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1.
Article in English | MEDLINE | ID: mdl-25497900

ABSTRACT

INTRODUCTION: Excitation of Acetylcholine-mediated (Ach) transmission (especially if irreversible) may pose life-threatening adverse events by increasing neuronal network activity. Unfortunately, adequate detection of this liability during early drug development is hampered, because published ex vivo electrophysiological models are very insensitive to this regard. For example, Eserine, which reversibly inhibits acetylcholinesterase (AchE) in the double digit nM range, affects electrically evoked potentials in hippocampal slices only at ≥10µM. Here, a significantly more sensitive method for detecting Ach-mediated alternations is presented by analyzing spontaneous neuronal network activity in hippocampal slices. METHODS: The microelectrode array (MEA) technique with an 8×8 electrode grid was applied to analyze evoked and spontaneous extracellular field recordings in parallel from acute rat hippocampal slices. For evoked potentials, the Schaffer collateral CA3-CA1 pathway was electrically stimulated and the resulting field potential analyzed at the CA1 pyramidal layer. Spontaneous spike activity was detected as negative inflections from the 100Hz high pass filtered signal. Spike frequency was analyzed within the whole CA1 region. RESULTS: Modification of Ach-mediated neuronal transmission via carbachol, Eserine, or Diisopropylfluorophosphate (DFP) does not induce any effects on evoked field potentials at physiologically relevant concentrations. Similar to previous reports, subtle effects were detectable at very high concentrations. By contrast, spontaneous spike frequency was already increased within the expected concentration range. Eserine-induced effects can also be reversed by atropine and washout. On the contrary, effects by the irreversible AchE-blocker DFP could not be washed out. DISCUSSION: Compared to evoked field potentials, spontaneous spike activity in the hippocampal CA1 region appears to be a significantly more sensitive parameter for functional electrophysiological analysis of drug induced Ach-mediated effects. This finding may supplement existing models for detection and prediction of drug-related adverse effects like seizure liability already during early development stages.


Subject(s)
Acetylcholine/pharmacology , Hippocampus/drug effects , Hippocampus/physiology , Animals , Hippocampus/cytology , Male , Microelectrodes , Neurons/drug effects , Neurons/metabolism , Rats , Rats, Wistar
2.
Psychopharmacology (Berl) ; 231(12): 2547-58, 2014 Jun.
Article in English | MEDLINE | ID: mdl-24419272

ABSTRACT

RATIONALE: The recently approved antidepressant vilazodone, a serotonin (5-HT)1A receptor partial agonist/selective 5-HT reuptake inhibitor offers new possibilities to study the underlying mechanisms of depression pharmacotherapy and of 5-HT augmenting antidepressants. OBJECTIVE: The role of the 5-HT1A receptor with respect to the regulation of 5-HT output in the mechanism of action of vilazodone. METHOD: We measured 5-HT levels in two subregions of the rat prefrontal cortex by microdialysis, and 5-hydroxytryptophan (5-HTP) accumulation and tissue 5-HT concentrations ex vivo. RESULTS: Vilazodone-induced maximal 5-HT levels were similar in the medial and the lateral cortex and were up to sixfold higher than those induced by paroxetine, citalopram, or fluoxetine tested in parallel. Depolarization/autoreceptor-insensitive 5-HT release by vilazodone could be excluded. The citalopram (1 µM, locally infused)-induced increase of 5-HT was further increased by vilazodone (1 mg/kg i.p.), but not by citalopram (10 mg/kg i.p.). Unlike fluoxetine, vilazodone-induced extracellular 5-HT output was not potentiated by cotreatment with the 5-HT1A receptor blocker N-[2-(4-{2-methoxyphenyl}-1-piperazinyl)-ethyl]-N-2-pyridinylcyclohexanecarboxamide (WAY 100635). In contrast to fluoxetine, vilazodone exhibited intrinsic 5-HT1A agonist activity: it reduced, similar to (±)-8-hydroxy-2-(dipropylamino)-tetralin (8-OH-DPAT), 5-HTP accumulation in striatum and n. raphe of reserpinized rats. Hence, vilazodone's agonistic actions must be 5-HT1A receptor-related since endogenous 5-HT is lacking in the reserpine-depleted animal. CONCLUSIONS: In spite of high intrinsic 5-HT1A activity in reserpinized rats, the net effect of vilazodone at release-regulating 5-HT1A autoreceptors must be inhibitory, leading to markedly increased 5-HT output. Another possibility is that vilazodone rapidly desensitizes autoinhibitory 5-HT1A receptors by an unknown mechanism.


Subject(s)
Antidepressive Agents/pharmacology , Benzofurans/pharmacology , Brain/drug effects , Brain/metabolism , Indoles/pharmacology , Piperazines/pharmacology , Receptor, Serotonin, 5-HT1A/metabolism , Serotonin/metabolism , 5-Hydroxytryptophan/metabolism , 8-Hydroxy-2-(di-n-propylamino)tetralin/pharmacology , Animals , Citalopram/pharmacology , Dose-Response Relationship, Drug , Drug Interactions , Extracellular Space/metabolism , Fluoxetine/pharmacology , Male , Paroxetine/pharmacology , Pyridines/pharmacology , Rats, Wistar , Serotonin 5-HT1 Receptor Agonists/pharmacology , Serotonin Antagonists/pharmacology , Serotonin Receptor Agonists/pharmacology , Selective Serotonin Reuptake Inhibitors/pharmacology , Vilazodone Hydrochloride
3.
J Biol Chem ; 282(39): 28501-28513, 2007 Sep 28.
Article in English | MEDLINE | ID: mdl-17686765

ABSTRACT

The human gene RSC1A1 codes for a 67-kDa protein named RS1 that mediates transcriptional and post-transcriptional regulation of Na(+)-D-glucose cotransporter SGLT1. The post-transcriptional regulation occurs at the trans-Golgi network (TGN). We identified two tripeptides in human RS1 (Gln-Cys-Pro (QCP) and Gln-Ser-Pro (QSP)) that induce posttranscriptional down-regulation of SGLT1 at the TGN leading to 40-50% reduction of SGLT1 in plasma membrane. For effective intracellular concentrations IC(50) values of 2.0 nM (QCP) and 0.16 nm (QSP) were estimated. Down-regulation of SGLT1 by tripeptides was attenuated by intracellular monosaccharides including non-metabolized methyl-alpha-D-glucopyranoside and 2-deoxyglucose. In small intestine post-transcriptional regulation of SGLT1 may contribute to glucose-dependent regulation of liver metabolism and intestinal mobility. QCP and QSP are transported by the H(+)-peptide cotransporter PepT1 that is colocated with SGLT1 in small intestinal enterocytes. Using coexpression of SGLT1 and PepT1 in Xenopus oocytes or polarized Caco-2 cells that contain both transporters we demonstrated that the tripeptides were effective when applied to the extracellular compartment. After a 1-h perfusion of intact rat small intestine with QSP, glucose absorption was reduced by 30%. The data indicate that orally applied tripeptides can be used to down-regulate small intestinal glucose absorption, e.g. in diabetes mellitus.


Subject(s)
Glucose/metabolism , Intestinal Absorption/drug effects , Monosaccharide Transport Proteins/metabolism , Oligopeptides/pharmacology , Protein Processing, Post-Translational/drug effects , Sodium-Glucose Transporter 1/metabolism , Animals , Antimetabolites/pharmacology , Biological Transport, Active/drug effects , Biological Transport, Active/physiology , Caco-2 Cells , Deoxyglucose/pharmacology , Diabetes Mellitus/drug therapy , Diabetes Mellitus/genetics , Diabetes Mellitus/metabolism , Female , Gastrointestinal Motility/drug effects , Gastrointestinal Motility/physiology , Gene Expression , Humans , Intestinal Absorption/physiology , Liver/metabolism , Male , Methylglucosides/pharmacology , Monosaccharide Transport Proteins/chemistry , Monosaccharide Transport Proteins/genetics , Oligopeptides/chemistry , Oligopeptides/therapeutic use , Oocytes/cytology , Peptide Transporter 1 , Protein Processing, Post-Translational/physiology , Rats , Rats, Wistar , Sodium-Glucose Transporter 1/genetics , Symporters/genetics , Symporters/metabolism , Xenopus laevis , trans-Golgi Network/metabolism
4.
ChemMedChem ; 1(2): 245-55, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16892357

ABSTRACT

The discovery of a novel class of highly potent and selective 5-HT2A antagonists is reported herein. Selectivity for the serotonin 5-HT2A receptor was optimized, decreasing the affinity of these antagonists toward the adrenergic alpha1 and dopaminergic D2 receptors, and especially to the 5-HT2C receptor. A series of corresponding 7-substituted indoles is described for the first time as serotonergic ligands. The enantiomer R-(+)-1-(4-fluorophenyl)-1-{1-[2-(4-fluorophenyl)ethyl]piperidin-4-yl} ethanol (R-(+)-74) was identified to have superior affinity for the serotonergic 5-HT2A receptor [IC50=0.37 nM] and selectivity toward the dopaminergic D2- [IC50=2300 nM], adrenergic alpha1- [IC50=1000 nM] and 5-HT2C receptors [IC50=490 nM].


Subject(s)
Piperidines/pharmacology , Serotonin 5-HT2 Receptor Antagonists , Serotonin Antagonists/pharmacology , Animals , Magnetic Resonance Spectroscopy , Male , Piperidines/chemistry , Rats , Receptor, Serotonin, 5-HT2A/chemistry , Serotonin Antagonists/chemistry , Spectrometry, Mass, Electrospray Ionization
5.
Bioorg Med Chem Lett ; 14(23): 5817-22, 2004 Dec 06.
Article in English | MEDLINE | ID: mdl-15501047

ABSTRACT

Neutral chlorothiophenecarboxamides bearing an amino acid and a substituted aniline were synthesized and investigated for their factor Xa inhibitory activity in vitro. From selected 2-methylphenyl morpholinones the solution properties were determined. The most soluble and active compounds were then investigated in different animal species to compare the pharmacokinetic parameters. This led to a potent, water soluble and orally bioavailable candidate for further development: EMD 495235.


Subject(s)
Factor Xa Inhibitors , Serine Proteinase Inhibitors/chemistry , Thiophenes/chemistry , Animals , Dogs , Factor Xa/metabolism , Female , Macaca fascicularis , Male , Rats , Rats, Wistar , Serine Proteinase Inhibitors/metabolism , Serine Proteinase Inhibitors/pharmacology , Structure-Activity Relationship , Thiophenes/metabolism , Thiophenes/pharmacology
6.
Bioorg Med Chem ; 12(18): 4843-52, 2004 Sep 15.
Article in English | MEDLINE | ID: mdl-15336263

ABSTRACT

The dual serotonin (5-HT) re-uptake inhibitor and 5-HT(1A) receptor agonist vilazodone was found to increase central serotonin levels in rat brain. In the course of structural modifications of vilazodone 3-[4-[4-(2-oxo-2H-1-benzopyran-6-yl)-1-piperazinyl]-butyl]-1H-indole-5-carbonitrile 8i and its fluorine analogue 6-[4-[4-(5-fluor-3-indolyl)-butyl]-1-piperazinyl]-2H-1-benzopyran-2-one have been identified. These unsubstituted chromenones are equally potent at the 5-HT(1A) receptor and 5-HT transporter. The implementation of nitrogen functionalities in position 3 of the chromenones resulted in compounds acting as agonists at the 5-HT(1A) receptor and as 5-HT re-uptake inhibitors like vilazodone. Ex vivo 5-HT re-uptake inhibition and in vitro 5-HT agonism were determined in the PCA- and GTPgammaS-assay, respectively. The potential of these chromenones to increase central 5-HT levels was measured in microdialysis studies and especially the derivatives 3-[4-[4-(3-amino-2-oxo-2H-chromen-6-yl)-piperazin-1-yl]-butyl]-1H-indole-5-carbonitrile 8f, ethyl (6-[4-[4-(5-cyano-1H-indol-3-yl)-butyl]-piperazin-1-yl]-2-oxo-2H-chromen-3-yl)-carbamate 8h and N-(6-[4-[4-(5-cyano-1H-indol-3-yl)-butyl]-piperazin-1-yl]-2-oxo-2H-chromen-3-yl)-acetamide 8k give rise to rapid development of increased serotonin levels in rat brain cortex, lasting longer than 3h.


Subject(s)
Benzofurans/pharmacology , Indoles/pharmacology , Piperazines/pharmacology , Selective Serotonin Reuptake Inhibitors/pharmacology , Serotonin 5-HT1 Receptor Agonists , Serotonin Receptor Agonists/pharmacology , Animals , Benzofurans/chemistry , Benzofurans/metabolism , Benzopyrans/chemistry , Benzopyrans/metabolism , Benzopyrans/pharmacology , Butylamines/chemistry , Butylamines/metabolism , Butylamines/pharmacology , Drug Combinations , Indoles/chemistry , Indoles/metabolism , Male , Molecular Structure , Piperazines/chemistry , Piperazines/metabolism , Protein Binding/drug effects , Protein Binding/physiology , Rats , Receptor, Serotonin, 5-HT1A/metabolism , Serotonin Receptor Agonists/chemistry , Serotonin Receptor Agonists/metabolism , Selective Serotonin Reuptake Inhibitors/chemistry , Selective Serotonin Reuptake Inhibitors/metabolism , Vilazodone Hydrochloride
7.
J Med Chem ; 47(19): 4677-83, 2004 Sep 09.
Article in English | MEDLINE | ID: mdl-15341483

ABSTRACT

A series of new 1-[4-(indol-3-yl)butyl]-4-arylpiperazines was prepared to identify highly selective and potent 5-HT(1A) agonists as potential pharmacological tools in studies of mood disorders. The combination of structural elements (indole-alkyl-amine and aryl-piperazine) known to introduce 5-HT(1A) receptor affinity and the proper selection of substituents (R on the indole moiety and R' on the aryl moiety) led to compounds with high receptor specificity and affinity. In particular, the introduction of the methyl ether or the unsubstituted carboxamide as substituents in position 5 of the indole (R) guaranteed serotonergic 5-HT(1A) affinity compared to the unsubstituted analogue. Para-substituted arylpiperazines (R') decreased dopaminergic D(2) binding and increased selectivity for the 5-HT(1A) receptor. Agonistic 5-HT(1A) receptor activity was confirmed in vivo in the ultrasonic vocalization test, and the results suggest that the introduction of the carboxamide residue leads to better bioavailability than the corresponding methyl ether. 3-[4-[4-(4-Carbamoylphenyl)piperazin-1-yl]butyl]-1H-indole-5-carboxamide 54 was identified as a highly selective 5-HT(1A) receptor agonist [GTPgammaS, ED(50) = 4.7 nM] with nanomolar 5-HT(1A) affinity [IC(50) = 0.9 nM] and selectivity [D(2), IC(50) > 850 nM]. 3-[4-[4-(4-Methoxyphenyl)piperazin-1-yl]butyl]-1H-indole-5-carboxamide 45 is one of the most potent and selective 5-HT(1A) agonists known [5-HT(1A), IC(50) = 0.09 nM; D(2), IC(50) = 140 nM].


Subject(s)
Butylamines/chemistry , Butylamines/pharmacology , Indoles/chemistry , Serotonin 5-HT1 Receptor Agonists , Animals , Biological Availability , Butylamines/administration & dosage , Butylamines/pharmacokinetics , Hippocampus/drug effects , Hippocampus/metabolism , Humans , Indoles/pharmacology , Inhibitory Concentration 50 , Male , Molecular Structure , Oxindoles , Piperidines/chemistry , Piperidines/pharmacology , Pyridines/chemistry , Pyridines/pharmacology , Rats , Receptor, Serotonin, 5-HT1A/metabolism , Structure-Activity Relationship , Substrate Specificity
8.
J Med Chem ; 47(19): 4684-92, 2004 Sep 09.
Article in English | MEDLINE | ID: mdl-15341484

ABSTRACT

Systematic structural modifications of indolealkylphenylpiperazines led to improved selectivity and affinity within this class of 5-HT(1A) receptor agonists. Introduction of electron-withdrawing groups in position 5 on the indole raises serotonin transporter affinity, and the cyano group proved to be the best substituent here. 5-Fluoro and 5-cyano substituted indoles show comparable results in in vitro and in vivo tests, and bioisosterism between these substituents was supported by calculation of the molecular electrostatic potentials and dipole moments. Compounds showing promising in vitro data were further examined in ex vivo (p-chloroamphetamine assay) and in vivo (ultrasonic vocalization) tests. Optimization of the arylpiperazine moiety indicated that the 5-benzofuranyl-2-carboxamide was best suited to increase 5-HT transporter and 5-HT(1A) receptor affinity and to suppress D(2) receptor binding. 5-[4-[4-(5-Cyano-3-indolyl)butyl]-1-piperazinyl]benzofuran-2-carboxamide 29 (vilazodone, EMD 68843) was identified as a highly selective 5-HT(1A) receptor agonist [GTPgammaS, ED(50) = 1.1 nM] with subnanomolar 5-HT(1A) affinity [IC(50) = 0.2 nM] and as a subnanomolar 5-HT reuptake inhibitor [RUI = 0.5 nM] showing a great selectivity to other GPCRs (e.g., D(2), IC(50) = 666 nM).


Subject(s)
Indoles/chemistry , Piperazines/chemistry , Piperazines/pharmacology , Selective Serotonin Reuptake Inhibitors/chemistry , Selective Serotonin Reuptake Inhibitors/pharmacology , Serotonin 5-HT1 Receptor Agonists , Serotonin/metabolism , Animals , Biological Transport/drug effects , Hippocampus/drug effects , Hippocampus/metabolism , Inhibitory Concentration 50 , Molecular Structure , Piperazine , Piperazines/chemical synthesis , Rats , Receptors, Serotonin, 5-HT1/metabolism , Selective Serotonin Reuptake Inhibitors/chemical synthesis , Structure-Activity Relationship
9.
Eur J Pharmacol ; 473(2-3): 229-30, 2003 Jul 25.
Article in English | MEDLINE | ID: mdl-12892843

ABSTRACT

The 5-HT2A receptor ligand 7-[4-[2-(4-fluoro-phenyl)-ethyl]-piperazine-1-carbonyl]-1H-indole-3-carbonitrile HCl (EMD 281014) selectively binds to human (h) and rat 5-HT2A receptors (IC50 values 0.35 and 1 nM, respectively; vs. 1334 nM for h5-HT2C) and inhibited 5-HT-stimulated [35S]guanosine 5'-O-3-thiotriphosphate (GTPgammaS)-accumulation in h5-HT2A transfected Chinese hamster ovary cells (IC50 9.3 nM). EMD 28014 counteracted the N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ)-induced decrease of [3H]ketanserin binding in rat frontal cortex (ID50 0.4 mg/kg p.o.) and R-(-)-1-(2,5-dimethoxy-4-iodophenyl)-aminopropane (DOI)-induced head-twitch behaviour in mice (ID50 0.01 mg/kg s.c., 0.06 mg/kg p.o.), demonstrating unique selectivity and efficacy.


Subject(s)
Indoles/pharmacology , Piperazines/pharmacology , Psychotropic Drugs/pharmacology , Serotonin 5-HT2 Receptor Antagonists , Animals , Brain/metabolism , CHO Cells , Cricetinae , Humans , Indoles/administration & dosage , Male , Mice , Piperazines/administration & dosage , Psychotropic Drugs/administration & dosage , Radioligand Assay
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