Dynamics of a three-variable nonlinear model of vasomotion: comparison of theory and experiment.

The effects of pharmacological interventions that modulate Ca(2+) homeodynamics and membrane potential in rat isolated cerebral vessels during vasomotion (i.e., rhythmic fluctuations in arterial diameter) were simulated by a third-order system of nonlinear differential equations. Independent control variables employed in the model were [Ca(2+)] in the cytosol, [Ca(2+)] in intracellular stores, and smooth muscle membrane potential. Interactions between ryanodine- and inositol 1,4,5-trisphosphate-sensitive intracellular Ca(2+) stores and transmembrane ion fluxes via K(+) channels, Cl(-) channels, and voltage-operated Ca(2+) channels were studied by comparing simulations of oscillatory behavior with experimental measurements of membrane potential, intracellular free [Ca(2+)] and vessel diameter during a range of pharmacological interventions. The main conclusion of the study is that a general model of vasomotion that predicts experimental data can be constructed by a low-order system that incorporates nonlinear interactions between dynamical control variables.

Differential activation of ion channels by inositol 1,4,5-trisphosphate (IP3)- and ryanodine-sensitive calcium stores in rat basilar artery vasomotion.

Spontaneous, rhythmical contractions, or vasomotion, can be recorded from cerebral vessels under both normal physiological and pathophysiological conditions. Using electrophysiology to study changes in membrane potential, the ratiometric calcium indicator Fura-2 AM to study changes in [Ca(2+)](i) in both the arterial wall and in individual smooth muscle cells (SMCs), and video microscopy to study changes in vessel diameter, we have investigated the cellular mechanisms underlying vasomotion in the juvenile rat basilar artery. During vasomotion, rhythmical oscillations in both membrane potential and [Ca(2+)](i) were found to precede rhythmical contractions. Nifedipine depolarized SMCs and abolished rhythmical contractions and depolarizations. [Ca(2+)](i) oscillations in the arterial wall became reduced and irregular, while [Ca(2+)](i) oscillations in adjacent SMCs were no longer synchronized. BAPTA-AM, thapsigargin and U73122 hyperpolarized SMCs, relaxed the vessel, decreased basal calcium levels and abolished vasomotion. Chloride substitution abolished rhythmical activity, depolarized SMCs, increased basal calcium levels and constricted the vessel, while niflumic acid and DIDS abolished vasomotion. Ryanodine, charybdotoxin and TRAM-34, but not iberiotoxin, 4-aminopyridine or apamin, each depolarized SMCs and increased the frequency of rhythmical depolarizations and [Ca(2+)](i) oscillations. We conclude that vasomotion in the basilar artery depends on the release of intracellular calcium from IP(3) (inositol 1,4,5,-trisphosphate)-sensitive stores which activates calcium-dependent chloride channels to depolarize SMCs. Depolarization in turn activates voltage-dependent calcium channels, synchronizing contractions of adjacent cells through influx of extracellular calcium. Subsequent calcium-induced calcium release from ryanodine-sensitive stores activates an intermediate conductance potassium channel, hyperpolarizing the SMCs and providing a negative feedback pathway for regeneration of the contractile cycle.

Voltage independence of vasomotion in isolated irideal arterioles of the rat.

The cellular mechanisms underlying vasomotion of irideal arterioles from juvenile rats have been studied using electrophysiological methods, ratiometric calcium measurements and video microscopy. Vasomotion was not affected by removal of the endothelium. Spontaneous contractions were preceded by spontaneous depolarizations. Both were abolished by the intracellular calcium chelator, BAPTA AM (20 microM), but not by ryanodine (10 microM), suggesting a dependence on the cyclical release of calcium from intracellular stores, other than those operated by ryanodine receptors. Oscillations were little changed when the membrane potential of short segments of arteriole was either depolarized or hyperpolarized. When the segments were voltage clamped, oscillating inward currents were recorded, indicating that the changes in membrane potential were voltage independent. Vasomotion was preceded by intracellular calcium oscillations and both were abolished by inhibitors of phospholipase C (U73122, 10 microM), phospholipase A(2) (AACOCF(3), 30 microM) and protein kinase C (chelerythrine chloride, 5 microM, and myristoylated protein kinase C peptide, 10 microM). Inhibition of vasomotion by the dual lipoxygenase and cyclo-oxygenase inhibitor, NDGA (10 microM), the lipoxygenase inhibitor, ETI (1 microM) but not by the cyclo-oxygenase inhibitors, aspirin (10 microM) and indomethacin (10 microM), or the cytochrome P450 inhibitor 17-ODYA (10 microM), suggested an involvement of the lipoxygenase pathway. The observations suggest that vasomotion of iris arterioles is voltage independent and results from the cyclical release of calcium from IP(3)-sensitive stores which are activated by cross talk between the phospholipase C and phospholipase A(2) pathways in vascular smooth muscle.

Sibutramine pharmacokinetics in young and elderly healthy subjects.

OBJECTIVE: To investigate the pharmacokinetics of the pharmacologically active metabolites of sibutramine (metabolites 1 and 2) in healthy young and elderly volunteers following a single oral dose of sibutramine. METHODS: This was an open, parallel-group study completed by 12 young (six male, six female; mean age 24.0 years) and 12 elderly (six male, six female; mean age 70.3 years) healthy volunteers. Blood samples were taken at intervals up to 48 h post-dose. Plasma concentrations of metabolites were determined using HPLC-MS. Model-independent pharmacokinetic parameters of the two metabolites were compared for the two age groups. RESULTS: The similarity of the plasma profiles of the two desmethyl metabolites showed that despite the possibility of reduced hepatic function due to age, the rate and extent of formation of these was the same in both young and elderly, i.e. sibutramine metabolism was not impaired in elderly subjects. There were also no significant differences in elimination of metabolite 2 between groups, although the elderly group showed a slight trend for a reduction in k(el). CONCLUSIONS: The pharmacokinetics of the two pharmacologically active metabolites of sibutramine (metabolites 1 and 2) were not significantly different between the young and elderly groups in this study. Based on this information, a similar dosing regimen would be appropriate for both the young and elderly.

Metabolism and disposition of 14C-granisetron in rat, dog and man after intravenous and oral dosing.

1. The disposition and metabolic fate of 14C-granisetron, a novel 5-HT3 antagonist, was studied in rat, dog, and male human volunteers after intravenous and oral administration. 2. Complete absorption occurred from the gastrointestinal tract following oral dosing, but bioavailability was reduced by first-pass metabolism in all three species. 3. There were no sex-specific differences observed in radiometabolite patterns in rat or dog and there was no appreciable change in disposition with dose between 0.25 and 5 mg/kg in rat and 0.25 and 10 mg/kg in dog. Additionally, there were no large differences in disposition associated with route of administration in rat, dog and man. 4. In rat and dog, 35-41% of the dose was excreted in urine and 52-62% in faeces, via the bile. Metabolites were largely present as glucuronide and sulphate conjugates, together with numerous minor polar metabolites. In man, about 60% of dosed radioactivity was excreted in urine and 36% in faeces after both intravenous and oral dosing. Unchanged granisetron was only excreted in urine (5-25% of dose). 5. The major metabolites were isolated and identified by MS spectroscopy and nmr. In rat, the dominant routes of biotransformation after both intravenous and oral dosing were 5-hydroxylation and N1-demethylation, followed by the formation of conjugates which were the major metabolites in urine, bile and plasma. In dog and man the major metabolite was 7-hydroxy-granisetron, with lesser quantities of the 6,7-dihydrodiol and/or their conjugates.

The effect of selective serotonin re-uptake inhibitors on cytochrome P4502D6 (CYP2D6) activity in human liver microsomes.

Inhibition of human cytochrome P4502D6 (CYP2D6)-catalysed metabolism can lead to clinically significant alterations in pharmacokinetics. Since there is evidence that the selective serotonin reuptake inhibitor (SSRI) class of antidepressant drugs might inhibit CYP2D6, the effects of five SSRIs on human liver microsomal CYP2D6 activity were compared with each other and with three tricyclic antidepressant drugs. On a molar basis, paroxetine was the most potent of the SSRIs at inhibiting the CYP2D6-catalysed oxidation of sparteine (Ki = 0.15 microM), although fluoxetine (0.60 microM) and sertaline (0.70 microM) had Ki values in the same range. Fluvoxamine (8.2 microM) and citalopram (5.1 microM) also inhibited CYP2D6 activity. The major circulating metabolites of paroxetine in man produced negligible inhibition. In contrast, norfluoxetine the active metabolite of fluoxetine, was a potent CYP2D6 inhibitor (0.43 microM). CYP2D6 activity was also diminished by the tricyclic antidepressant drugs clomipramine (2.2 microM), desipramine (2.3 microM) and amitriptyline (4.0 microM). These findings suggest that compounds with SSRI activity are likely to interact with human CYP2D6 in vivo with the potential of causing drug interactions.

The role of cytochrome P4502D6 in the metabolism of paroxetine by human liver microsomes.

Paroxetine is a selective serotonin reuptake inhibitor possessing anti-depressant activity. Demethylenation of the methylenedioxy phenyl group is the initial step in its metabolism, the liberated carbon appearing in vitro as formate. A radioassay involving [14C-methylenedioxy] paroxetine was developed and used to examine the role of cytochrome P4502D6 in paroxetine metabolism by human liver microsomes. The rate of formate production was much higher in microsomes from an extensive metaboliser of debrisoquine than from a poor metaboliser. Also, demethylenation of paroxetine was inhibited by the quinidine and quinine isomer pair in microsomes from the extensive metaboliser only. These observations strongly suggested that the process was catalysed by the enzyme cytochrome P4502D6. Metabolism could not be completely inhibited by quinidine, the residual activity representing the contribution of at least one other enzyme. The ability of microsomes from a poor metaboliser of debrisoquine to demethylenate paroxetine provided further evidence for the involvement of an enzyme distinct from P4502D6. This was confirmed by kinetic analysis of the process in microsomes from both poor and extensive metabolisers. It is concluded that, in man, the initial step of paroxetine metabolism is performed by at least two enzymes, one of which is cytochrome P4502D6.

A review of the metabolism and pharmacokinetics of paroxetine in man.

Paroxetine is well absorbed from the gastrointestinal tract, and appears to undergo first-pass metabolism which is partially saturable. Consistent with its lipophilic amine character, paroxetine is extensively distributed into tissues. Its plasma protein binding at therapeutically relevant concentrations is about 95%. Paroxetine is eliminated by metabolism involving oxidation, methylation, and conjugation. All of these factors lead to wide interindividual variation in the pharmacokinetics of paroxetine. Renal clearance of the compound is negligible. The major metabolites of paroxetine are conjugates which do not compromise its selectivity nor contribute to the clinical response. Ascending single-dose studies reveal that the pharmacokinetics of paroxetine are non-linear to a limited extent in most subjects and to a marked degree in only a few. Also, steady-state pharmacokinetic parameters are not predictable from single-dose data. In many subjects, daily administration of 20-50 mg of paroxetine leads to little or no disproportionality in plasma levels with dose, although in a few subjects this phenomenon is evident. Steady-state plasma concentrations are generally achieved within 7 to 14 days. The terminal half-life is about one day, although there is a wide intersubject variability (e.g. with 30 mg, a range of 7-65 hours was observed in a group of 28 healthy young subjects). In elderly subjects there is wide interindividual variation in steady-state pharmacokinetic parameters, with statistically significantly higher plasma concentrations and slower elimination than in younger subjects, although there is a large degree of overlap in the ranges of corresponding parameters. In severe renal impairment higher plasma levels of paroxetine are achieved than in healthy individuals after single dose. In moderate hepatic impairment the pharmacokinetics after single doses are similar to those of normal subjects. Paroxetine is not a general inducer or inhibitor of hepatic oxidation processes, and has little or no effect on the pharmacokinetics of other drugs examined. Its metabolism and pharmacokinetics are to some degree affected by the induction or inhibition of drug metabolizing enzyme(s). From a pharmacokinetic standpoint, drug interactions involving paroxetine are considered unlikely to be a frequent occurrence. Data available have failed to reveal any correlation between plasma concentrations of paroxetine and its clinical effects (either efficacy or adverse events).

Metabolism of nabumetone (BRL 14777) by various species including man.

Radiotracer methodology was used to study the metabolic fate of 4-(6-methoxy-2-naphthyl)-butan-2-one (nabumetone) after oral administration to rats, mice, rabbits, dogs, rhesus monkeys and healthy human subjects. Parent compound was not detected in plasma and urine and the major circulating metabolite in all species was identified as 6-methoxy-2-naphthylacetic acid, a compound known to possess anti-inflammatory activity. Metabolites were mainly excreted in urine from which four principal metabolites were isolated and identified by mass spectrometry and independent synthesis. Pathways involving O-demethylation, reduction of the ketone group and oxidation of the butanone side-chain to acetic acid occurred in all species, but the ratios of the metabolic end-products tended to be species dependent. In the rat about half of the administered nabumetone was oxidized to the pharmacologically active acid metabolite.

Penetration of nabumetone into inflammatory exudates in the rat.

The penetration of radioactivity into inflamed sites induced by subcutaneously implanted cotton pellets was studied in the rat after oral administration of [14C]nabumetone. Equilibration between inflamed site and plasma concentrations was slow, maximum concentrations in the pellet granuloma being later than those in the plasma. Nabumetone itself was found only in very low concentrations at the inflamed site and was not found in plasma. The major component (70%) of the radioactivity at both sites was the metabolite 6-methoxy-2-naphthylacetic acid, a compound which has anti-inflammatory properties. Elimination of radioactivity from the plasma and inflamed site was virtually complete within 24 h of dosing. Daily dosing with [14C]nabumetone confirmed that there was no progressive accumulation of radioactivity at the inflamed site.

Biotransformation of phenprocoumon in the rat.

The metabolic fate of phenprocoumon [3-(alpha-ethylbenzyl)-4-hydroxycoumarin] in the rat is described. The major metabolites, 4',-6-,7-, and 8-hyproxyphenprocoumon, have been identified yb mass spectrometry, TLC, and uv and compared with authentic smaples. Metabolites are mainly excreted via the feces. The results are compared with those previously reported for warfarin.