MG cells with 10 μM and 30 μM (p < 0.05) respectively. The effect on viability was partially dependent on PPARγ activation in U87MG cells but not U251MG cells, whereby PPARγ blockade with GW9662 had a synergistic effect. We conclude that PPARγ agonists may be therapeutically beneficial in the treatment of gliomas and furthermore suggest a novel role for these agents in the treatment of tumour associated seizures through the reduction in extracellular glutamate.

Defects in the genes coding for the skeletal muscle ryanodine receptor (RYR1) and alpha 1 subunit of the dihydropyridine receptor (CACNA1S) have been identified as causative for malignant hyperthermia (MH). Sixty-two MH susceptible individuals presenting to the same diagnostic centre had copy deoxyribonucleic acid, derived from muscle ribonucleic acid, sequenced to identify variants with the potential to be responsible for the MH phenotype in both RYR1 and CACNA1S. These genetic findings were combined with clinical episode details and in vitro contracture test results to improve our understanding of the Australian MH cohort. Twelve novel variants were identified in RYR1 and six in CACNA1S. Known RYR1 causative mutations were identified in six persons and novel variants in RYR1 and CACNA1S in a further 17 persons. Trends indicated higher mutation identification in those with more definitive clinical episodes and stronger in vitro contracture test responses.

Neurons release neurotransmitters such as glutamate to communicate with each other and to coordinate brain functioning. As increased glutamate release is indicative of neuronal maturation and activity, a system that can measure glutamate levels over time within the same tissue and/or culture system is highly advantageous for neurodevelopmental investigation. To address such challenges, we develop for the first time a convenient method to realize functionalized borosilicate glass capillaries with nanostructured texture as an electrochemical biosensor to detect glutamate release from cerebral organoids generated from human embryonic stem cells (hESC) that mimic various brain regions. The biosensor shows a clear catalytic activity toward the oxidation of glutamate with a sensitivity of 93 ± 9.5 nA·µM−1·cm−2. It was found that the enzyme-modified microelectrodes can detect glutamate in a wide linear range from 5 µM to 0.5 mM with a limit of detection (LOD) down to 5.6 ± 0.2 µM. Measurements were performed within the organoids at different time points and consistent results were obtained. This data demonstrates the reliability of the biosensor as well as its usefulness in measuring glutamate levels across time within the same culture system.

Tramadol is used in children aged

The genetic basis of malignant hyperthermia (MH) is not fully characterised and likely involves more than just the currently classified mutations in the gene encoding the skeletal muscle ryanodine receptor ( RYR1) and the gene encoding the α1 subunit of the dihydropyridine receptor ( CACNA1S). In this paper we sequence other genes involved in calcium trafficking within skeletal muscle in patients with positive in vitro contracture tests, searching for alternative genes associated with MH. We identified four rare variants in four different genes ( CACNB1, CASQ1, SERCA1 and CASQ2) encoding proteins involved in calcium handling in skeletal muscle in a cohort of 30 Australian MH susceptible probands in whom prior complete sequencing of RYR1 and CACNA1S had yielded no rare variants. These four variants have very low minor allele frequencies and while it is tempting to speculate that they have a role in MH, they remain at present variants of unknown significance. Nevertheless they provide the basis for a new set of functional studies, which may indeed identify novel players in MH.

We compared plasma and cerebrospinal fluid (CSF) pharmacokinetics of paracetamol after intravenous (IV) and oral administration to determine dosing regimens that optimize CSF concentrations.Twenty-one adult patients were assigned randomly to 1 g IV, 1 g oral or 1.5 g oral paracetamol. An IV cannula and lumbar intrathecal catheter were used to sample venous blood and CSF, respectively, over 6 hours. The plasma and CSF maximum concentrations (Cmax), times to maximum concentrations (Tmax), and area under the plasma and CSF concentration-time curves (AUCs) were calculated using noncompartmental techniques. Significance was defined by P.0167 (Bonferroni correction for 3 comparisons for each parameter). Probability (XY) (p″) with Bonferroni corrected 95% confidence intervals (CIs) were calculated (CIs including 0.5 meet the null hypothesis). Results are presented as median (range) or p″ (CI). P values are listed as 1 g IV vs 1 g orally, 1 g IV vs 1.5 g orally and 1 g orally vs 1.5 g orally, respectively.Wide variation in measured paracetamol concentrations was observed, especially in the oral groups. The median plasma Cmax in the 1 g IV group was significantly greater than the oral groups. In contrast, the median CSF Cmax was not different between groups. The median plasma Tmax in the 1 g IV group was 105 and 75 minutes earlier than in the 1 and 1.5 g oral groups. The median CSF Tmax was not significantly different between groups. The median plasma AUC (total) was not significantly different between groups; however, in the first hour, the median plasma AUC was significantly greater in the IV group than in the oral groups. In the second hour, there was no difference between groups. The median CSF AUC (total) did not significantly differ between groups; however, in the first hour, the median CSF AUC was significantly greater in the IV compared with the orally groups. In the second hour, there was no difference between groups. Our analysis indicated that the median Cmax, Tmax, and AUC values lacked precision because of small sample sizes.Peak plasma concentrations were greater and reached earlier after IV than oral dosing. Evidence for differences in CSF Cmax and Tmax was lacking because of the small size of this study.

Nineteen adult patients of either gender received intravenous infusions of propofol, scaled to estimated lean body mass (LBM), for 150 minutes as part of a balanced anaesthetic. Arterial blood was assayed for whole blood propofol. The first subject received propofol at a fixed rate of 0.058 mg.min-1.kgLBM-1. Subsequent groups received variable rate infusions based on the ratio of the infusion rate to the propofol concentration at each sampling point in the previous group, multiplied by the target concentration. After groups of one, two, five and 11 subjects, the median weighted residual was 0.040 and median absolute weighted residual was 0.153. Population pharmacokinetic analysis of the final group of six females and five males, aged 29 to 70 years and of 16.5 to 44.2% body fat, resulted in a two compartment pharmacokinetic model with coefficients and standard errors of V1=0.102 (0.0155) l/kgLBM, V2=0.257 (0.079) l/kgLBM, k10=0.423 (0.069) /min, k12=0.222 (0.051) /min, k21=0.084 (0.02) /min and clearance=0.0418 (0.0023) l.min-1.kgLBM-1. The only significant covariate was LBM. Within infusion data improved prediction when compared with data derived in previous studies from random observations.

Advances in analysis of the RYR1 gene (which encodes the skeletal muscle ryanodine receptor) show that genetic examination is a useful adjunct to the in vitro contracture test in the diagnosis of malignant hyperthermia, as defects in RYR1 have been shown to be responsible for malignant hyperthermia susceptibility. DNA from 34 malignant hyperthermia susceptible individuals and four malignant hyperthermia equivocal subjects was examined using direct sequencing of ‘hotspots’ in the RYR1 gene to identify mutations associated with malignant hyperthermia. Seven different causative mutations (as defined by the European Malignant Hyperthermia Group) in nine malignant hyperthermia susceptible individuals were identified. In another six malignant hyperthermia susceptible individuals, five different published but as yet functionally uncharacterised mutations were identified. A further three as yet unpublished and functionally uncharacterised (novel) mutations were identified in three malignant hyperthermia susceptible samples. If the novel and previously published mutations prove to be functionally associated with calcium homeostasis, then this method of analysis achieved a mutation detection rate of 47%. Based on the number of relatives presenting to our unit in the study period, the muscle biopsy rate would have decreased by 25%. That we only identified a genetic defect in RYR1 in 47% of in vitro contracture test positive individuals suggests that there are other areas in RYR1 where pathogenic mutations may occur and that RYR1 may not be the sole gene associated with malignant hyperthermia. It may also reflect a less than 100% specificity of the in vitro contracture test.

Background We tested the hypothesis that there is greater suppression of autonomic reflexes during general anaesthesia when fentanyl is administered epidurally than when it is given intravenously. Methods Ten volunteers were anaesthetized with desflurane. Noxious stimuli of variable intensity were then delivered by tetanic electrical stimuli. Heart rate, arterial pressure, and pupillary dilation in response to these stimuli defined nociception. Seven of these volunteers participated twice using a crossover design: they received i.v. fentanyl on one study day and epidurally on the other. Autonomic responses to alternative tetanic stimuli at L4 and C5 dermatomes were measured every 5 min for 3 h after fentanyl administration. Results After a brief redistribution period, plasma fentanyl concentrations were virtually identical on both days. After stimulation of the L4 dermatome only, block of pupillary reflex dilation was greater by 47 (22)% after epidural fentanyl compared with i.v. fentanyl. Time to maximal depression of reflex dilation after L4 stimulation was 41 (13) min. Arterial pressure and heart rate decreased after fentanyl by either route but there were no differences observed between L4 and C5 stimulations. Conclusion We conclude that during general anaesthesia, epidural fentanyl enhances antinociception by a spinal mechanism which can be detected by pupillary dilation but not by changes in arterial pressure or heart rate.

Reactive oxygen species (ROS) have been linked with both depressed Na(+),K(+)-pump activity and skeletal muscle fatigue. This study investigated N-acetylcysteine (NAC) effects on muscle Na(+),K(+)-pump activity and potassium (K(+)) regulation during prolonged, submaximal endurance exercise. Eight well-trained subjects participated in a double-blind, randomised, crossover design, receiving either NAC or saline (CON) intravenous infusion at 125 mg kg(-1) h(-1) for 15 min, then 25 mg kg(-1) h(-1) for 20 min prior to and throughout exercise. Subjects cycled for 45 min at 71% , then continued at 92% until fatigue. Vastus lateralis muscle biopsies were taken before exercise, at 45 min and fatigue and analysed for maximal in vitro Na(+),K(+)-pump activity (K(+)-stimulated 3-O-methyfluorescein phosphatase; 3-O-MFPase). Arterialized venous blood was sampled throughout exercise and analysed for plasma K(+) and other electrolytes. Time to fatigue at 92% was reproducible in preliminary trials (c.v. 5.6 +/- 0.6%) and was prolonged with NAC by 23.8 +/- 8.3% (NAC 6.3 +/- 0.5 versus CON 5.2 +/- 0.6 min, P < 0.05). Maximal 3-O-MFPase activity decreased from rest by 21.6 +/- 2.8% at 45 min and by 23.9 +/- 2.3% at fatigue (P < 0.05). NAC attenuated the percentage decline in maximal 3-O-MFPase activity (%Deltaactivity) at 45 min (P < 0.05) but not at fatigue. When expressed relative to work done, the %Deltaactivity-to-work ratio was attenuated by NAC at 45 min and fatigue (P < 0.005). The rise in plasma [K(+)] during exercise and the Delta[K(+)]-to-work ratio at fatigue were attenuated by NAC (P < 0.05). These results confirm that the antioxidant NAC attenuates muscle fatigue, in part via improved K(+) regulation, and point to a role for ROS in muscle fatigue.

Background There are few data describing levobupivacaine pharmacokinetics in infants ( Methods An open-label study was undertaken to examine the pharmacokinetics of levobupivacaine 2.5 mg ml −1 , 2 mg kg −1 in children aged less than 3 months after single-shot caudal epidural administration. Plasma concentrations were determined at intervals from 0.5 to 4 h after injection. A population pharmacokinetic analysis of levobupivacaine time–concentration profiles (84 observations) from 22 infants with mean postnatal age (PNA) 2.0 (range 0.6–2.9) months was undertaken using non-linear mixed effects models (NONMEM). Time–concentration profiles were analysed using a one-compartment model with first-order input and first-order elimination. Estimates were standardized to a 70 kg adult using allometric size models. Results Population parameter estimates (between-subject variability) for total levobupivacaine were clearance (CLt) 12.8 [coefficient of variation (CV) 50.6%] litre h −1 70 kg −1 , volume of distribution (Vt) 202 (CV 31.6%) litre 70 kg −1 , absorption half-life (Tabs) 0.323 (CV 18.6%) h 70 kg −1 . Estimates for the unbound drug were clearance (CLfree) 104 (CV 43.5%) litre h −1 70 kg −1 , volume of distribution (Vfree) 1700 (CV 44.9%) litre 70 kg −1 , absorption half-life (Tabsfree) 0.175 (CV 83.7%) h 70 kg −1 . There was no effect attributable to PNA on CL or V. Time to peak plasma concentration (Tmax) was 0.82 (CV 18%) h. Peak plasma concentration (Cmax) was 0.69 (CV 25%) µg ml −1 for total levobupivacaine and 0.09 (CV 37%) µg ml −1 for unbound levobupivacaine. Conclusions Clearance in infants is approximately half that described in adults, suggesting immaturity of P450 CYP3A4 and CYP1A2 enzyme isoforms that metabolize levobupivacaine in infants. This lower clearance delays Tmax, which was noted to occur approximately 50 min after administration of caudal epidural levobupivacaine.

Areas of insensibility produced by neuraxial anesthesia or peripheral nerve blocks can be detected during general anesthesia by failure of noxious stimulation to trigger pupillary reflex dilation. We examined the latency of pupillary reflex dilation and the effect of fentanyl on the latency of reflex dilation during anesthesia in nine volunteers. We hypothesized that the reflex was generated by slowly conducting C nociceptive fibers and would be significantly delayed if a distal dermatome (L4) was stimulated compared with a proximal dermatome (C5). We also hypothesized that fentanyl would prolong the latency and alter the shape of the reflex. After induction of general anesthesia, pupillary reflex dilation was measured with an infrared pupillometer every 5 min after stimulations of the L4 and C5 dermatomes. Fentanyl (3 μg/kg) was then given intravenously. Pupillary reflex dilation latencies were calculated by examining each individual measurement. After 3 h, naloxone (400 μg) was given intravenously; anesthesia was then discontinued. Pupillary reflex dilation had a long latency and consisted of distinct early and late phases. No differences were found between latencies of reflex dilation after simulation of L4 and C5 dermatomes either before or after fentanyl administration. Fentanyl at high concentrations essentially eliminated pupillary reflex dilation; but over the 180-min observation period, first early and then late dilation returned. Fentanyl produced a small increase in the latency of the initial early dilation. We conclude that pupillary reflex dilation during anesthesia is not initiated by slowly conducting C fibers and that fentanyl depresses the reflex in a stereotypical manner.

Recent studies have indicated that despite stringent sterilization processes, the reusable silicone laryngeal mask airway (LMA) has the potential for disease transmission through residual biological debris. As a result, a polyvinyl chloride (PVC) disposable LMA has been introduced. This randomized trial involved 138 spontaneously ventilating adult patients who underwent elective surgery requiring airway management with an LMA to determine whether there is a clinical difference between the disposable Portex® LMA (PLMA) and the standard reusable LMA in terms of ease of insertion, intra-operative cuff pressures and postoperative incidence of sore throat. There was no significant difference in first attempt insertion success rates (79% vs 84%) or difference in the incidence of postoperative sore throat observed between the two groups. Cuff pressure increases with nitrous oxide anaesthesia were significantly larger with the reusable LMA. The disposable PLMA provided a suitable airway in spontaneously ventilating patients without the risk of disease transmission inherent in a reusable device.

Background and Purpose— Hypothermia might prove to be therapeutically beneficial in stroke victims; however, even mild hypothermia provokes vigorous shivering. Meperidine and dexmedetomidine each linearly reduce the shivering threshold (triggering core temperature) with minimal sedation. We tested the hypothesis that meperidine and dexmedetomidine synergistically reduce the shivering threshold without producing substantial sedation or respiratory depression. Methods— We studied 10 healthy male volunteers (18 to 40 years) on 4 days: (1) control (no drug); (2) meperidine (target plasma level 0.3 μg/mL); (3) dexmedetomidine (target plasma level 0.4 ng/mL); and (4) meperidine plus dexmedetomidine (target plasma levels of 0.3 μg/mL and 0.4 ng/mL, respectively). Lactated Ringer’s solution (≈4°C) was infused through a central venous catheter to decrease tympanic membrane temperature by ≈2.5°C/h; mean skin temperature was maintained at 31°C. An increase in oxygen consumption >25% of baseline identified the shivering threshold. Sedation was evaluated by using the Observer’s Assessment of Sedation/Alertness scale. Two-way repeated-measures ANOVA was used to identify interactions between drugs. Data are presented as mean±SD; P Results— The shivering thresholds on the study days were as follows: control, 36.7±0.3°C; dexmedetomidine, 36.0±0.5°C ( P P P P =0.19). There was trivial sedation with either drug alone or in combination. Respiratory rate and end-tidal P co 2 were well preserved on all days. Conclusions— Dexmedetomidine and meperidine additively reduce the shivering threshold; in the small doses tested, the combination produced only mild sedation and no respiratory toxicity.

Background The authors evaluated a device designed to provide conscious sedation with propofol (propofol-air), or propofol combined with 50% nitrous oxide (N2O; propofol-N2O). An element of this device is the automated responsiveness test (ART), a method for confirming that patients remain conscious. The authors tested the hypotheses that the ART predicts loss of consciousness and that failure to respond to the ART precedes sedation-induced respiratory or hemodynamic toxicity. Methods The protocol consisted of sequential 15-min cycles in 20 volunteers. After a 15-min control period, propofol was infused to an initial target effect-site concentration of 0.0 microg/ml with N2O or 1.5 microg/ml with air. Subsequently, the propofol target effect-site concentration was increased by a designated increment (0.25 and 0.5 microg/ml) and the process repeated. This sequence was continued until loss of consciousness, as defined by an Observer's Assessment of Alertness/Sedation (OAA/S) score of 10/20 or less, or until an adverse physiologic event was detected. Results The OAA/S score at which only 50% of the volunteers were able to respond to the ART (P50) during propofol-N2O was 11.1 of 20 (95% confidence interval [CI]: 10.6-11.8); the analogous P50 was 11.8 of 20 (95% CI: 11.4-12.3) with propofol-air. Failure to respond to the ART occurred at a plasma propofol concentration of 0.7 +/- 0.6 microg/ml with propofol-N2O and 1.6 +/- 0.6 microg/ml with propofol-air, whereas loss of consciousness occurred at 1.2 +/- 0.8 microg/ml and 1.9 +/- 0.7 microg/ml, respectively. There were no false-normal ART responses. Conclusion The ART can guide individual titration of propofol because failure to respond to responsiveness testing precedes loss of consciousness and is not susceptible to false-normal responses. The use of N2O with propofol for conscious sedation decreases the predictive accuracy of the ART.

Background Epidural analgesia is frequently associated with hyperthermia during labor and in the postoperative period. The conventional assumption is that hyperthermia is caused by the technique, although no convincing mechanism has been proposed. However, pain in the "control" patients is inevitably treated with opioids, which themselves attenuate fever. Fever associated with infection or tissue injury may then be suppressed by opioids in the "control" patients while being expressed normally in patients given epidural analgesia. The authors therefore tested the hypothesis that fever in humans is manifested normally during epidural analgesia, but is suppressed by low-dose intravenous opioid. Methods The authors studied eight volunteers, each on four study days. Fever was induced each day by 150 IU/g intravenous interleukin 2. Volunteers were randomly assigned to: (1) a control day when no opioid or epidural analgesia was given; (2) epidural analgesia using ropivacaine alone; (3) epidural analgesia using ropivacaine in combination with 2 microg/ml fentanyl; or (4) intravenous fentanyl at a target plasma concentration of 2.5 ng/ml. Results Fentanyl halved the febrile response to pyrogen, decreasing integrated core temperature from 7.0 +/- 3.2 degrees C. h on the control day, to 3.8 +/- 3.0 degrees C. h on the intravenous fentanyl day. In contrast, epidural ropivacaine and epidural ropivacaine-fentanyl did not inhibit fever. The fraction of core-temperature measurements that exceeded 38 degrees C was halved by intravenous fentanyl, and the fraction exceeding 38.5 degrees C was reduced more than fivefold. Conclusions These data support the authors' proposed mechanism for hyperthermia during epidural analgesia. Fever during epidural analgesia should thus not be considered a complication of the anesthetic technique per se.

Inhalation induction with sevoflurane was compared with propofol or sevoflurane/propofol in 60 unpremedicated adults. Target concentrations for the three groups (with 60% nitrous oxide) were 3% end-tidal sevoflurane, 12 mg/l propofol and 1.5% sevoflurane/6 mg/l propofol respectively, prior to insertion of a laryngeal mask airway (LMA) at 10 minutes. Induction of anaesthesia was satisfactory in each group, but movement response to LMA insertion was observed in 20 patients (least in the sevoflurane group). Cardiovascular responses were similar except for a lower heart rate in the sevoflurane group. EEG bispectral index suggested a greater depth of anaesthesia in the inhalation induction group. A bispectral index of 60 separated patients responding to LMA insertion from nonresponders (P=0.006), and had a sensitivity of 68% and specificity 70%. Movement response was not predicted by cardiovascular changes.

The analgesic tramadol inhibits the neuronal reuptake of norepinephrine and 5-hydroxytryptamine, facilitates 5-hydroxytryptamine release, and activates mu-opioid receptors. Each of these actions is likely to influence thermoregulatory control. We therefore tested the hypothesis that tramadol inhibits thermoregulatory control. Eight volunteers were evaluated on four study days, on which they received no drugs, tramadol 125 mg, tramadol 250 mg, and tramadol 250 mg with naloxone, respectively. Skin and core temperatures were gradually increased until sweating was observed and then decreased until vasoconstriction and shivering were detected. The core temperature triggering each response defined its threshold. Tramadol decreased the sweating threshold by -1.03 +/- 0.67 degrees C microgram-1.mL (r2 = 0.90 +/- 0.12). Tramadol also decreased the vasoconstriction threshold by -3.0 +/- 4.0 degrees C microgram-1.mL (r2 = 0.94 +/- 0.98) and the shivering threshold by -4.2 +/- 4.0 degrees C microgram-1.mL(r2 = 0.98 +/- 0.98). The sweating to vasoconstriction interthreshold range nearly doubled from 0.3 +/- 0.4 degree C to 0.7 +/- 0.6 degree C during the administration of large-dose tramadol (P = 0.04). The addition of naloxone only partially reversed the thermoregulatory effects of tramadol. The thermoregulatory effects of tramadol thus most resemble those of midazolam, another drug that slightly decreases the thresholds triggering all three major autonomic thermoregulatory defenses. In this respect, both drugs reduce the "setpoint" rather than produce a generalized impairment of thermoregulatory control. Nonetheless, tramadol nearly doubled the interthreshold range at a concentration near 200 ng/mL. This indicates that tramadol slightly decreases the precision of thermoregulatory control in addition to reducing the setpoint.The authors evaluated the effects of the analgesic tramadol on the three major thermoregulatory responses: sweating, vasoconstriction, and shivering. Tramadol had only slight thermoregulatory effects. Its use is thus unlikely to provoke hypothermia or to facilitate fever.

Background Estimation of the mu-agonist opioid effect in anesthetized and paralyzed patients is often imprecise and can be obscured by concomitant administration of drugs that affect the sympathetic nervous system, such as beta-adrenergic blocking agents. As an alternative to hemodynamic measures of opioid effect, the authors tested the hypothesis that the pupillary light reflex or pupillary reflex dilation correlated with alfentanil concentrations during isoflurane anesthesia. Methods Six volunteers were anesthetized on 4 days with 0.8% isoflurane. Alfentanil was administered intravenously to target total plasma concentrations of 0, 25, 50, and 100 ng/ml. A 5-s tetanic electrical stimulus was applied to the skin. Pupil size and the pupillary light reflex were recorded before and after alfentanil administration, and before and for 8 min after the stimulus. Results Alfentanil exponentially impaired reflex pupillary dilation, decreasing the maximum response amplitude from 5 mm at 0 ng/ml, to 2.3 mm at 25 ng/ml, to 1.0 mm at 50 ng/ml, and finally to 0.2 mm at 100 ng/ml. In contrast, only the highest concentration of alfentanil depressed the dilation of the pupil in the first 2 s after the stimulus. Alfentanil administration had no effect on the pupillary light reflex. Conclusions Dilation of the pupil in response to a noxious stimulus is a measure of opioid effect in isoflurane-anesthetized volunteers. In contrast, the pupillary light reflex is unaffected by alfentanil during isoflurane anesthesia. These data suggest that stimulus-induced pupillary dilation may be used to evaluate the analgesic component of a combined volatile and opioid anesthetic.

To target appropriate drug concentrations and to facilitate comparisons between drugs, the potency of propofol must be firmly established. We therefore determined the arterial blood propofol concentration preventing movement in 50% of patients after skin incision and the ability of arterial blood pressure and heart rate to predict movement after incision. Fifteen healthy women scheduled for breast surgery were randomly assigned to computer-targeted propofol blood concentrations. No other drugs were administered. Fifteen minutes after starting the propofol infusion, a 5-cm skin incision was made. Patients were observed for gross purposeful movement for 1 min. Arterial blood was sampled for propofol to confirm steady-state blood concentrations. Arterial blood pressure and heart rate were measured noninvasively. Logistic regression was used to calculate the propofol blood concentrations and arterial blood pressures at which 50% and 95% of patients did not move after skin incision (CP50 and CP95, MABP50 and MABP95, respectively). The CP50 and CP95 values for propofol were 14.3 +/- 1.6 microg/mL (mean +/- SE) and 20.6 microg/mL, respectively. The MABP50 and MABP95 values were 63 +/- 4 mm Hg and 43 mm Hg, respectively. Heart rate did not differ significantly in patients who moved and who did not move. Propofol blood concentrations required to prevent movement in most patients resulted in significant arterial hypotension.

Background Meperidine administration is a more effective treatment for shivering than equianalgesic doses of other opioids. However, it remains unknown whether meperidine also profoundly impairs other thermoregulatory responses, such as sweating or vasoconstriction. Proportional inhibition of vasoconstriction and shivering suggests that the drug acts much like alfentanil and anesthetics but possesses greater thermoregulatory than analgesic potency. In contrast, disproportionate inhibition would imply a special antishivering mechanism. Accordingly, the authors tested the hypothesis that meperidine administration produces a far greater concentration-dependent reduction in the shivering than vasoconstriction threshold. Methods Nine volunteers were each studied on three days: 1) control (no opioid); 2) a target total plasma meperidine concentration of 0.6 microgram/ml (40 mg/h); and 3) a target concentration of 1.8 micrograms/ml (120 mg/h). Each day, skin and core temperatures were increased to provoke sweating and then subsequently reduced to elicit vasoconstriction and shivering. Core-temperature thresholds (at a designated skin temperature of 34 degrees C) were computed using established linear cutaneous contributions to control sweating (10%) and vasoconstriction and shivering (20%). The dose-dependent effects of unbound meperidine on thermoregulatory response thresholds was then determined using linear regression. Results are presented as means +/- SDs. Results The unbound meperidine fraction was approximately 35%. Meperidine administration slightly increased the sweating threshold (0.5 +/- 0.8 degree C.microgram-1.ml; r2 = 0.51 +/- 0.37) and markedly decreased the vasoconstriction threshold (-3.3 +/- 1.5 degrees C.microgram-1.ml; r2 = 0.92 +/- 0.08). However, meperidine reduced the shivering threshold nearly twice as much as the vasoconstriction threshold (-6.1 +/- 3.0 degrees C.microgram-1.ml; r2 = 0.97 +/- 0.05; P = 0.001). Conclusions The special antishivering efficacy of meperidine results at least in part from an uncharacteristically large reduction in the shivering threshold rather than from exaggerated generalized thermoregulatory inhibition. This pattern of thermoregulatory impairment differs from that produced by alfentanil, clonidine, propofol, and the volatile anesthetics, all which reduce the vasoconstriction and shivering thresholds comparably.

Non-steroidal anti-inflammatory drugs (NSAIDs) provide effective analgesia after orthopaedic surgery and reduce opioid requirements. The need for parenteral NSAIDs with peripheral surgery is controversial. In this study 10 patients were treated with oral tenoxicam 20 mg preoperatively, and at 4 hours and 28 hours after knee ligament reconstruction surgery. Plasma concentrations of tenoxicam, an NSAID with a long elimination half-life, were measured for 10 days. All patients received patient-controlled intravenous morphine postoperatively, which delayed absorption of the second and third tenoxicam doses. However, plasma concentrations of tenoxicam were achieved and maintained for the five-day surgical admission above the level considered to produce effective analgesia. Oral analgesic administration is a simple and feasible option in the perioperative period.

Background The transversus abdominis plane (TAP) block is a regional anaesthetic technique that blocks abdominal wall somatic afferent nerves. We conducted a prospective observational study to evaluate the venous plasma concentrations of ropivacaine during a continuous TAP infusion. Methods Twenty patients who were planned to undergo intra-abdominal cavity surgery requiring a mid-line laparotomy incision were enrolled. Patients were excluded if they had a history of chronic pain, opioid tolerance, renal or hepatic impairment, or contraindication to study medications. Subjects received a standardized general anaesthetic, and at the completion of surgery, ultrasound-guided subcostal or posterior TAP blocks and catheters. A TAP infusion of 2 mg ml −1 ropivacaine was administered for 72 h after operation. Data collection during the 72 h included morphine requirements, pain scores, and plasma ropivacaine levels. Results TAP blocks and catheters were successfully inserted in all recruited subjects. The fourth subject experienced neurological symptoms attributed to local anaesthetic toxicity, but did not have high plasma ropivacaine concentrations. However, the protocol was amended for the subsequent 16 subjects, to a weight-based dosing regimen. The range of total plasma ropivacaine concentrations was 0.98–3.41 mg litre −1 for posterior infusions and 0.96–3.48 mg litre −1 for subcostal infusions. Four subjects had total ropivacaine levels >3.4 mg litre −1 . The range of unbound plasma ropivacaine concentrations was 0.022–0.135 mg litre −1 for posterior infusions and 0.031–0.120 mg litre −1 for subcostal infusions. Conclusion Given the potential for high plasma concentrations from a bilateral TAP infusion technique, attention should be paid to individualized dosing strategies.

Background The transversus abdominis plane (TAP) block involves injecting a large volume of local anaesthetic between the muscles of the abdominal wall. Plasma concentrations of ropivacaine after gynaecological laparotomy are potentially high enough to result in systemic toxicity, and there are pharmacokinetic reasons why pregnancy may increase susceptibility to local anaesthetic toxicity. Methods Adult female patients (n=30) undergoing elective Caesarean section under spinal anaesthesia received bilateral ultrasound-guided TAP blocks after wound closure (2.5 mg kg−1 of ropivacaine diluted to 40 ml). Venous blood samples were collected at 10, 20, 30, 45, 60, 90, 120, 180 and 240 min following the block. Blood samples were assayed for total and free ropivacaine concentrations. Patients were assessed for symptoms of local anaesthetic toxicity. Results The mean [standard deviation ( sd )] peak total concentration of ropivacaine occurred at 30 min post-injection and was 1.82 (0.69) μg ml−1. The maximum detected concentration in any patient was 3.76 μg ml−1 (at 10 min post-injection). Three patients reported symptoms of mild neurotoxicity, and the mean ( sd ) peak levels were elevated in these patients, 2.70 (0.46) µg ml−1. Conclusions TAP blocks can result in elevated plasma ropivacaine concentrations in patients undergoing Caesarean section, which may be associated with neurotoxicity.

Perioperative hypothermia usually results largely from pharmacologic inhibition of normal thermoregulatory control.Midazolam is a commonly used sedative and anesthetic adjuvant whose thermoregulatory effects are unknown. We therefore tested the hypothesis that midazolam administration impairs thermoregulatory control. Eight volunteers were studied on 2 days each, once without drug and once at a target total plasma midazolam concentration of 0.3 micro gram/mL (corresponding to administration of approximate equals 40 mg over approximate equals 4 h). Each day, skin and core temperatures were increased sufficiently to provoke sweating, and then reduced to elicit peripheral vasoconstriction and shivering. We mathematically compensated for changes in skin temperature using the established linear cutaneous contributions to control of each response. From these calculated thresholds (core temperatures triggering responses at a designated skin temperature of 34 degrees C), we determined the thermoregulatory effects of midazolam. The sweating threshold was decreased approximate equals 0.3 degrees C by midazolam administration: 37.3 +/- 0.2 degrees C vs 37.0 +/- 0.3 degrees C (P = 0.0004, paired t-test). Midazolam decreased the core temperature that triggered vasoconstriction somewhat more: 37.1 +/- 0.2 degrees C vs 36.3 +/- 0.5 degrees C (P = 0.0002). Similarly, midazolam decreased the shivering threshold: 35.9 +/- 0.3 degrees C vs 35.3 +/- 0.6 degrees C (P = 0.03). The sweating-to-vasoconstriction (interthreshold) range, therefore, increased from 0.2 +/- 0.1 degrees C to 0.7 +/- 0.3 degrees C (P = 0.002). Although statistically significant, this relatively small increase contrasts markedly with the 3-5 degrees C interthreshold ranges produced by clinical doses of volatile anesthetics, propofol, and opioids. Thus, plasma concentrations of midazolam far exceeding those used routinely produce relatively little impairment of thermoregulatory control. (Anesth Analg 1995;81:393-8)

Background The inability of edrophonium to rapidly reverse a deep nondepolarizing neuromuscular block may be due to inadequate dosage or a ceiling effect to antagonism of neuromuscular block by edrophonium. A ceiling effect means that only a certain level of neuromuscular block could be antagonized by edrophonium. Neuromuscular block greater than this could not be completely antagonized irrespective of the dose of edrophonium administered. The purpose of this study was to determine whether a ceiling effect occurred for antagonism of an atracurium-induced neuromuscular block by edrophonium and, if so, the maximum level of block that could be antagonized by edrophonium. Methods In 30 adult patients, atracurium was administered to maintain a constant neuromuscular block. The level of block varied between patients. Evoked adductor pollicis twitch tension was monitored. Incremental doses of edrophonium were administered while the infusion of atracurium continued. Increments were given until adequate recovery occurred, as defined by a train-of-four (TOF) ratio > or = 70%, or until no further antagonism of the block could be achieved. The probability of being able to effect adequate recovery by antagonism with edrophonium was determined using a logistic regression model. Cumulative dose-response curves were constructed using the logit transformation of the neuromuscular effect versus the logarithm of the cumulative dose of edrophonium. Results In 14 patients with a block of 25-77% depression of the first twitch response, antagonism by edrophonium to a TOF ratio > or = 70% was possible, whereas in 16 patients with a 60-92% depression of T1, a TOF ratio > or = 70% was not achievable, indicating that a ceiling effect for antagonism by edrophonium occurred. A block of 67 +/- 3% (mean +/- SE) had a 50% probability of adequate antagonism. In patients in whom block was antagonized to a TOF ratio < 70%, 95% of the peak antagonistic effect occurred with an edrophonium dose of 0.8 +/- 0.33 mg.kg-1 (mean +/- SD). Conclusions There is a maximum level of neuromuscular block that can be antagonized by edrophonium to effect adequate recovery. The level corresponds approximately to the reappearance of the fourth response to TOF stimulation. It is probably safest to wait until this level of block occurs before edrophonium is given for reversal. Earlier administration will not hasten recovery.

Interpleural infusions (24 hr) were given to 14 adults (29-81 yr) having surgery via right subcostal incisions. Patient-controlled analgesia with morphine was also available for 72 hours following surgery. An infusion of 0.25% bupivacaine with adrenaline 1/400,000 was commenced at 0.1 ml/kg/hr at the conclusion of surgery after an initial 20 ml bolus. The 24 hr cumulative morphine dose increased by 62% (P < 0.01) on the second postoperative day after the bupivacaine infusion was ceased, but the pain score did not change. There was no clinical evidence of bupivacaine toxicity although the maximum bupivacaine concentration ranged from 0.74-6.52 mg/l, mean 2.90 (SEM 0.44) mg/l and vascular uptake was rapid in two patients. The concentration of S-bupivacaine was consistently greater than R-bupivacaine (P < 0.05) and total body clearance was less. The unbound bupivacaine fraction also changed from 3.6% (SEM 0.6) at the start of the infusion to 2.0% (SEM 0.3) at 24 hr (P < 0.005). The total body clearance of each enantiomer was reduced during the 24 hours (P < 0.001). No relationship between pharmacokinetic parameters and weight, age or sex was found (P>0.25 for each). Variation between patients was reduced when parameters were estimated for the free (unbound) bupivacaine (P < 0.001).

Background The transversus abdominis plane block is a novel technique involving injection of local anaesthetic between the internal oblique and the transversus abdominis muscles of the abdominal wall. It is possible that injection of a large dose of local anaesthetic into a relatively vascular plane may result in toxic concentrations. One previously published study examined plasma lidocaine concentrations after transversus abdominus plane block and showed potentially toxic plasma concentrations. Although ropivacaine is most commonly used for this technique, plasma concentrations of ropivacaine after this block have not been reported previously. Methods Adult female patients undergoing elective open gynaecological surgery received bilateral ultrasound-guided transverse abdominal plane blocks before surgical incision (3 mg kg−1 of ropivacaine diluted to 40 ml). Venous blood was collected each 15 min for the first hour, each 30 min for the second hour, and then at 3, 4, 12, and 24 h post-block. Results Twenty-eight patients were recruited. The mean (sd) peak total ropivacaine concentration occurred 30 min post-injection and was 2.54 (sd 0.75) µg ml−1. The highest measured concentration was 4.00 µg ml−1, also 30 min post-injection. Mean total concentrations remained above 2.20 µg ml−1 for up to 90 min post-injection. The mean unbound peak venous concentration was 0.14 (0.05) µg ml−1, and the peak was 0.25 µg ml−1. Conclusions Transversus abdominus plane block using 3 mg kg−1 of ropivacaine produces venous plasma concentrations that are potentially neurotoxic, although broadly consistent with plasma levels found after injection at other comparable sites.

The dosage, haemodynamic and respiratory effects of propofol for laryngeal mask airway (LMA) insertion were investigated. Fifty patients (ASA I-II) were randomly assigned one of four induction doses of propofol (1.5–2.5 mg/kg) delivered over 30 seconds and the first attempt at LMA insertion was made at 90 seconds. The LMA was inserted at 90 seconds in 35 patients and by 300 seconds in 13 others (mean plasma concentration at 90 seconds was 7.7 mcg/ml (no delay) versus 5.2 mcg/ml (insertion delayed), P < 0.01). Insertion was less successful after 1.5 mcg/kg (failed at 90 seconds in 6 of 12 patients), but did not vary with the other doses. Additional propofol (0.5 mg/kg/30s) was required in 22 patients for LMA insertion or to prevent movement, resulting in propofol concentrations at 120–180 seconds above 7 mcg/ml. Respiratory effects were minor, but MAP decreased by 18±1.4 mmHg at 90 seconds. Cardiovascular effects did not differ significantly between dosage groups or with the use of additional propofol.

In previous studies, we showed that failure to respond to automated responsiveness monitor (ARM) precedes potentially serious sedation-related adversities associated with loss of responsiveness, and that the ARM was not susceptible to false-positive responses. It remains unknown, however, whether loss and return of response to the ARM occur at similar sedation levels. We hypothesized that loss and return of response to the ARM occur at similar sedation levels in individual subjects, independent of the propofol effect titration scheme.Twenty-one healthy volunteers aged 20-45 yr underwent propofol sedation using an effect-site target-controlled infusion system and two different dosing protocol schemes. In all, we increased propofol effect-site concentration (Ce) until loss of response to the ARM occurred. Subsequently, the propofol Ce was decreased either by a fixed percentage (20%, 30%, 40%, 50%, 60%, and 70%; fixed percentage protocol, n = 10) or by a linear deramping (0.1, 0.2, and 0.3 microg x mL(-1) x min(-1); deramping protocol, n = 11) until the ARM response returned. Consequently, the propofol Ce was maintained at the new target for a 6-min interval (Ce plateau) during which arterial samples for propofol determination were obtained, and a clinical assessment of sedation (Observer's Assessment of Alertness/Sedation [OAA/S] score) performed. Each participant in the two protocols experienced each percentage or deramping rate of Ce decrease in random order. The assumption of steady state was tested by plotting the limits of agreement between the starting and ending plasma concentration (Cp) at each Ce plateau. The probability of response to the ARM as a function of propofol Ce, Bispectral Index (BIS) of the electroencephalogram, and OAA/S score was estimated, whereas the effect of the protocol type on these estimates was evaluated using the nested model approach (NONMEM). The combined effect of propofol Ce and BIS on the probability for ARM response was also evaluated using a fractional probability model (P(BIS/Ce)).The measured propofol Cp at the beginning and the end of the Ce plateau was almost identical. The Ce50 of propofol for responding to the ARM was 1.73 (95% confidence interval: 1.55-2.10) microg/mL, whereas the corresponding BIS50 was 75 (71.3-77). The OAA/S50 probability for ARM response was 12.5/20 (12-13.4). A fractional probability (P(BIS/Ce)) model for the combined effect of BIS and Ce fitted the data best, with an estimated contribution for BIS of 63%. Loss and return of ARM response occurred at similar sedation levels in individual subjects.Reproducible ARM dynamics in individual subjects compares favorably with clinical and electroencephalogram sedation end points and suggests that the ARM could be used as an independent instrumental guide of drug effect during propofol-only sedation.

We have assessed, in 200 patients, the determinants of the reversal time of competitive neuromuscular block by anticholinesterase when alcuronium and atracurium neuromuscular block were antagonized by neostigmine 0.04 and 0.08 mg kg-1 and edrophonium 0.5 and 1.0 mg kg-1. A biexponential relationship was found between the reversal time (time from injection of anticholinesterase to a train-of-four ratio of 70%) and the degree of neuromuscular block at reversal (all groups; F ratio, P less than 0.05). Reversal time was determined by two processes: direct antagonism by the anticholinesterase and spontaneous recovery of the neuromuscular blocking agent, with the latter becoming the major determinant at profound levels of neuromuscular block (0-10% of control twitch height). Neostigmine, in the doses studied, appeared to have a higher "ceiling" of neuromuscular block which it completely antagonized, although edrophonium had a more rapid onset of action. The reversal time for alcuronium became progressively longer relative to atracurium as neuromuscular block increased because of the slower spontaneous recovery rate. Avoidance of profound neuromuscular block at the completion of surgery is required to ensure reliable antagonism of the block within 5-10 min by an anticholinesterase. Neostigmine 0.08 mg kg-1 was found to be the most effective agent in antagonizing profound neuromuscular block.

The pharmacokinetics of atracurium were investigated by a model-independent method during continuous infusion under propofol anaesthesia. Following an intubating dose of suxamethonium, atracurium was infused according to a predetermined profile which continually set the infusion rate to maintain stable muscle paralysis and a target steady state plasma concentration when equilibrium between the bio-phase and plasma had occurred. Atracurium was infused for the first 1 h to maintain a target steady state plasma concentration of 1.0 μg ml−1. Thereafter, the target plasma concentration was adjusted to maintain 90% muscle paralysis. The maintenance infusion rate required to maintain 90% paralysis was 4.25 (SD) 1.11 μg kg−1 min−1, with an estimated steady state plasma concentration of atracurium required to maintain 90% paralysis (CpSS 90) of 1.13 (0.24) μg ml−1. The clearances of atracurium, estimated by the constant infusion rate required to maintain the steady state plasma concentration, at 50–60 min and during estimation of CpSS 90 were 3.8 (1.0) and 3.9 (1.1) ml kg−1 min−1 (ns), respectively. The volume of distribution at steady state of atracurium after 1 h of infusion, calculated using the clearance and the area under the plasma concentration-time curve to 1 h, was 130 (50) ml kg−1. These estimates of the pharmacokinetic parameters of atracurium are markedly different from those derived from pharmacokinetic analysis of single bolus dose data. Normalization of the pharmacokinetic parameter estimates by lean body mass decreased interpatient variability and improved precision in comparison with the un-weighted data and normalization by total body weight.

For normal anaesthetic practice, monitoring of neuromuscular blockade is best performed by stimulation of the ulnar nerve at the wrist with a peripheral nerve stimulator and evaluation of the response of the thumb. Determination of the initial threshold for stimulation in the awake patient to allow estimation of the current required for supramaximal stimulation is an important set-up procedure to improve accuracy. The degree of paralysis of specific muscle groups such as the diaphragm can be inferred from their sensitivity to neuromuscular blocking agents relative to adductor pollicis. Monitoring with different stimulation patterns allows a wide spectrum of muscle paralyses to be evaluated.

We have assessed in 20 patients the accuracy and precision of an infusion profile for atracurium, which continually set the infusion rate to maintain stable muscle paralysis and a target steady state plasma concentration, when equilibrium between the biophase and plasma had occurred. Muscle paralysis was stable after 20 min, with a mean absolute drift in muscle paralysis in the succeeding 40 min of 0.13 (SD 0.07)% T1/Tc (height of first twitch/height of control twitch) per min. The plasma samples after 30 min, which were assessed empirically as being in equilibrium with the biophase, had an overall mean bias of 8.0 (SEM 3.7)% (P less than 0.05) and an overall mean absolute prediction error of 16.4 (SEM 2.5)% from the target steady state concentration being delivered by the infusion. The profile was then used to estimate the steady state plasma concentration of atracurium required to maintain 90% paralysis (Cpss90), by manually adjusting the delivered target concentration of the infusion until muscle paralysis was stable at 88-92% inhibition of T1/Tc for 15-20 min, with three plasma samples taken over the next 10 min. Measurements were completed within 60-90 min. The mean Cpss90 of atracurium with propofol was 1.039 (SD 0.224) microgram ml-1 (n = 10), with thiopentone 1.334 (0.378) microgram ml-1 (n = 10), and with opioid anaesthesia 0.915 (0.221) microgram ml-1 (n = 10). These differences in the Cpss90 explain some of the variability in response which occurs with neuromuscular blocking drugs. The technique enables the Cpss90 of a myoneural blocker to be determined by a simple model-independent method.

The blood concentration associated with loss of response (LOR) to command in 50% of subjects (CP50(LOR)) is an important measure of anesthetic potency. We therefore determined the CP50(LOR) in 40 healthy surgical patients, aged 18-60 yr old, receiving propofol alone or propofol with 67% nitrous oxide (N2O). Patients were randomized to receive 100% oxygen or 67% N2O in oxygen via facemask. Three minutes later, a target-controlled propofol infusion was commenced at a concentration determined by the response of the previous patient in the same group. Fifteen minutes later, response to command was assessed by a blinded observer. Arterial blood samples were taken for propofol assay, and the bispectral index (BIS) was monitored continuously. At testing for response to command, both the measured and target propofol concentrations were significantly larger and BIS values significantly smaller in the propofol-alone group compared with the propofol-N2O group. The CP50(LOR) of propofol in the propofol-alone group was 4.58 mug/mL (95% confidence interval [CI], 1.14-15.36) and 2.67 microg/mL (95% CI, 2.28-3.17) in the propofol-N2O group. The BIS value when 50% of patients responded to command was 60 (95% CI, 55-65) in the propofol-alone group and 75 (95% CI, 73-83) in the propofol-N2O group.

Background Evidence suggests that the rate at which intravenous anesthetics are infused may influence their plasma-effect site equilibration. The authors used five different rates of propofol administration to test the hypothesis that different sedation endpoints occur at the same effect site propofol concentration, independent of the infusion rate. The authors concurrently evaluated the automated responsiveness monitor (ARM) against other sedation measures and the propofol effect site concentration. Methods With Human Studies Committee approval, 18 healthy volunteers received five consecutive target-controlled propofol infusions. During each infusion, the effect site concentration was increased by a rate of 0.1, 0.3, 0.5, 0.7, or 0.9 microg . ml . min. The Bispectral Index and ARM were recorded at frequent intervals. The times of syringe drop and loss and recovery of responsiveness were noted. Pharmacokinetic and pharmacodynamic modeling was performed using NONMEM. Results When the correct rate of plasma-effect site equilibration was determined for each individual (plasma-effect site equilibration = 0.17 min, time to peak effect = 2.7 min), the effect site concentrations associated with each clinical measure were not affected by the rate of increase of effect site propofol concentration. ARM correlated with all clinical measures of drug effect. Subjects invariably stopped responding to ARM at lower effect site propofol concentrations than those associated with loss of responsiveness. Conclusions : Population-based pharmacokinetics, combined with real-time electroencephalographic measures of drug effect, may provide a means to individualize pharmacodynamic modeling during target-controlled drug delivery. ARM seems useful as an automated measure of sedation and may provide the basis for automated monitoring and titration of sedation for a propofol delivery system.

We tested the efficacy of pethidine and cutaneous warming to prevent shivering during percutaneous cooling in unanaesthetized patients. Ten patients scheduled for cranial neurosurgery received pethidine infusion and skin warming. The Setpoint internal heat-exchanging catheter was inserted and cooling to 33.5 degrees C was started. Clonidine and chlorpromazine were given as "rescue medication" to treat shivering. General anaesthesia was planned to be induced after cooling was complete. Rewarming was initiated at dural closure. Three patients successfully completed the protocol, cooling to 33.8 degrees C at a median rate of 3.6 (range: 3.4-3.8) degrees C/h. Two patients cooled to 33.8 degrees C but required treatment for shivering (cooling rate: 2.9 [2.8-3.1] degrees C/h). Four patients failed to cool adequately because of refractory shivering (cooling rate: 20 [1.5-2.9] degrees C/h). One patient did not shiver and yet failed to cool adequately (cooling rate: 0.76 degrees C/h). Rewarming was at a rate of 26 (1.2-4.3) degrees C/h. There were no significant complications arising from catheter placement. The combination of skin warming and pethidine was not reliable enough to be recommended for use during endovascular cooling in unanaesthetized patients.

The production of reactive oxygen species in skeletal muscle is linked with muscle fatigue. This study investigated whether the antioxidant compound N-acetylcysteine (NAC) augments time to fatigue during prolonged, submaximal cycling exercise. Seven men completed a double-blind, crossover study, receiving NAC or placebo before and during cycling exercise, comprising 45 min at 70% of peak oxygen consumption (V̇o2 peak) and then to fatigue at 90% V̇o2 peak. NAC was intravenously infused at 125 mg·kg-1·h-1 for 15 min and then 25 mg·kg-1·h-1 for 20 min before and throughout exercise, which was continued until fatigue. Arterialized venous blood was analyzed for NAC concentration, hematology, and plasma electrolytes. NAC induced no serious adverse reactions and did not affect hematology, acid-base status, or plasma electrolytes. Time to fatigue was reproducible in preliminary trials (coefficient of variation 7.4 ± 1.2%) and was not augmented by NAC (NAC 14.6 ± 4.5 min; control 12.8 ± 5.4 min). However, time to fatigue during NAC trials was correlated with V̇o2 peak ( r = 0.78; P < 0.05), suggesting that NAC effects on performance may be dependent on training status. The rise in plasma K+ concentration at fatigue was attenuated by NAC ( P < 0.05). The ratio of rise in K+ concentration to work and the percentage change in time to fatigue tended to be inversely related ( r = -0.71; P < 0.07). Further research is required to clarify a possible training status-dependent effect of NAC on muscle performance and K+ regulation.

Alfentanil and propofol total intravenous anaesthesia was assessed in 25 patients undergoing coronary artery bypass graft surgery with cardiopulmonary bypass (CPB). A manually controlled alfentanil infusion, calculated from estimated lean body mass and published pharmacokinetic data, was effective in achieving target plasma concentrations, while the “Diprifusor” system was used to vary propofol target concentrations according to changes in haemodynamics and anaesthetic requirement. The effects of CPB on alfentanil plasma concentrations were offset by changes in protein binding and free-fraction of the drug. With the use of only two target plasma concentrations for alfentanil (changed after CPB), a pre-determined infusion profile ensured effective plasma concentrations during surgery and concentrations unlikely to inhibit extubation within six hours of sternal closure.

Infusion of the antioxidant N-acetylcysteine (NAC) reduces fatigability in electrically evoked human muscle contraction, but due to reported adverse reactions, no studies have investigated NAC infusion effects during voluntary exercise in humans. We investigated whether a modified NAC-infusion protocol (125 mg · kg−1· h−1for 15 min, then 25 mg · kg−1· h−1) altered blood redox status and enhanced performance during intense, intermittent exercise. Eight untrained men participated in a counterbalanced, double-blind, crossover study in which they received NAC or saline (control) before and during cycling exercise, which comprised three 45-s bouts and a fourth bout that continued to fatigue, at 130% peak oxygen consumption. Arterialized venous blood was analyzed for glutathione status, hematology, and plasma electrolytes. NAC infusion induced no severe adverse reactions. Exercise decreased the reduced glutathione ( P < 0.005) and increased oxidized glutathione concentrations ( P < 0.005); NAC attenuated both effects ( P < 0.05). NAC increased the rise in plasma K+concentration-to-work ratio ( P < 0.05), indicating impaired K+regulation, although time to fatigue was unchanged (NAC 102 ± 45 s; saline 107 ± 53 s). Thus NAC infusion altered blood redox status during intense, intermittent exercise but did not attenuate fatigue.

After institutional approval and with written informed consent, eight surgical patients were infused intravenously with remifentanil at 250 ng.kg lean body mass (LBM) -1 .min -1 for 30 min. Cardiovascular and respiratory parameters were recorded and arterial blood samples were taken at regular intervals. In each patient, the same protocol was repeated 40 min later during propofol infused to a target concentration of 3.0 μg.ml -1. Blood concentrations of remifentanil and propofol were assayed using capillary gas chromatography and high performance liquid chromatography techniques respectively. The number of subjects enrolled was determined by testing the successive areas under the remifentanil time-concentration curve (AUC) for significant difference or non-difference using sequential analysis. The median measured propofol concentration was 3.5 (range: 2.6–4.5) μg.ml -1 which did not change significantly during the second remifentanil infusion. The median AUC during propofol infusion was greater than control in all subjects, although there was considerable variation of 94.4 (64.3–129.6) versus 64.6 (34.8–126.9) ng.ml -1 .min; P=0.008, n=8. After 30 min, there was no significant difference in remifentanil concentration during propofol infusion when compared with remifentanil alone of 4.6 (3.2–5.7) versus 3.8 (1.6–4.9) ng.ml -1; P=0.73, n=8. Co-administration of propofol and remifentanil may result in greater remifentanil concentrations than when remifentanil is infused alone.

INTRODUCTION: Glioblastoma multiforme (GBM) cells release glutamate through expression of system xc−, which exchanges intracellular glutamate for extracellular cysteine. Lack of glutamate uptake through low excitatory amino acid transporter 2 (EAAT2) expression permits high extracellular glutamate levels, causing damaging excitotoxicty to surrounding parenchyma and epileptogenesis at the peritumoural edge. PPARγ agonists upregulate EAAT2 expression and prevent glutamate mediated excitotoxicity in stroke models. We investigated PPARγ agonists in GBM glutamate regulation. METHOD: Glioma cell lines U87MG and U251, and Glioma stem cells (GSCs) isolated from freshly resected tumour specimens were exposed to increasing concentrations of Pioglitazone. Effects on cell viability, protein expression and extracellular glutamate levels were evaluated with an ATP assay, Western Blotting and HPLC respectively. RESULTS: EAAT2 expression was upregulated in a dose and time dependent manner in both U87MG and U251 cells. Glutamate levels were reduced with the addition of pioglitazone, where statistical significance was reached in U251 cells at a concentration of 30 µM pioglitazone (n = 3, p < 0.05). Pioglitazone significantly reduced the cell viability of U87MG and U251 cells at concentrations of ≥ 30 µM (n = 4, p < 0.05) and 100 µM (n = 6, p < 0.05) respectively. Photomicrographs revealed morphological changes in the glioma cell lines and inability of the GSCs to form neurospheres with increasing pioglitazone concentrations. CONCLUSION: This study provides preliminary evidence of PPARγ-mediated regulation of EAAT2 expression in human glioblastoma cells and suggests a novel treatment strategy for glioma associated seizures.

Mild hypothermia (i.e., 34 degrees C) may prove therapeutic for patients with stroke, but it usually provokes shivering. We tested the hypothesis that the combination of buspirone (a serotonin 1A partial agonist) and meperidine synergistically reduces the shivering threshold (triggering tympanic membrane temperature) to at least 34 degrees C while producing little sedation or respiratory depression. Eight volunteers each participated on four randomly-assigned days: 1) large-dose oral buspirone (60 mg); 2) large-dose IV meperidine (target plasma concentration of 0.8 microg/mL); 3) the combination of buspirone (30 mg) and meperidine (0.4 microg/mL); and 4) a control day without drugs. Core hypothermia was induced by infusion of lactated Ringer's solution at 4 degrees C. The control shivering threshold was 35.7 degrees C +/- 0.2 degrees C. The threshold was 35.0 degrees C +/- 0.8 degrees C during large-dose buspirone and 33.4 degrees C +/- 0.3 degrees C during large-dose meperidine. The threshold during the combination of the two drugs was 33.4 degrees C +/- 0.7 degrees C. There was minimal sedation on the buspirone and combination days and mild sedation on the large-dose meperidine day. End-tidal PCO2 increased approximately 10 mm Hg with meperidine alone. Buspirone alone slightly reduced the shivering threshold. The combination of small-dose buspirone and small-dose meperidine acted synergistically to reduce the shivering threshold while causing little sedation or respiratory toxicity.Mild hypothermia may be an effective treatment for acute stroke, but it usually triggers shivering, which could be harmful. Our results indicate that the combination of small-dose buspirone and small-dose meperidine acts synergistically to reduce the shivering threshold while causing little sedation or respiratory toxicity. This combination may facilitate the induction of therapeutic hypothermia in stroke victims.

The special antishivering action of meperidine may be mediated by its kappa or anticholinergic actions. We therefore tested the hypotheses that nalbuphine or atropine decreases the shivering threshold more than the vasoconstriction threshold. Eight volunteers were each evaluated on four separate study days: 1) control (no drug), 2) small-dose nalbuphine (0.2 microg/mL), 3) large-dose nalbuphine (0.4 microg/mL), and 4) atropine (1-mg bolus and 0.5 mg/h). Body temperature was increased until the patient sweated and then decreased until the patient shivered. Nalbuphine produced concentration-dependent decreases (mean +/- SD) in the sweating (-2.5 +/- 1.7 degrees C. microg(-1). mL; r(2) = 0.75 +/- 0.25), vasoconstriction (-2.6 +/- 1.7 degrees C. microg(-1). mL; r(2) = 0.75 +/- 0.25), and shivering (-2.8 +/- 1.7 degrees C. microg(-1). mL; r(2) = 0.79 +/- 0.23) thresholds. Atropine significantly increased the thresholds for sweating (1.0 degrees C +/- 0.4 degrees C), vasoconstriction (0.9 degrees C +/- 0.3 degrees C), and shivering (0.7 degrees C +/- 0.3 degrees C). Nalbuphine reduced the vasoconstriction and shivering thresholds comparably. This differs markedly from meperidine, which impairs shivering twice as much as vasoconstriction. Atropine increased all thresholds and would thus be expected to facilitate shivering. Our results thus fail to support the theory that activation of kappa-opioid or central anticholinergic receptors contribute to meperidine's special antishivering action.