0 Schaddelee, M. P. Groenendaal, D. DeJongh, J. Cleypool, C. G. I. Jzerman AP De Boer, A. G. Danhof, M. 2004 J Pharmacol Exp Ther 311 1138-46 2004/08/05 Dec 0022-3565 (Print) 15292459 Adenosine/*analogs , &, derivatives/*pharmacokinetics, Algorithms, Animals, Bayes Theorem, Blood Proteins/metabolism, Blood-Brain Barrier/*physiology, Deoxyadenosines/*pharmacology, Humans, Injections, Intravenous, Linear Models, Male, Microdialysis, Mod A population pharmacokinetic model is proposed for estimation of the brain distribution clearance of synthetic A1 receptor agonists in vivo. Rats with permanent venous and arterial cannulas in combination with a microdialysis probe in the striatum received intravenous infusions of 8-methylamino-N6-cyclopentyladenosine (MCPA) and 2'-deoxyribose-N6-cyclopentyladenosine (2'-dCPA) (10 mg kg(-1)). The clearance for transport from blood to the brain was estimated by simultaneous analysis of the blood and extracellular fluid concentrations using a compartmental pharmacokinetic model. The proposed pharmacokinetic model consists of three compartments describing the time course of the concentration in blood in combination with three compartments for the brain extracellular fluid concentrations. The blood clearance was 7.4 +/- 0.5 for MCPA and 7.2 +/- 1.4 ml min(-1) for 2'-dCPA. The in vivo microdialysis recoveries determined by the dynamic-no-net-flux method were independent of time with values of 0.21 +/- 0.02 and 0.22 +/- 0.01 for MCPA and 2'-dCPA, respectively. The values of the intercompartmental clearance for the distribution from blood to brain were 1.9 +/- 0.4 versus 1.6 +/- 0.3 mul min(-1) for MCPA and 2'-dCPA, respectively. It is concluded that on basis of the novel six-compartment model precise estimates of the rate of brain distribution are obtained that are independent of eventual differences in systemic exposure. The low brain distribution rates of MCPA and 2'-dCPA were consistent with in vitro tests. Furthermore, a slow elimination from the brain compartment was observed, indicating that the duration of central nervous system effects may be much longer than expected on the basis of the terminal half-life in blood. Schaddelee, M PGroenendaal, DDeJongh, JCleypool, C G JIJzerman, A PDe Boer, A GDanhof, MResearch Support, Non-U.S. Gov'tUnited StatesThe Journal of pharmacology and experimental therapeuticsJ Pharmacol Exp Ther. 2004 Dec;311(3):1138-46. Epub 2004 Aug 3. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15292459 0 Bueters, T. J. I. Jzerman AP van Helden, H. P. Danhof, M. 2003 Toxicol Appl Pharmacol 192 86-94 2003/10/14 Oct 1 0041-008X (Print) 14554106 Acetylcholine/metabolism, Algorithms, Animals, Area Under Curve, Behavior, Animal/drug effects, Blood-Brain Barrier/drug effects, Brain/*metabolism, Brain Chemistry/drug effects, Chemical Warfare Agents/*poisoning, Deoxyadenosines/administration , &, dos The objective of the present study was to determine (1) the influence of sarin poisoning (144 microg/kg s.c.) on the pharmacokinetics and brain distribution of the adenosine A1 receptor partial agonist 2'-deoxy-N6-cyclopentyladenosine (2'dCPA), and (2) the effect of 2'dCPA (20 mg/kg i.v.) on the central acetylcholine (ACh) release and protection against sarin toxicity. A five-compartment model successfully described the pharmacokinetic profile of 2'dCPA in blood and brain microdialysate. A covariate analysis revealed that the volume of distribution of 2'dCPA in blood was different in sarin-poisoned rats, 177 +/- 7 versus 148 +/- 8 ml in control rats. However, the transport of 2'dCPA from blood to the brain was unaffected as reflected by the values of the intercompartmental transport clearances, 0.21 +/- 0.02 and 0.21 +/- 0.04 microl/min in control and sarin-poisoned rats, respectively. Also the area-under-curve (AUC) ratios of brain microdialysate and blood were identical with values of 0.02 +/- 0.001 and 0.02 +/- 0.002, respectively, demonstrating the restricted transport of 2'dCPA into the brain in both treatment groups. Treatment of sarin-poisoned rats by 2'dCPA did not adequately prevent the accumulation of ACh in the central nervous system. 2'dCPA delayed the emergence of concomitant symptoms compared to untreated rats, but eventually only 29% of the animals survived 24 h. In conclusion, the pharmacokinetic profile of 2'dCPA in blood was slightly changed by sarin, but not the distribution of 2'dCPA into the brain. The therapeutic efficacy of 2'dCPA against sarin was limited, presumably due to insufficient quantities of 2'dCPA reaching the brain. Bueters, Tjerk J HIJzerman, Ad Pvan Helden, Herman P MDanhof, MeindertUnited StatesToxicology and applied pharmacologyToxicol Appl Pharmacol. 2003 Oct 1;192(1):86-94. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14554106 0 Bueters, T. J. van Duivenvoorde, L. M. Danhof, M. I. Jzerman AP van Helden, H. P. 2003 Eur J Pharmacol 481 141-6 2003/12/04 Nov 28 0014-2999 (Print) 14642778 Acetylcholine/*antagonists , &, inhibitors/metabolism/*secretion, Adenosine/*analogs , &, derivatives/chemistry/*pharmacology, Animals, Corpus Striatum/*drug effects/metabolism/secretion, Dose-Response Relationship, Drug, Male, Rats, Rats, Wistar, Synap The application of adenosine A(1) receptor agonists in regard to cerebral disorders is hampered by serious cardiovascular side effects. This problem might be circumvented by using low-efficacy agonists (partial agonists). The objective of the present study was to characterize the effects of the full agonist N(6)-cyclopentyladenosine (CPA) and its low-efficacy derivatives 3'-deoxy-CPA (3-DCPA), 8-propylamino-CPA (8-PCPA) and 8-butylamino-CPA (8-BCPA) on the 4-aminopyridine (4AP)-evoked release of [3H]-acetylcholine in a rat striatal synaptosomal system. The reason for studying these partial agonists in particular was their established low cardiovascular side effect profile. CPA reached a concentration-dependent maximal inhibition of the evoked acetylcholine release of 38+/-3%. 3-DCPA and 8-PCPA inhibited the acetylcholine release by 29+/-5% and 38+/-3%, respectively. On the other hand, 8-BCPA only diminished the acetylcholine release by 19+/-3%. This inhibitory effect was reversible upon coadministration of the nonselective adenosine antagonist theophylline, but not by the selective adenosine A(2A) receptor antagonist 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5- c]pyrimidine (SCH 58261). It is concluded that some partial adenosine A(1) receptor agonists behave as full agonists with respect to the inhibition of acetylcholine release, while lacking profound cardiovascular side effects. These preliminary results encourage further investigation of their tissue selectivity and therapeutic potential in vivo. Bueters, Tjerk J Hvan Duivenvoorde, Leonie MDanhof, MeindertIJzerman, Ad Pvan Helden, Herman P MNetherlandsEuropean journal of pharmacologyEur J Pharmacol. 2003 Nov 28;481(2-3):141-6. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14642778 0 Bueters, T. J. van Helden, H. P. I. Jzerman AP Danhof, M. 2003 Neurosci Lett 343 57-61 2003/05/17 May 29 0304-3940 (Print) 12749997 Acetylcholine/*pharmacokinetics, Adenosine/*analogs , &, derivatives/*pharmacology, Animals, Dose-Response Relationship, Drug, Male, Microdialysis/methods, Neostriatum/*drug effects/*metabolism, Rats, Rats, Wistar, Receptor, Adenosine A2B, Receptors, Pur The objective of this study was to characterize the effects of the adenosine A(1) receptor agonist N(6)-cyclopentyladenosine (CPA) and its low efficacy derivatives 2'-deoxy-CPA (2DCPA), 3'-deoxy-CPA (3DCPA), 8-ethylamino-CPA (8ECPA) and 8-butylamino-CPA (8BCPA) on the release of acetylcholine (ACh) using intrastriatal microdialysis. These low efficacy agonists exhibited lower effects on the cardiovascular system than CPA. A concentration-dependent inhibition of ACh release was observed with a maximum of 60.5+/-2.4% for CPA, 42.5+/-2.3% for 2DCPA, 45.3+/-5.8% for 3DCPA, 57.1+/-1.4% for 8ECPA and 93.1+/-10.9% for 8BCPA, respectively. This effect was counteracted by the adenosine A(1) receptor antagonist 8-cyclopentyltheophylline. These findings show that low efficacy adenosine A(1) agonists inhibit striatal ACh release equally effective as CPA, suggesting that central nervous system-selective actions can be obtained with these compounds. Bueters, Tjerk J Hvan Helden, Herman P MIJzerman, Adriaan PDanhof, MeindertComparative StudyIrelandNeuroscience lettersNeurosci Lett. 2003 May 29;343(1):57-61. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12749997 0 Bueters, T. J. van Helden, H. P. Danhof, M. I. Jzerman AP 2002 Eur J Pharmacol 454 177-82 2002/11/08 Nov 15 0014-2999 (Print) 12421644 Acetylcholine/metabolism/*secretion, Allosteric Regulation/drug effects/physiology, Animals, Corpus Striatum/*drug effects/metabolism/secretion, Dose-Response Relationship, Drug, Male, Rats, Rats, Wistar, Receptors, Purinergic P1/agonists/*antagonists , & The objective of the present study was to characterize the adenosine A(1) receptor allosteric enhancing and antagonistic actions of (2-amino-4,5,6,7-tetrahydrobenzo[b]thiophen-3-yl)(3,4-dichlorophenyl)metha none (LUF 5484) and (2-amino-4,5-dimethyl-3-thienyl)-[3-(trifluoromethyl)phenyl]methanone (PD 81,723) on striatal acetylcholine release. Upon local administration in conscious rats, LUF 5484 or PD 81,723 caused a concentration-dependent increase of extracellular acetylcholine levels of approximately 40%, which was similar to that obtained by the selective adenosine A(1) receptor antagonists 8-cyclopentyl-1,3-dimethylxanthine (8CPT) and N(6)-cyclopentyl-9-methyladenine (N0840). In interaction experiments, LUF 5484 or PD 81,723 did not change the inhibition of acetylcholine release by the adenosine A(1) receptor agonist N(6)-cyclopentyladenosine (CPA), whereas 8CPT caused an eightfold rightward shift. Acetylcholine concentrations were diminished with 62+/-3%, 48+/-11% and 56+/-9% by CPA, CPA+LUF 5484 and CPA+PD 81,723, respectively. In conclusion, the antagonistic action of LUF 5484 and PD 81,723 seems to counteract the putative allosteric actions with respect to the reduction of striatal acetylcholine release. Bueters, Tjerk J Hvan Helden, Herman P MDanhof, MeindertIJzerman, Ad PNetherlandsEuropean journal of pharmacologyEur J Pharmacol. 2002 Nov 15;454(2-3):177-82. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12421644 0 Bueters, T. J. Groen, B. Danhof, M. I. Jzerman AP Van Helden, H. P. 2002 Arch Toxicol 76 650-6 2002/11/05 Nov 0340-5761 (Print) 12415428 Acetylcholine/secretion, Adenosine/*analogs , &, derivatives/*therapeutic use, Animals, Cholinesterase Inhibitors/*poisoning, Cholinesterases/metabolism, Corpus Striatum/enzymology, Male, Microdialysis, Organothiophosphorus Compounds/*poisoning, Parathio The objective of the present study was to investigate whether reduction of central acetylcholine (ACh) accumulation by adenosine receptor agonists could serve as a generic treatment against organophosphate (OP) poisoning. The OPs studied were tabun ( O-ethyl- N-dimethylphosphoramidocyanidate), sarin (isopropylmethylphosphonofluoridate), VX ( O-ethyl- S-2-diisopropylaminoethylmethylphosphonothiolate) and parathion ( O, O-diethyl- O-(4-nitrophenyl)phosphorothioate). The efficacy of the adenosine A(1) receptor agonist N(6)-cyclopentyladenosine (CPA) against an OP intoxication was examined on the basis of the occurrence of clinical symptoms that are directly associated with such intoxication. CPA (1-2 mg/kg) effectively attenuated the cholinergic symptoms and prevented mortality in lethally tabun- or sarin-intoxicated rats. In contrast, CPA (2 mg/kg) proved to be ineffective against VX or parathion intoxication. Intracerebral microdialysis studies revealed that survival of sarin-poisoned and CPA-treated animals coincided with a minor elevation of extracellular ACh concentrations in the brain relative to the baseline value, whereas an 11-fold increase in transmitter levels was observed in animals not treated with CPA. In VX-intoxicated rats, however, the ACh amounts increased 18-fold, irrespective of treatment with CPA. The striatal acetylcholinesterase (AChE) activity following a lethal sarin intoxication was completely abolished in the vehicle-treated animals, whereas 10% and 60% AChE activity remained in animals treated with 2 mg/kg CPA 1 min after or 2 min prior to the poisoning, respectively. In VX-intoxicated animals the AChE activity in the brain was strongly reduced (striatum 10%, hippocampus 1%) regardless of the CPA treatment. These results demonstrate that CPA is highly effective against tabun or sarin poisoning, but fails to protect against VX or parathion. Survival and attenuation of clinical signs in tabun- or sarin-poisoned animals are associated with a reduction of ACh accumulation and with protection of AChE activity in the brain. Bueters, Tjerk J HGroen, BasDanhof, MeindertIJzerman, Ad PVan Helden, Herman P MGermanyArchives of toxicologyArch Toxicol. 2002 Nov;76(11):650-6. Epub 2002 Aug 21. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12415428 0 van Schaick, E. A. Tukker, H. E. Roelen, H. C. I. Jzerman AP Danhof, M. 1998 Br J Pharmacol 124 607-18 1998/07/01 Jun 0007-1188 (Print) 9647488 Adenosine/*analogs , &, derivatives/pharmacokinetics/pharmacology, Animals, Blood Pressure/drug effects, Fatty Acids, Nonesterified/blood, Half-Life, Heart Rate/drug effects, Hemodynamics/*drug effects, Lipolysis/*drug effects, Male, Rats, Rats, Wistar, 1. A1 adenosine receptor agonists with reduced intrinsic activity may be therapeutically useful as result of an increased selectivity of action. In this study the tissue selectivity of three 8-alkylamino substituted analogues of N6-cyclopentyladenosine (CPA) was investigated for haemodynamic and anti-lipolytic effects using an integrated pharmacokinetic-pharmacodynamic approach. 2. Chronically instrumented male Wistar rats received intravenous infusions of 4.0 mg kg(-1) 8-methylaminoCPA (8MCPA), 12.0 mg kg(-1) 8-ethylaminoCPA (8ECPA), 20.0 mg kg(-1) 8-butylaminoCPA (8BCPA) or vehicle during 15 min. During experimentation, serial arterial blood samples were drawn for the determination of agonist concentrations and plasma non-esterified fatty acid (NEFA) levels. Blood pressure and heart rate were monitored continuously. In addition to the CPA analogues, each rat received a rapid bolus infusion of CPA to determine the maximal effects of the full agonist. 3. The concentration-time profiles of the CPA analogues could be described by a bi-exponential function. Values for clearance, volume of distribution at steady state and elimination half-life were 44+/-5, 48+/-6 and 39+/-2 ml min(-1) kg(-1), 0.97+/-0.09, 0.84+/-0.10 and 1.05+/-0.07 1 kg(-1) and 25+/-2, 28+/-2 and 40+/-2 min for 8MCPA, 8ECPA and 8BCPA, respectively (mean+/-s.e.mean, n=6-8). 4. Different models were used to derive the concentration-effect relationships for heart rate and NEFA, yielding estimates of potency (EC50) and intrinsic activity (Emax) for both effects of the compounds in vivo. On heart rate the compounds acted as partial agonists, with Emax values of -173+/-14, -131+/-11 and -71+/-6 beats min(-1) for 8MCPA, 8ECPA and 8BCPA, respectively. These Emax values were significantly lower than the maximal effect of CPA (-208+/-8 beats min(-1)). With regard to the anti-lipolytic effect all three compounds were full agonists and lowered NEFA levels to the same extent as CPA (69%). The estimated Emax values were 63+/-5, 63+/-4 and 68+/-2%, respectively. 5. Furthermore, the compounds were more potent in causing anti-lipolytic than cardiovascular effects. The EC50 values for the NEFA and heart rate lowering effects were 37+/-15, 68+/-22 and 659+/-108 ng ml(-1) and 164+/-22, 341+/-76 and 975+/-190 ng ml(-1) for 8MCPA, 8ECPA and 8BCPA, respectively (mean+/-s.e.mean, n=6-8). 6. This study demonstrates that partial agonists for the A1 adenosine receptor have increased selectivity of action in vivo. The 8-alkylamino analogues of CPA may be useful anti-lipolytics with less pronounced haemodynamic side effects. van Schaick, E ATukker, H ERoelen, H CIJzerman, A PDanhof, MComparative StudyResearch Support, Non-U.S. Gov'tEnglandBritish journal of pharmacologyBr J Pharmacol. 1998 Jun;124(3):607-18. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9647488 0 van Schaick, E. A. de Greef, H. J. Langemeijer, M. W. Sheehan, M. J. I. Jzerman AP Danhof, M. 1997 Br J Pharmacol 122 525-33 1997/11/14 Oct 0007-1188 (Print) 9351510 3-Hydroxybutyric Acid, Adenosine/*analogs , &, derivatives/blood/pharmacokinetics/pharmacology, Animals, Antilipemic Agents/blood/*pharmacokinetics/pharmacology, Blood Pressure/drug effects, Blood Proteins/metabolism, Chromatography, High Pressure Liquid 1. The purpose of this study was to develop and validate an integrated pharmacokinetic-pharmacodynamic model for the anti-lipolytic effects of the adenosine A1-receptor agonist N6-(p-sulphophenyl)adenosine (SPA). Tissue selectivity of SPA was investigated by quantification of haemodynamic and anti-lipolytic effects in individual animals. 2. After intravenous infusion of SPA to conscious normotensive Wistar rats, arterial blood samples were drawn for determination of blood SPA concentrations, plasma non-esterified fatty acid (NEFA) and beta-hydroxybutyrate levels. Blood pressure and heart rate were monitored continuously. 3. The relationship between the SPA concentrations and the NEFA lowering effect was described by the indirect suppression model. Administration of SPA at different rates and doses (60 microg kg[-1] in 5 min and 15 min, and 120 microg kg[-1] in 60 min) led to uniform pharmacodynamic parameter estimates. The averaged parameters (mean+/-s.e., n=19) were Emax: -80+/-2% (% change from baseline), EC50: 22+/-2 ng ml(-1), and Hill factor: 2.2+/-0.2. 4. In another group, given 400 microg kg(-1) SPA in 15 min, pharmacodynamic parameters for both heart rate and anti-lipolytic effect were derived within the same animal. The reduction in heart rate was directly related to blood concentration on the basis of the sigmoidal Emax model. SPA inhibited lipolysis at concentrations lower than those required for an effect on heart rate. The EC50 values (mean+/-s.e., n=6) were 131+/-31 ng ml(-1) and 20+/-3 ng ml(-1) for heart rate and NEFA lowering effect, respectively. 5. In conclusion, the relationship between blood SPA concentrations and anti-lipolytic effect was adequately described by the indirect suppression model. For SPA a 6 fold difference in potency was observed between the effects on heart rate and NEFAs, indicating some degree of tissue selectivity in vivo. van Schaick, E Ade Greef, H JLangemeijer, M WSheehan, M JIJzerman, A PDanhof, MN01MH30003/MH/NIMH NIH HHS/United StatesResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, P.H.S.EnglandBritish journal of pharmacologyBr J Pharmacol. 1997 Oct;122(3):525-33. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9351510 0 Bol, C. J. I. Jzerman AP Danhof, M. Mandema, J. W. 1997 J Pharm Sci 86 822-6 1997/07/01 Jul 0022-3549 (Print) 9232523 Adrenergic alpha-Agonists/*blood/metabolism, Adrenergic alpha-Antagonists/metabolism, Animals, Binding, Competitive, Cattle, Clonidine/*metabolism, Idazoxan/analogs , &, derivatives/metabolism, Imidazoles/*blood/metabolism, Magnesium/pharmacology, Male, This paper describes the development and implementation of a sensitive radioreceptor assay (RRA) for determining concentrations of dexmedetomidine, an alpha-2 adrenergic agonist with anesthetic properties, in rat plasma. Calf retina membranes were selected as the alpha-2 adrenergic receptor source, and the alpha-2 antagonist [3H]RX821002 and the alpha-2 agonist [3H]clonidine were evaluated as radioligands. We optimized the binding conditions for both radioligands and chose a radioligand for implementation in the RRA based on the characteristics of the inhibition binding curves with dexmedetomidine. The final method is based on competition between the radioligand [3H]clonidine and dexmedetomidine for high-affinity binding sites present in calf retina membranes. The assay has a coefficient of variation of 8% in the range 23.7-592 pg for 0.2 mL of plasma. This assay can be applied to pharmacokinetic-pharmacodynamic studies of dexmedetomidine. Bol, C JIJzerman, A PDanhof, MMandema, J WGM 51309/GM/NIGMS NIH HHS/United StatesComparative StudyResearch Support, U.S. Gov't, P.H.S.United statesJournal of pharmaceutical sciencesJ Pharm Sci. 1997 Jul;86(7):822-6. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9232523 0 van Schaick, E. A. Kulkarni, C. von Frijtag Drabbe Kunzel, J. K. Mathot, R. A. Cristalli, G. I. Jzerman AP Danhof, M. 1997 Naunyn Schmiedebergs Arch Pharmacol 356 827-37 1998/02/07 Dec 0028-1298 (Print) 9453470 Adenosine/*analogs , &, derivatives/chemistry/pharmacokinetics/pharmacology, Animals, Biological Availability, Blood Pressure/drug effects, Electroencephalography/*drug effects, Half-Life, Heart Rate/drug effects, Male, Metabolic Clearance Rate, Rats, Ra In this study we investigated the relationship between the pharmacokinetics and the cardiovascular and electroencephalogram (EEG) effects of three adenosine agonists with differing lipophilicity. Conscious normotensive rats received either 600 microg/kg N6-(p-sulphophenyl) adenosine (SPA), 200 microg/kg N6-cyclopentyladenosine (CPA) or 600 microg/kg 1-deaza-2-chloro-N6-cyclopentyladenosine (DCCA) in a 5-min intravenous infusion. Changes in haemodynamics and EEG were monitored in conjunction with arterial blood sampling to determine blood concentrations of the compounds. The three adenosine agonists showed large differences in pharmacokinetic properties, resulting in terminal half-lives of 66 +/- 10, 8.2 +/- 0.4 and 24 +/- 1 min (mean +/- SEM) for SPA, CPA, and DCCA respectively. SPA had a significantly lower blood clearance relative to CPA and DCCA, whereas DCCA had the largest volume of distribution and degree of plasma protein binding. The relationship between concentration and heart rate could be described adequately by the sigmoidal Emax model. For SPA, CPA, and DCCA the EC50 values based on free drug concentrations were 423 +/- 92, 1.8 +/- 0.4 and 9.5 +/- 1.1 nM respectively. These in vivo values correlated closely with the affinity of the compounds for the adenosine A1 receptor as determined in radioligand binding studies, with corresponding Ki values of 1410 +/- 220, 4.7 +/- 0.6 and 102 +/- 74 nM (mean +/- SEM) respectively. In the EEG, only CPA produced a small decrease in the amplitude of beta waves. This study demonstrates that the three adenosine analogues have large differences in pharmacokinetics, which complicates comparison of their cardiovascular and central responses simply on the basis of dose. The application of an integrated PK/PD approach permits estimates of potency and activity which are independent of underlying dose and pharmacokinetics. van Schaick, E AKulkarni, Cvon Frijtag Drabbe Kunzel, J KMathot, R ACristalli, GIJzerman, A PDanhof, MComparative StudyResearch Support, Non-U.S. Gov'tGermanyNaunyn-Schmiedeberg's archives of pharmacologyNaunyn Schmiedebergs Arch Pharmacol. 1997 Dec;356(6):827-37. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9453470 0 Van Schaick, E. A. Jacobson, K. A. Kim, H. O. I. Jzerman AP Danhof, M. 1996 Eur J Pharmacol 308 311-4 1996/07/25 Jul 25 0014-2999 (Print) 8858305 Adenosine/*analogs , &, derivatives/pharmacology, Animals, Blood Pressure/*drug effects, Heart Rate/*drug effects, Histamine/blood, Histamine Release/*drug effects, Male, Rats, Rats, Wistar, Receptors, Purinergic P1/*agonists The hemodynamic effects of the novel, selective adenosine A3 receptor agonist 2-chloro-N6-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide (2-Cl-IB-MECA) were investigated in conscious rats. Intravenous administration of 200 micrograms/kg 2-Cl-IB-MECA resulted in a short-lasting hypotension, which was accompanied by a 50-100-fold increase in plasma histamine concentrations. Administration of a second dose of 2-Cl-IB-MECA did not elicit any hemodynamic effects. Also no histamine release was observed after the second dose. The observation of rapid tachyphylaxis in combination with histamine release suggests that mast cell mediator release plays a key role in the hemodynamic effects of adenosine A3 receptor agonists. Van Schaick, E AJacobson, K AKim, H OIJzerman, A PDanhof, MNetherlandsEuropean journal of pharmacologyEur J Pharmacol. 1996 Jul 25;308(3):311-4. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8858305 0 van der Wenden, E. M. von Frijtag Drabbe Kunzel, J. K. Mathot, R. A. Danhof, M. I. Jzerman AP Soudijn, W. 1995 J Med Chem 38 4000-6 1995/09/29 Sep 29 0022-2623 (Print) 7562934 Adenosine/*analogs , &, derivatives/pharmacology, Animals, Blood Pressure/drug effects, Guanosine Triphosphate/pharmacology, Heart Rate/drug effects, Male, Rats, Rats, Wistar, Receptors, Purinergic P1/*agonists, Ribose, Structure-Activity Relationship We have adopted a practical three-step route for the synthesis of 2'- and 3'-deoxy analogues of N6-substituted adenosines: protection of the hydroxyl groups, replacement of the N6-amino by a better leaving group, and combined deprotection and N6-amination in the last step. This route was used to synthesize deoxy analogues of CPA, CHA, and R- and S-PIA. The compounds were tested on the adenosine A1 and A2a receptors in our search for partial agonists for these receptors. The GTP shift was used as an in vitro measure for the intrinsic activity of these compounds; the in vivo intrinsic activities of the deoxy analogues of CPA and R-PIA were determined in the rat cardiovascular system. Thus, it was shown that the hydroxyl groups are determinants for the affinity and intrinsic activity of these analogues. Removal of the 2'- and 3'-hydroxyl groups affects affinity and intrinsic activity, whereas removal of the 5'-hydroxyl group decreases only affinity. van der Wenden, E Mvon Frijtag Drabbe Kunzel, J KMathot, R ADanhof, MIJzerman, A PSoudijn, WResearch Support, Non-U.S. Gov'tUnited statesJournal of medicinal chemistryJ Med Chem. 1995 Sep 29;38(20):4000-6. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7562934 0 Mathot, R. A. Van der Wenden, E. M. Soudijn, W. I. Jzerman AP Danhof, M. 1995 Br J Pharmacol 116 1957-64 1995/10/01 Oct 0007-1188 (Print) 8640332 Adenosine/administration , &, dosage/*analogs , &, derivatives/pharmacokinetics/pharmacology, Animals, Blood Pressure/drug effects, Bradycardia/chemically induced, Deoxyribose/analogs , &, derivatives, Dose-Response Relationship, Drug, Heart Rate/drug e 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. Mathot, R AVan der Wenden, E MSoudijn, WIJzerman, A PDanhof, MIn VitroEnglandBritish journal of pharmacologyBr J Pharmacol. 1995 Oct;116(3):1957-64. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8640332 0 Mathot, R. A. Cleton, A. Soudijn, W. I. Jzerman AP Danhof, M. 1995 Br J Pharmacol 114 761-8 1995/02/01 Feb 0007-1188 (Print) 7773536 Adenosine/*analogs , &, derivatives/blood/pharmacokinetics/pharmacology, Animals, Antihypertensive Agents/*pharmacokinetics/pharmacology, Blood Pressure/*drug effects, Blood Pressure Determination, Chromatography, High Pressure Liquid, Dose-Response Rela 1. The aim of the present investigation was to determine the relationship between the blood concentration and haemodynamic effects of the adenosine A2a receptor agonist, CGS 21680C (the sodium salt of 2-p-(2-carboxyethyl)phenylethylamino-5'-N-ethylcarboxamidoadeno sin e) in conscious normotensive rats. 2. Chronically cannulated rats were randomly assigned to three groups which received 300, 1000 or 3000 micrograms kg-1 (0.56, 1.9 or 5.6 mumol kg-1) of CGS 21680C intravenously over 15 min. The mean arterial blood pressure (MAP) and heart rate (HR) were monitored continuously during the experiment and serial arterial blood samples were taken for analysis of drug concentration. The ratio MAP/HR was also calculated, which may reflect changes in total peripheral resistance on the assumption that no changes in stroke volume occur. 3. For each individual rat the reduction in mean arterial pressure was related to the blood concentration according to the sigmoidal Emax model. The concentration-effect relationships were consistent for the different treatment groups. The potency based on free drug concentrations (EC50,u) was 5.8 ng ml-1 (11 nM) (mean +/- s.e.; n = 19) and correlated well with the reported adenosine A2a receptor affinity (Ki 19 nM). In comparison with the reduction in blood pressure, CGS 21680C exhibited a greater potency for the reduction of the ratio MAP/HR. 4. It is concluded that estimates can be obtained for the potency and intrinsic activity of adenosine A2a receptor agonists in vivo by pharmacokinetic-pharmacodynamic analysis of mean arterial pressure data in a rat model. In future studies, total peripheral resistance may also be useful as a pharmacodynamic parameter for A24 activation, provided that possible changes of the stroke volume are also assessed.Keywords: Adenosine A24 receptor; cardiovascular effects; CGS 21680 (2-p-(2-carboxyethyl)phenylethylamino-5'-N-ethylcarboxamidoadenosine);phar macokinetic-pharmacodynamic modelling Mathot, R ACleton, ASoudijn, WIJzerman, A PDanhof, MEnglandBritish journal of pharmacologyBr J Pharmacol. 1995 Feb;114(4):761-8. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7773536 0 Van der Wenden, E. M. Hartog-Witte, H. R. Roelen, H. C. von Frijtag Drabbe Kunzel, J. K. Pirovano, I. M. Mathot, R. A. Danhof, M. Van Aerschot, A. Lidaks, M. J. I. Jzerman AP et al., 1995 Eur J Pharmacol 290 189-99 1995/08/15 Aug 15 0014-2999 (Print) 7589213 Adenosine/*analogs , &, derivatives/chemistry/*pharmacology, Adenylate Cyclase/metabolism, Animals, Cells, Cultured, Heart Rate/drug effects, Kinetics, Rats, Receptors, Purinergic P1/*agonists, Theophylline/*analogs , &, derivatives/chemistry/*pharmacol A series of 8-substituted adenosine and theophylline-7-riboside analogues (28 and 9 compounds, respectively) was tested on adenosine A1 and A2A receptors as an extensive exploration of the adenosine C8-region. Alkylamino substituents at the 8-position cause an affinity decrease for adenosine analogues, but an affinity increase for theophylline-7-riboside derivatives. The affinity decrease is probably due to a direct steric hindrance between the C8-substituent and the binding site as well as to electronic effects, not to a steric influence on the ribose moiety to adopt the anti conformation. The 8-substituents increase the affinity of theophylline-7-riboside analogues probably by binding to a lipophilic binding site. The intrinsic activity was tested in vitro for some 8-substituted adenosine analogues, by determining the GTP shift in receptor binding studies and the inhibition of adenylate cyclase in a culture of rat thyroid FRTL-5 cells, and in vivo in the rat cardiovascular system for 8-butylaminoadenosine. Thus, it was shown that 8-ethyl-, 8-butyl-, and 8-pentylamino substituted analogues of adenosine may be partial agonists in vitro, and that 8-butylaminoadenosine is a partial agonist for the rat cardiovascular A1 receptor in vivo. Van der Wenden, E MHartog-Witte, H RRoelen, H Cvon Frijtag Drabbe Kunzel, J KPirovano, I MMathot, R ADanhof, MVan Aerschot, ALidaks, M JIJzerman, A PIn VitroNetherlandsEuropean journal of pharmacologyEur J Pharmacol. 1995 Aug 15;290(3):189-99. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7589213 0 Appel, S. Mathot, R. A. Langemeijer, M. W. I. Jzerman AP Danhof, M. 1995 Br J Pharmacol 115 1253-9 1995/08/01 Aug 0007-1188 (Print) 7582553 Adenosine/*analogs , &, derivatives/blood/pharmacokinetics/pharmacology, Animals, Blood Pressure/drug effects, Blood Proteins/metabolism, Heart Rate/drug effects, Male, Models, Chemical, Protein Binding, Rats, Rats, Wistar, Receptors, Purinergic P1/agoni 1. The purpose of this investigation was to develop a pharmacokinetic-pharmacodynamic model for the interaction between an adenosine A1 receptor agonist and antagonist in vivo. The adenosine A1 receptor agonist, N6-cyclopentyladenosine (CPA) and the antagonist, 8-cyclopentyltheophylline (CPT) were used as model drugs. The CPA-induced reduction in mean arterial pressure and heart rate were used as measurements of effect. 2. Four groups of eight rats each received 200 micrograms kg-1 of CPA i.v. in 5 min during a steady-state infusion of CPT at a rate of 0, 57, 114 or 228 micrograms kg-1 h-1. The haemodynamic parameters were continuously measured and frequent blood samples were taken to determine the pharmacokinetics of the drugs. 3. CPT had no influence on the pharmacokinetics of CPA and the baseline values of the haemodynamic variables. Furthermore, no clear antagonism by CPT was observed of the CPA-induced reduction in mean arterial pressure. However, CPT antagonized the effect on heart rate, and with increasing CPT concentrations, a parallel shift of the CPA concentration-effect relationship to the right was observed. 4. An agonist-antagonist interaction model was used to characterize the interaction quantitatively. On the basis of this model, the pharmacodynamic parameters of both CPA and CPT could be estimated. For CPA the values were (mean +/- s.e.): Emax = 198 +/- 11 b.p.m., EC50 = 2.1 +/- 0.7 ng ml-1, Hill factor = 2.3 +/- 0.6 and for CPT: EC50 = 3.7 +/- 0.3 ng ml-1 and Hill factor = 3.1 +/- 0.1. 5. It is concluded that the competitive agonist-antagonist interaction model may be of value to characterize quantitatively the pharmacodynamic interactions between adenosine A1 receptor ligands in vivo.Keywords: N6-cyclopentyladenosine; 8-cyclopentyltheophylline; pharmacokinetics; pharmacodynamics; interaction in vivo; mean arterial pressure; heart rate; modelling Appel, SMathot, R ALangemeijer, M WIJzerman, A PDanhof, MEnglandBritish journal of pharmacologyBr J Pharmacol. 1995 Aug;115(7):1253-9. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7582553 0 I. Jzerman AP van der Wenden, E. M. von Frijtag Drabbe Kunzel, J. K. Mathot, R. A. Danhof, M. Borea, P. A. Varani, K. 1994 Naunyn Schmiedebergs Arch Pharmacol 350 638-45 1994/12/01 Dec 0028-1298 (Print) 7708121 Animals, Cells, Cultured, Forskolin/pharmacology, Guanosine Triphosphate, Rats, Receptors, Purinergic P1/*agonists/drug effects, Temperature, Theophylline/*agonists/pharmacology, Xanthine, Xanthines/chemistry/*pharmacology Theophylline-7-riboside was evaluated as a partial agonist for rat adenosine receptors. Radioligand binding experiments were performed on both A1 and A2a adenosine receptors, using several methodologies to discriminate between agonists and antagonists. Mainly from thermodynamic data it was concluded that on A1 receptors theophylline-7-riboside had characteristics intermediate between full agonists, such as N6-cyclopentyladenosine, and full antagonists, such as the xanthines. The partial agonistic behaviour of theophylline-7-riboside was further explored in second messenger studies in intact cells. In FRTL-5 rat thyroid cells theophylline-7-riboside behaved as a partial agonist for A1 receptors, slightly inhibiting forskolin-stimulated cyclic AMP levels. The implications of these biochemical findings were further analysed in in vivo pharmacology. The infusion of theophylline-7-riboside in conscious, normotensive rats led to marked changes in cardiovascular parameters, although less outspoken than observed with full agonists for either A1 or A2a receptors. The concomitant determination of the blood concentrations of theophylline-7-riboside and its metabolite theophylline allowed the estimation of in vivo pharmacokinetic and pharmacodynamic parameters. Thus, the EC50 value of theophylline-7-riboside for lowering the mean arterial pressure was 47 +/- 12 micrograms/ml blood. The short duration of action of theophylline-7-riboside makes it improbable that its metabolite theophylline interferes with its effects. In conclusion, theophylline-7-riboside is one of the first partial agonists for adenosine receptors. It may serve as a tool in further investigations of adenosine receptor partial agonism. IJzerman, A Pvan der Wenden, E Mvon Frijtag Drabbe Kunzel, J KMathot, R ADanhof, MBorea, P AVarani, KGermanyNaunyn-Schmiedeberg's archives of pharmacologyNaunyn Schmiedebergs Arch Pharmacol. 1994 Dec;350(6):638-45. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7708121 0 Irth, H. Cleton, A. Mathot, R. A. Danhof, M. I. Jzerman AP Tjaden, U. R. van der Greef, J. 1994 J Chromatogr B Biomed Appl 658 207-12 1994/08/05 Aug 5 1572-6495 (Print) 7952127 Acetates, Adenosine/*analogs , &, derivatives/blood/pharmacokinetics, Animals, *Boronic Acids, Chromatography, High Pressure Liquid/*methods/statistics , &, numerical data, Drug Monitoring, Hydrogen-Ion Concentration, Male, Phenethylamines/*blood/pharma An analytical method is described for the selective determination of A1 or A2 adenosine receptor agonists in blood. By implementing solid-phase extraction using immobilized-phenylboronic acid (PBA) in sample pretreatment, all adenosine derivatives are retained via their intact cis-diol group. On-line desorption of the analytes from the PBA support to the C18 analytical column is performed by injection of a small plug of perchloric acid. Fluorescence and UV detection are employed for the different adenosine derivatives. The method is applied to the determination of 2-[p-(2-carboxyethyl)phenylethylamino]-5'-N- ethylcarboxyamidoadenosine (CGS 21680, I) in blood using fluorescence detection. The only off-line sample handling step is the extraction of blood with ethyl acetate and subsequent evaporation of the extraction solvent. The detection limit of the method was 0.25 ng (signal-to-noise ratio 3:1) and the determination limit for I in blood (pretreatment of 100 microliters) was 5 ng/ml. The method was validated and used to study the pharmacokinetics of I in rats. Irth, HCleton, AMathot, R ADanhof, MIJzerman, A PTjaden, U Rvan der Greef, JNetherlandsJournal of chromatography. B, Biomedical applicationsJ Chromatogr B Biomed Appl. 1994 Aug 5;658(1):207-12. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7952127 0 Mathot, R. A. Appel, S. van Schaick, E. A. Soudijn, W. I. Jzerman AP Danhof, M. 1993 J Chromatogr 620 113-20 1993/10/22 Oct 22 0021-9673 (Print) 8106577 Adenosine/*analogs , &, derivatives/analysis/pharmacokinetics, Animals, Chromatography, High Pressure Liquid/*methods, Drug Stability, Hydrogen-Ion Concentration, Kinetics, Ligands, Male, Rats, Rats, Wistar, Receptors, Purinergic P1/antagonists , &, inh This report describes a rapid and sensitive analysis for the simultaneous detection of the adenosine A1 receptor ligands N6-cyclopentyladenosine (CPA) and 8-cyclopentyltheophylline (CPT) in rat blood. The method involved alkaline extraction of the compounds and internal standard N6-cyclohexyladenosine (CHA) with ethyl acetate, followed by isocratic reversed-phase high-performance liquid chromatography on a 3-microns MicroSphere C18 column with UV detection at 269 nm. The mobile phase consisted of a mixture of 10 mM acetate buffer (pH 4.0)-methanol-acetonitrile (56:40:4, v/v/v) with a flow-rate of 0.50 ml/min. The total run time was ca. 19 min. For CPA and CPT extraction yields were greater than 77 and 66% in the concentration range of 0.010-0.75 microgram/ml and 0.025-15 micrograms/ml, respectively, with intra- and inter-assay variations less than 9%. In 100 microliter blood samples the corresponding limits of detection were 3.3 and 6.2 ng/ml (signal-to-noise ratio = 3). CPA was found to be degraded in rat blood in vitro with a half-life of 24 min at 37 degrees C. The utility of the analytical method was established by analyzing blood samples from rats which had received an intravenous administration of 200 micrograms/kg CPA or 12 mg/kg CPT. Due to its rapidity and sensitivity this method is concluded to be particularly useful in pharmacokinetic studies with CPA and CPT. Mathot, R AAppel, Svan Schaick, E ASoudijn, WIJzerman, A PDanhof, MNetherlandsJournal of chromatographyJ Chromatogr. 1993 Oct 22;620(1):113-20. http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8106577