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G Lambrecht

Publications and source records attributed to G Lambrecht.

At least 37 records · Page 2Linked to original sources
PubMed1997

Evidence for muscarinic M4 receptors mediating nonadrenergic noncholinergic relaxations in rabbit anococcygeus muscle.

The aim of the present study was to characterize the muscarinic receptor subtype mediating nonadrenergic noncholinergic (NANC) relaxations in the rabbit anococcygeus muscle (RAM) by the use of muscarinic receptor agonists and a battery of key muscarinic antagonists. In addition, experiments were carried out to identify the NANC neurotransmitter(s) involved in the inhibitory NANC neurotransmission. In preparations with histamine-raised tone, the nonselective muscarinic agonists (pD2 values) (+)-muscarine (5.23), cis-dioxolane (5.16), oxotremorine M (4.95) and carbachol (4.06) produced concentration-dependent relaxations corresponding to 72.6-85.0% of the histamine-induced precontraction. In contrast, the subtype-preferring (M1/M4 over M2 and M3 receptors) agonists 4-(3-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium chloride (McN-A-343), (S)-4-(dimethylamino)-1-methyl-2-butynyl-N-(3-chlorophenyl)carbamate methobromide [(S)-BN 228], (R)- and (S)-N-methyl-N-(1-methyl-4-pyrrolidino-2-butynyl)acetamide [(R)- and (S)-BM 5] showed no or rather low [(S)-BN 228] muscarinic activity. The low potencies, together with the ineffectiveness of some agonists, indicated a low effective receptor reserve associated with the relaxant response. No contractile responses to (+)-muscarine were observed neither in unstimulated nor in precontracted preparations suggesting that the existence of an excitatory postjunctional muscarinic receptor may be excluded. The nicotinic antagonist hexamethonium had no influence on relaxant responses to (+)-muscarine, but abolished relaxations elicited by (-)-nicotine. This demonstrates that the RAM contains also nicotinic acetylcholine receptors (AchRs) mediating inhibitory NANC responses. Relaxations induced by the stimulation of muscarinic and nicotinic AchRs as well as by electrical field stimulation (EFS) were completely abolished by tetrodotoxin and were also sensitive to the nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine (L-NOARG), indicating that NO plays an important role as an inhibitory NANC transmitter in RAM. All muscarinic antagonists investigated did not influence the histamine-induced precontraction, but shifted the concentration-response curve of (+)-muscarine to the right in a parallel fashion. Schild analysis yielded regression lines of unit slope, indicating competitive antagonism. The following rank order of antagonist potencies (pA2 values) was found: 11-([4-[4-(diethylamino)-butyl]-1-piperidinyl]-acetyl-5,11-dihydro-6H-py rido (2,3-b) (1,4)-benzodiazepine-6-one hydrochloride (AQ-RA 741; 8.08) = himbacine (8.03) > or = tripitramine (7.69) > or = p-fluorohexahydro-sila-difenidol (p-F-HHSiD; 7.48) > or = methoctramine (7.30) > or = pirenzepine (7.18) > or = guanylpirenzepine (6.24). A comparison of the pA2 values determined in the RAM with published data from binding studies at muscarinic M1-M4 and m5 receptors suggests that the functional muscarinic receptor mediating NANC relaxation in the RAM is of the M4 subtype. Taken together, the results obtained in the present study provide convincing evidence that relaxant responses elicited by muscarinic stimuli in RAM are nitrergic in nature and mediated by muscarinic M4 receptors located somadendritically on NANC neurons. Thus, the isolated RAM may serve as a functional muscarinic M4 receptor model.

Animals↗
PubMed1997

Characterization of postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle.

The present study was designed to characterize the postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle by means of a series of muscarinic agonists and subtype-preferring key muscarinic antagonists. Cumulative addition of muscarinic agonists elicited concentration-dependent contractions with the following rank order of potency (pD2 values): (+)-muscarine (6.36) > or = oxotremorine M (6.21) > or = arecaidine propargyl ester (APE) (6.18) > carbachol (5.68) = (+/-)-methacholine (5.65) > 4-(4-chlorophenyl-carbamoyloxy)-2-butynyltrimethylammonium chloride (4-Cl-McN-A-343) (4.28) > 4-(3-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium chloride (McN-A-343) (3.89). (+)-Muscarine, oxotremorine M, carbachol and (+/-)-methacholine behaved as full agonists, whereas APE, 4-Cl-McN-A-343 and McN-A-343 displayed partial agonism. The contractile responses of the rat anococcygeus muscle to (+/-)-methacholine were competitively antagonized by pirenzepine (pA2 = 6.92), 11-[[4-[4-(diethylamino)butyl]-1-piperidinyl]acetyl] 5,11-dihydro-6H-pyrido(2,3-b) (1,4)-benzodiazepine-6-one (AQ-RA 741; pA2 = 6.75), himbacine (pA2 = 7.11), (+/-)-p-fluoro-hexahydro-sila-difenidol (p-F-HHSiD; pA2 = 7.68) and the (R)- and (S)-enantiomers of hexahydro-difenidol [(R)-HHD: pA2 = 8.52; (S)-HHD: pA2 = 6.06]. A comparison of the pA2 values derived from studies of contraction in rat anococcygeus muscle with literature binding (pKi values) and functional affinities (pA2 values) obtained at native M1-M4 receptors strongly suggests that the postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle are of the M3 subtype.

Animals↗
PubMed1997

Vasoconstrictor responses via P2X-receptors are selectively antagonized by NF023 in rabbit isolated aorta and saphenous artery.

1. The effects of NF023, the symmetrical 3'-urea of 8-(benzamido)naphthalene-1,3,5-trisulphonic acid), and its parent compound suramin were investigated on vasoconstrictor responses to alpha, beta-methylene ATP in rabbit isolated saphenous artery and vasodilator responses to ATP in noradrenaline-precontracted rabbit isolated thoracic aorta. 2. In rabbit isolated saphenous artery, alpha, beta-methylene ATP-induced vasoconstrictor responses via P2X-receptors were concentration-dependently and competitively antagonised by NF023 (30-300 microM; pA2 = 5.69 +/- 0.04). Suramin (100-1000 microM) also competitively blocked vasoconstrictor responses to alpha, beta-methylene ATP, albeit with lower potency (pA2 = 4.79 +/- 0.05). In contrast, NF023 (100 microM) did not significantly affect contractile responses to noradrenaline or histamine in the saphenous artery. 3. In noradrenaline-precontracted rabbit isolated thoracic aorta preparations, ATP (3-3000 microM) concentration-dependently induced relaxations via endothelium-dependent or smooth muscle P2Y-receptor subtypes. NF023 (30-300 microM) failed to block relaxant responses to ATP at endothelium-dependent P2Y-receptors, whereas suramin (100-1000 microM) did antagonise endothelium-dependent vasodilator responses to ATP. Neither NF023 (100 microM) nor suramin (300 microM) influenced vasorelaxant responses to ATP via endothelium-independent P2Y-receptors. 4. In conclusion, this study outlines the selectivity of NF023 as an effective P2X-receptor antagonist in rabbit isolated blood vessels without affecting endothelium-dependent or endothelium-independent P2Y-receptor subtypes, adrenoceptors or histamine receptors.

Adenosine Triphosphate↗
PubMed1996

Stereoselective interaction of uncharged esters at four muscarinic receptor subtypes.

We investigated the binding and pharmacological properties of the esters of 3,3-dimethylbutan-1-ol (the carbon analogue of choline) with either diphenylglycolic acid, (R)-phenylcyclohexylglycolic acid, or (S)-phenylcyclohexylglycolic acid [BS-6181, (R)-BS-7826 and (S)-BS-7826, respectively] at muscarinic M1, M2, M3 (Hm3) and M4 receptors. The three uncharged compounds were muscarinic receptor antagonists, with pA2 or pKi values between 7.9 and 5.6. The achiral ester BS-6181 displayed highest affinity for M1, M3 (Hm3) and M4 receptors (pA2 or pKi = 7.2-7.6) and lower affinity for M2 receptors (pA2 or pKi = 6.7 and 6.8). The four muscarinic receptor subtypes were able to distinguish between the two enantiomers of the cyclohexyl derivative of BS-6181 [(R)- and (S)-BS-7826], with a preference for the (R)-isomer (up to 79-fold). Interestingly, the (S)-enantiomer of BS-7826, being the distomer, was found to be M4 selective (pKi/M4 = 6.9; pA2 or pKi/M1-M3 (Hm3) = 5.6-6.2). These results indicate that uncharged compounds may (stereo)selectively bind to muscarinic receptors via hydrophobic interactions. Thus, an ionic bond between muscarinic ligands and an anionic site of the receptor is not absolutely necessary for recognition of muscarinic receptors.

Animals↗
PubMed1996

Design and pharmacology of selective P2-purinoceptor antagonists.

1. The symmetrical 3'-urea of 8-(benzamido)naphthalene-1,3,5-trisulfonic acid (NF023) and pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) were investigated for their ability to antagonize responses mediated via P2-purinoceptors. In addition, putative inhibitory effects of these compounds on ecto-nucleotidase activity were studied. 2. In rabbit vas deferens, PPADS antagonized contractions to alpha, beta-methylene ATP (alpha, beta-mATP) in a pseudoirreversible manner (pA2 = 6.34). This P2X-antagonism by PPADS has been confirmed in certain vascular and visceral smooth muscles (pA2 = 6.02-6.41). PPADS inhibited P2Y-receptor-mediated relaxant responses to ADP-beta-S or 2-methylthio ATP (2-MeSATP) in guinea-pig taenia coli (pA2 = 4.59 and 5.26, respectively) as well as in rat duodenum (pA2 = 5.09) and mesenteric artery (pA2 = 5.46). Experiments on the rat mesenteric arterial bed demonstrated the ineffectiveness of PPADS at P2U-purinoceptors. P2T-purinoceptor-mediated platelet aggregation was affected only at high concentrations of PPADS (> 100 microM). PPADS (100 microM) was also weakly active in inhibiting ecto-nucleotidase activity in guinea-pig taenia coli. 3. In rabbit vas deferens, NF023 antagonized P2X-purinoceptor-mediated contractions to alpha, beta-mATP in a competitive manner. The estimated pA2 value of 5.68 was very similar to that obtained at P2X-receptors in rat mesenteric (pA2' = 5.54) and rabbit saphenous artery (pA2 = 5.69). In rat duodenum and guinea-pig taenia coli, NF023 competitively inhibited relaxant responses to the P2Y-selective agonists, ADP-beta-S and 2-MeSATP (pA2 = 4.00-4.25). In rat mesenteric arterial bed, NF023 antagonized vasodilator responses mediated by P2Y-purinoceptors (pA2 = 4.94), but had no effect on vasodilator responses to the P2U-selective agonist, UTP. NF023 was found to inhibit ectonucleotidase activity in guinea-pig taenia coli and rabbit vas deferens (pIC40 = 3.52 and 4.12, respectively). 4. At 100 microM, PPADS and NF023 did not interact with alpha 1- and alpha 2-adrenoceptors, adenosine A1- and A2-, histamine H1- and muscarinic M1-, M2- and M3-receptors. 5. In conclusion, the results demonstrate that PPADS and NF023 are specific P2-receptor antagonists and useful tools in the study of multiple subtypes of P2-purinoceptors.

Animals↗
PubMed1996

Influence of monovalent cations on the binding of a charged and an uncharged ('carbo'-)muscarinic antagonist to muscarinic receptors.

1. The effect of the buffer concentration on binding of [3H]-N-methylscopolamine to muscarinic receptors M2 was tested in rat heart. Tracer binding was of low affinity in a 20 mM imidazole buffer (pKD 8.3), inhibited by an increase from 10 to 100 mM of the sodium phosphate buffer concentration (pKD 9.92 to 9.22), slightly inhibited by an increase of the Tris/HC1 buffer concentration from 20 to 100 mM (pKD 9.70 to 9.47) and unaffected by an increase of the histidine/HC1 buffer concentration from 20 to 100 mM (pKD 9.90 to 9.82). We chose the last buffer to analyse the effect of ions on antagonists binding to cardiac M2 receptors and to transiently expressed wild-type and (Y533-->F) mutant m3 muscarinic receptors in COS-7 cells. 2. Equilibrium [3H]-N-methylscopolamine binding to cardiac M2 receptors was inhibited, apparently competitively, by monovalent salts (LiCl > or = NaCl > or = KCl). In contrast, binding of the uncharged 3,3-dimethylbutan-1-ol ester of diphenylglycolic acid (BS-6181) was facilitated by addition of monovalent salts (LiCl > or = NaCl > or = KCl) to the binding buffer. This cation binding pattern is consistent with interaction with a large, negative field strength binding site, such as, for instance, a carboxylic acid. 3. In the presence of 100 mM NaCl, [3H]-N-methylscopolamine had a similar affinity for the wild-type m3 receptor (pKD 9.85) and for a (Y533-->F) mutant m3 receptor (pKD 9.68). However, in the absence of added salts, the tracer had a significantly lower affinity for the mutated (pKD 10.19) as compared to the wild-type (pKD 10.70) m3 receptor. BS-6181 had a significantly lower affinity for the (Y533-->F) mutant m3 muscarinic receptor, as compared to the wild-type m3 receptor, both in the absence (pKD 6.19-6.72) in the presence (pKD 6.48-7.40) of 100 mM NaCl. The effects of NaCl on binding of the uncharged ester and of [3H]-N-methylscopolamine to the m3 receptor were decreased by the mutation. 4. Taken together, these results support the hypothesis that monovalent cations from the buffer may interact with the cation binding site of the receptors (an aspartate residue in the third transmembrane helix of muscarinic receptors). Buffer cations may inhibit competitively the binding of (charged) muscarinic ligands having a tertiary amine or ammonium group, while facilitating the receptor recognition by uncharged, isosteric 'carbo-analogues'. Mutation of the (Y533-->F) of the m3 receptor decreased the affinity of the receptor for positive charges, including the sodium ion.

Animals↗
PubMed1996

Stereoselective recognition of the enantiomers of phenglutarimide and of six related compounds by four muscarinic receptor subtypes.

1. We have compared the binding properties of the enantiomers of phenglutarimide (1) and of six related compounds to M1 receptors in NB-OK-1 cells, M2 receptors in rat heart, M3 receptors in rat pancreas and the M4 receptors of rat striatum, with their functional (antimuscarinic) properties in rabbit vas deferens (M1/M4-like), guinea-pig atria (M2) and guinea-pig ileum (M3) receptors. The binding properties of the enantiomers of three of the compounds were also measured on cloned human m1-m4 receptors expressed by CHO cells, using [3H]-N-methylscopolamine ([3H]-NMS) as radioligand. 2. The high affinity enantiomers behaved as competitive antagonists in binding and pharmacological studies. (S)-phenglutarimide (pKi-M1 = 9.0/9.3) and (R)-thienglutarimide (pKi-M1 = 8.6/9.2) recognized selectively the native M1 > M4 > M3 > M2 receptors in tissues as well as the respective cloned receptors. 3. The pA2 values at the inhibitory heteroreceptors in the rabbit vas deferens, and at the guinea-pig atria and ileum for the seven more potent enantiomers were compatible with the previous classification of these receptors as M1/M4-like, M2 and M3, respectively. 4. Replacement of the phenyl by a thienyl ring or of the diethylamino by a piperidino group in the phenglutarimide molecule did not affect markedly the potencies of the high affinity enantiomer. In contrast, replacement of the phenyl by a cyclohexyl ring decreased 20 fold the active enantiomers potency. Methylation of the piperidine-2,6-dione nitrogen also reduced markedly the eutomers' affinities, more on the M1 than on the other subtypes. 5. The selectivity profiles (recognition of four receptor subtypes) of six of the seven less active enantiomers were different from the corresponding more active enantiomers selectivity profiles, suggesting that the preparations used in this study were pure. However, we cannot not exclude the hypothesis that the batch of (S)-thienglutarimide used in this study was contaminated by less than 0.02% of the eutomer. 6. In contrast with the eutomer binding site, replacement of the phenyl ring by a thienyl or cyclohexyl ring did not affect binding of the low affinity enantiomers to the muscarinic receptor or the [3H]-NMS-receptor complex. The replacement of the diethylamino group by a piperidine ring, and N-methylation of the piperidine-2,6 dione moiety increased slightly these enantiomers' potencies. 7. The muscarinic receptors were extremely stereoselective, and had up to 20000 fold lower affinity for the less active enantiomers. However, the stereochemical requirements of the muscarinic receptor subtypes were different for the enantiomers of compounds 1-7, being most stringent at M1 receptors. 8. The weaker enantiomers behaved as competitive antagonists in pharmacological studies, at least in the concentration-range investigated.

Animals↗
PubMed1995

The (S)-(+)-enantiomer of dimethindene: a novel M2-selective muscarinic receptor antagonist.

The present study was designed to determine to determine the in vitro affinity profile of (R)-(-)-dimethindene and (S)-(+)-dimethindene at muscarinic receptor subtypes using both functional and binding assays. In addition, the racemate was investigated in functional studies. The functional muscarinic receptors studied were putative M1 receptors in rabbit vas deferens and rat duodenum, M2 receptors in guinea-pig left atria and rabbit vas deferens, as well as M3 receptors in guinea-pig ileum and trachea. Furthermore, the histamine H1 antagonism by (R)-(-)- and (S)-(+)-dimethindene has been examined in guinea-pig ileum. Muscarinic binding selectivity was assessed in homogenates from human neuroblastoma NB-OF 1 cells (M1), rat heart (M2), pancreas (3) and striatum (M4). The results demonstrate that (S)-(+)-dimethindene is a potent M2-selective muscarinic receptor antagonist (pA2 = 7.86/7.74; pKi = 7.78) with lower affinities for the muscarinic M1 (pA2 = 6.83/6.36; pKi = 7.08), the M3 (pA2 = 6.92/6.96; pKi = 6.70) and the M4 receptors (pKi = 7.00), respectively. The (S)-(+)-enantiomer was more potent (up to 41-fold) than the (R)-(-)-enantiomer in all muscarinic assays. In contrast, the stereoselectivity was inverse at histamine H1 receptors, the (R)-(-)-enantiomer being the eutomer (pA2 = 9.42; pA2/(S)-isomer = 7.48). In conclusion, (S)-(+)-dimethindene is a useful tool to investigate muscarinic receptor heterogeneity. In addition, this lipophilic compound might become the starting point for the development of M2-selective muscarinic receptor antagonists useful as diagnostic tools for quantifying muscarinic M2 receptors in the central nervous system with positron emission tomography imaging, and to test the hypothesis that muscarinic M2 receptor antagonists show beneficial effects in the treatment of cognitive disorders.

Animals↗
PubMed1995

Regioisomeric 5(3)-aminomethyl-3(5)-phenylisoxazoles: synthesis, spectroscopic discrimination, and muscarinic activity.

The regioselective synthesis of isomeric 5(3)-aminomethyl-3(5)-phenyl isoxazoles using different methods is described. Spectroscopic data, especially mass spectrometric fragmentation, were used to identify and characterize the regioisomers. The muscarinic activity of these isoxazoles was assayed on isolated guinea-pig ileum and atria as well as on isolated rabbit vas deferens.

Animals↗
PubMed1995

The design and pharmacology of novel selective muscarinic agonists and antagonists.

The muscarinic pharmacology of C1-methyl-substituted chiral compounds related to McN-A-343 and of (R)- and (S)-dimethindene has been studied. Among the McN-A-343 analogues, the (S)-enantiomers were more potent and had higher affinity than the (R)-isomers. The quaternary compound (S)-BN 228 was found to be the most potent M1-selective agonist known today (pEC50: M1/rabbit vas deferens = 7.83; M2/guinea-pig atria = 6.35; M3/guinea-pig ileum = 6.29). In both the atria and ileum the tertiary carbamate, (S)-4-F-MePyMcN, was a competitive antagonist (pA2 value = 7.39 and 6.82, respectively). In contrast, in rabbit vas deferens (S)-4-F-MePyMcN was a potent partial agonist (pEC50 = 7.22; apparent efficacy = 0.83). These results indicate that (S)-4-F-MePyMcN might be a useful tool to study M1 receptor-mediated effects involved in central cholinergic function. (S)-Dimethindene was a potent M2-selective antagonist (pA2 = 7.86/atria; pKi = 7.8/rat heart) with lower affinities for the M1 (pA2 = 6.36/rat duodenum; pKi = 7.1/NB-OK 1 cells), M3 (pA2 = 6.92/guinea-pig ileum; pKi = 6.7/rat pancreas) and M4 receptors (pKi = 7.0/rat striatum). It was more potent (up to 41-fold) than the (R)-isomer. In contrast, the stereoselectivity was inverse at ileal H1 receptors (pA2: (R)-isomer = 9.42; (S)-isomer = 7.48). Thus, (S)-dimethindene could be a valuable agent to test the hypothesis that M2 antagonists show beneficial effects in the treatment of cognitive disorders. It might also become the starting point for the development of diagnostic tools for quantifying M2 receptors in the CNS with PET imaging.

(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethy↗
PubMed1995

Inhibitory action of PPADS on relaxant responses to adenine nucleotides or electrical field stimulation in guinea-pig taenia coli and rat duodenum.

1. The effect of pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) on the relaxant response to adenine nucleotides was examined in the carbachol-contracted guinea-pig taenia coli and rat duodenum, two tissues possessing P2y-purinoceptors. In addition, in the taenia coli PPADS was investigated for its effect on relaxations evoked by adenosine, noradrenaline and electrical field stimulation. In order to assess the selectivity of PPADS between P2-purinoceptor blockade and ectonucleotidase activity, its influence on ATP degradation was studied in guinea-pig taenia coli. 2. The resulting rank order of potency for the adenine nucleotides in guinea-pig taenia coli was: 2-methylthio ATP >> ATP > alpha,beta-methylene ATP with the respective pD2-values 7.96 +/- 0.08 (n = 23), 6.27 +/- 0.12 (n = 21) and 5.88 +/- 0.04 (n = 24). 3. In guinea-pig taenia coli, PPADS (10-100 microM) caused a consistent dextral shift of the concentration-response curve (CRC) of 2-methylthio ATP and ATP resulting in a biphasic Schild plot. A substantial shift was only observed at 100 microM PPADS, the respective pA2-values at this particular concentration were 5.26 +/- 0.16 (n = 5) and 5.15 +/- 0.13 (n = 6). Lower concentrations of PPADS (3-30 microM) antagonized the relaxant effects to alpha,beta-methylene ATP in a surmountable manner. An extensive shift of the CRC was produced only by 30 microM PPADS (pA2 = 5.97 +/- 0.08, n = 6), and the Schild plot was again biphasic. 4. The relaxant responses to electrical field stimulation (80 V, 0.3 ms, 5 s, 0.5-16 Hz) in guinea-pigtaenia coli were concentration-dependently inhibited by PPADS (10-100 microM).5. In guinea-pig taenia coli, the potency of ATP in inducing relaxation appeared to be independent of its rate of degradation by ecto-nucleotidases, since the Km-value (366 microM) obtained in the enzyme assay was much higher than the functional EC50-value (0.45 microM) of ATP. PPADS (3-100 microM) was only weakly active in inhibiting ecto-nucleotidase activity leaving a residual activity of 81.8 +/- 5.1% at 100 microM.Enzyme inhibition by PPADS was concentration-independent and non-competitive.6. In rat duodenum, the rank order of potency was: 2-methylthio ATP >ATP> >alpha,beta-methylene ATP,the respective pD2-values being 6.98 +/- 0.04 (n = 76), 6.26 +/- 0.02 (n = 6) and 4.83 +/- 0.02 (n = 6). Among these agonists, 2-methylthio ATP displayed the lowest apparent efficacy.7. The CRC of 2-methylthio ATP in rat duodenum was shifted to the right by PPADS (10-100 microM) ina concentration-dependent manner, and Schild analysis gave a pA2-value of 5.09 +/- 0.06 (slope = 1.02,n=14).8 PPADS was without any effect on the carbachol-induced contraction in guinea-pig taenia coli or rat duodenum and on the relaxation to noradrenaline or adenosine in guinea-pig taenia coli.9 In conclusion, the antagonistic properties of PPADS at the taenia coli and rat duodenum P2y-purinoceptors were different from those recently described at the P2x-subtype: inhibition of P2y-purinoceptor-mediated responses was observed at higher concentrations (3-100 microM vs. 1-10 (30) microM).Furthermore, we conclude that in addition to the classical P2y-subtype, which is largely PPADS-resistant,the guinea-pig taenia coli may be endowed with a distinct relaxation-mediating P2-purinoceptor subtype which is sensitive to PPADS.

Adenosine↗
PubMed1995

Two different inhibitory effects of pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid on adenosine diphosphate-induced human platelet aggregation.

In the present study, the novel compound pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), which has been shown to inhibit P2x-purinoceptor-mediated contractions in smooth muscle, was investigated for its antagonistic effects on adenosine diphosphate (ADP)-induced human platelet aggregation in platelet-rich plasma (PRP) and in washed platelets, respectively. Suramin served as reference compound. In PRP, suramin (1 mmol/l) was inactive whereas PPADS (1 mmol/l) considerably reduced the extent of aggregation. In contrast, both suramin (1 mmol/l) and PPADS (500 mumol/l) markedly depressed the aggregation of washed platelets. In addition, there was a peculiarity in washed platelets: a delay of onset of platelet aggregation up to 15 min in the presence of PPADS (10-500 mumol/l) and suramin (0.1-1 mmol/l). Thus, in the present study washed platelets were more suited to detect an influence of PPADS and suramin as inhibitors of ADP-induced aggregation, and a delay of onset of aggregation was the most sensitive parameter in this regard. Comparing the effective threshold concentration of PPADS at P2x (1 mumol/l)- and the platelet P2t-purinoceptor, PPADS proves to be P2x-selective.

Adenosine Diphosphate↗
PubMed1995

Aliphatic and heterocyclic analogues of arecaidine propargyl ester. Structure-activity relationships of mono- and bivalent ligands at muscarinic M1 (M4), M2 and M3 receptor subtypes.

The pharmacological profiles of tertiary and quaternary monovalent (1b-6b) and bivalent ligands (7a-12b), closely related to arecaidine propargyl ester (CAS 35516-99-5, APE, 1a), were evaluated at muscarinic receptors in rat superior cervical ganglia (M1), rabbit was deferens (M1/M4-like), guinea-pig atria (M2) and guinea-pig ileum (M3). In the monovalent ligand series (1a-6b) APE (1a) displayed the highest potency at all three muscarinic receptors [M2 (-log EC50 = 8.12) > or = M3 (-log EC50 = 7.77) = M1/M4 (-log EC50/vas deferens = 7.72)], whereas in the bivalent ligand series (7a-12b) arecaidine 2-butyne-1,4-diyl bisester (bisABE, 7a) was the most potent agonist with functional selectivity for M1/M4 (-log EC50/vas deferens = 6.94) and M2 receptors (-log EC50 = 7.10) over M3 receptors (-log EC50 = 6.27). On ganglia bisABE elicited M2 receptor-mediated hyperpolarisations, which were followed by long-lasting pirenzepine-sensitive depolarisations. However, the potency at M1 receptors in ganglia of APE (-log EC50 = 6.96) and bisABE (-log EC50 = 5.69) was lower than that in rabbit vas deferens. All bivalent molecules exhibited decreased potencies when compared with their monovalent analogues. However, a change in potency profiles was often obtained. The quaternary isonicotinic acid 2-butyne-1,4-diyl bisester (10b) displayed some functional selectivity for M2 receptors (-log EC50 = 5.78) [6- to 9-fold over M1/M4 (-log EC50/vas deferens = 5.03) and M3 receptors (-log EC50 = 4.83)] without showing nicotinic effects.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Characterization of the prejunctional muscarinic receptors mediating inhibition of evoked release of endogenous noradrenaline in rabbit isolated vas deferens.

The aim of the present study was to characterize the prejunctional modulation of evoked release of endogenous noradrenaline in rabbit vas deferens by the use of muscarinic receptor agonists and subtype-preferring antagonists. Vasa deferentia of the rabbit were stimulated electrically by trains of 120 pulses delivered at 4 Hz or trains of 30 pulses at 1 Hz. The inhibition by muscarinic agonists of the stimulation-evoked overflow of endogenous noradrenaline in the absence and presence of antagonists was used to determine affinity constants for antagonists. These values were compared with those observed at putative M1 receptors inhibiting neurogenic twitch contractions in the rabbit vas deferens and with affinity data obtained at M1(m1)-M4(m5) receptors in functional studies and binding experiments. The evoked overflow of noradrenaline from sympathetic nerves was enhanced by the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), the P2 purinoceptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) and indomethacin, indicating a tonic inhibition by endogenous A1 and P2 purinoceptor agonists and prostanoids, respectively. The stimulation-evoked overflow at 4 Hz was not sensitive to inhibition by the muscarinic agonists methacholine or 4-(4-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium iodide (4-Cl-McN-A-343). In contrast, at a stimulation frequency of 1 Hz the evoked noradrenaline release was decreased by muscarinic agonists (EC50): arecaidine propargyl ester (0.062 microM), 4-Cl-McN-A-343 (0.32 microM), 4-(4-fluorophenylcarbamoyloxy)-2-butynyl-N-methyl-pyrrolidinium tosylate (4-F-PyMcN+; 0.48 microM) and methacholine (0.86 microM). The affinity constants of most of the muscarinic antagonists [atropine: pKB = 9.47; (R)-trihexyphenidyl: pKB = 9.18; pirenzepine: pA2 = 7.68; methoctramine: pKB = 6.90] are consistent with estimates of these antagonists at M1(m1) receptors determined in various functional and binding studies. The high antagonistic potency of pirenzepine and (R)-trihexyphenidyl and the agonistic activity of 4-F-PyMcN+ argue for the involvement of M1, and against that of M2 and M3 receptors in the inhibition of evoked noradrenaline overflow. However, the high apparent pKB of 8.30 for himbacine is not in accordance with an M1 receptor; by contrast, it would be compatible with the presence of M2 or M4 receptors. The potencies of the tested muscarinic agonists and antagonists largely agree with those obtained for the inhibition of neurogenic twitch responses (0.05 Hz) in the rabbit vas deferens.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Pharmacokinetic and pharmacodynamic aspects of an ophthalmic pilocarpine nanoparticle-delivery-system.

The regional pharmacokinetics as well as the pharmacodynamics of pilocarpine-loaded nanoparticles for the treatment of glaucoma were investigated and compared to a solution of this drug. Polybutylcyanoacrylate nanoparticles were prepared by an emulsion polymerization process. Formulations with different drug concentrations (2-6%) as well as different particle concentrations were investigated and analyzed for size and drug loading. Drug binding to the particles was achieved at a level of 10-18% of the total drug content. The colloidal nanoparticles were sufficiently small (diameter: 100-300 nm) for a non-irritating application to the eye. All preparations were applied to the eyes of New Zealand white rabbits which were treated with betamethasone before to create an elevated intraocular pressure (IOP). Pilocarpine concentrations, assayed from aqueous humor using gaschromatography, increased by 23% (AUC) for nanoparticle suspensions compared to aqueous reference solutions. Additionally, t1/2 was prolonged and the elimination coefficient was significantly decreased. Pharmacodynamic effects such as miosis and IOP reduction were investigated. tmax values of aqueous humor concentration were observed to be in a similar time range as miosis tmax readings. It was found that at lower drug contents a more pronounced prolongation of miosis was achieved with nanoparticles versus a standard solution. The IOP-reduction was significantly prolonged with nanoparticles preparations; whereas maximum reduction was obtained with a reference solution after 1-2 hours, it was reached with nanoparticles at about 2-3 hours. Differences between nanoparticles and aqueous solutions were most pronounced at lower drug concentrations.

Animals↗
PubMed1994

Binding and functional properties of hexocyclium and sila-hexocyclium derivatives to muscarinic receptor subtypes.

1. We have compared the binding properties of several hexocyclium and sila-hexocyclium derivatives to muscarinic M1 receptors (in rat brain, human neuroblastoma (NB-OK 1) cells and calf superior cervical ganglia), rat heart M2 receptors, rat pancreas M3 receptors and M4 receptors in rat striatum, with their functional antimuscarinic properties in rabbit vas deferens (M1/M4-like), guinea-pig atria (M2), and guinea-pig ileum (M3) muscarinic receptors. 2. Sila-substitution (C/Si exchange) of hexocyclium (-->sila-hexocyclium) and demethyl-hexocyclium (-->demethyl-sila-hexocyclium) did not significantly affect their affinities for muscarinic receptors. By contrast, sila-substitution of o-methoxy-hexocyclium increased its affinity 2 to 3 fold for all the muscarinic receptor subtypes studied. 3. The p-fluoro- and p-chloro-derivatives of sila-hexocyclium had lower affinities than the parent compound at the four receptor subtypes, in binding and pharmacological studies. 4. In binding studies, o-methoxy-sila-hexocyclium (M1 = M4 > or = M3 > or = M2) had a much lower affinity than sila-hexocyclium for the four receptor subtypes, and discriminated the receptor subtypes more poorly than sila-hexocyclium (M1 = M3 > M4 > M2). This is in marked contrast with the very clear selectivity of o-methoxy-sila-hexocyclium for the prejunctional M1/M4-like heteroreceptors in rabbit vas deferens. 5. The tertiary amines demethyl-hexocyclium, demethyl-sila-hexocyclium and demethyl-o-methoxy-sila-hexocyclium had 10 to 30 fold lower affinities than the corresponding quaternary ammonium derivatives.

Animals↗