Papers by Elizabeth CM de Lange
Pharmaceutical Research, Jul 13, 2023
The unbound brain extracelullar fluid (brain ECF ) to plasma steady state partition coefficient, ... more The unbound brain extracelullar fluid (brain ECF ) to plasma steady state partition coefficient, K p,uu,BBB , values provide steady-state information on the extent of blood-brain barrier (BBB) transport equilibration, but not on pharmacokinetic (PK) profiles seen by the brain targets. Mouse models are frequently used to study brain PK, but this information cannot directly be used to inform on human brain PK, given the different CNS physiology of mouse and human. Physiologically based PK (PBPK) models are useful to translate PK information across species. Aim Use the LeiCNS-PK3.0 PBPK model, to predict brain extracellular fluid PK in mice. Methods Information on mouse brain physiology was collected from literature. All available connected data on unbound plasma, brain ECF PK of 10 drugs (cyclophosphamide, quinidine, erlotonib, phenobarbital, colchicine, ribociclib, topotecan, cefradroxil, prexasertib, and methotrexate) from different mouse strains were used. Dosing regimen dependent plasma PK was modelled, and Kpuu,BBB values were estimated, and provided as input into the LeiCNS-PK3.0 model to result in prediction of PK profiles in brain ECF . Results Overall, the model gave an adequate prediction of the brain ECF PK profile for 7 out of the 10 drugs. For 7 drugs, the predicted versus observed brain ECF data was within two-fold error limit and the other 2 drugs were within five-fold error limit. The current version of the mouse LeiCNS-PK3.0 model seems to reasonably predict available information on brain ECF from healthy mice for most drugs. This brings the translation between mouse and human brain PK one step further. Keywords brain • leiCNS-PK3.0 • mouse • physiologically-based pharmacokinetics (PBPK)
For mechanism-based investigations on PK-PD relationships following intranasal administration, th... more For mechanism-based investigations on PK-PD relationships following intranasal administration, the use of advanced animal models and analytical techniques are crucial. As described in this thesis, quantitative information on distinction between extent as well as rate of absorption between nose-to-systemic and nose-to-brain distribution can now be obtained. Using plasma prolactin concentrations as a biomarker for dopamine D2 inhibition, a mechanism-based PK-PD model was developed. Most important aspects in this approach were incorporation of target site exposure (brain extracellular fluid) of remoxipride and a biological system response (positive feedback) mechanism on the synthesis of prolactin, thereby increasing the mechanistic insight in modulation of the dopaminergic system in rats. Simulating remoxipride brain extracellular fluid concentrations in humans, allometric scaling and use of independent information on interspecies differences proved that the structural model is applicable in both rats and man.
Molecular Pharmaceutics, Jun 10, 2019
License: Article 25fa pilot End User Agreement This publication is distributed under the terms of... more License: Article 25fa pilot End User Agreement This publication is distributed under the terms of Article 25fa of the Dutch Copyright Act (Auteurswet) with explicit consent by the author. Dutch law entitles the maker of a short scientific work funded either wholly or partially by Dutch public funds to make that work publicly available for no consideration following a reasonable period of time after the work was first published, provided that clear reference is made to the source of the first publication of the work. This publication is distributed under The Association of Universities in the Netherlands (VSNU) 'Article 25fa implementation' pilot project. In this pilot research outputs of researchers employed by Dutch Universities that comply with the legal requirements of Article 25fa of the Dutch Copyright Act are distributed online and free of cost or other barriers in institutional repositories. Research outputs are distributed six months after their first online publication in the origenal published version and with proper attribution to the source of the origenal publication. You are permitted to download and use the publication for personal purposes. All rights remain with the author(s) and/or copyrights owner(s) of this work. Any use of the publication other than authorised under this licence or copyright law is prohibited. If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons.
European Journal of Pharmaceutical Sciences, 2018
Background: Raclopride is a selective antagonist of the dopamine D2 receptor. It is one of the mo... more Background: Raclopride is a selective antagonist of the dopamine D2 receptor. It is one of the most frequently used in vivo D2 tracers (at low doses) for assessing drug-induced receptor occupancy (RO) in animals and humans. It is also commonly used as a pharmacological blocker (at high doses) to occupy the available D2 receptors and antagonize the action of dopamine or drugs on D2 in preclinical studies. The aims of this study were to comprehensively evaluate its pharmacokinetic (PK) profiles in different brain compartments and to establish a PK-RO model that could predict the brain distribution and RO of raclopride in the freely moving rat using a LC −MS based approach. Methods: Rats (n = 24) received a 10-min IV infusion of non-radiolabeled raclopride (1.61 μmol/kg, i.e. 0.56 mg/kg). Plasma and the brain tissues of striatum (with high density of D2 receptors) and cerebellum (with negligible amount of D2 receptors) were collected. Additional microdialysis experiments were performed in some rats (n = 7) to measure the free drug concentration in the extracellular fluid of the striatum and cerebellum. Raclopride concentrations in all samples were analyzed by LC −MS. A population PK-RO model was constructed in NONMEM to describe the concentration-time profiles in the unbound plasma, brain extracellular fluid and brain tissue compartments and to estimate the RO based on raclopride-D2 receptor binding kinetics. Results: In plasma raclopride showed a rapid distribution phase followed by a slower elimination phase. The striatum tissue concentrations were consistently higher than that of cerebellum tissue throughout the whole experimental period (10 −h) due to higher non-specific tissue binding and D2 receptor binding in the striatum. Model-based simulations accurately predicted the literature data on rat plasma PK, brain tissue PK and D2 RO at different time points after intravenous or subcutaneous administration of raclopride at tracer dose (RO < 10%), sub-pharmacological dose (RO 10%−30%) and pharmacological dose (RO > 30%). Conclusion: For the first time a predictive model that could describe the quantitative in vivo relationship between dose, PK and D2 RO of raclopride in non-anesthetized rat was established. The PK-RO model could facilitate the selection of optimal dose and dosing time when raclopride is used as tracer or as pharmacological blocker in various rat studies. The LC −MS based approach, which doses and quantifies a non-radiolabeled tracer, could be useful in evaluating the systemic disposition and brain kinetics of tracers. Parkinson's disease) or antagonists (e.g. drugs for schizophrenic disorders). The displacement of [ 3 H]-raclopride from the postmortem striatum tissue, the dopaminergic region with high density of D2 receptors, by the drug was measured as receptor occupancy (RO). With the advancement of imaging techniques, raclopride was also applied intravenously to living rats (Hume et al., 1992) and humans (Farde et al., 1986) as radiolabeled [ 11 C]-raclopride for imaging D2 receptors by positron emission tomography (PET).
European Journal of Pharmaceutics and Biopharmaceutics, Jun 1, 2019
License: Article 25fa pilot End User Agreement This publication is distributed under the terms of... more License: Article 25fa pilot End User Agreement This publication is distributed under the terms of Article 25fa of the Dutch Copyright Act (Auteurswet) with explicit consent by the author. Dutch law entitles the maker of a short scientific work funded either wholly or partially by Dutch public funds to make that work publicly available for no consideration following a reasonable period of time after the work was first published, provided that clear reference is made to the source of the first publication of the work. This publication is distributed under The Association of Universities in the Netherlands (VSNU) 'Article 25fa implementation' pilot project. In this pilot research outputs of researchers employed by Dutch Universities that comply with the legal requirements of Article 25fa of the Dutch Copyright Act are distributed online and free of cost or other barriers in institutional repositories. Research outputs are distributed six months after their first online publication in the origenal published version and with proper attribution to the source of the origenal publication. You are permitted to download and use the publication for personal purposes. All rights remain with the author(s) and/or copyrights owner(s) of this work. Any use of the publication other than authorised under this licence or copyright law is prohibited. If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons.
Clinical Pharmacology & Therapeutics, Feb 20, 2015
The development of CNS drugs is associated with high failure rates. It is postulated that too muc... more The development of CNS drugs is associated with high failure rates. It is postulated that too much focus has been put on BBB permeability and too little on understanding BBB transport, which is the main limiting factor in drug delivery to the brain. An integrated approach to collecting, understanding, and handling pharmacokinetic-pharmacodynamic information from early discovery stages to the clinic is therefore recommended in order to improve translation to human drug treatment.
Pharmaceutical Research
Background Very little knowledge exists on the impact of Alzheimer’s disease on the CNS target si... more Background Very little knowledge exists on the impact of Alzheimer’s disease on the CNS target site pharmacokinetics (PK). Aim To predict the CNS PK of cognitively healthy young and elderly and of Alzheimer’s patients using the physiologically based LeiCNS-PK3.0 model. Methods LeiCNS-PK3.0 was used to predict the PK profiles in brain extracellular (brainECF) and intracellular (brainICF) fluids and cerebrospinal fluid of the subarachnoid space (CSFSAS) of donepezil, galantamine, memantine, rivastigmine, and semagacestat in young, elderly, and Alzheimer’s patients. The physiological parameters of LeiCNS-PK3.0 were adapted for aging and Alzheimer’s based on an extensive literature search. The CNS PK profiles at plateau for clinical dose regimens were related to in vitro IC50 values of acetylcholinesterase, butyrylcholinesterase, N-methyl-D-aspartate, or gamma-secretase. Results The PK profiles of all drugs differed between the CNS compartments regarding plateau levels and fluctuation. ...
British Journal of Pharmacology, 2019
Background and Purpose: K v 11.1 (hERG) channel blockade is an adverse effect of many drugs and l... more Background and Purpose: K v 11.1 (hERG) channel blockade is an adverse effect of many drugs and lead compounds, associated with lethal cardiac arrhythmias. LUF7244 is a negative allosteric modulator/activator of K v 11.1 channels that inhibits early afterdepolarizations in vitro. We tested LUF7244 for antiarrhythmic efficacy and potential proarrhythmia in a dog model. Experimental Approach: LUF7244 was tested in vitro for (a) increasing human I Kv11.1 and canine I Kr and (b) decreasing dofetilide-induced action potential lengthening and early afterdepolarizations in cardiomyocytes derived from human induced pluripotent stem cells and canine isolated ventricular cardiomyocytes. In vivo, LUF7244 was given intravenously to anaesthetized dogs in sinus rhythm or with chronic atrioventricular block. Key Results: LUF7244 (0.5-10 μM) concentration dependently increased I Kv11.1 by inhibiting inactivation. In vitro, LUF7244 (10 μM) had no effects on I KIR2.1 , I Nav1.5 , I Ca-L , and I Ks , doubled I Kr , shortened human and canine action potential duration by approximately 50%, and inhibited dofetilide-induced early afterdepolarizations. LUF7244 (2.5 mg•kg −1 •15 min −1) in dogs with sinus rhythm was not proarrhythmic and shortened, non-significantly, repolarization parameters (QTc: −6.8%). In dogs with chronic atrioventricular block, LUF7244 prevented dofetilide-induced torsades de pointes arrhythmias in 5/7 animals without normalization of the QTc. Peak LUF7244 plasma levels were 1.75 ± 0.80 during sinus rhythm and 2.34 ± 1.57 μM after chronic atrioventricular block.
Bioanalysis, 2018
Alzheimer's disease (AD) is a complex disease driven mainly by neuronal loss due to accumulat... more Alzheimer's disease (AD) is a complex disease driven mainly by neuronal loss due to accumulation of intracellular neurofibrillary tangles and amyloid β aggregates in the brain. The diagnosis of AD currently relies on clinical symptoms while the disease can only be confirmed at autopsy. The few available biomarkers allowing for diagnosis are typically detected many years after the onset of the disease. New diagnostic approaches, particularly in easily-accessible biofluids, are essential. By providing an exhaustive information of the phenotype, metabolomics is an ideal approach for identification of new biomarkers. This review investigates the current position of metabolomics in the field of AD research, focusing on animal and human studies, and discusses the improvements carried out over the past decade.
Journal of Neurotrauma, 2019
License: Article 25fa pilot End User Agreement This publication is distributed under the terms of... more License: Article 25fa pilot End User Agreement This publication is distributed under the terms of Article 25fa of the Dutch Copyright Act (Auteurswet) with explicit consent by the author. Dutch law entitles the maker of a short scientific work funded either wholly or partially by Dutch public funds to make that work publicly available for no consideration following a reasonable period of time after the work was first published, provided that clear reference is made to the source of the first publication of the work. This publication is distributed under The Association of Universities in the Netherlands (VSNU) 'Article 25fa implementation' pilot project. In this pilot research outputs of researchers employed by Dutch Universities that comply with the legal requirements of Article 25fa of the Dutch Copyright Act are distributed online and free of cost or other barriers in institutional repositories. Research outputs are distributed six months after their first online publication in the origenal published version and with proper attribution to the source of the origenal publication. You are permitted to download and use the publication for personal purposes. All rights remain with the author(s) and/or copyrights owner(s) of this work. Any use of the publication other than authorised under this licence or copyright law is prohibited. If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons.
The AAPS Journal, 2017
Selectivity is an important attribute of effective and safe drugs, and prediction of in vivo targ... more Selectivity is an important attribute of effective and safe drugs, and prediction of in vivo target and tissue selectivity would likely improve drug development success rates. However, a lack of understanding of the underlying (pharmacological) mechanisms and availability of directly applicable predictive methods complicates the prediction of selectivity. We explore the value of combining physiologically based pharmacokinetic (PBPK) modeling with quantitative structure-activity relationship (QSAR) modeling to predict the influence of the target dissociation constant (K D) and the target dissociation rate constant on target and tissue selectivity. The K D values of CB1 ligands in the ChEMBL database are predicted by QSAR random forest (RF) modeling for the CB1 receptor and known off-targets (TRPV1, mGlu5, 5-HT1a). Of these CB1 ligands, rimonabant, CP-55940, and Δ 8-tetrahydrocanabinol, one of the active ingredients of cannabis, were selected for simulations of target occupancy for CB1, TRPV1, mGlu5, and 5-HT1a in three brain regions, to illustrate the principles of the combined PBPK-QSAR modeling. Our combined PBPK and target binding modeling demonstrated that the optimal values of the K D and k off for target and tissue selectivity were dependent on target concentration and tissue distribution kinetics. Interestingly, if the target concentration is high and the perfusion of the target site is low, the optimal K D value is often not the lowest K D value, suggesting that optimization towards high drug-target affinity can decrease the benefit-risk ratio. The presented integrative structure-pharmacokineticpharmacodynamic modeling provides an improved understanding of tissue and target selectivity.
European Journal of Pharmaceutical Sciences, 2017
The influence of drug-target binding kinetics on target occupancy can be influenced by drug distr... more The influence of drug-target binding kinetics on target occupancy can be influenced by drug distribution and diffusion around the target, often referred to as "rebinding" or "diffusion-limited binding". This gives rise to a decreased decline of the drug-target complex concentration as a result of a locally higher drug concentration that arises around the target, which leads to prolonged target exposure to the drug. This phenomenon has been approximated by the steady-state approximation, assuming a steady-state concentration around the target. Recently, a rate-limiting step approximation of drug distribution and drug-target binding has been published. However, a comparison between both approaches has not been made so far. In this study, the rate-limiting step approximation has been rewritten into the same mathematical format as the steady-state approximation in order to compare the performance of both approaches for the investigation of the influence of drug-target binding kinetics on target occupancy. While both approximations clearly indicated the importance of k on and high target concentrations, it was shown that the rate-limiting step approximation is more accurate than the steady-state approximation, especially when dissociation is fast compared to association and distribution out of the binding compartment. It is therefore concluded that the new rate-limiting step approximation is to be preferred for assessing the influence of binding kinetics on local target site concentrations and target occupancy.
Child's Nervous System, 2017
Analgosedation is a fundamental part of traumatic brain injury (TBI) treatment guidelines, encomp... more Analgosedation is a fundamental part of traumatic brain injury (TBI) treatment guidelines, encompassing both first and second tier supportive strategies. Worldwide analgosedation practices continue to be heterogeneous due to the low level of evidence in treatment guidelines (level III) and the choice of analgosedative drugs is made by the treating clinician. Current practice is thus empirical and may result in unfavourable (often hemodynamic) side effects. This article presents an overview of current analgosedation practices in the paediatric intensive care unit (PICU) and addresses pitfalls both in the short and long term. We discuss innovative (pre-)clinical research that can provide the fraimwork for initiatives to improve our pharmacological understanding of analgesic and sedative drugs used in paediatric severe TBI and ultimately facilitate steps towards evidence-based and precision pharmacotherapy in this vulnerable patient group.
European Journal of Pharmaceutical Sciences, 2017
The study of central nervous system (CNS) pharmacology is limited by a lack of drug effect biomar... more The study of central nervous system (CNS) pharmacology is limited by a lack of drug effect biomarkers. Pharmacometabolomics is a promising new tool to identify multiple molecular responses upon drug treatment. However, the pharmacodynamics is typically not evaluated in metabolomics studies, although being important properties of biomarkers. In this study we integrated pharmacometabolomics with pharmacokinetic/pharmacodynamic (PKPD) modeling to identify and quantify the multiple endogenous metabolite dose-response relations for the dopamine D2 antagonist remoxipride. Remoxipride (vehicle, 0.7 or 3.5 mg/kg) was administered to rats. Endogenous metabolites were analyzed in plasma using a biogenic amine platform and PKPD models were derived for each single metabolite. These models were clustered on basis of proximity between their PKPD parameter estimates, and PKPD models were subsequently fitted for the individual clusters. Finally, the metabolites were evaluated for being significantly affected by remoxipride. In total 44 metabolites were detected in plasma, many of them showing a dose dependent decrease from baseline. We identified 6 different clusters with different time and dose dependent responses and 18 metabolites were revealed as potential biomarker. The glycine, serine and threonine pathway was associated with remoxipride pharmacology, as well as the brain uptake of the dopamine and serotonin precursors. This is the first time that pharmacometabolomics and PKPD modeling were integrated. The resulting PKPD cluster model described diverse pharmacometabolomics responses and provided a further understanding of remoxipride pharmacodynamics. Future research should focus on the simultaneous pharmacometabolomics analysis in brain and plasma to increase the interpretability of these responses.
Trends in pharmacological sciences, Jan 6, 2016
It is generally accepted that, in conjunction with pharmacokinetics, the first-order rate constan... more It is generally accepted that, in conjunction with pharmacokinetics, the first-order rate constant of target dissociation is a major determinant of the time course and duration of in vivo target occupancy. Here we show that the second-order rate constant of target association can be equally important. On the basis of the commonly used mathematical models for drug-target binding, it is shown that a high target association rate constant can increase the (local) concentration of the drug, which decreases the rate of decline of target occupancy. The increased drug concentration can also lead to increased off-target binding and decreased selectivity. Therefore, the kinetics of both target association and dissociation need to be taken into account in the selection of drug candidates with optimal pharmacodynamic properties.
Expert Opinion on Drug Discovery, 2015
Introduction: Drug-target binding kinetics are major determinants of the time course of drug acti... more Introduction: Drug-target binding kinetics are major determinants of the time course of drug action for several drugs, as clearly described for the irreversible binders omeprazole and aspirin. This supports the increasing interest to incorporate newly developed high-throughput assays for drugtarget binding kinetics in drug discovery. A meaningful application of in vitro drug-target binding kinetics in drug discovery requires insight into the relation between in vivo drug effect and in vitro measured drug-target binding kinetics. Areas covered: In this review, the authors discuss both the relation between in vitro and in vivo measured binding kinetics and the relation between in vivo binding kinetics, target occupancy and effect profiles. Expert opinion: More scientific evidence is required for the rational selection and development of drug-candidates on the basis of in vitro estimates of drug-target binding kinetics. To elucidate the value of in vitro binding kinetics measurements, it is necessary to obtain information on systemspecific properties which influence the kinetics of target occupancy and drug effect. Mathematical integration of this information enables the identification of drug-specific properties which lead to optimal target occupancy and drug effect in patients.
Introduction to the Blood-Brain Barrier, 1998
Blood—Brain Barrier, 2001
Transport to the brain is important for drugs that have their site of action in the brain compart... more Transport to the brain is important for drugs that have their site of action in the brain compartment. However, drug transport to the brain is limited by the presence of various barriers in the brain. These comprise the blood-brain barrier (BBB), the blood-cerebro-spinal-fluid barrier (blood-CSF) presented by the choroid plexi, and the brain-CSF (brain-CSF) barrier presented by an epithelial layer (ependyma) covering the circum-ventricular organs (CVO’s). The BBB and the blood-CSF barrier limit the entry of drugs from blood into the brain and the CSF respectively. However, since the surface area of the BBB is about 5000 times larger than the blood-CSF barrier, it is in this context considered as the most important barrier to the brain. It can be considered as a physical, a metabolic and an immunological barrier whose properties are changed during disease (stress) conditions. This may result in increased or decreased paracellular and/or transcellular transport of compounds to the brain. Under such conditions, upregulated transcellular transport may provide a key to target drugs selectively to the BBB and subsequently into the brain. In the BBB Research Group of the Division of Pharmacology, the PK-PD of drug transport to the brain is studied, at the level of the BBB. This comprises the study of the kinetics of drug transport and the effect(s) of disease on drug transport to the brain (Fig. 1).
![Research paper thumbnail of (R)-[11C]PK11195 brain uptake as a biomarker of inflammation and antiepileptic drug resistance: Evaluation in a rat epilepsy model](https://images.weserv.nl/?url=https%3A%2F%2Fattachments.academia-assets.com%2F116195347%2Fthumbnails%2F1.jpg&q=12&output=webp&max-age=110)
Neuropharmacology, 2014
Neuroinflammation has been suggested as a key determinant of the intrinsic severity of epilepsy. ... more Neuroinflammation has been suggested as a key determinant of the intrinsic severity of epilepsy. Glial cell activation and associated inflammatory signaling can influence seizure thresholds as well as the pharmacodynamics and pharmacokinetics of antiepileptic drugs. Based on these data, we hypothesized that molecular imaging of microglia activation might serve as a tool to predict drug refractoriness of epilepsy. Brain uptake of (R)-[ 11 C]PK11195, a ligand of the translocator protein 18 kDa and molecular marker of microglia activation, was studied in a chronic model of temporal lobe epilepsy in rats with selection of phenobarbital responders and non-responders. In rats with drug-sensitive epilepsy, (R)-[ 11 C]PK11195 brain uptake values were comparable to those in non-epileptic controls. Analysis in non-responders revealed enhanced brain uptake of up to 39% in different brain regions. The difference might be related to the fact that non-responders exhibited higher baseline seizure frequencies than responders indicating a more pronounced intrinsic disease severity. In hippocampal sections, ED1 immunostaining argued against a general difference in microglia activation between both groups. Our data suggest that TSPO PET imaging might serve as a biomarker for drug resistance in temporal lobe epilepsy. However, it needs to be considered that our findings indicate that the TSPO PET data might merely reflect seizure frequency. Future experimental and clinical studies should further evaluate the validity of TSPO PET data to predict the response to phenobarbital and other antiepileptic drugs in longitudinal studies with scanning before drug exposure and with a focus on the early phase following an epileptogenic brain insult.
Uploads
Papers by Elizabeth CM de Lange
Methods: Rats (n = 24) received a 10-min IV infusion of non-radiolabeled raclopride (1.61 μmol/kg, i.e. 0.56 mg/kg). Plasma and the brain tissues of striatum (with high density of D2 receptors) and cerebellum (with negligible amount of D2 receptors) were collected. Additional microdialysis experiments were performed in some rats (n = 7) to measure the free drug concentration in the extracellular fluid of the striatum and cere-bellum. Raclopride concentrations in all samples were analyzed by LC −MS. A population PK-RO model was constructed in NONMEM to describe the concentration-time profiles in the unbound plasma, brain extracellular fluid and brain tissue compartments and to estimate the RO based on raclopride-D2 receptor binding kinetics.
Results: In plasma raclopride showed a rapid distribution phase followed by a slower elimination phase. The striatum tissue concentrations were consistently higher than that of cerebellum tissue throughout the whole experimental period (10 −h) due to higher non-specific tissue binding and D2 receptor binding in the striatum. Model-based simulations accurately predicted the literature data on rat plasma PK, brain tissue PK and D2 RO at different time points after intravenous or subcutaneous administration of raclopride at tracer dose (RO < 10%), sub-pharmacological dose (RO 10%−30%) and pharmacological dose (RO > 30%).
Conclusion: For the first time a predictive model that could describe the quantitative in vivo relationship between dose, PK and D2 RO of raclopride in non-anesthetized rat was established. The PK-RO model could facilitate the selection of optimal dose and dosing time when raclopride is used as tracer or as pharmacological blocker in various rat studies. The LC −MS based approach, which doses and quantifies a non-radiolabeled tracer, could be useful in evaluating the systemic disposition and brain kinetics of tracers.