Pharmacokinetics and hemodynamic effects of long-term nisoldipine treatment in hypertensive patients

In six patients with essential hypertension, the pharmacokinetics of nisoldipine were investigated before, during, and after 4 weeks of treatment. On day 1, nisoldipine was infused intravenously (i.v. 2 mg in 2 h); on day 2, oral nisoldipine treatment (10-mg tablets twice daily) was started for 4 weeks. During this period, patients came to the hospital six times, on which occasions blood samples were taken for the determination of trough and peak concentrations of nisoldipine. After 4-week treatment, the infusion experiment was repeated. In the first infusion experiment, systemic clearance was 1.02 +/- 0.23 L/min (mean +/- SD), terminal half-life (t1/2) was 15.4 +/- 6.7 h, and volume of distribution was 5.9 +/- 1.8 L/kg. After 4 weeks, these parameters had not changed significantly. Nisoldipine lowered blood pressure (BP) in all patients, whereas forearm blood flow and heart rate (HR) increased. Neither were the hemodynamic changes different after the oral treatment period, although basal BP was lower than before oral treatment. No accumulation of nisoldipine occurred during the 4-weeks treatment period.     

Increased Sensitivity to the Anticonvulsant Effect of Valproate in Aging BN/BiRij Rats

The aim of the present investigations was to study the influence of increasing age on the pharmacodynamics of valproate in BN/BiRij rats, applying a threshold for electrically induced localized seizure activity as a measure of the anticonvulsant effect. Seven groups of healthy male BN/BiRij rats were used, aged 3, 6, 12, 19, 25, 31, and 37 months. Individual plasma concentration versus anticonvulsant effect relationships were determined during a continuous intravenous infusion of sodium valproate at a rate of 5.5 mg/min/kg. The infusion was terminated when the anticonvulsant effect intensity had reached the maximum attainable level or at a total infusion time of three hours. A nonlinear relationship between valproate concentration and anticonvulsant effect intensity was observed with no maximal effect in the concentration range up to 1200 mg · L^–1. With increasing age a parallel shift in the concentration versus anticonvulsant effect relationships toward lower concentrations occurred. Thus increasing age appears to be associated with an increased sensitivity to the anticonvulsant effect of valproate.Suzanne Hovinga: Deceased January 30, 1991.     

Pharmacodynamics of tolerance development to the anesthetic and anticonvulsant effects of phenobarbital in rats

In this study, the potential development of tolerance towards the anesthetic and anticonvulsant effects of phenobarbital after chronic administration to rats was investigated, differentiating between dispositional and functional tolerance. Chronic exposure to phenobarbital by daily ip injections of 20 or 100 mg/kg for 14 days caused a 30% increase in clearance within 1 week. No changes occurred in the total amounts of phenobarbital and para-hydroxyphenobarbital excreted into urine and feces. Pharmacodynamic effects were quantitated after 2 weeks of treatment. The anesthetic effect was measured by slow iv infusion of phenobarbital until onset of loss of righting reflex (LRR), followed by measurement of drug concentrations in serum (both total and free), brain, and cerebrospinal fluid (CSF). With a phenobarbital dose of 100 mg/kg daily, CSF concentrations at the onset of LRR significantly increased from 136 +/- 12 to 176 +/- 21 mg/L (p less than 0.001), indicating that functional tolerance developed for the anesthetic effect. Protection of phenobarbital against convulsions induced by pentylenetetrazol (PTZ), as measured by the elevation of the PTZ plasma threshold concentration necessary to elicit seizures at the EC50 of phenobarbital, was not altered. The discrepancy observed in the development of functional adaptation of the CNS demonstrates that different mechanisms of action are reflected in the different measures of the anesthetic and anticonvulsant effects of phenobarbital that were utilized.     

Deoxyribose analogues of N6-cyclopentyladenosine (CPA): partial agonists at the adenosine A1 receptor in vivo.

1. The purpose of the present study was to quantify the cardiovascular effects of the 2'-, 3'-, 5'-deoxyribose analogues of the selective adenosine A1 receptor agonist, N6-cyclopentyladenosine (CPA) in vivo. The blood concentration-effect relationships of the compounds were assessed in individual rats and correlated to their receptor binding characteristics. 2. The pharmacokinetics and pharmacodynamics of the compounds were determined after a single intravenous infusion of 0.80 mg kg (-1) (63 micromol kg(-1)of 2' dCPA. The heart rate (HR) and mean arterial blood pressure (MAP) were monitored continuously during the experiment and serial arterial blood samples were taken for analysis of drug concentration. 3. The relationship between blood concentrations and the reductions in both heart rate and blood pressure were described according to the sigmoidal Emax model. For the bradycardiac effect, the potencies based on free drug concentrations (EC50,u) of 5'dCPA, 3'dCPA, 2'dCPAin blood were 5.9 +/- 1.7, 18 +/- 4 and 260 +/- 70 ng ml (-1) (19 +/- 6, 56 +/- 11 and 830 +/- 210 nM), respectively, and correlated well with the adenosine A1 receptor affinity in vitro. The Emax value of 2'dCPA was significantly less than those of the other compounds, suggesting that this compound may be regarded as a partial agonist when compared to the other analogues. The rank order of the maximal reduction in heart rate of the compounds corresponded well with the order of the GTP-shifts, as determined in vitro. 4. It is concluded that deoxyribose derivatives of CPA may be partial agonists for the adenosine A1 receptor and may serve as tools for further investigation of adenosine receptor partial agonism in vivo.     

The unit impulse response procedure for the pharmacokinetic evaluation of drug entry into the central nervous system

The unit impulse response theory has been adapted to characterize the transport profile of drugs into the central nervous system (CNS). From the obtained input function, the cumulative plasma volume (V) cleared by transport into the CNS in time can be calculated. Simulation studies demonstrated that transport governed by passive diffusion resulted in a linear relationship between V and time, while the slope of the line, the blood- brain barrier (BBB) clearance, proved to be an adequate and model independent parameter to characterize drug transport into the CNS. The error in the result of the numerical procedure could be limited to less than 10% of the theoretically predicted value. Superposition of 5 or 10% random noise on simulated data did not result in significant differences between the calculated and theoretically predicted clearance values. Simulations of carrier-mediated transport resulted in nonlinear transport curves; the degree of nonlinearity, and thus the detectability, was dependent on the initial degree of saturation of the system, the rate of desaturation, as caused by drug elimination processes and the noise level on the data. In vivoexperiments in the rat were performed, using atenolol, acetaminophen, and antipyrine as model drugs. Linear transport relationships were obtained for all drugs, indicating that transport was dependent on passive diffusion or a low affinity carrier system. BBB- clearance values were 7±1 l/min for atenolol, 63±7 ul/min for acetaminiphen and 316±25 l/min for antipyrine. These experiments validate the applicability of the presented technique in in vivostudies.     

High-performance liquid chromatography of the neuroactive steroids alphaxalone and pregnanolone in plasma using dansyl hydrazine as fluorescent label: application to a pharmacokinetic-pharmacodynamic study in rats

This report describes a rapid and sensitive analytical method for the quantification of the neuroactive steroids alphaxalone and pregnanolone in rat plasma using derivatization with dansyl hydrazine as fluorescent label. The method involves protein precipitation, alkaline derivatization and extraction of the compounds and internal standard pregnenolone with dichloromethane, followed by isocratic reversed-phase high-performance liquid chromatography on a 3-microm Microsphere C18 column with fluorescence detection at wavelengths 332 nm and 516 nm for excitation and emission, respectively. The mobile phase consists of a mixture of 25 mM acetate buffer (pH 3.7)-acetonitrile (45:55, v/v for alphaxalone and 40:60, v/v for pregnanolone) with a flow-rate of 1 ml/min. The total run time was approximately 35 min. In the concentration range of 0.010-10 microg ml(-1), the intra- and inter-assay coefficients of variation were less than 17% for both methods. In 50 microl plasma samples the corresponding limits of detection were 10 ng ml(-1) (signal-to-noise ratio=3). The utility of the analytical method was established by analyzing plasma samples from rats, which had received an intravenous administration of 5 mg kg(-1) alphaxalone or pregnanolone. Values for clearance, volume of distribution at steady state and terminal half life were 71.9 ml min(-1) kg(-1), 814 mg kg(-1) and 13.5 min for alphaxalone and 69.2 ml min(-1) kg(-1), 1,638 ml kg(-1) and 27.8 min for pregnanolone, respectively. Due to its simplicity and sensitivity this method can be used on a routine basis for pharmacokinetic analysis of neuroactive steroids.     

The Use of Intracerebral Microdialysis to Determine Changes in Blood-Brain Barrier Transport Characteristics

The aim of this study was to determine whether changes in the transport of drugs into the brain could be determined by in vivo intracerebral microdialysis. Atenolol was used as a model drug to determine blood-brain barrier (BBB) transport characteristics. In rats, unilateral opening of the blood-brain barrier was achieved by infusion of hyperosmolar mannitol (25%, w/v) into the left internal carotid artery. BBB transport, expressed as the ratio of the area under the curve (AUC) of atenolol in brain extracellular fluid over plasma, was three times higher for the mannitol treated hemisphere as compared with the contralateral brain or after infusion of saline, being (mean ± SEM) 0.094 ± 0.024 (n = 16), 0.029 ± 0.007 (n = 12) and 0.030 ± 0.009 (n = 12) respectively. Further evaluation of the data indicated that for experiments performed in the morning the mannitol infusion had little effect on the extent of transport of atenolol into the brain, while in the afternoon BBB transport was about 10-fold higher than in the contralateral and saline group. The mean afternoon ratios ± SEM were 0.155 ± 0.038 (n = 8), 0.012 ± 0.003 (n = 6) and 0.018 ± 0.006 (n = 6) respectively. It is concluded that intracerebral microdialysis is capable of revealing changes in BBB transport and regional and time-dependent differences in drug levels can be demonstrated with the use of this technique.     

A combined specific target site binding and pharmacokinetic model to explore the non-linear disposition of draflazine

The capacity-limited high-affinity target site binding of draflazine to the nucleoside transporters located on the erythrocytes is a source of nonlinearity in the pharmacokinetics of the drug. An attractive feature of draflazine is that the specific target site binding characteristics can be determined easily by simultaneously measuring plasma and whole blood concentrations of the drug. Measured drug concentrations following various infusion rates and infusion durations were used to develop a model in which the interrelated blood-plasma distribution, elimination, and specific target site binding of draflazine were incorporated simultaneously. The estimated binding (dissociation) constant Kd was 0.57 ng/ml plasma and the maximal specific erythrocyte binding capacity (BmaxRBC) was 163 ng/ml RBC. The maximal specific binding capacity to the tissues (Bmaxtissue) was estimated to be about 1 mg. The estimated volume of the central compartment (Vplasma + tissue fluids) was 12.9 L and the total intrinsic CL was 645 ml/min. After validation, the model was used to further investigate the impact of the specific high-affinity target site binding of draflazine on its disposition in plasma. The time required to reach steady-state plasma concentrations of draflazine decreased with an increasing infusion rate. Time profiles of the plasma concentrations were not always representative for the time profiles of the specific target site (RBC) occupancy of draflazine, but the t1/2,z in plasma paralleled that of the drug at target sites. The apparent Vd and the t1/2,z decreased with increasing single doses whereas the total CL remained constant. The recovery of draflazine was also dose dependent and increased with increasing doses. Finally, the total CL and apparent Vd of the first dose were greater than those of the second dose of draflazine.     

Pharmacodynamic characterization of the electroencephalographic effects of thiopental in rats

We have developed a chronically instrumented rat model that uses changes in electroencephalographic wave forms to estimate continuously the degree of central nervous system (CNS) depression induced by thiopental. Such changes were subject to aperiodic signal analysis, a technique that breaks down the complex EEG into a series of discreet neurologic "events" which are then quantitated as waves/sec. We thus obtained a continuous measure of CNS drug effect. In addition we continuously recorded central arterial blood pressure and heart rate and monitored ventilatory status using arterial blood gas determinations. We also determined, with frequent arterial blood sampling, the distribution and elimination of thiopental in individual animals. The time lag occurring in the curve representing arterial concentration of thiopental vs. EEG effect suggests that arterial plasma is not kinetically equivalent to the EEG effect site. Application of semiparametric pharmacodynamic modeling techniques enabled us to estimate equilibration rate constant (Keo) for concentrations of thiopental between arterial plasma and the effect site. The half-life for equilibration of thiopental with the EEG (CNS) effect was less than 80 sec. Knowledge of the rate of equilibration permitted characterization of the relationship between the steady state plasma concentrations and CNS effect of thiopental, as measured by activation and slowing of the EEG. At concentrations of thiopental below 5 micrograms/ml, EEG activity was 180% higher than during the baseline awake state. Thiopental produced an activated EEG over more than 20% of the concentration-effect relationship. Further increases in the concentration of thiopental at the site of effect depressed EEG activity progressively until complete suppression of the EEG signal occurred (at which time, the concentration was approximately 80 micrograms/ml). This report describes our model and its application to the assessment of the pharmacodynamics of thiopental as manifested by changes on the EEG.     

Targeting liposomes with protein drugs to the blood-brain barrier in vitro.

In this study, we aim to target pegylated liposomes loaded with horseradish peroxidase (HRP) and tagged with transferrin (Tf) to the BBB in vitro. Liposomes were prepared with the post-insertion technique: micelles of polyethylene glycol (PEG) and PEG-Tf were inserted into pre-formed liposomes containing HRP. Tf was measured indirectly by measuring iron via atomic absorption spectroscopy. All liposomes were around 100 nm in diameter, contained 5-13 microg HRP per mumol phospholipid and 63-74 Tf molecules per liposome (lipo Tf) or no Tf (lipo C). Brain capillary endothelial cells (BCEC) were incubated with liposomes at 4 degrees C (to determine binding) or at 37 degrees C (to determine association, i.e. binding+endocytosis) and the HRP activity, rather than the HRP amount was determined in cell lysates. Association of lipo Tf was two- to three-fold higher than association of lipo C. Surprisingly, the binding of lipo Tf at 4 degrees C was four-fold higher than the association of at 37 degrees C. Most likely this high binding and low endocytosis is explained by intracellular degradation of endocytosed HRP. In conclusion, we have shown targeting of liposomes loaded with protein or peptide drugs to the BCEC and more specifically to the lysosomes. This is an advantage for the treatment of lysosomal storage disease. However, drug targeting to other intracellular targets also results in intracellular degradation of the drug. Our experiments suggest that liposomes release some of their content within the BBB, making targeting of liposomes to the TfR on BCEC an attractive approach for brain drug delivery.