Brain Distribution Studies Research Articles

Co-administration strategy to enhance brain accumulation of vandetanib by modulating P-glycoprotein (P-gp/Abcb1) and breast cancer resistance protein (Bcrp1/Abcg2) mediated efflux with m-TOR inhibitors

The objectives of this study were (i) to characterize the interaction of vandetanib with P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp1) in vitro and in vivo (ii) to study the modulation of P-gp and BCRP mediated efflux of vandetanib with specific transport inhibitors and m-TOR inhibitors, everolimus and temsirolimus. Cellular accumulation and bi-directional transport studies in MDCKII cell monolayers were conducted to delineate the role of efflux transporters on disposition of vandetanib. Brain distribution studies were conducted in male FVB wild-type mice with vandetanib administered intravenously either alone or in the presence of specific inhibitors and m-TOR inhibitors. In vitro studies suggested that vandetanib is a high affinity substrate of Bcrp1 but is not transported by P-gp. Interestingly, in vivo brain distribution studies in FVB wild type mice indicated that vandetanib penetration into the brain is restricted by both Bcrp1 and P-gp mediated active efflux at the blood brain barrier (BBB). Co-administration of elacridar, a dual P-gp/BCRP inhibitor increased the brain to plasma concentration ratio of vandetanib upto 5 fold. Of the two m-TOR pathway inhibitors examined; everolimus showed potent effect on modulating vandetanib brain penetration whereas no significant affect on vandetanib brain uptake was observed following temsirolimus co-administration. This finding could be clinically relevant as everolimus can provide synergistic pharmacological effect in addition to primary role of vandetanib efflux modulation at BBB for the treatment of brain tumors.

Quantitative determination and pharmacokinetic study of the novel anti-Parkinson's disease candidate drug FLZ in rat brain by high performance liquid chromatography–tandem mass spectrometry

FLZ (N-[2-(4-hydroxy-phenyl)-ethyl]-2-(2,5-dimethoxy-phenyl)-3-(3-methoxy-4-hydroxyphenyl)-acrylamide) is a novel anti-Parkinson's disease candidate drug. A sensitive and specific high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and validated for the quantification of FLZ in rat brain. Carbamazepine was selected as the internal standard. Sample preparation involved double liquid-liquid extraction by n-hexane and ethyl acetate with high extraction efficiency. The chromatographic separation was achieved on a Zorbax SB-C(18) column (100 mm × 2.1 mm, 3.5 μm) with an isocratic elution system comprised of acetonitrile and 0.3% aqueous acetic acid at a flow rate of 0.3 ml/min. The elutes were detected under positive electrospray ionization (ESI) and the target analytes were quantified by multiple reaction monitoring (MRM) mode. The method was sensitive with the lowest limit of quantification (LLOQ) at 1.0 ng/g brain tissue. Good linearity (r>0.99) was obtained over the range of 1.0-400 ng/g. The intra- and inter-day precision ranged from 0.68% to 12%, while the accuracy between 92.7% and 111%. In addition, the stability, recovery and matrix effect involved in this method were also validated. The method was used to investigate the pharmacokinetics of FLZ in rat brain successfully after intravenous administration. The brain distribution studies showed that the brain distribution of FLZ was limited with the penetration ratio less than 0.1 in rats, with no target effect in the seven collected regions. Inhibition of P-glycoprotein (P-gp) by zosuquidar·3HCl ((2R)-1-{4-[(1aR,10bS)-1,1-difluoro-1,1a,6,10b-tetrahydrodibenzo[a,e]cyclopropa[c][7]annulen-6-yl]-1-piperazinyl}-3-(5-quinolinyloxy)-2-propanol trihydrochloride) resulted in a significant increase in brain-to-plasma ratio, while no significant increase by inhibition of breast cancer resistance protein (BCRP) by ko143 (2-methyl-2-propanyl 3-[(3S,6S,12aS)-6-isobutyl-9-methoxy-1,4-dioxo-1,2,3,4,6,7,12,12a-octahydropyrazino[1',2':1,6]pyrido[3,4-b]indol-3-yl]propanoate). The results indicated that FLZ had poor penetration to the brain due to the P-gp transport system.

Determination of cediranib in mouse plasma and brain tissue using high-performance liquid chromatography–mass spectrometry

A new high performance liquid chromatography coupled with tandem mass spectrometry (HPLC–MS/MS) assay for cediranib, a tyrosine kinase inhibitor for VEGFRs, was developed and validated, for the determination of plasma and brain levels of cediranib in small specimen volumes. Tyrphostin (AG1478) was used as internal standard. Mouse plasma and brain homogenate samples were prepared using liquid–liquid extraction. The assay was validated for a 2.5–2500 ng/mL concentration range for plasma, and for 1–2000 ng/mL range for brain homogenate. For these calibration ranges, within-assay variabilities were 1.1–14.3% for plasma and 1.5–9.4% for brain homogenate; between-assay variabilities were 2.4–9.2% for plasma, and 4.9–10.2% for brain homogenate. Overall accuracy ranged from 101.5 to 107.0% for plasma and 96.5 to 100.2% for brain homogenate, for all target concentrations. The developed assay has been successfully applied for a brain distribution study in mice at an oral dose of 5 mg/kg.

Brain Distribution Study of Imperatorin in Rats after Oral Administration Assessed by HPLC

Imperatorin (IMP) is a biologically active ingredient isolated from a traditional Chinese medicine (TCM), Angelica dahurica. To obtain the brain distribution data of IMP in rats, the concentrations of IMP in cortex, cerebellum, diencephalon, brain stem, striatum and hippocampus were measured by a simple and sensitive HPLC–UV method. The analytes were prepared by a liquid–liquid extraction method and the separation of IMP was performed on a Hypersil BDS C18 column (250 mm × 4.6 mm i.d., 5 μm) using acetonitrile–water (60:40, v/v) as mobile phase which was delivered at 1.0 mL min−1. Ultraviolet detection was performed at 300 nm. Using a weighted (1/c2) least square method, linear calibration curves for the six regions were obtained (r ≥ 0.9990) with a lower limit of quantification (LLOQ) of 0.075 μg g−1 for cortex, cerebellum, diencephalon and brain stem or 0.15 μg g−1 for striatum and hippocampus, and the recovery was greater than 90% for each tissue sample. The within- and between-day precisions (expressed as the relative standard deviation, RSD) were less than 10%. The validated method has been successfully applied to the brain distribution study in rats. The results showed that IMP could pass through the blood–brain barrier easily. And the higher concentration in striatum and hippocampus compared with the others might indicate that they were the target regions of IMP in rat brain.

Poly(ethylene oxide/propylene oxide) copolymer thermo-reversible gelling system for the enhancement of intranasal zidovudine delivery to the brain

The purpose of this study was to investigate the olfactory transfer of zidovudine (ZDV) after intranasal (IN) administration and to assess the effect of thermoreversible gelling system on its absorption and brain uptake. The nasal formulation was prepared by dissolving ZDV in pH 5.5 phosphate buffer solution comprising of 20% polyethylene oxide/propylene oxide (Poloxamer 407, PLX) as thermoreversible gelling agent and 0.1% n-tridecyl-β- d-maltoside (TDM) as permeation enhancer. This formulation exhibited a sufficient stability and an optimum gelation profile at 27–30 °C. The in vitro permeation studies across the freshly excised rabbit nasal mucosa showed a 53% increase in the permeability of ZDV from the formulation. For in vivo evaluation, the drug concentrations in the plasma, cerebrospinal fluid (CSF) and six different regions of the brain tissues, i.e. olfactory bulb (OB), olfactory tract (OT), anterior, middle and posterior segments of cerebrum (CB), and cerebellum (CL) were determined by LC/MS method following IV and IN administration in rabbits at a dose of 1 mg/kg. The IN administration of Poloxamer 407 and TDM based formulation showed a systemic bioavailability of 29.4% while exhibiting a 4 times slower absorption process ( t max = 20 min) than control solution ( t max = 5 min). The CSF and brain ZDV levels achieved after IN administration of the gelling formulation were approximately 4.7–56 times greater than those attained after IV injection. The pharmacokinetic and brain distribution studies revealed that a polar antiviral compound, ZDV could preferentially transfer into the CSF and brain tissue via an alternative pathway, possibly olfactory route after intranasal administration.

Microemulsion-based novel transdermal delivery system of tetramethylpyrazine: preparation and evaluation in vitro and in vivo

ObjectiveTo deliver 2,3,5,6-tetramethylpyrazine (TMP) in a relatively large dose through a transdermal route and facilitate the practical application of microemulison in transdermal drug delivery.MethodsThe pseudo-ternary phase diagram for microemulsion regions was constructed using isopropyl myristate as oil phase, Labrasol® as surfactant, and Plurol® Oleique CC 497 as cosurfactant. A uniform experimental design was applied for formulation optimization. In vitro skin permeation experiments of six formulations were undertaken with TMP transdermal patch (EUDRAGIT® E100 as matrix) and TMP saturated solution as controls. We prepared TMP-oil dispersed in water-ethylene vinyl acetate-transdermal therapeutic system (TMP-O/W-EVA-TTS) with microemulsion as reservoir and EVA membrane as release liner; pharmacokinetic and brain distribution studies in rats were conducted with TMP transdermal patches as control.ResultsThe skin fluxes of TMP from microemulsions were 8.2- to 26.7-fold and 0.9- to 4.7-fold higher than those of TMP transdermal patch and TMP saturated solution, respectively, and were strongly affected by the microemulsion composition. The improvement in TMP solubility as well as the skin permeation enhancement effect of microemulsion components contributed mainly to transdermal delivery facilitation. In the pharmacokinetic study, the relative bioavailability of TMP-O/W-EVA-TTS was 350.89% compared with the TMP transdermal patch. Higher and more stable TMP contents in rat plasma were obtained after administration of TMP-O/WEVA- TTS than after application of TMP transdermal patch. In the brain distribution study, higher rate and extent of TMP distribution to brain, and lower rate of TMP clearance from brain were observed after transdermal administration of TMP-O/W-EVA-TTS than after application of TMP transdermal patch.ConclusionThe novel transdermal delivery system prepared in this study showed a remarkable skin permeation improvement of microemulsion and facilitated its practical application in transdermal drug delivery. With this system as a vehicle, a relatively large dose of TMP could enable successful drug delivery via the transdermal route.

Functionalized gold nanoparticles: a detailed in vivo multimodal microscopic brain distribution study

In the present study, the in vivo distribution of polyelectrolyte multilayer coated gold nanoparticles is shown, starting from the living animal down to cellular level. The coating was designed with functional moieties to serve as a potential nano drug for prion disease. With near infrared time-domain imaging we followed the biodistribution in mice up to 7 days after intravenous injection of the nanoparticles. The peak concentration in the head of mice was detected between 19 and 24 h. The precise particle distribution in the brain was studied ex vivo by X-ray microtomography, confocal laser and fluorescence microscopy. We found that the particles mainly accumulate in the hippocampus, thalamus, hypothalamus, and the cerebral cortex.

P-glycoprotein and Breast Cancer Resistance Protein Influence Brain Distribution of Dasatinib

The novel tyrosine kinase inhibitor dasatinib (Sprycel; BMS-354825) is approved for use in imatinib (Gleevec; STI 571)-resistant or -intolerant chronic myelogenous leukemia and may be useful for other tumors in the central nervous system (CNS). The objective of this study was to investigate the role of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in modulating the CNS penetration of dasatinib. Results from the in vitro studies indicate that cellular delivery of dasatinib is significantly limited by active efflux due to both P-gp and BCRP. Permeability studies indicated greater permeability in the basolateral-to-apical direction than in the apical-to-basolateral direction due to active efflux by P-gp or BCRP. Selective inhibitors of P-gp and BCRP, such as (R)-4-((1aR,6R,10bS)-1,2-difluoro-1,1a,6,10b-tetrahydrodibenzo-(a,e)cyclopropa(c) cycloheptan-6-yl)-alpha-((5-quinoloyloxy)methyl)-1-piperazineethanol, trihydrochloride (zosuquidar; LY335979) and 3-(6-isobutyl-9-methoxy-1,4-dioxo-1,2,3,4,6,7,12,12alpha-octahydropyrazino1',2': 1,6pryrido3,4-bindol-3-yl)-propionic acid tert-butyl ester (Ko143), were able to restore the intracellular accumulation and abolish the directionality in net flux of dasatinib. In vivo brain distribution studies showed that the CNS distribution of dasatinib is limited, with the brain-to-plasma concentration ratios less than 0.12 in wild-type mice, which increased approximately 8-fold in Mdr1a/b(-/-) Bcrp1(-/-) mice. Dasatinib brain distribution was significantly increased in Mdr1a/b(-/-) mice and when wild-type mice were pretreated with LY335979. Simultaneous inhibition of P-gp and BCRP by elacridar [N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide] (GF120918) resulted in a 5-fold increase in brain concentration. These in vitro and in vivo studies demonstrate that dasatinib is a substrate for the important efflux transporters p-glycoprotein and BCRP. These transport systems play a significant role in limiting the CNS delivery of dasatinib and may have direct implications in the treatment of primary and metastatic brain tumors.

Synthesis and biological evaluation of one novel technetium-99m-labeled nitroquipazine derivative as an imaging agent for serotonin transporter

Imaging of serotonin transporter (SERT) by positron emission tomography (PET) or single-photon emission-computed tomography (SPECT) in humans would provide useful information in diagnosis and therapy of several neurodegenerative and neuropsychiatric disorders. 6-Nitroquipazine is a highly potent and selective inhibitor of the SERT. For the development of new 99mTc-labeled 6-nitroquipazine derivatives as SERT imaging agents, novel [ N-[2-((3-(4-(6-nitroquinolin-2-yl)piperazin-1-yl)propyl)(2-mercaptoethyl)amino]-acetyl-2-aminoethanethiolato] [ 99mTc]technetium (V) oxide ( 99mTc-MAMA-3-PQ) and its rhenium analog were synthesized and characterized. 99mTc-MAMA-3-PQ displayed high initial brain uptake (0.52% ID/organ at 2 min post-injection (pi)) and relatively fast washout in mice (0.09% ID/organ at 60 min pi). The regional brain distribution studies in rats showed high-specific binding ratios at 60 min pi. Maximum regional contrast ratio observed for thalamus/cerebellum was 2.94, followed by 2.62 for hypothalamus/cerebellum. These encouraging results lead us to further explore its derivatives as new imaging agents for the SERT in the brain.

Evaluation of [11C]rofecoxib as PET tracer for cyclooxygenase 2 overexpression in rat models of inflammation

Overexpression of cyclooxygenase type 2 (COX-2) is triggered by inflammatory stimuli, but it also plays a prominent role in the initiation and progression of various diseases. This study aims to investigate [(11)C]rofecoxib as a positron emission tomography (PET) tracer for COX-2 expression. [(11)C]Rofecoxib was prepared by methylation of its sulphinate precursor. Regional brain distribution and specific binding of [(11)C]rofecoxib in healthy rats was studied by ex vivo biodistribution and autoradiography. Regional brain distribution and PET imaging studies were also performed on rats with severe encephalitis, caused by nasal infection with herpes simplex virus (HSV). Finally, ex vivo biodistribution and blocking studies were carried in rats with a sterile inflammation, induced by intramuscular turpentine injection. [(11)C]rofecoxib brain uptake in control animals corresponded with the known distribution of COX-2. Pretreatment with NS398 significantly reduced tracer uptake in the cingulate/frontopolar cortex, whereas the reduction in hippocampus approached significance. Ex vivo autoradiography also revealed preferential tracer uptake in hippocampus and cortical areas that could be blocked by NS398. In HSV-infected animals, [(11)C]rofecoxib uptake was moderately increased in all brain regions, but it could not be blocked with indomethacin. Yet, some PET images revealed increased tracer uptake in brain areas with microglia activation. In turpentine-injected animals, [(11)C]rofecoxib uptake in inflamed muscle was not higher than in control muscle and could not be blocked with NS398. Indomethacin caused a slight reduction in muscle uptake. Despite the apparent correlation between [(11)C]rofecoxib uptake and COX-2 distribution in healthy rats, [(11)C]rofecoxib could not unambiguously detect COX-2 overexpression in two rat models of inflammation.

P-glycoprotein-Mediated Active Efflux of the Anti-HIV1 Nucleoside Abacavir Limits Cellular Accumulation and Brain Distribution

P-glycoprotein (P-gp)-mediated efflux at the blood-brain barrier has been implicated in limiting the brain distribution of many anti-HIV1 drugs, primarily protease inhibitors, resulting in suboptimal concentrations in this important sanctuary site. The objective of this study was to characterize the interaction of abacavir with P-gp and determine whether P-gp is an important mechanism in limiting abacavir delivery to the central nervous system (CNS). In vitro and in vivo techniques were employed to characterize this interaction. Abacavir stimulated P-gp ATPase activity at high concentrations. The cellular accumulation of abacavir was significantly decreased by approximately 70% in Madin-Darby canine kidney II (MDCKII)-MDR1 monolayers compared with wild-type cells and was completely restored by the P-gp inhibitors ((R)-4-((1aR,6R,10bS)-1,2-difluoro-1,1a,6,10b-tetrahydrodibenzo(a,e)cyclopropa(c)cycloheptan-6-yl)-alpha-((5-quinoloyloxy)methyl)-1-piperazineethanol, trihydrochloride) (LY335979) and N-[4-[2-(6,7-dimethoxy-3,4-dihydro-1H-isoquinolin-2-yl)ethyl]phenyl]-5-methoxy-9-oxo-10H-acridine-4-carboxamide (GF120918). Directional flux experiments indicated that abacavir had greater permeability in the basolateral-to-apical direction (1.58E-05 cm/s) than in the apical-to-basolateral direction (3.44E-06 cm/s) in MDR1-transfected monolayers. The directionality in net flux was abolished by both LY335979 and GF120918. In vivo brain distribution studies showed that the AUC(plasma) in mdr1a(-/-) CF-1 mutant mice was approximately 2-fold greater than the AUC(plasma) in the wild type, whereas the AUC(brain) in the mutant was 20-fold higher than that in the wild type. Therefore, the CNS drug targeting index, defined as the ratio of AUC brain-to-plasma for mutant over wild type, was greater than 10. These data are the first in vitro and in vivo evidence that a nucleoside reverse transcriptase inhibitor is a P-gp substrate. The remarkable increase in abacavir brain distribution in P-gp-deficient mutant mice over wild-type mice suggests that P-gp may play a significant role in restricting the abacavir distribution to the CNS.

Preparation and biodistribution of 99mTc-tricarbonyl complex with 4-[(2-methoxyphenyl)piperazin-1-yl]-dithioformate as a potential 5-HT 1A receptor imaging agent

The goal of this study is to develop a novel 5-HT 1A receptor imaging agent. 4-[(2-methoxyphenyl)piperazin-1-yl]-dithioformate (MPPDTF) was labeled with 99mTc-tricarbonyl core via dithioformate moiety in high yield (>96% by HPLC). 99mTc(CO) 3–MPPDTF is a neutral and lipophilic complex, which was confirmed by paper electrophoresis and octanol/water partition coefficient ( P=27.0±1.4, n=3), respectively. In vivo biodistribution indicated that this complex had moderate brain uptake (0.53±0.10% ID/g at 5 min and 0.42±0.02% ID/g at 120 min) and good retention (about 80% of the activity was retained in the brain at 120 min post-injection). Regional brain distribution study showed that hippocampus, where the 5-HT 1A receptor density is high, had the highest uptake (0.60±0.02% ID/g at 5 min p.i.) and the cerebellum, where the 5-HT 1A receptor density is low, had the lowest uptake (0.10±0.02% ID/g at 5 min p.i.). After blocking with 8-OH-DPAT, the hippocampus uptake was decreased obviously while the cerebellum uptake was increased slightly. This result indicates that 99mTc(CO) 3–MPPDTF complex has specific binding to 5-HT 1A receptor.

Transport, Metabolism, and in Vivo Population Pharmacokinetics of the Chloro Benztropine Analogs, a Class of Compounds Extensively Evaluated in Animal Models of Drug Abuse

Recently, extensive behavioral research has been conducted on the benztropine (BZT) analogs with the goal of developing successful therapeutics for cocaine abuse. The present study was conducted to characterize the contribution of dispositional factors in mediating the behavioral differences among the chloro BZT analogs and to identify cytochrome P450 enzymes involved in their metabolism. Bidirectional transport and efflux studies of four of the chloro BZT analogs were conducted. Screening with a panel of human and rat Supersomes was performed for 4',4''-diCl BZT. In addition, pharmacokinetic and brain distribution studies for 4'-Cl and 4',4''-diCl BZT in Sprague-Dawley rats were conducted. The permeability of the chloro analogs ranged from 8.26 to 32.23 and from 1.37 to 21.65 x 10(-6) cm/s, whereas the efflux ratios ranged from 2.1 to 6.9 and from 3.3 to 28.4 across Madin-Darby canine kidney-multidrug resistance 1 (MDCK-MDR1) and Caco-2 monolayers, respectively. The P-glycoprotein (P-gp) inhibitor verapamil reduced the efflux ratios and enhanced the absorptive transport of the chloro BZT analogs. 4',4''-diCl BZT was a substrate of human CYP2D6 and 2C19 and rat 2C11 and 3A1. The brain uptake for 4'-Cl and 4',4''-diCl BZT was comparable and higher than previously reported for cocaine (brain-to-plasma partition coefficient = 4.6-4.7 versus 2.1 for cocaine). The rank order for t(1/2) was 4',4''-diCl BZT >> 4'-Cl BZT > cocaine and for steady-state volume of distribution was 4'-Cl BZT > 4',4''-diCl BZT >> cocaine. In conclusion, the chloro analogs differ significantly in their clearance and duration of action, which correlates to their behavioral profiles and abuse liability. Furthermore, these results suggest that the distinctive behavioral profile of these analogs is not due to limited brain exposure.

Radiosynthesis and mouse brain distribution studies of [ 11C] CP-126,998: a PET ligand for in vivo study of acetylcholinesterase

The selective, reversible acetylcholinesterase inhibitor 5,7-Dihydro-7-methyl-3- [2-[1-(phenylmethyl]-4-piperidinyl]ethyl]-6 H-pyrrolo[3,2- f]-1,2-benzisoxazol3–6-one (CP-126,998) was labeled with C-11 iodomethane via base-promoted alkylation of the lactam nitrogen. [ 11C] CP-126,998 was synthesized in good radiochemical yield (13–29% non-decay corrected) and high specific radioactivity (177–418 GBq/μmol). In vivo mouse biodistribution studies reveal [ 11C] CP-126,998 to localize preferentially in striatal tissue, a region known to be rich in acetylcholinesterase. Competitive blocking studies using a variety of acetylcholinesterase inhibitors (diisopropylfluorophosphate, tacrine, CP-118,954) verified the specificity of the PET radiotracer for brain acetylcholinesterase.

New radiolabeled CCK-8 analogues [Tc-99m-GH-CCK-8 and Tc-99m-DTPA-CCK-8]: preparation and biodistribution studies in rats and rabbits

The aim of this study is to label CCK-8 with Tc-99m and to investigate its radiopharmaceutical potential. CCK-8 was labeled with Tc-99m using GH and DTPA as bifunctional chelating agents. Labeling efficiency was higher than 99%. Complex was stable more than 5 hours at room temperature. 37 MBq Tc-99m-GH-CCK-8 or Tc-99m-DTPA-CCK-8 was administered intravenously to rabbits for biodistribution experiments. Dynamic and static images were obtained from anterior projection using a Camstar XC/T gamma camera. For quantitative evaluation, regions of interest were drawn on organs and time-activity curves were generated. The highest accumulation occurred in brain within 10 and 30 minutes after injection. Renal and hepatobiliary excretion were observed. Brain distribution studies in rats showed the highest activity was in hypothalamus. Results demonstrated that Tc-99m-GH-CCK-8 and Tc-99m-DTPA-CCK-8 analogs may be a useful new class of receptor-binding peptides for diagnosis and therapy of brain diseases related with CCK-B receptor-expressing tumors.

A strategy for increasing the brain uptake of a radioligand in animals: use of a drug that inhibits plasma protein binding

A positron-emitter labeled radioligand for the glycine-binding site of the N-methyl- d-aspartate (NMDA) receptor, [ 11C]L-703,717, was examined for its ability to penetrate the brain in animals by simultaneous use with drugs having high-affinity separate binding sites on human serum albumin. [ 11C]L-703,717 has poor blood–brain barrier (BBB) permeability because it binds tightly to plasma proteins. Co-injection of warfarin (50–200 mg/kg), a drug that binds to albumin and resembles L-703,717 in structure, dose-dependently enhanced the penetration by [ 11C]L-703,717 in mice, resulting in a five-fold increase in the brain radioactivity at 1 min after the injection. Drugs structurally unrelated to L-703,717, salicylate, phenol red, and L-tryptophan, were less effective or ineffective in increasing the uptake of [ 11C]L-703,717. These results suggest that the simultaneous use of a drug that inhibits the binding of a radioligand to plasma proteins is a useful way to overcome the poor BBB permeability of the radioligand triggered by its tight binding to plasma proteins. In brain distribution studies in rodents, it was found that, after the increase in brain uptake with warfarin, much of the glycine site antagonist accumulates in the cerebellum but its pharmacological specificity did not match the glycine site of NMDA receptors.

Rapid and differential losses of in vivo dopamine transporter (DAT) and vesicular monoamine transporter (VMAT2) radioligand binding in MPTP-treated mice.

The dose- and time-dependent changes of in vivo radioligand binding to the neuronal membrane dopamine transporter (DAT) and vesicular monoamine transporter type 2 (VMAT2) were examined in mouse brain after MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) administrations. Regional brain distribution studies were done in male C57BL/6 mice using simultaneous injections of d-threo-[(3)H]methylphenidate (DAT) and (+)-alpha-[(11)C]dihydrotetrabenazine (VMAT2). Single (55 mg/kg i.p. ) or multiple (4 x 10 mg/kg i.p., 1-hour intervals) administration of MPTP caused significant reductions in [(3)H]methylphenidate and [(11)C]dihydrotetrabenazine specific striatal binding, measured 14 days later. The single high dose of MPTP produced greater losses of [(11)C]dihydrotetrabenazine binding than did the multiple MPTP dosing regimen. Using the single high dose of MPTP, changes of in vivo binding of the two radioligands were determined at 1, 3, and 14 days after neurotoxin injection. At 1 day, there are large losses of [(3)H]methylphenidate binding (DAT) but no changes in [(11)C]dihydrotetrabenazine binding to the VMAT2 site in the striatum. At 3 and 14 days, there were >50% losses of binding of both bot radioligands, but significantly (P < 0.001) greater losses of VMAT2 binding of [(11)C]dihydrotetrabenazine. These studies indicate that the losses of the neuronal membrane and vesicular transporters are not always equal, and do not occur in the same time frame, after administration of the neurotoxin MPTP.

Exploratory pharmacokinetics and brain distribution study of a neuropeptide FF antagonist by liquid chromatography/atmospheric pressure ionization tandem mass spectrometry.

Dansyl-Pro-Gln-Arg-NH(2), an N-terminally modified tripeptide amide and a putative neuropeptide FF antagonist, was amenable to both positive-ion ESI and APCI. The protonated molecule yielded several fragment ions upon collision-induced dissociation in a quadrupole ion trap instrument for the development of LC/MS/MS assay methods. ESI clearly outperformed APCI in limits of detection, and was the method of choice for coupling with narrow-bore reversed-phase liquid chromatography to assess the pharmacokinetic profile and brain concentration of the neuropeptide FF antagonist in experimental animals. While plasma could be analyzed after rapid sample preparation, brain tissue required cleanup (solid phase extraction) and preconcentration before injection, and the assay was prone to matrix interference. This study indicated a rapid disappearance of dansyl-Pro-Gln-Arg-NH(2) from the plasma and the brain, and modest CNS bioavailability after intravenous administration to rats.

Synthesis and brain distribution of carbon-11 labeled analogs of antagonists for the NMDA receptor coupled PCP-binding site

Two phencyclidine (PCP) analogs, 3 and 4, and one thienylcyclohexyl-piperidine (TCP) analog, (±)6, were labeled with positron emitter carbon-11. These compounds displayed higher in vitro binding affinities than PCP itself for the PCP-binding site located inside the ion channel on the N-methyl-D-aspartate (NMDA) receptors. Brain distribution studies in mice showed different uptake characteristics between the PCP and TCP analogs, indicating their different in vivo interactions with the brain components including the PCP-binding site probably due to the different physicochemical properties of the molecules. © 1998 John Wiley & Sons, Ltd.

Brain and spinal cord distribution of biphalin: correlation with opioid receptor density and mechanism of CNS entry.

Biphalin [(Tyr-D-Ala-Gly-Phe-NH)2] is a bivalent, opioid peptide containing two pharmacophores linked by a hydrazine bridge. When administered intracerebroventricularly, it has been shown to be more potent than morphine and etorphine at eliciting antinociception. Biphalin has also been shown to cross both the blood-brain and blood-cerebrospinal fluid barriers. To understand the basis of biphalin's potency, regional brain and spinal cord distribution studies with [125I-Tyr1]biphalin were performed 5, 20, and 40 min after intravenous bolus injections. A statistically greater amount of [125I-Tyr1]biphalin was detected in the nucleus accumbens compared with other brain regions (p < 0.05). This correlates with the high density of delta- and mu-opioid receptor mRNA and binding sites shown to be expressed in the nucleus accumbens. Also, a statistically greater amount of [125I-Tyr1] biphalin was detected in two other circumventricular organs, the choroid plexus and pituitary, when compared with other brain regions. These studies provide evidence that biphalin can reach not only brain sites, but also spinal sites to elicit antinociception. The overall CNS distribution of [125I-Tyr1]biphalin was decreased with naloxone, D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2, or naltrindole pretreatment, showing that biphalin detected in the brain and spinal cord is binding to delta- and mu-opioid receptors. Additional in situ brain perfusion experiments identified a saturable component contributing to CNS entry of [125I-Tyr1]biphalin, which could be described by Michaelis-Menten kinetics with a Km of 2.6 +/- 4.8 microM, Vmax of 14.6 +/- 2.89 pmol(-1) x min(-1) x g(-1), and Kd of 0.568 +/- 0.157 microl x min(-1) x g(-1). Brain entry of [125I-Tyr1]biphalin was sensitive to 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid and L-phenylalanine, suggesting use of the large neutral amino acid carrier. This work provides evidence that biphalin is a promising, potent analgesic that has a unique mechanism for reaching both spinal and supraspinal opioid receptor sites.