Centre for Anaesthesia and Pain Management Research, University of Sydney at Royal North Shore Hospital, St Leonards, NSW 2065, Australia*Corresponding author
Accepted for publication: August 31, 2001
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Abstract |
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Methods. Group 1 animals were infused with i.v. ketamine for 5 min; in group 2, constant low plasma concentrations of alfentanil were maintained by computer-controlled infusion; in group 3, the treatments were combined. Serial plasma and terminal tissue concentrations were measured by high performance liquid chromatography or gas chromatography-mass spectrometry.
Results. In the presence of alfentanil, the mean plasma ketamine concentrationtime area under the curve (AUC) value was significantly lower (by 13%, P<0.05), while clearance (ClT) and volume of distribution (VSS) were significantly higher (by 16 and 28%, respectively, both P<0.05). Tissue:plasma distribution coefficients for ketamine in the presence of alfentanil were significantly higher in forebrain (by 128%, P<0.005), hindbrain (by 207%, P<0.01), gut (by 254%, P<0.005), and fat (by 344%, P<0.0001). Mean AUC values for alfentanil did not differ significantly in the presence of ketamine, but alfentanil tissue concentrations were significantly lower in forebrain (by 77%, P<0.0001), hindbrain (by 28%, P<0.01), heart (by 33%, P<0.01), lung (30%, P<0.05), and gut (by 21%, P<0.05). Corresponding tissue:plasma distribution coefficients were significantly lower for forebrain (by 69%, P<0.0001) alone.
Conclusions. The finding that the distribution of ketamine into the brain was increased by low plasma concentrations of alfentanil could have important clinical applications for pain management.
Br J Anaesth 2002; 88: 94100
Keywords: anaesthetics i.v., ketamine; analgesics opioid, alfentanil; pharmacokinetics, tissue uptake; pain, management; rat
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Introduction |
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This study was designed to determine the whole body pharmacokinetics and tissue distribution of ketamine and of its pharmacologically active metabolite, norketamine, when administered alone and in the presence of constant low-level plasma alfentanil concentrations, in rats.
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Materials and methods |
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Drugs
Ketamine hydrochloride (KetalarTM, Parke-Davis Australia Pty Ltd, Sydney, Australia) was diluted to a concentration of 20 mg ml1 (17.3 mg ml1 as base); alfentanil hydrochloride (RapifenTM, Astra-Zeneca Australia Pty Ltd, Sydney, Australia) was diluted to a concentration of 100 µg ml1 (92 µg ml1 as base). Both drug solutions were diluted with de-ionized water; alfentanil contained 5 units ml1 heparin.
Experimental design
Studies were performed in three groups of animals 24 h after cannulation, and were carried out between 14:00 and 19:00 h. Infusion and sampling lines (75 and 45 cm, respectively) were attached to the venous and arterial indwelling cannulae, and the animals were placed in a study chamber and allowed to settle for 30 min before commencement of the study. The relevant drug infusions (see below) were delivered by a syringe driver (Harvard Apparatus, Model 22) from a gas tight syringe (Hamilton, 5 ml). In groups 1 and 3 (described below), immediately after loss of righting reflex, the animals were removed from the study chamber, a rectal thermal probe was inserted, and body temperature was maintained by a heating lamp; no loss of righting reflex occurred in group 2. Once motor function began to return the rectal probe was removed and the animals were returned to the study chamber. Arterial blood samples (100 µl) were collected into heparinized (50 units) 1.5 ml polyethylene microfuge tubes for drug analysis; these were replaced with 3 volumes of 0.9% saline as described previously.3 After the final blood sample was collected, the animals were killed by carbon dioxide asphyxiation. Central nervous system (CNS=regional brain and spinal cord), heart, lung, liver, kidney, gut, adductor muscle, and epididymal fat pad tissues were sampled for the determination of tissue:plasma drug distribution. The tissue: plasma distribution coefficients of ketamine and alfentanil were determined after each of the following infusion paradigms.
Group 1. An infusion of ketamine was administered at a constant rate of 10 mg kg1 min1 over 5 min; serial arterial blood samples were taken before, then at 1.5, 3, 5, 7, 10, 15, 20, 30, 40, 60, 80, 100, 120, 140, 160, and 180 min after commencement of the infusion.
Group 2. An infusion of alfentanil, computer-controlled by Stanpump software,4 was used to maintain a constant plasma concentration of alfentanil (target value of 100 µg ml1) that would be antinociceptive, without causing significant ventilatory impairment or EEG changes, based on previous animal experiments.58 Pharmacokinetic parameters, describing a three-compartment mamillary model, were obtained from previously published data9 and used to implement the infusion algorithm. Serial arterial blood samples were collected before, then at 1.5, 3, 5, 10, 20, 40, 60, 80, 100, 120, 140, 160, and 180 min for the duration of the infusion.
Group 3. An infusion of ketamine was administered over 5 min as in group 1, the infusion line for ketamine was then removed and a new infusion line primed with alfentanil was attached; 7 min later (i.e. at 12 min after the start of the ketamine infusion), alfentanil was delivered by the Stanpump infusion paradigm as in group 2. Blood samples were collected as before in group 1: the first blood sample to contain alfentanil was taken at 15 min (i.e. 3 min after commencement of the alfentanil infusion). Ketamine and alfentanil plasma concentrations were measured in the same blood samples.
Analytical procedures
In group 1, concentrations of ketamine and norketamine were measured as the sum of individual R()- and S(+)-ketamine enantiomer concentrations determined by HPLC following chiral separation on a micro chiral-AGP column.10 This method was not suitable for the concurrent determination of ketamine and alfentanil in groups 2 and 3, due to marked differences in their chromatographic characteristics on this column type and inadequate sensitivity for the determination of alfentanil concentrations. In these groups the plasma and tissue concentrations of alfentanil, ketamine, and norketamine were determined by adaptation of previously published gas chromatography-mass spectrometry (GC-MS) procedures,11 12 as described below.
Plasma. After the addition of cyproheptadine (50 µl, 1 µg ml1, internal standard) and Na3PO4 (100 µl, 0.5 M), aliquots of plasma (50 µl) in polypropylene tubes (1.5 ml) were briefly vortex-mixed, then extracted by shaking (5 min) with cyclohexane containing iso-amyl alcohol (1 ml, 2% v.v1). Tubes were centrifuged (3000 r.p.m., 5 min) and the organic phase transferred to a fresh polypropylene tube (1.5 ml) with a Pasteur pipette, dried in a bench rotatory vacuum centrifuge, reconstituted in toluene (100 µl), and transferred to polypropylene inserts immediately before analysis.
Tissues. Homogenates of tissue (100 mg ml1) in NaH2PO4 (100 mM) were frozen and resuspended after thawing immediately before analysis. Aliquots of homogenate (1 ml) were transferred to polypropylene tubes (10 ml), before the addition of cyproheptadine (100 µl, 1 µg ml1, internal standard) and NaOH (250 µl, 1 M). The tubes were briefly vortex-mixed, then extracted by shaking (5 min) with cyclohexane containing iso-amyl alcohol (3 ml, 2% v.v1). After centrifugation (3000 r.p.m., 5 min), the organic phase was transferred to a fresh polypropylene tube (10 ml) with a Pasteur pipette, and HCl (400 µl, 0.5 M) was added. Tubes were shaken (5 min) and centrifuged as before, the organic phase was decanted and discarded, and NaOH (60 µl, 5 M) was added. After brief vortex mixing, cyclohexane containing iso-amyl alcohol (1 ml, 2% v.v1) was added and the tubes shaken and centrifuged as before. The organic phase was transferred to a polypropylene tube (1.5 ml) with a Pasteur pipette, dried in a bench rotatory vacuum centrifuge, reconstituted in toluene (100 µl), and transferred to polypropylene inserts immediately before analysis.
GC-MS methodology. Toluene extracts were analysed on a Hewlett Packard 5972 MS/5890 Group II GC system, operating in electron impact mode (70 eV), after split-less injection onto a capillary column (HP-5MS, 30 m length x 0.25 mm ID, 0.25-µm film thickness). The oven temperature was held at 110°C initially for 0.5 min then increased at 25°C min1 to 280°C, while the injector and transfer lines were maintained at 280°C. Helium was used as the carrier gas at a flow rate of 1 ml min1. A run time of 10 min and an injection volume of 1 µl were used for the analysis of ketamine and norketamine, while for alfentanil a run time of 15 min and an injection volume of 5 µl was used. Mass fragments (and retention times) for ketamine and norketamine were monitored with a dwell time of 50 ms at m/z 180 (6.1 min) and m/z 166 (6.0 min), respectively; alfentanil and cyproheptadine were monitored with a dwell time of 100 ms at m/z 289 (12.7 min) and m/z 287 (8.2 min), respectively.
Data analysis
Area under the curve (AUC) values for each plasma concentrationtime data set were determined by the linear trapezoid method. Distribution coefficients were calculated as the ratio of tissue to plasma analyte concentrations, uncorrected for residual tissue blood volume. Between group comparisons of the tissue distribution coefficients for ketamine and norketamine were performed on log-transformed values, and the antilogs of the mean log-transformed values were subsequently referred to as the log-normalized values of the distribution coefficients. Regional CNS tissue drug concentrations and distribution coefficients were compared by one-way analysis of variance (ANOVA); individual post-hoc comparisons between mean values were performed by the method of Least Significant Difference (Statistix for Windows®, Analytical Software, Tallahassee, FL, USA). Tissue drug concentrations and distribution coefficients between the two infusion paradigms were compared by Students two sample t-tests. The pharmacokinetic parameters for ketamine were determined by fitting polyexponential decay equations to the concentrationtime washout curve, and the mean total body clearance (ClT), initial dilution volume (VC), volume of distribution at steady state (VSS), terminal half-life (t1/2), and mean residence time in the body (MRT) were calculated by conventional means.13 Data are expressed as mean (SEM) unless specified otherwise.
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Results |
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Alfentanil arterial plasma concentrations from the Stanpump-controlled infusions in group 2 and 3 animals are shown in Figure 024F2. In group 2 animals, where alfentanil infusion was given alone, the animals displayed episodic sleep-like behaviour, but did not demonstrate other potential alfentanil-induced effects such as loss of righting reflex, ataxia, increased muscle tone, or convulsions. The mean AUC value from 20 to 180 min for alfentanil was not significantly different between the infusions in group 2 (7625 (134) ng min ml1) and group 3 (7194 (634) ng min ml1), but plasma alfentanil concentrations were significantly lower (P=0.0012) at 20 min in group 3. Tissue concentrations of alfentanil at 180 min following infusions in groups 2 and 3 are shown in Table 4. Significant regional differences in CNS alfentanil concentrations were apparent in both group 2 (F2,21=22.76, P<0.0001; forebrain > hindbrain and spinal cord: P=0.001) and group 3 (F2,21=6.48, P=0.0064; forebrain < hindbrain and spinal cord: P=0.02). Concentrations of alfentanil were significantly lower in forebrain (P<0.0001) and hindbrain (P<0.01), when alfentanil infusion followed ketamine administration. Similarly, there were significant differences in the corresponding regional CNS distribution coefficients following alfentanil infusions in both group 2 (F2,21=19.28, P<0.0001; forebrain > hindbrain and spinal cord: P=0.001) and group 3 (F2,21=4.45, P=0.024; forebrain < hindbrain and spinal cord: P=0.05). Alfentanil distribution coefficient for the forebrain was significantly lower (P<0.0001) when alfentanil infusion followed ketamine administration. Although concentrations of alfentanil were significantly lower in heart (P<0.01), lung (P<0.0001), and gut (P<0.0001) following infusions in group 3, the corresponding distribution coefficients did not differ significantly.
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Discussion |
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Alfentanil has a very short duration of action following a brief duration of infusion, mainly due to its weak tissue binding.19 Although there is evidence for alfentanil to exert a longer half-life and duration of effect with longer durations of infusion,2022 it is well-suited to i.v. infusion delivery, especially for intraoperative analgesia in surgical patients. Opioids such as alfentanil reproducibly modify the quantitative electroencephalogram (EEG), especially in the delta (0.54.5 Hz) frequency band. Alfentanil plasma concentrations used in the present study were in the range reported as having little to no direct EEG effect. Other studies on antinociceptive effects of alfentanil have found an EC50 of 24 ng ml1 for suppression of tooth pulp evoked potential (TPEP) changes in the EEG.8 Thus, the present study used alfentanil plasma concentrations within the antinociceptive range, but lower than values associated with significant direct EEG changes (EC50 approximately 200 ng ml1).5 7
Mean values for ketamine ClT and VSS (Table 1) were consistent with values reported by others;23 however, the significantly lower plasma ketamine concentrationtime AUC values for group 3 compared with group 1 animals reflect the greater ClT and VSS for ketamine during alfentanil infusion. Arterial blood pH and PCO2 were not measured in this study but, for several reasons, we believe that a ventilatory depressant effect of alfentanil was unlikely to account for this difference. There were no obvious effects of alfentanil in animals when alfentanil infusions were given alone. Plasma alfentanil concentrations less than 100 ng ml1 have previously been shown to have no effect on arterial PCO2.8 Moreover, as the pKa of ketamine is 7.5,24 a decreased arterial blood pH due to ventilatory depression would increase the ionized state fraction of ketamine, thereby retarding its extravascular distribution and resulting in lower values for VSS and possibly ClT. Similarly, cardiovascular differences between the two groups are also unlikely to account for the different pharmacokinetic values for ketamine in the presence of alfentanil. Attenuation by alfentanil of ketamines sympathomimetic-cardiovascular effects could also be expected to retard distribution and reduce VSS and possibly ClT. Both the VSS and ClT of ketamine; however, were significantly increased in the presence of alfentanil.
Alfentanil appeared to facilitate the uptake of ketamine into both gut and adipose tissue. Opioids tend to reduce sympathetic tone and enhance parasympathetic tone. These effects, along with a possible direct vascular effect, can result in dilation of arterial and venous vascular beds,25 and possibly account for the greater uptake into gut and adipose tissue, and the higher clearance, found for ketamine in the presence of alfentanil infusion. In the lung, however, tissue uptake of ketamine was less suggesting that, following i.v. infusion of alfentanil, high local concentrations of alfentanil may have displaced ketamine from tissue binding sites. This suggestion is consistent with the known higher affinity of alfentanil than ketamine for plasma proteins,23 the high degree of ketamine uptake into the lungs,26 and saturability of binding of such basic drugs in lung tissue.27
Tissue norketamine concentrations were significantly higher in liver (52%), kidney (40%), and gut (53%), when ketamine infusion was followed by alfentanil. The lack of significant difference in the corresponding distribution coefficients for norketamine in these tissues most likely reflects significant local metabolism of ketamine to norketamine in these tissues.10 Significantly higher concentrations of norketamine in adipose tissue, however, probably reflects redistribution from the gut, as local metabolic conversion of ketamine to norketamine would be unlikely. However, the mean AUC value for norketamine was significantly higher (by 25%) in group 1 than group 3 due, mainly, to significant divergence in plasma norketamine concentrations from 7 (to 90) min, 2 min after cessation of the ketamine infusion (alfentanil infusions in group 3 did not begin until 12 min after the ketamine infusion had commenced). Norketamine in group 1 samples was quantified by spectrophotometric detection following chiral stationary phase HPLC separation, but in group 3 norketamine was quantified by the more specific technique of mass-selective detection following gas chromatographic separation. Norketamine, after its formation from ketamine by N-demethylation, may undergo further oxidative metabolism to 6-hydroxynorketamine.28 It is possible that such an additional metabolite contributed to the AUC values for plasma norketamine in group 1, because norketamine concentrations in liver, kidney, and gut (the tissues that contributed to circulating norketamine concentrations) were in fact higher in group 3.
Plasma alfentanil concentrations were significantly lower at 20 min when Stanpump-controlled alfentanil infusion followed ketamine infusion. Ketamine concentrations in plasma were three orders of magnitude higher than those of alfentanil, and competitive displacement from plasma protein binding sites may have facilitated tissue uptake in the early stages of the alfentanil infusion.23 The subsequent redistribution of ketamine into more poorly perfused tissues appeared to offset the initial facilitated tissue uptake of alfentanil following ketamine infusion. Although tissue concentrations of alfentanil were significantly lower in heart, lung, and gut, when alfentanil infusion followed ketamine administration, the distribution coefficients indicated that tissue uptake of alfentanil by these tissues was not affected by ketamine infusion.
In the CNS, tissue uptake of ketamine into both the forebrain and hindbrain was significantly greater in the presence of alfentanil, although ketamine significantly reduced tissue uptake of alfentanil into the forebrain, possibly suggesting local competitive displacement of alfentanil by ketamine from tissue binding sites. Recently, ketamine has been found to exert analgesic effects supraspinally, through activation of inhibitory monoaminergic pathways descending to the spinal cord, and antihyperalgesic effects at the level of the spinal cord through inhibition of NMDA receptor function.29 Moreover, opioids also exert analgesic effects through activation of monoaminergic pathways that originate in the brain stem and descend to the spinal cord.30 Plasma alfentanil concentrations of approximately 50 ng ml1, maintained over the duration of this investigation, were approximately double the EC50 found for suppression of TPEP8 and therefore likely to be associated with analgesia. Others have reported that ketamine allays the development of antinociceptive tolerance to alfentanil.31 This finding that the distribution of ketamine into the brain was increased by constant low plasma concentrations of alfentanil could therefore have implications for the interpretation of such pharmacological studies and have important clinical applications for pain management.
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Acknowledgements |
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References |
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2 Persson J, Scheinin H, Hellstrom G, Bjorkman S, Gotharson E, Gustafsson LL. Ketamine antagonises alfentanil-induced hypoventilation in healthy male volunteers. Acta Anaesthesiol Scand 1999; 43: 74452[ISI][Medline]
3 Mather LE, Edwards SR, Duke CC. Electroencephalographic effects of thiopentone and its enantiomers in the rat. Life Sci 2000; 66: 105114[ISI][Medline]
4 Shafer SL, Gregg KM. Algorithms to rapidly achieve and maintain stable drug concentrations at the site of drug effect with a computer-controlled infusion pump. J Pharmacokin Biopharm 1992; 20: 14769[ISI][Medline]
5 Cox EH, Jeanine GN, Van Hemert JG, Tukker EJ, Danhof M. Pharmacokinetic-pharmacodynamic modelling of the EEG effect of alfentanil in rats. J Pharmacol Toxicol Methods 1997; 38: 99108[ISI][Medline]
6 Cox EH, Kerbusch T, Van der Graaf PH, Danhof M. Pharmaco kinetic-pharmacodynamic modeling of the electroencephalogram effect of synthetic opioids in the rat: correlation with the interaction at the mu-opioid receptor. J Pharmacol Exp Ther 1998; 284: 1095103
7 Cox EH, Kuipers JA, Danhof M. Pharmacokinetic-pharmacodynamic modelling of the EEG effect of alfentanil in rats: assessment of rapid functional adaptation. Br J Pharmacolol 1998; 124: 153440[Abstract]
8 Cox EH, Langemeijer MWE, Danhof M. Pharmacokinetic-pharmacodynamic modelling of the analgesic effect of alfentanil in the rat using tooth pulp evoked potentials. J Pharmacol Toxicol Methods 1998; 39: 1927[ISI][Medline]
9 Mandema JW, Wada DR. Pharmacodynamic model for acute tolerance development to the electroencephalographic effects of alfentanil in the rat. J Pharmacol Exp Ther 1995; 275: 118593[Abstract]
10 Edwards SR, Mather LE. Tissue uptake of ketamine and norketamine enantiomers in the rat: indirect evidence for extahepatic metabolic inversion. Life Sci 2001; 69: 205166[ISI][Medline]
11 Feng N, Vollenweider FX, Minder EI, Rentsch K, Grampp T, Vonderschmitt DJ. Development of a gas chromatography-mass spectrometry method for determination of ketamine in plasma and its application to human samples. Ther Drug Monit 1995; 17: 95100[ISI][Medline]
12 Mautz DS, Labroo R, Kharasch ED. Determination of alfentanil and noralfentanil in human plasma by gas chromatography-mass spectrometry. J Chromatogr B 1994; 658: 14953[ISI]
13 Sebaldt RJ, Kreeft JH. Efficient pharmacokinetic modeling of complex clinical dosing regimens: the universal elementary dosing regimen and computer algorithm EDFAST. J Pharm Sci 1987; 76: 93100[ISI][Medline]
14 Church J, Lodge D. N-methyl-D-aspartate antagonism is central to the actions of ketamine and other phencyclidine receptor ligands. In: Domino EF, ed. Status of Ketamine in Anesthesiology. Ann Arbor: NPP Books, 1990; 50119
15 Maurset A, Skoglund LA, Hustveit O, Oye I. Comparison of ketamine and pethidine in experimental and postoperative pain. Pain 1989; 36: 3741[ISI][Medline]
16 Oye I, Paulsen O, Maurset A. Effects of ketamine on sensory perception: evidence for a role of NMDA receptors. J Pharmacol Exp Ther 1992; 260: 120913[Abstract]
17 Shimoyama M, Shimoyama N, Gorman AL, Elliott KJ, Inturrisi CE. Oral ketamine is antinociceptive in the rat formalin test: role of the metabolite, norketamine. Pain 1999; 81: 8593[ISI][Medline]
18 Ebert B, Mikkelsen S, Thorkildsen C, Borgbjerg FM. Norketamine, the main metabolite of ketamine, is a non-competitive NMDA receptor antagonist in the rat cortex and spinal cord. Eur J Pharmacol 1997; 333: 99104[ISI][Medline]
19 Mather LE. Clinical pharmacokinetics of fentanyl and its newer derivatives. Clin Pharmacokinet 1983; 8: 42246[ISI][Medline]
20 Hughes MA, Glass PS, Jacobs JR, Context-sensitive half-time in multicompartment pharmacokinetic models for intravenous anesthetic drugs. Anesthesiology 1992; 76: 33441[ISI][Medline]
21 Kapila A, Glass PS, Jacobs JR, et al. Measured context-sensitive half-times of remifentanil and alfentanil. Anesthesiology 1995; 83: 96875[ISI][Medline]
22 Castor G, Altmayer P, Venitz J, Einfluss der Infusionsdauer auf das Eliminationsverhalten von Alfentanil. [Effect of duration of infusion on elimination rate of alfentanil.] Anaesthesiol Reanim 1995; 20: 97100[Medline]
23 Björkman S, Redke F. Clearance of fentanyl, alfentanil, methohexitone, thiopentone, and ketamine in relation to estimated hepatic blood flow in several animal species: application to prediction of clearance in man. J Pharm Pharmacol 2000; 52: 106574[ISI][Medline]
24 White PF. Ketamine-its pharmacology and therapeutic uses. Anesthesiology 1982; 56: 119136.[ISI][Medline]
25 Bailey PL. Opioids in operative anesthesia. In: Herz A, ed. Handbook of Experimental Pharmacology Vol. 104(II): Opioids II. New York: Springer, 1993; 74558.
26 Shiue CY, Vallabhahosula S, Wolf AP, et al. Carbon-11 labelled ketamine-synthesis, distribution in mice and PET studies in baboons. Nuclear Med Biol 1997; 24: 14550[ISI]
27 Boer F, Engbers FH, Bovill JG, Burm AG, Hak A. First-pass pulmonary retention of sufentanil at three different background blood concentrations of the opioid. Br J Anaesth 1995; 74: 505
28 Woolf TF, Adams JD. Biotransformations of ketamine, (Z)-6-hydroxyketamine, and (E)-6-hydroxyketamine by rat, rabbit, and human liver microsomal preparations. Xenobiotica 1987; 17: 83947[ISI][Medline]
29 Kawamata T, Omote K, Sonoda H, Kawamata M, Namiki A. Analgesic mechanisms of ketamine in the presence and absence of peripheral inflammation. Anesthesiology 2000; 93: 5208[ISI][Medline]
30 Fields HL. Brainstem mechanisms of pain modulation: anatomy and physiology. In: Herz A, ed. Handbook of Experimental Pharmacology Vol. 104(II): Opioids II. New York: Springer, 1993; 320
31 Kissin I, Bright CA, Bradley EL jr. The effect of ketamine on opioid-induced acute tolerance: can it explain reduction of opioid consumption with ketamine-opioid analgesic combinations? Anesth Analg 2000; 91: 14838