Methadone disposition and pharmacodynamics are highly susceptible to interactions with antiretroviral drugs. Methadone clearance and drug interactions have been attributed to cytochrome P4503A4 (CYP3A4), but actual mechanisms are unknown. Drug interactions can be both clinically and mechanistically informative. This investigation assessed effects of the protease inhibitor indinavir on methadone pharmacokinetics and pharmacodynamics, hepatic and intestinal CYP3A4/5 activity (using alfentanil), and intestinal transporter activity (using fexofenadine).
Twelve healthy volunteers underwent a sequential crossover. On three consecutive days they received oral alfentanil plus fexofenadine, intravenous alfentanil, and intravenous plus oral (deuterium-labeled) methadone. This was repeated after 2 weeks of indinavir. Plasma and urine analytes were measured by mass spectrometry. Opioid effects were measured by miosis.
Indinavir significantly inhibited hepatic and first-pass CYP3A activity. Intravenous alfentanil systemic clearance and hepatic extraction were reduced to 40-50% of control, apparent oral clearance to 30% of control, and intestinal extraction decreased by half, indicating 50% and 70% inhibition of hepatic and first-pass CYP3A activity. Indinavir increased fexofenadine area under the plasma concentration-time curve 3-fold, suggesting significant P-glycoprotein inhibition. Indinavir had no significant effects on methadone plasma concentrations, methadone N-demethylation, systemic or apparent oral clearance, renal clearance, hepatic extraction or clearance, or bioavailability. Methadone plasma concentration-effect relationships were unaffected by indinavir.
Despite significant inhibition of hepatic and intestinal CYP3A activity, indinavir had no effect on methadone N-demethylation and clearance, suggesting little or no role for CYP3A in clinical disposition of single-dose methadone. Inhibition of gastrointestinal transporter activity had no influence of methadone bioavailability.
Intravenous delivery of volatile fluorinated anesthetics has a number of potential advantages when compared to the current inhalation method of administration. We reported previously that the IV delivery of sevoflurane can be achieved through an emulsion composed of a linear fluorinated diblock copolymer, a stabilizer, and the anesthetic. However, this original emulsion was subject to particle size growth that would limit its potential clinical utility. We hypothesized that the use of bulkier fluorous groups and smaller poly(ethylene glycol) moieties in the polymer design would result in improved emulsion stability while maintaining anesthetic functionality.
The authors prepared emulsions incorporating sevoflurane, perfluorooctyl bromide as a stabilizing agent, and combinations of linear fluorinated diblock copolymer and a novel dibranched fluorinated diblock copolymer. Emulsion stability was assessed using dynamic light scattering. The ability of the emulsions to induce anesthesia was tested in vivo by administering them intravenously to fifteen male Sprague-Dawley rats and measuring loss of the forepaw righting reflex.
20% (volume/volume) sevoflurane emulsions incorporating mixtures of dibranched- and linear diblock copolymers had improved stability, with those containing an excess of the dibranched polymers displaying stability of particle size for over one year. The ED50s for loss of forepaw righting reflex were all similar, and ranged between 0.55 and 0.60 ml/kg body weight.
Hemifluorinated dibranched polymers can be used to generate exceptionally stable sevoflurane nanoemulsions, as required of formulations intended for clinical use. Intravenous delivery of the emulsion in rats resulted in induction of anesthesia with rapid onset and smooth and rapid recovery.
Whether decreasing the local anesthetic concentration during a continuous femoral nerve block results in less quadriceps weakness remains unknown.
Preoperatively, bilateral femoral perineural catheters were inserted in patients undergoing bilateral knee arthroplasty (n = 36) at a single clinical center. Postoperatively, right-sided catheters were randomly assigned to receive perineural ropivacaine of either 0.1% (basal 12 mL/h; bolus 4 mL) or 0.4% (basal 3 mL/h; bolus 1 mL), with the left catheter receiving the alternative concentration/rate in an observer- and subject-masked fashion. The primary endpoint was the maximum voluntary isometric contraction of the quadriceps femoris muscles the morning of postoperative day 2. Equivalence of treatments would be concluded if the 95% confidence interval for the difference fell within the interval of −20% to 20%. Secondary endpoints included active knee extension, passive knee flexion, tolerance to cutaneous electrical current applied over the distal quadriceps tendon, dynamic pain scores, opioid requirements, and ropivacaine consumption.
Quadriceps maximum voluntary isometric contraction for limbs receiving 0.1% ropivacaine was a mean (SD) of 13 (8) N·m, versus 12 (8) N·m for limbs receiving 0.4% [intra-subject difference of 3 (40) percentage points; 95% CI −10 to 17; p = 0.63]. Because the 95% confidence interval fell within prespecified tolerances, we conclude that the effect of the two concentrations were equivalent. Similarly, there were no statistically significant differences in secondary endpoints.
For continuous femoral nerve blocks, we found no evidence that local anesthetic concentration and volume influence block characteristics, suggesting that local anesthetic dose (mass) is the primary determinant of perineural infusion effects.
During extended storage, erythrocytes undergo functional changes. These changes reduce the viability of erythrocytes leading to release of oxyhemoglobin, a potent scavenger of nitric oxide. We hypothesized that transfusion of ovine packed erythrocytes (PRBC) stored for prolonged periods would induce pulmonary vasoconstriction in lambs, and that reduced vascular nitric oxide concentrations would increase this vasoconstrictor effect.
We developed a model of autologous stored blood transfusion in lambs (n=36). Leukoreduced blood was stored for either 2 days (fresh PRBC) or 40 days (stored PRBC). Fresh or stored PRBC were transfused into donors instrumented for awake hemodynamic measurements. Hemodynamic effects of PRBC transfusion were also studied after infusion of NG-nitro-L-arginine methyl-ester (25 mg/kg) or during inhalation of nitric oxide (80 ppm).
Cell-free hemoglobin levels were higher in the supernatant of stored PRBC than in supernatant of fresh PRBC (Mean±SD, 148±20 versus 41±13 mg/dl, respectively, P<0.001). Pulmonary artery pressure during transfusion of stored PRBC transiently increased from 13±1 to 18±1 mmHg (P<0.001) and was associated with increased plasma hemoglobin concentrations. NG-nitro-L-arginine methyl-ester potentiated the increase in pulmonary arterial pressure induced by transfusing stored PRBC, whereas inhalation of nitric oxide prevented the vasoconstrictor response.
Our results suggest that patients with reduced vascular nitric oxide levels due to endothelial dysfunction may be more susceptible to adverse effects of transfusing blood stored for prolonged periods. These patients might benefit from transfusion of fresh PRBC, when available, or inhaled nitric oxide supplementation to prevent the pulmonary hypertension associated with transfusion of stored PRBC.
Structural MRI is used to longitudinally monitor the progression of Alzheimer's disease from its presymptomatic to symptomatic phases. Using magnetic resonance imaging data from the Alzheimer's Disease Neuroimaging Initiative (ADNI), we tested the hypothesis that surgery would affect brain parameters associated with progression of dementia.
Materials and Methods
Brain images from the neuroimaging initiative database were used to study normal volunteer subjects and patients with mild cognitive impairment for the age group 55 to 90 inclusive. We compared changes in regional brain anatomy for three visits that defined two inter-visit intervals for a surgical cohort (n=41) and a propensity matched non-surgical control cohort (n = 123). The first interval for the surgical cohort contained the surgical date. Regional brain volumes were determined with Freesurfer and quantitatively described with J-image software (University of California at San Francisco). Statistical analysis used Repeated Measures ANCOVA (SPSS, v.18.0; Chicago, IL).
We found that surgical patients, during the first follow-up interval (5–9 months), but not subsequently, had increased rates of atrophy for cortical gray matter and hippocampus, and lateral ventricle enlargement, as compared to non-surgical controls. A composite score of five cognitive tests during this interval showed reduced performance for surgical patients with mild cognitive impairment.
Elderly subjects after surgery experienced an increased rate of brain atrophy during the initial evaluation interval, a time associated with enhanced risk for postoperative cognitive dysfunction. Although there was no difference in atrophy rate by diagnosis, subjects with mild cognitive impairment suffered greater subsequent cognitive effects.
Propofol exposure to neurons during synaptogenesis results in apoptosis leading to cognitive dysfunction in adulthood. Previous work from our laboratory showed that isoflurane neurotoxicity occurs through p75 neurotrophin receptor (p75NTR) and subsequent cytoskeleton depolymerization. Given that isoflurane and propofol both suppress neuronal activity, we hypothesized that propofol also induces apoptosis in developing neurons through p75NTR.
DIV5-7 neurons were exposed to propofol (3 µM) for 6 hr and apoptosis was assessed by cleaved caspase-3 (Cl-Csp3) immunoblot and immunofluorescence microscopy. Primary neurons from p75NTR−/− mice or wild-type neurons were treated with propofol, with or without pretreatment with TAT-Pep5 (10 µM, 15 min), a specific p75NTR inhibitor. P75NTR−/− neurons were transfected for 72 h with a lentiviral vector containing the synapsin driven p75NTR gene (Syn-p75NTR) or control vector (Syn-GFP) prior to propofol. To confirm our in vitro findings, wild type mice and p75NTR−/− mice (PND5) were pre-treated with either TAT-Pep5 or TAT-ctrl followed by propofol for 6 h.
Neurons exposed to propofol showed a significant increase in Cl-Csp3, an effect attenuated by TAT-Pep5 and hydroxyfasudil. Apoptosis was significantly attenuated in p75NTR−/− neurons. In p75NTR−/− neurons transfected with Syn-p75NTR, propofol significantly increased Cl-Csp3 in comparison to Syn-GFP transfected p75NTR−/− neurons. Wild type mice exposed to propofol exhibited increased Cl-Csp3 in the hippocampus, an effect attenuated by TAT-Pep5. By contrast, propofol did not induce apoptosis in p75NTR−/− mice.
These results demonstrate that propofol induces apoptosis in developing neurons in vivo and in vitro and implicate a role for p75NTR and the downstream effector ROCK.
Exposure of rhesus macaque fetuses for 24 h, or neonates for 9 h, to ketamine anesthesia causes neuroapoptosis in the developing brain. The present study further clarifies the minimum exposure required for, and the extent and spatial distribution of, ketamine-induced neuroapoptosis in rhesus fetuses and neonates.
Ketamine was administered by intravenous infusion for 5 h to postnatal day 6 rhesus neonates, or to pregnant rhesus females at 120 days gestation (full term = 165 days). Three hours later, fetuses were delivered by caesarian section, and the fetal and neonatal brains were studied for evidence of apoptotic neurodegeneration, as determined by activated caspase-3 staining.
Both the fetal (n = 3) and neonatal (n = 4) ketamine-exposed brains had a significant increase in apoptotic profiles compared to drug-naive controls (fetal n = 4; neonatal n = 5). Loss of neurons due to ketamine exposure was 2.2 times greater in fetuses than in neonates. The pattern of neurodegeneration in fetuses was different from that in neonates, and all subjects exposed at either age had a pattern characteristic for that age.
The developing rhesus macaque brain is sensitive to the apoptogenic action of ketamine at both a fetal and neonatal age, and exposure duration of 5 h is sufficient to induce a significant neuroapoptosis response at either age. The pattern of neurodegeneration induced by ketamine in fetuses was different from that in neonates, and loss of neurons attributable to ketamine exposure was 2.2 times greater in the fetal than neonatal brains.
Low-dose ketamine is used as analgesic for acute and chronic pain. It is metabolized in the liver to norketamine via cytochrome P450 enzymes. There are few human data on the involvement of CYP enzymes on the elimination of norketamine and its possible contribution to analgesic effect. The aim of this study was to investigate the effect of cytochrome P450 enzyme induction by rifampicin on the pharmacokinetics of S-ketamine and its major metabolite, S-norketamine, in healthy volunteers.
Twenty healthy male subjects received 20 mg/70kg/h (n = 10) or 40 mg/70kg/h (n = 10) intravenous S-ketamine for 2-h following either 5 days of oral rifampicin (once daily 600 mg) or placebo treatment. During and 3-h following drug infusion arterial plasma concentrations of S-ketamine and S-norketamine were obtained at regular intervals. The data were analyzed with a compartmental pharmacokinetic model consisting of three compartments for S-ketamine, three sequential metabolism compartments and two S-norketamine compartments using the statistical package NONMEM version VII.
Rifampcin caused a 10% and 50% reduction in the area-under-the-curve of the plasma concentrations of S-ketamine and S-norketamine, respectively. The compartmental analysis indicated a 13% and 200% increase in S-ketamine and S-norketamine elimination from their respective central compartments by rifampicin.
A novel observation is the large effect of rifampicin on S-norketamine concentrations and indicates that rifampicin induces the elimination of S-ketamine’s metabolite, S-norketamine, probably via induction of the CYP3A4 and/or CYP2B6 enzymes.
The long-lasting high affinity opioid buprenorphine has complex pharmacology including ceiling effects with respect to analgesia and respiratory depression. Plasma concentrations of the major buprenorphine metabolites norbuprenorphine, buprenorphine-3-glucuronide, and norbuprenorphine-3-glucuronide approximate or exceed those of the parent drug. Buprenorphine glucuronide metabolites pharmacology is undefined. This investigation determined binding and pharmacological activity of the two glucuronide metabolites, and in comparison with buprenorphine and norbuprenorphine.
Competitive inhibition of radioligand binding to human mu, kappa, delta opioid and nociceptin receptors was used to determine glucuronide binding affinities for these receptors. Common opiate effects were assessed in vivo in Swiss Webster mice. Antinociception was assessed using a tail-flick assay, respiratory effects were measured using unrestrained whole-body plethysmography, and sedation was assessed by inhibition of locomotion measured by open-field testing.
Buprenorphine-3-glucuronide had high affinity for human mu (Ki = 4.9±2.7 pM), delta (Ki = 270±0.4 nM), and nociceptin (Ki = 36±0.3 μM) but not kappa receptors. Norbuprenorphine-3-glucuronide had affinity for human kappa (Ki = 300±0.5 nM) and nociceptin (Ki= 18±0.2 μM) but not mu or delta receptors. At the dose tested, buprenorphine-3-glucuronide had a small antinociceptive effect. Neither glucuronide had significant effects on respiratory rate, but norbuprenorphine-3-glucuronide decreased tidal volume. Norbuprenorphine-3-glucuronide also caused sedation.
Both glucuronide metabolites of buprenorphine are biologically active at doses relevant to metabolite exposures which occur after buprenorphine. Activity of the glucuronides may contribute to the overall pharmacology of buprenorphine.
Methadone is frequently used in adult anesthesia and pain treatment. Methadone pharmacokinetics in adults are well characterized, including the perioperative period. Methadone is also used in children. There is, however, no information on methadone pharmacokinetics in children of any age. The purpose of this investigation was to determine the pharmacokinetics of intravenous methadone in children undergoing surgery. Perioperative opioid-sparing effects were also assessed.
Eligible subjects were children 5–18 yr undergoing general anesthesia and surgery, with an anticipated postoperative inpatient stay exceeding 3d. Three groups of 10–11 patients each received intravenous methadone HCl after anesthetic induction in ascending dose groups of 0.1, 0.2, and 0.3 mg/kg (up to 20 mg). Anesthetic care was not otherwise changed. Venous blood was obtained for 4d, for stereoselective determination of methadone and metabolites. Pain assessments were made each morning. Daily and total opioid consumption was determined. Perioperative opioid consumption and pain was determined in a second cohort, which was matched to age, sex, race, ethnicity, surgical procedure, and length of stay, but not receiving methadone.
The final methadone study cohort was 31 adolescents (14 ± 2 yr, range 10–18) undergoing major spine surgery for a diagnosis of scoliosis. Methadone pharmacokinetics were linear over the dose range 0.1–0.3 mg/kg. Disposition was stereoselective. Methadone administration did not dose-dependently affect postoperative pain scores, and did not dose-dependently decrease daily or total postoperative opioid consumption in spinal fusion patients.
Methadone enantiomers disposition in adolescents undergoing surgery was similar to that in healthy adults.
Etomidate is a sedative–hypnotic that is often given as a single intravenous bolus but rarely as an infusion because it suppresses adrenocortical function. Methoxycarbonyl etomidate and (R)-ethyl 1-(1-phenylethyl)-1H-pyrrole-2-carboxylate (carboetomidate) are etomidate analogs that do not produce significant adrenocortical suppression when given as a single bolus. However, the effects of continuous infusions on adrenocortical function are unknown. In this study, we compared the effects of continuous infusions of etomidate, methoxycarbonyl etomidate, and carboetomidate on adrenocortical function in a rat model.
A closed-loop system using the electroencephalographic burst suppression ratio as the feedback was used to administer continuous infusions of etomidate, methoxycarbonyl etomidate, or carboetomidate to Sprague–Dawley rats. Adrenocortical function was assessed during and after infusion by repetitively administering adrenocorticotropic hormone 1–24 and measuring serum corticosterone concentrations every 30 min.
The sedative–hypnotic doses required to maintain a 40% burst suppression ratio in the presence of isoflurane, 1%, and the rate of burst suppression ratio recovery on infusion terminationvaried(methoxycarbonyletomidate>carboetomidate > etomidate). Serum corticosterone concentrations were reduced by 85% and 56% during 30-min infusions of etomidate and methoxycarbonyl etomidate, respectively. On infusion termination, serum corticosterone concentrations recovered within 30 min with methoxycarbonyl etomidate but persisted beyond an hour with etomidate. Carboetomidate had no effect on serum corticosterone concentrations during or after continuous infusion.
Our results suggest that methoxycarbonyl etomidate and carboetomidate may have clinical utility as sedative–hypnotic maintenance agents when hemodynamic stability is desirable.
Procedural sedation is frequently performed in spontaneously breathing patients but hypnotics and opioids decrease respiratory drive and place the upper airway at risk for collapse.
In a randomized, controlled, cross-over, pharmaco-physiological study in 12 rats, we conducted acute experiments to compare breathing, and genioglossus electromyogram activity at equianesthetic concentrations of ketamine, a non-competitive N-methyl-D-aspartate receptor antagonist that combines potent analgesic with hypnotic action effects, versus propofol. In 10 chronically instrumented rats resting in a plethysmograph, we measured these variables as well as electroencephalography during 5 conditions: quiet wakefulness, non-rapid-eye-movement sleep, rapid eye movement sleep, low-dose (60mg/kg intraperitoneally) and high-dose ketamine anesthesia (125mg/kg intraperitoneally).
Ketamine anesthesia was associated with markedly increased genioglossus activity (1.5 to 5-fold higher values of genioglossus electromyogram) compared with sleep and propofol induced unconsciousness. Plethysmography revealed a respiratory stimulating effect: higher values of flow rate, respiratory rate, and duty-cycle (effective inspiratory time, 1.5-to-2-fold higher values). During wakefulness and normal sleep, the beta (f=6.51, p=0.04) electroencephalogram power spectrum was an independent predictor of genioglossus activity, indicating an association between electroencephalographic determinants of consciousness and genioglossus activity. Following ketamine administration, electroencephalogram power spectrum and genioglossus electroencephalogram was dissociated (p=0.9 for the relationship between delta/theta power spectrum and genioglossus electromyogram).
Ketamine is a respiratory stimulant that abolishes the coupling between loss-of-consciousness and upper airway dilator muscle dysfunction in a wide dose-range. Ketamine compared with propofol might help stabilize airway patency during sedation and anesthesia.
Propofol produces its major actions via γ-aminobutyric acid type A (GABAA) receptors. At low concentrations, propofol enhances agonist-stimulated GABAA receptor activity, and high propofol concentrations directly activate receptors. Etomidate produces similar effects, and there is convincing evidence that a single class of etomidate sites mediate both agonist modulation and direct GABAA receptor activation. It is unknown if the propofol binding site(s) on GABAA receptors that modulate agonist-induced activity also mediate direct activation.
GABAA α1β2γ2L receptors were heterologously expressed in Xenopus oocytes and activity was quantified using voltage clamp electrophysiology. We tested whether propofol and etomidate display the same linkage between agonist modulation and direct activation of GABAA receptors by identifying equi-efficacious drug solutions for direct activation. We then determined whether these drug solutions produce equal modulation of GABA-induced receptor activity. We also measured propofol-dependent direct activation and modulation of low GABA responses. Allosteric coagonist models similar to that established for etomidate, but with variable numbers of propofol sites, were fitted to combined data.
Solutions of 19 μM propofol and 10 μM etomidate were found to equally activate GABAA receptors. These two drug solutions also produced indistinguishable modulation of GABA-induced receptor activity. Combined electrophysiological data behaved in a manner consistent with allosteric co-agonist models with more than one propofol site. The best fit was observed when the model assumed three equivalent propofol sites.
Our results support the hypothesis that propofol, like etomidate, acts at GABAA receptor sites mediating both GABA modulation and direct activation.
Severe autonomic failure occurs in about 1 in 1000 people. Such patients are remarkable for the striking and sometimes paradoxical responses they manifest to a variety of physiologic and pharmacologic stimuli. Orthostatic hypotension is often the finding most commonly noted by physicians, but a myriad of additional and less well-understood findings also occur. These include supine hypertension, altered drug sensitivity, hyperresponsiveness of blood pressure to hypo/hyperventilation, sleep apnea and other neurological disturbances are also frequently encountered.
In this article we will review the clinical pathophysiology that underlies autonomic failure, with a particular emphasis on those aspects most relevant to the care of such patients in the perioperative settings. Strategies used by clinicians to diagnose and treat these patients, and the impact of these interventions on the pre-operative, intra-operative, and post-operative care that these patients undergo is a crucial element in the optimized management of care in these patients.
Perioperative myocardial infarction is a common and potentially fatal complication after noncardiac surgery, particular among patients with cardiovascular risk factors. β-blockers have been considered a mainstay in prevention and treatment of perioperative myocardial infarction, yet recent evidence suggests that β-blockers may have an unfavorable risk profile in this setting and the use has become controversial. What seems conspicuously absent from the current discussion is the appreciation of how much interindividual genetic variation influences the clinical response to β-blocker therapy. Genetic variation in the adrenergic signaling pathway is common, and has a major impact on adrenergic receptor function and β-blocker efficacy in other cardiovascular diseases such as heart failure and hypertension. Genetic variation in the cytochrome P450 2D6 enzyme, which is responsible for the metabolism of most β-blockers, is also important and can lead to poor metabolizing of β-blockers (potential toxicity) or their ultra-rapid degradation (decreased efficacy). Here, we review the molecular, cellular and physiologic consequences of polymorphisms in the adrenergic signaling pathway and CYP2D6 gene, and show that these are likely relevant factors influencing efficacy, safety and toxicity of β-blocker therapy in prevention and treatment of perioperative myocardial infarction.
Prescription opioid abuse is a significant concern in treating chronic pain, yet few studies examine how neuropathic pain alters the abuse liability of commonly abused prescription opioids.
Normal and spinal nerve ligated (SNL) rats were implanted with electrodes into the left ventral tegmental area (VTA). Rats were trained to lever press for intracranial electrical stimulation (VTA ICSS), and the effects of methadone, fentanyl, hydromorphone, and oxycodone on facilitation of VTA ICSS were assessed. A second group of neuropathic rats were implanted with intrathecal catheters and the effects of intrathecal clonidine, adenosine, and gabapentin on facilitation of VTA ICSS were assessed. The effects of electrical stimulation of the VTA on mechanical allodynia were assessed in SNL rats.
Responding for VTA ICSS was similar in control and SNL rats. Methadone, fentanyl, and hydromorphone were less potent in facilitating VTA ICSS in SNL rats. Oxycodone produced a significant facilitation of VTA ICSS in control (maximum shift 24.10 ± 6.19 Hz) but not SNL rats (maximum shift 16.32 ± 7.49 Hz), but also reduced maximal response rates in SNL rats. Intrathecal administration of clonidine, adenosine, and gabapentin failed to facilitate VTA ICSS in SNL rats, and electrical stimulation of the VTA did not alter mechanical allodynia following nerve injury.
The present data suggests that the positive reinforcing effects of commonly abused prescription opioids are diminished following nerve injury. Additionally, alleviation of mechanical allodynia with nonopioid analgesics does not appear to stimulate limbic dopamine pathways originating from the VTA in SNL rats.
Surgery often causes prolonged post-operative pain for which the mechanisms are unknown. Here we investigate the role of p38, a pain-associated Mitogen Activated Protein Kinase, in induction and maintenance of such pain.
Male rats were subjected to the Skin Muscle Incision Retraction procedure at the saphenous region that causes ~4 weeks of secondary tactile hyperalgesia in the ipsilateral plantar region, indicating central sensitization. Spinal cord was sectioned from L3 and L4+5 vertebral segments and stained for activated p38 (P-p38) at post-operative day 3 (POD3), just as secondary hyperalgesia develops, at POD10-12, the time of maximum hyperalgesia, and at POD35, after the resolution of hyperalgesia. Some sections were co-stained for microglia, astrocytes and neurons. Intrathecal injections of a P-p38 inhibitor, occurred at POD2 or POD9, and subsequent changes in pain monitored.
Skin Muscle Incision Retraction increased the numbers of dorsal horn P-p38 positive cells in L3 by ~3-fold and in L4+5 by ~7-fold, from POD3 to POD11-12. This increase was accompanied by a shift from microglia to neurons, resulting in a ~20-fold increase in P-p38 positive neurons in L4−5 over this time. No P-p38 was detected in astrocytes. A P-p38 inhibitor given at POD2 prevented development of secondary hypersensitivity, but when given at POD9 the same dose gave weak relief of pain for <3h.
Spinal P-p38 Mitogen Activated Protein Kinase, activated after incision-retraction, is important for the induction of prolonged pain, but despite increased levels near the time of maximum pain, its functional importance for the maintenance of pain is not great.
Synaptic receptors of the nicotinic receptor gene family are pentamers of subunits. This modular structure creates problems in studies of drug actions, related to the number of copies of a subunit which are present and their position. A separate issue concerns the mechanism of action of many anesthetics, which involves potentiation of responses to neurotransmitter. Potentiation requires an interaction between transmitter and potentiator, mediated through the target receptor. We have studied the mechanism by which neurosteroids potentiate transmitter responses, using concatemers of covalently linked subunits to control the number and position of subunits in the assembled receptor and to selectively introduce mutations into positionally defined copies of a subunit. We found that the steroid need interact with only one site to produce potentiation, that the native sites for steroid interaction have indistinguishable properties, and that steroid potentiation appears to result from a global effect on receptor function.
The γ-aminobutyric acid type A receptor is the major transmitter-gated inhibitory channel in the central nervous system. The receptor is a target for anesthetics, anticonvulsants, anxiolytics and sedatives whose actions facilitate the flow of chloride ions through the channel and enhance the inhibitory tone in the brain. Both the kinetic and structural aspects of the actions of modulators of the γ-aminobutyric acid type A receptor are of great importance to understanding the molecular mechanisms of general anesthesia. In this review, we describe the structural rearrangements that take place in the γ-aminobutyric acid type A receptor during channel activation and modulation, focusing on data obtained using voltage-clamp fluorometry. Voltage-clamp fluorometry entails the binding of an environmentally-sensitive fluorophore molecule to a site of interest in the receptor, and measurement of changes in the fluorescence signal resulting from activation- or modulation-elicited structural changes. Detailed investigations can provide a map of structural changes that underlie or accompany the functional effects of modulators.
Protein kinase C (PKC) is a family of serine/threonine kinases that contains more than 10 isozymes. Evidence suggests that PKC may play important roles in pain modulation, but the isozyme-specific effects of PKC on different aspects of pain modulation are not fully understood. We hypothesize that different PKC isozymes play different roles in different aspects of pain modulation.
The nociceptive behaviors of mice with deletion of PKC α, β, γ, or δ in multiple pain models were compared with their respective wild type littermates. Also, the morphine analgesia and the development of morphine tolerance in mice with deletion of PKC γ were compared with their respective wild type littermates.
Thermal hyperalgesia induced by complete Freund’s adjuvant injection was significantly attenuated by the deletion of PKC β, γ or δ, but not PKC α. Deletion of PKC γ significantly attenuated neuropathic mechanical allodynia induced by spared nerve injury, whereas deletion of PKC α enhanced this allodynia. Baseline thermal and mechanical sensitivity, nociceptive behaviors induced by formalin, mechanical allodynia induced by complete Freund’s adjuvant injection, were not altered by deletion of PKC α, β, γ or δ. Finally, morphine analgesia and the development of morphine tolerance were not altered in PKC γ-deficient mice.
PKC plays isozyme-specific effects in pain modulation.
The metabotropic glutamate receptor 5 noncompetitive antagonist fenobam is analgesic in rodents. Future development of fenobam as an analgesic in humans will require a favorable long-term treatment profile and a lack of significant deleterious side effects. This study aimed to determine if tolerance to fenobam’s analgesic effects developed over 14 days and to assess for side effects in mice.
Mouse models of pain, locomotor behavior, and coordination were used. Fenobam or vehicle (n = 8 or 11 per group) was administered for 14 days and analgesic tolerance to fenobam was assessed using the formalin test. Histopathology examination and serum chemistry analysis post-14-day fenobam administration were also assessed (n = 12 or 9). The effects of fenobam on locomotor activity were assessed in the open field and elevated zero maze (n = 8 or 7). Coordination was assessed using ledge crossing and vertical pole descent tasks (n = 11 or 10).
Tolerance to fenobam’s analgesic effect did not develop after 14 days. Chronic fenobam administration resulted in statistically significantly less weight gain compared to vehicle controls, but did not cause any physiologically or statistically significant hematological abnormalities, altered organ function, or abnormal histopathology of the liver, brain, or testes. Fenobam administration resulted in a metabotropic glutamate receptor 5-dependent increase in exploratory behavior but does not impair motor coordination at analgesic doses.
Analgesic tolerance to repeat fenobam dosing does not develop. Chronic dosing of up to 14-days is well tolerated. Fenobam represents a promising candidate for the treatment of human pain conditions.
Volatile general anesthetics inhibit neurotransmitter release by a mechanism not fully understood. Genetic evidence in C. elegans has shown that a major mechanism of action of volatile anesthetics acting at clinical concentrations in this animal is presynaptic inhibition of neurotransmission. To define additional components of this presynaptic volatile anesthetic mechanism, C. elegans mutants isolated as phenotypic suppressors of a mutation in syntaxin, an essential component of the neurotransmitter release machinery, were screened for anesthetic sensitivity phenotypes.
Sensitivity to isoflurane concentrations was measured in locomotion assays on adult C. elegans. Sensitivity to the acetylcholinesterase inhibitor aldicarb was used as an assay for the global level of C. elegans acetylcholine release. Comparisons of isoflurane sensitivity (measured by the EC50) were made by simultaneous curve-fitting and F-test.
Among the syntaxin suppressor mutants, js127 was the most isoflurane resistant with an EC50 more than three-fold wild type. Genetic mapping, sequencing, and transformation phenocopy showed that js127 was an allele of acy-1, which encodes an adenylate cyclase expressed throughout the C. elegans nervous system and in muscle. js127 behaved as a gain-of-function mutation in acy-1 and had increased levels of cyclic adenosine monophosphate. Testing of single and double mutants along with selective tissue expression of the js127 mutation revealed that acy-1 acts in neurons within a G□s – PKA – UNC-13-dependent pathway to regulate behaviour and isoflurane sensitivity.
Activation of neuronal adenylate cyclase antagonizes isoflurane inhibition of locomotion in C. elegans.
A pharmacogenomic approach was used to further localize the genetic region responsible for previously observed enhanced cardiovascular sensitivity to propofol in Dahl Salt Sensitive (SS) vs. control Brown Norway (BN) rats.
Propofol infusion levels that decreased blood pressure by 50% were measured in BN.13SS rats (substitution of SS chromosome 13 into BN) and in 5 congenic (partial substitution) strains of SS.13BN. The effect of superfused 2,6 diisopropylphenol on small mesenteric arterial vascular smooth muscle transmembrane potential was measured in congenic strains before and during superfusion with Rp-cAMPS and Rp-8-pCPT-cGMPS, inhibitors of protein kinase A and G respectively. The genetic locus and potential role of the renin gene in mediating VSM sensitivity to propofol were determined in three selected sub-congenic SS.BN13 strains.
A 30 – 32% smaller propofol infusion rate reduced blood pressure by 50% in BN.13SS compared to BN and the SS.13BN congenic containing a 80 BN gene substitution. Compared to the latter, SS exhibited greater protein kinase A dependent vascular smooth muscle hyperpolarization in response to propofol. Using sub-congenics, the increased propofol-induced cardiovascular sensitivity and hyperpolarization was further localized to an 8-gene region (containing the BN renin gene). Blockade of angiotensin (AT1) receptors with losartan in this sub-congenic, elevated propofol-induced hyperpolarization by 3 fold, to that observed in SS.
Enhanced cardiovascular sensitivity to propofol in SS (compared to BN) is caused by an altered renin gene. Through modified second messenger function, this differentially regulates VSM contractile state and reduces vascular tone exacerbating cardiovascular depression by propofol.
Anesthesiologists are increasingly confronting the difficult problem of management of patients with sepsis both in the operating room and in the intensive care unit. Sepsis occurs in over 750,000 patients in the United States annually and is responsible for more than 210,000 deaths. Approximately 40% of all intensive care unit patients either have sepsis on admission to the intensive care unit or develop sepsis during their stay in the intensive care unit. There have been significant advances in both understanding of the pathophysiology of the disorder as well as in therapy. Although deaths due to sepsis remain stubbornly high, new treatment algorithms have lead to a reduction in overall mortality. Thus, it is important for anesthesiologists and critical care practitioners to be aware of these new therapeutic regimens. The goal of this review is to include both practical points on important advances in current therapy of sepsis as well as provide a vision of future new immunotherapeutic approaches.
Low tidal volumes have been associated with improved outcomes in patients with established acute lung injury. The role of low tidal volume ventilation in patients without lung injury is still unresolved. We hypothesized that such a strategy in patients undergoing elective surgery would reduce ventilator-associated lung injury and that this improvement would lead to a shortened time to extubation
A single-center randomized controlled trial was undertaken in 149 patients undergoing elective cardiac surgery. Ventilation with 6 versus 10 ml/kg tidal volume was compared. Ventilator settings were applied immediately after anesthesia induction and continued throughout surgery and the subsequent intensive care unit stay. The primary endpoint of the study was time to extubation. Secondary endpoints included the proportion of patients extubated at 6 h and indices of lung mechanics and gas exchange as well as patient clinical outcomes.
Median ventilation time was not significantly different in the low tidal volume group; a median (interquartile range) of 450 (264–1,044) min was achieved compared with 643 (417–1,032) min in the control group (P = 0.10). However, a higher proportion of patients in the low tidal volume group was free of any ventilation at 6 h: 37.3% compared with 20.3% in the control group (P = 0.02). In addition, fewer patients in the low tidal volume group required rein-tubation (1.3 vs. 9.5%; P = 0.03).
Although reduction of tidal volume in mechanically ventilated patients undergoing elective cardiac surgery did not significantly shorten time to extubation, several improvements were observed in secondary outcomes. When these data are combined with a lack of observed complications, a strategy of reduced tidal volume could still be beneficial in this patient population.