Ventilatory responses of healthy subjects to intravenous combinations of morphine and oxycodone under imposed hypercapnic and hypoxaemic conditions.

AIMS: Previous isobolographic analysis revealed that coadministration of morphine and oxycodone produces synergistic antinociception in laboratory rodents. As both opioids can produce ventilatory depression, this study was designed to determine whether their ventilatory effects were synergistic when coadministered to healthy human subjects. METHODS: A placebo-controlled, randomized, crossover study was performed in 12 male volunteers. Ventilatory responses to hypoxaemia and hypercapnia were determined from 1-h intravenous infusions of saline ('placebo'), 15 mg morphine sulphate (M), 15 mg oxycodone hydrochloride (O), and their combination in the dose ratios of 1:2, 1:1, 2:1. Drug and metabolite concentrations in serial peripheral venous blood samples were measured by high-performance liquid chromatography-MS/MS. RESULTS: 'Placebo' treatment was without significant ventilatory effects. There were no systematic differences between active drug treatments on either the slopes or intercepts of the hypoxaemic and hypercapnia ventilation responses. During drug treatment, the mean minute ventilation at PetCO(2) = 55 mmHg (V(E55)) decreased to 74% of the subjects' before treatment values (95% confidence interval 62, 87), 68% (57, 80), 69% (59, 79), 68% (63, 73), and 61% (52, 69) for M15, M10/O5, M7.5/O7.5, M5/O10 and O15, respectively. Recovery was more prolonged with increasing oxycodone doses, corresponding to its greater potency and lower clearance compared with morphine. CONCLUSIONS: Although adverse ventilatory effects of these drugs were found as expected, no unexpected or disproportionate effects of any of the morphine and oxycodone treatments were found that might impede their use in combination for pain management.

Hepatic bipolar radiofrequency ablation creates coagulation zones close to blood vessels: a finite element study.

Radiofrequency (RF) ablation has become an important means of treatment of non-resectable primary and metastatic liver tumours. Recurrence of treated tumours is associated with cancer cell survival next to blood vessels. The paper examines the performance of classical monopolar, and two configurations of bipolar, RF ablation using a LeVeen ten-prong catheter. Finite element method models of monopolar and bipolar configurations were created at 5 mm distance from a vessel of the size of a typical portal vein (10 mm diameter). In one bipolar configuration, the probes were oriented in the same axial direction (asymmetric configuration); in the second bipolar configuration, the two probes were facing each other (symmetric configuration). The distribution of temperature and current density was analysed for three different flow conditions: normal flow, reduced flow due to portal hypertension and high flow. For normal flow, the distance between the formed coagulation zone and the blood vessel was 1.8 mm for monopolar, 1 mm for asymmetric bipolar, and 0.2 mm for symmetric bipolar, configurations. Symmetric bipolar RF ablation creates coagulation zones significantly closer to blood vessels compared with monopolar RF ablation. This may reduce tumour cell survival next to blood vessels and reduce recurrence rates.

Hydromorphone-3-glucuronide: a more potent neuro-excitant than its structural analogue, morphine-3-glucuronide.

In humans, hydromorphone (HMOR) is metabolised principally by conjugation with glucuronic acid to form hydromorphone-3-glucuronide (H3G), a close structural analogue of morphine-3-glucuronide (M3G), the major metabolite of morphine. In a previous study we described the biochemical synthesis of H3G together with a preliminary evaluation of its pharmacology which revealed that it is a neuroexcitant in rats in a manner analogous to M3G. Thus the aims of the current study were to quantify the neuro-excitatory behaviours evoked by intracerebroventricular (icv) H3G in the rat and to define its potency relative to M3G. Groups of adult male Sprague-Dawley rats received icv injections (1 microL) of H3G (1 - 3 microg), M3G (2 - 7 microg) or vehicle via a stainless steel guide cannula that had been implanted stereotaxically seven days prior to drug administration. Behavioural excitation was monitored by scoring fifteen different behaviours (myoclonic jerks, chewing, wet-dog-shakes, rearing, tonic-clonic-convulsions, explosive motor behaviour, grooming, exploring, general activity, eating, staring, ataxia, righting reflex, body posture, touch evoked agitation) immediately prior to icv injection and at the following post-dosing times: 5, 15, 25, 35, 50, 65 and 80 min. H3G produced dose-dependent behavioural excitation in a manner analogous to that reported previously for M3G by our laboratory and reproduced herein. H3G was found to be approximately 2.5-fold more potent than M3G, such that the mean (+/- S.D.) ED50 values were 2.3 (+/- 0.1) microg and 6.1 (+/- 0.6) microg respectively. Thus, our data clearly imply that if H3G crosses the BBB with equivalent efficiency to M3G, then the myoclonus, allodynia and seizures observed in some patients dosed chronically with large systemic doses of HMOR, are almost certainly due to the accumulation of sufficient H3G in the central nervous system, to evoke behavioural excitation.

Sex-related differences in antinociception and tolerance development following chronic intravenous infusion of morphine in the rat: modulatory role of testosterone via morphine clearance.

This study investigated possible sex-related differences in levels of antinociception and the rate of development of tolerance to the antinociceptive effects following prolonged (48 h) intravenous (i.v.) morphine administration in the rat. Groups of adult intact male, castrated male, female, and testosterone-pretreated female Sprague-Dawley rats received prolonged (48 h) infusions of i.v. morphine (5 or 10 mg/day) plus intra-arterial (i.a.) saline or i.v. morphine (5 mg/day) plus i.a. chloramphenicol (300 mg/day). Antinociception was quantified using the hotplate test. Serum concentrations of morphine and morphine-3-glucuronide (M3G) were assayed using high performance liquid chromatography with electrochemical detection, whereas the serum testosterone concentrations were quantified using an enzyme-linked immunosorbent assay method. Consistent with our previous findings in intact male rats, prolonged coinfusion of chloramphenicol with morphine produced a marked increase in the extent and duration of morphine antinociception in all experimental groups. Additionally, female and castrated male rats developed tolerance more slowly than either intact male or testosterone-pretreated female rats, when coinfused with parenteral morphine plus chloramphenicol. However, mean levels of antinociception were not significantly correlated with either the mean serum morphine or M3G concentrations, but were significantly inversely correlated with the mean values of the M3G/morphine serum molar concentration ratio. Testosterone pretreatment of female rats for 1 week before chronic morphine infusion abolished antinociception and markedly reduced both the serum morphine and M3G concentrations. These latter findings imply that testosterone modulates antinociception evoked by prolonged morphine infusion in rats via a mechanism that appears to involve modulation of morphine metabolism.

Systemic coadministration of chloramphenicol with intravenous but not intracerebroventricular morphine markedly increases morphine antinociception and delays development of antinociceptive tolerance in rats.

Chloramphenicol, an in vitro inhibitor of the glucuronidation of morphine to its putative antianalgesic metabolite, morphine-3-glucuronide (M3G), was coadministered with morphine in adult male Sprague-Dawley rats to determine whether it inhibited the in vivo metabolism of morphine to M3G, thereby enhancing morphine antinociception and/or delaying the development of antinociceptive tolerance. Parenteral chloramphenicol was given acutely (3-h studies) or chronically (48-h studies). Morphine was administered by the i.v. or i.c.v. route. Control rats received chloramphenicol and/or vehicle. Antinociception was quantified using the hotplate latency test. Coadministration of chloramphenicol with i.v. but not i.cv. morphine increased the extent and duration of morphine antinociception by approximately 5.5-fold relative to rats that received i.v. morphine alone. Thus, the mechanism through which chloramphenicol enhances i.v. morphine antinociception in the rat does not directly involve supraspinal opioid receptors. Acutely, parenteral coadministration of chloramphenicol and morphine resulted in an approximately 75% increase in the mean area under the serum morphine concentration-time curve but for chronic dosing there was no significant change in this curve, indicating that factors other than morphine concentrations contribute significantly to antinociception. Antinociceptive tolerance to morphine developed more slowly in rats coadministered chloramphenicol, consistent with our proposal that in vivo inhibition of M3G formation would result in increased antinociception and delayed development of tolerance. However, our data also indicate that chloramphenicol inhibited the biliary secretion of M3G. Whether chloramphenicol altered the passage of M3G and morphine across the blood-brain barrier remains to be investigated.

Cerebrospinal fluid and plasma concentrations of morphine, morphine-3-glucuronide, and morphine-6-glucuronide in patients before and after initiation of intracerebroventricular morphine for cancer pain management.

UNLABELLED: Twenty-three patients treated with intracerebroventricular (ICV) morphine in this study not only obtained excellent pain relief without rapid increases in dose, but also experienced a reduction in morphine-related side effects. By 24 h after initiation of ICV morphine, the mean trough cerebrospinal fluid (CSF) morphine concentration (approximately 20 microM) was 50-fold higher than the baseline concentration (approximately 0.4 microM), and the CSF concentration of morphine-6-glucuronide (M6G) was undetectable (<0.01 microM). The mean CSF concentration of morphine-3-glucuronide (M3G) decreased 90%, from a baseline concentration of 1 microM to 0.1 microM by Day 7 postventriculostomy. Thereafter, the mean trough CSF M3G concentration remained relatively constant while ICV morphine was continued, although the concomitant M3G plasma concentrations were undetectable (<0.01 microM). The large increase in the CSF morphine concentration in patients receiving ICV morphine strongly suggests that increased CSF morphine levels are unlikely to be the primary cause of analgesic tolerance or undesirable excitatory side effects (hyperalgesia, myoclonus, seizures) experienced by some patients receiving chronic large-dose systemic morphine. IMPLICATIONS: After initiation of intracerebroventricular morphine, cancer patients experienced excellent pain relief. Although the mean morphine concentration in cerebrospinal fluid increased 50-fold relative to preventriculostomy levels, rapid dose increases did not occur, which suggests that increased cerebrospinal fluid morphine levels are unlikely to be the main cause of analgesic tolerance.

Solid-phase extraction method with high-performance liquid chromatography and electrochemical detection for the quantitative analysis of oxycodone in human plasma.

A sensitive and reproducible solid-phase extraction (SPE) method for the quantification of oxycodone in human plasma was developed. Varian Certify SPE cartridges containing both C8 and benzoic acid functional groups were the most suitable for the extraction of oxycodone and codeine (internal standard), with consistently high (> or =80%) and reproducible recoveries. The elution mobile phase consisted of 1.2 ml of butyl chloride-isopropanol (80:20, v/v) containing 2% ammonia. The quantification limit for oxycodone was 5.3 pmol on-column. Within-day and inter-day coefficients of variation were 1.2% and 6.8% respectively for 284 nM oxycodone and 9.5% and 6.2% respectively for 28.4 nM oxycodone using 0.5-ml plasma aliquots.

Hydromorphone-3-glucuronide: biochemical synthesis and preliminary pharmacological evaluation.

Hydromorphone-3-glucuronide (H3G) was synthesized biochemically using rat liver microsomes, uridine-5'-diphosphoglucuronic acid (UDPGA) and the substrate, hydromorphone. Initially, the crude putative H3G product was purified by ethyl acetate precipitation and washing with acetonitrile. Final purification was achieved using semi-preparative high-performance-liquid-chromatography (HPLC) with ultraviolet (UV) detection. The purity of the final H3G product was shown by HPLC with electrochemical and ultraviolet detection to be > 99.9% and it was produced in a yield of = 60% (on a molar basis). The chemical structure of the putative H3G was confirmed by enzymatic hydrolysis of the glucuronide moiety using beta-glucuronidase, producing a hydrolysis product with the same HPLC retention time as the hydromorphone reference standard. Using HPLC with tandem mass spectrometry (HPLC-MS-MS) in the positive ionization mode, the molecular mass (M+1) was found to be 462 g/mol, in agreement with H3G's expected molecular weight of 461 g/mol. Importantly, proton-NMR indicated that the glucuronide moiety was attached at the 3-phenolic position of hydromorphone. A preliminary evaluation of H3G's intrinsic pharmacological effects revealed that following i.c.v. administration to adult male Sprague-Dawley rats in a dose of 5 microg, H3G evoked a range of excitatory behavioural effects including chewing, rearing, myoclonus, ataxia and tonic-clonic convulsions, in a manner similar to that reported previously for the glucuronide metabolites of morphine, morphine-3-glucuronide and normorphine-3-glucuronide.

Improved one-step solid-phase extraction method for morphine, morphine-3-glucuronide, and morphine-6-glucuronide from plasma and quantitation using high-performance liquid chromatography with electrochemical detection.

This communication describes an improved one-step solid-phase extraction method for the recovery of morphine (M), morphine-3-glucuronide (M3G), and morphine-6-glucuronide (M6G) from human plasma with reduced coextraction of endogenous plasma constituents, compared to that of the authors' previously reported method. The magnitude of the peak caused by endogenous plasma components in the chromatogram that eluted immediately before the retention time of M3G has been reduced (approximately 80%) significantly (p < 0.01) while achieving high extraction efficiencies for the compounds of interest, viz morphine, M6G, and M3G (93.8 +/- 2.5, 91.7 +/- 1.7, and 93.1 +/- 2.2%, respectively). Furthermore, when the improved solid-phase extraction method was used, the extraction cartridge-derived late-eluting peak (retention time 90 to 100 minutes) reported in our previous method, was no longer present in the plasma extracts. Therefore the combined effect of reducing the recovery of the endogenous components of plasma that chromatographed just before the retention time of M3G and the removal of the late-eluting, extraction cartridge-derived peak has resulted in a decrease in the chromatographic run-time to 20 minutes, thereby increasing the sample throughput by up to 100%.

Directed case study method for teaching human anatomy and physiology.

A mastery of human anatomy and physiology requires a familiarity with a vast number of details about the human body. A directed method of case analysis is described that helps students deepen and solidify their understanding of anatomical and physiological facts, concepts, and principles. The successful case had four distinctive features as follows: clear learning objectives, a concise and informative scenario, straightforward and didactic questions, and an emphasis on information readily available to the student. A directed case study is presented, and its salient features are described. A procedure for integrating case analyses into an undergraduate anatomy and physiology course is outlined. Student response to this type of case study suggests that this method improves the ease of learning, the depth of learning, and an appreciation of the relevance of and a curiosity about anatomy and physiology. The addition of case analyses to a two-semester integrated course in anatomy and physiology was also associated with an improvement in exam performance. The regular use of directed case analysis is a valuable addition to the traditional methods of lecture, textbook reading, and laboratory for the teaching of human anatomy and physiology.

Quantitation of morphine, morphine-3-glucuronide, and morphine-6-glucuronide in plasma and cerebrospinal fluid using solid-phase extraction and high-performance liquid chromatography with electrochemical detection.

An original, sensitive, and specific high-performance liquid chromatographic (HPLC) assay was developed for the quantitation of morphine and its two major metabolites, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G), in human plasma and cerebrospinal fluid (CSF) and in rat plasma, using hydromorphone as the internal standard. Solid-phase extraction was used to separate morphine and its glucuronide metabolites from plasma constituents. Extraction efficiencies of morphine, M3G, and M6G from human plasma samples (0.5 ml) were 84, 87, and 88%, respectively. Extraction efficiencies of morphine, M3G, and M6G did not differ significantly (p > 0.05) between human plasma and CSF or rat plasma. Morphine, M3G, M6G, and hydromorphone were separated on a 10 mu C8 Resolve radially compressed cartridge using a mobile phase comprising methanol:acetonitrile:phosphate buffer, (0.0125M pH 7.5; 10:10:80), in which 11 mg/L of cetyltrimethylammonium bromide (cetrimide) was dissolved. Quantitation was achieved using a single electrochemical detector at ambient temperature (23 degrees C). Standard curves were linear over the ranges 0.020-2.190, 0.027-2.709, and 0.027-0.542 microM for morphine, M3G, and M6G, respectively. Lower limits of detection for morphine, M3G, and M6G in human plasma and CSF samples (0.5 ml) were 0.020, 0.027, and 0.027 microM, respectively. Corresponding lower limits of detection in rat plasma (0.1 ml) were 0.102, 0.135, and 0.135 microM, respectively. Intraassay precision for low and high concentrations of morphine, M3G, and M6G were < 23 and < 8% respectively. Similarly, interassay accuracy for low and medium concentrations of morphine, M3G, and M6G were < 17% and were < 9% for high concentrations.

Determination of the serum protein binding of oxycodone and morphine using ultrafiltration.

Protein binding of oxycodone and morphine in human serum was determined in vitro using ultrafiltration. Binding studies were also performed using both purified human serum albumin and human alpha 1-acid glycoprotein (AAG). Albumin was found to be the major binding protein for both oxycodone and morphine. The serum protein binding of both oxycodone and morphine was independent of drug concentration in the therapeutic range (5-100 ng/ml), but was dependent on protein concentration. In addition, bound fractions of oxycodone and morphine increased with increasing concentrations of both albumin and AAG. At physiological pH and temperature, the mean (+/- SD) serum protein binding of oxycodone was 45.1% (+/- 0.4%) and that of morphine was 35.3% (+/- 0.2%) A decrease in temperature from 37 to 23 degrees C significantly increased the serum protein binding of oxycodone and morphine by 8-9% (p < 0.0001) and 7-10% (p < 0.0001), respectively, indicating the importance of maintaining the temperature at 37 degrees C during protein binding experiments. A reduction in pH from 7.75-8.85 to 7.4 significantly reduced serum protein binding of both oxycodone and morphine by 4-5% (p < 0.0001) and 4-7% (p < 0.0001), respectively. Serum samples, to which known concentrations of oxycodone had been added and which were stored at -20 degrees C, showing a gradual but significant decline (p < 0.0001) in serum protein binding of oxycodone from approximately 45 to 39% during the 4-week storage period.(ABSTRACT TRUNCATED AT 250 WORDS)

Cimetidine: an inhibitor and an inducer of rat liver microsomal cytochrome P-450.

1. Cimetidine pretreatment of male Sprague-Dawley rats caused a significant increase in the specific content of total hepatic cytochrome P-450, supporting the hypothesis that this H2-receptor antagonist has monooxygenase induction effects. 2. Quantitative ultrastructural studies of liver of cimetidine-pretreated animals also supported this hypothesis in showing a significant proliferation of smooth endoplasmic reticulum. These ultrastructural changes were qualitatively similar to those produced by treatment of rats with phenobarbital, a well-characterized monooxygenase-inducing agent whose effects were studied for comparative purposes. 3. Competitive inhibition of metoprolol alpha-hydroxylation by cimetidine in liver microsomes prepared from untreated animals (Ki = 18.8 microM) was also demonstrated. 4. These results allowed testing of the hypothesis (Burnet et al. 1986) that inhibition of a defined monooxygenase should lead to induction of the synthesis of the relevant cytochrome P-450 isozyme. 5. The finding that metoprolol alpha-hydroxylase activity of liver microsomes was lowered, not elevated, by pretreatment of animals with cimetidine argues against the concept of a causal link between monooxygenase inhibition and induction.

Familial dementia of adult onset with pathological findings of a 'non-specific' nature.

A family is described in which 4 of 10 siblings developed a dementing illness that culminated in death within five to six years of onset. The pathological findings in 3 members were strikingly similar, and consisted of widespread nerve cell loss and astrocytosis within the cerebral cortex, status spongiosus within the outer cortical layers and, in 2, nerve cell loss and astrocytosis within the dorsomedial nucleus of the thalamus. It is concluded that the disorder described in this report does not conform precisely to any of the currently recognized categories of familial dementing disease.

A comprehensive scoring system for evaluating Noonan syndrome.

A multidisciplinary team assessed 23 patients with various manifestations of the Noonan syndrome, including pulmonary valve stenosis (with leaflet dysplasia), "typical" facial appearance (including hypertelorism, epicanthic folds, flat nasal bridge, and apparently low-set ears), short stature, and mental retardation. Seven patients had a family history of the syndrome. A comprehensive scoring system was devised on the basis of frequency and severity of manifestations and results of invasive and noninvasive tests in these patients and those reported in the literature. The scoring system was condensed into a score card for clinical use and validated by "blind" application to patients with isolated pulmonary valve stenosis or suspected Noonan syndrome. Use of a scoring system to diagnose a syndrome for which there is no specific diagnostic test facilitates accuracy and decreases observer bias. In the case of unusual congenital disorders it is particularly valuable for a pediatrician in general practice.