Biomedical subjects

J S Ward

Publications and source records attributed to J S Ward.

At least 37 records · Page 2Linked to original sources

PubMed1997

In vivo pharmacology of butylthio[2.2.2] (LY297802 / NNC11-1053), an orally acting antinociceptive muscarinic agonist.

Butylthio[2.2.2] (LY297802 / NNC11-1053) is a mixed muscarinic cholinergic receptor agonist/antagonist that produces antinociception in mice and rats. As such, butylthio[2.2.2] may have therapeutic utility in the treatment of pain. Butylthio[2.2.2] was fully efficacious in the mouse grid shock, writhing, tail-flick and hot plate tests with ED50 values ranging from 1.5 to 12.2 mg/kg after oral administration. In contrast, the ED50 values for morphine ranged from 7.3 to 72 mg/kg after oral administration. Scopolamine was a competitive antagonist of the antinociceptive effects of butylthio[2.2.2]. Butylthio[2.2.2] did not produce either salivation or tremor at therapeutic doses; rather, there was a 50- to >100-fold separation between therapeutic doses and doses which produced side-effects. Butylthio[2.2.2] had high affinity for muscarinic receptors, but little if any affinity for other neurotransmitter receptors or uptake sites. In isolated tissues, butylthio[2.2.2] was an agonist with high affinity at M1 receptors in rabbit vas deferens, an antagonist at M2 receptors in guinea pig atria as well as an antagonist at M3 receptors in guinea pig urinary bladder. Although it has been suggested that M1 receptors mediate the antinociceptive effects of muscarinic agonists, M1 efficacy is not a requirement for antinociception, and, in vivo, the antinociceptive effects of muscarinic agonists are blocked by the intrathecal administration of pertussis toxin, indicating the involvement of m2 or m4 receptors. Since butylthio[2.2.2] is an M2 antagonist, antinociception is therefore most likely mediated by m4 receptors. Butylthio[2.2.2] is currently undergoing clinical development as a novel analgesic.

Administration, Oral↗

PubMed1997

Contrast-enhanced abdominal MR angiography: optimization of imaging delay time by automating the detection of contrast material arrival in the aorta.

PURPOSE: To improve gadolinium-enhanced magnetic resonance (MR) angiogram quality by automatically synchronizing acquisition of central k-space image data with the arterial phase of contrast material bolus infusion. MATERIALS AND METHODS: A spin-echo pulse sequence with orthogonal 90 degrees and 180 degrees pulses was used to monitor signal in a single 4 x 4 x 12-cm voxel that encompassed a segment of aorta. An increase in signal that corresponded to the arrival of gadolinium was used to trigger three-dimensional, spoiled gradient-echo abdominal MR angiography in 50 adult patients. RESULTS: Arterial signal intensity increased 28-fold with automatic compared to 19-fold with manual triggering (P < .05) at an approximate dose of 0.3 mmol/kg. Automatic triggering with a lower dose (approximately 0.2 mmol/kg) resulted in 20-fold arterial enhancement, which is comparable with enhancement after manual triggering at the high dose. In addition, venous enhancement was less (1.5-fold) with automatic than with manual (3.5-fold) triggering at the same dose (P < .05). CONCLUSION: Automatic triggering results in improved arterial-tovenous contrast. It increases arterial enhancement or enables MR angiograms to be obtained with less contrast material. The authors now routinely use this technique for aortorenal imaging with a gadolinium-based contrast material dose of 20 mmol (40 mL) in patients who weigh more than 50 kg and 10 mmol (20 mL) in patients who weigh less than 50 kg.

Abdomen↗

PubMed1997

Hemodynamically significant atherosclerotic renal artery stenosis: MR angiographic features.

PURPOSE: To identify magnetic resonance (MR) angiographic features of hemodynamically significant renal artery stenosis. MATERIALS AND METHODS: Forty-seven patients underwent MR angiography of the renal arteries, including T1-weighted spin-echo and three-dimensional gadolinium-enhanced spoiled gradient-echo and three-dimensional phase-contrast pulse sequences, followed by renal revascularization. Thirty-five patients (52 arteries) were identified who benefited from renal revascularization, which indicated that they had hemodynamically significant renal artery stenoses. Kidney length, cortical thickness, parenchymal enhancement, and poststenotic dilatation were measured. Arteries were also examined for signal drop-out (dephasing) on phase-contrast angiograms; dephasing was considered severe if the stenotic artery appeared occluded on phase-contrast angiograms. RESULTS: Poststenotic dilatation of greater than 20% was present in 36 (59%) of 52 hemodynamically significant renal artery stenoses, and severe dephasing was present in 45 (87%) of 52. In patients with unilateral hemodynamically significant stenosis or occlusion, mean ischemic kidney length was reduced to 9.3 cm compared with 10.7 cm for the contralateral normal kidney (P = .009), mean parenchymal thickness was reduced (1.2 vs 1.7 cm; P < .001), and mean parenchymal enhancement was 15% less on the ischemic side (P = .05). Severe dephasing on phase-contrast angiograms was present in nine (75%) of 12 unilateral hemodynamically significant stenoses but in only one contralateral normal renal artery (P < .001). CONCLUSION: MR angiography depicts features of renal artery stenosis that are markers of hemodynamic significance.

Adult↗

PubMed1997

M1 receptor agonist activity is not a requirement for muscarinic antinociception.

The analgesic effects of a series of muscarinic agonists were investigated by use of the mouse acetic acid writhing, grid-shock, hot-plate and tail-flick tests. The compounds tested were oxotremorine, pilocarpine, arecoline, aceclidine, RS86 and four 3-3(substituted-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahy-dro-1 -methyl pyridines (substituted TZTP), these being propoxy-TZTP, 3-Cl-propylthio-TZTP, xanomeline (hexyloxy-TZTP) and hexylthio-TZTP. These agonists were also assayed for their ability to displace [3H]oxotremorine-M and [3H]pirenz-epine binding and for their functional selectivity at pharmaco-logic M1, M2 and M3 receptors. These compounds all produced dose-dependent antinociceptive effects in all of the mouse analgesia tests. The effects of oxotremorine in the writhing test were fully antagonized by the muscarinic antagonist scopolamine (0.1 mg/kg), but only partially antagonized by methsco-polamine (10 mg/kg) and unaffected by the opioid antagonist naltrexone. 3-Cl-propylthio-TZTP and propoxy-TZTP had virtually no effect at the M1 receptor subtype as measured by the human m1 clone expressed in baby hamster kidney cells or the rabbit vas deferens assay. These compounds, however, were more potent in the analgesia tests than the selective M1 agonists xanomeline and hexylthio-TZTP. These data suggest that muscarinic analgesia is mediated by central muscarinic receptors. However, activity at the M1 receptor subtype is not a requirement for antinociceptive activity.

Analgesics↗

PubMed1997

Butylthio[2.2.2] (NNC 11-1053/LY297802): an orally active muscarinic agonist analgesic.

Butylthio[2.2.2] ((+)-(S)-3-(4-(Butylthio)-1,2,5-thiadiazol-3-yl)-1-azabicyclo[2.2. 2] octane) is an agonist/antagonist at muscarinic receptors. The analgesic potential of butylthio[2.2.2] was assessed in the mouse by use of the grid-shock, tail-flick, hotplate and writhing tests. The ED50 values ranged from 0.19 to 1.47 mg/kg and 1.51 to 12.23 mg/kg 30 min after s.c. and p.o. administration, respectively, yielding p.o./s.c. ratios ranging from 7 to 27. The ED50 values for salivation and tremor were > 30 and 12.31 mg/kg s.c., and > 60 and > 60 mg/kg p.o., yielding therapeutic windows > 130 and 54, and, > 40 and > 40, after s.c. and p.o. administration, respectively. Motor impairment or lethality were only seen at doses 116 and 254 times higher than the antinociceptive doses. Butylthio[2.2.2] was equieffective to, and 3- to 24-fold more potent than morphine. The duration of action was similar to that of morphine. The dose-response curve was shifted dose dependently to the right by the muscarinic antagonist scopolamine but not by the opioid antagonist naltrexone. The antinociceptive effect of butylthio[2.2.2] was reversed by the centrally acting muscarinic antagonist scopolamine but not by the peripherally acting muscarinic antagonist methscopolamine. After 6.5 days repeated dosing in mice, morphine produced marked tolerance, whereas butylthio[2.2.2] produced minimal, if any, tolerance. In the rat grid-shock test, ED50 values of 0.26 mg/kg s.c. and 25.28 mg/kg p.o. were obtained. These data show that butylthio[2.2.2] is a potent and efficacious antinociceptive with a very favorable therapeutic window after s.c. and p.o. administration in mice, and with good efficacy in rats.

Administration, Oral↗

PubMed1997

Pharmacology of butylthio[2.2.2] (LY297802/NNC11-1053): a novel analgesic with mixed muscarinic receptor agonist and antagonist activity.

Butylthio[2.2.2], ((+)-(S)-3-(4-butylthio-1,2,5-thiadiazol-3-yl)-1-azabicyclo[2.2.2] octane; LY297802/NNC11-1053) is a muscarinic receptor ligand which is equiefficacious to morphine in producing antinociception. In vitro, butylthio[2.2.2] had high affinity for muscarinic receptors in brain homogenates, but had substantially less or no affinity for several other neurotransmiter receptors and uptake sites. In isolated tissues, butylthio[2.2.2] was an agonist with high affinity for M1 receptors in the rabbit vas deferens (IC50 = 0.33 nM), but it was an antagonist at M2 receptors in guinea pig atria (pA2 = 6.9) and at M3 receptors in guinea pig urinary bladder (pA2 = 7.4) and a weak partial agonist in guinea pig ileum, which contains a heterogeneous population of muscarinic receptors. In vivo, butylthio[2.2.2] was without effect on acetylcholine, dopamine and serotonin levels in rat brain. Moreover, butylthio[2.2.2] did not decrease charcoal meal transit in mice, nor did it significantly alter heart rate in rats. Further, butylthio[2.2.2] did not produce parasympathomimetic effects such as salivation or tremor in mice, but it antagonized salivation and tremor produced by the nonselective muscarinic agonist oxotremorine. The present data demonstrate that butylthio[2.2.2] is a novel muscarinic receptor mixed agonist/antagonist and its pharmacological profile suggests that it may have clinical utility in the management of pain as an alternative to opioids.

Analgesics↗

PubMed1995

Functionally selective M1 muscarinic agonists. 3. Side chains and azacycles contributing to functional muscarinic selectivity among pyrazinylazacycles.

In an attempt to improve upon the M1 agonist activity of the selective M1 agonist xanomeline and related compounds, the M1 muscarinic efficacies and potencies of 3- and 6-substituted pyrazinylazacycles were varied by changing both the 3- and 6-substituents as well as the azacycle. Significant improvements in efficacy and potency over the previously prepared [3-(hexyloxy)pyrazinyl]tetrahydropyridine 19 were obtained with the [3-(hexyloxy)pyrazinyl]-quinuclidine 5i. The M1 activity of 5i showed some enantioselectivity with (S)-5i being ca. 4-fold more potent than (R)-5i. Like 19 and xanomeline, 5i was a functionally selective M1 agonist that showed greater functional selectivity than widely studied pyrazinylquinuclidine 5n (L-689,660). The improved functional selectivity of 5i over 5n could be attributed to the additional binding interactions between the hexyloxy side chain of 5i and the M1 receptor that are not available to 5n. Although 5i may show M1 functional selectivity comparable to xanomeliine, 5i is a less efficacious and potent M1 agonist than xanomeline.

Animals↗

PubMed1995

Muscarinic agonists as analgesics. Antinociceptive activity versus M1 activity: SAR of alkylthio-TZTP's and related 1,2,5-thiadiazole analogs.

Alkylthio-TZTPs (3-(3-alkylthio-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1-met hylpyridines) and corresponding azabicyclic analogs were tested for m1 efficacy in cloned human m1 receptors and for antinociceptive activity in the mouse grid shock assay. The m1 (%PI) SAR were distinctly different from the analgesia and the salivation SAR, suggesting that analgesia is mediated by neither m1 nor M3 muscarinic receptors.

Analgesics↗

PubMed1995

The muscarinic M1 agonist xanomeline increases soluble amyloid precursor protein release from Chinese hamster ovary-m1 cells.

The functionally selective M1 agonist xanomeline, which is currently undergoing clinical trials as a therapy for Alzheimer's disease, was compared to the muscarinic agonist carbachol for effects on secretion of soluble amyloid precursor protein (APPs) from Chinese hamster ovary cells transfected with the human m1 receptor (CHO-m1). Release of APPs from CHO-m1 cells was increased maximally (4-10 fold) by 100 microM carbachol (EC50 = 11 microM) and by 100 nM xanomeline (EC50 = 10 nM). Stimulation of APPs secretion by xanomeline and carbachol was blocked by preincubation with 1 microM atropine. Carbachol did not stimulate APPs secretion from non-transfected CHO cells. Pilocarpine at 1 mM also increased APPs release. The efficacy of carbachol, xanomeline and pilocarpine for stimulating APPs secretion did not differ significantly. Activation of protein kinase C (PKC) in m1 transfected cell lines by 1 microM phorbol dibutyrate (PDBu) increased APPs release, and this was inhibited 97% by the PKC inhibitor bisindolemalemide. The PKC inhibitor decreased xanomeline and carbachol-stimulated APPs secretion by only 25-30%. These results demonstrate that xanomeline increased APPs release by activation of m1 muscarinic receptors and support the possibility that cholinergic replacement therapy for Alzheimer's Disease may reduce amyloid deposition.

Amyloid beta-Protein Precursor↗

PubMed1994

Xanomeline: a novel muscarinic receptor agonist with functional selectivity for M1 receptors.

Xanomeline [3(3-hexyloxy-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1- methylpyridine] has been evaluated as a muscarinic receptor agonist. In vitro, xanomeline had high affinity for muscarinic receptors in brain homogenates, but had substantially less or no affinity for a number of other neurotransmitter receptors and uptake sites. In cells stably expressing genetic m1 receptors, xanomeline increased phospholipid hydrolysis in CHO, BHK and A9 L cells to 100, 72 and 55% of the nonselective agonist carbachol. In isolated tissues, xanomeline had high affinity for M1 receptors in the rabbit vas deferens (IC50 = 0.006 nM), low affinity for M2 receptors in guinea pig atria (EC50 = 3 microM), was a weak partial agonist in guinea pig ileum and was neither an agonist nor antagonist in guinea pig bladder. In vivo, xanomeline increased striatal levels of dopamine metabolites, presumably by acting at M1 heteroreceptors on dopamine neurons to increase dopamine release. In contrast, xanomeline had only a relatively small effect on acetylcholine levels in brain, indicating that it is devoid of actions at muscarinic autoreceptors. In the gastrointestinal tract, xanomeline inhibited small intestinal and colonic motility, but increased small intestinal transmural potential difference. In contrast to the nonselective muscarinic agonist oxotremorine, xanomeline did not produce salivation, tremor nor hypothermia; it did, however, increase heart rate. The present data are consistent with the interpretation that xanomeline is a novel muscarinic receptor agonist with functional selectivity for M1 muscarinic receptors both in vitro and in vivo.

Animals↗

PubMed1994

Neurochemical effects of the M1 muscarinic agonist xanomeline (LY246708/NNC11-0232).

Xanomeline [3(3-hexyloxy-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1-me thylpyridine)] was evaluated in vivo in rat brain for effects on neurotransmitter turnover and inhibition of ex vivo binding of muscarinic radioligands. Xanomeline produced dose-related increases in the metabolite of dopamine, dihydroxyphenylacetic acid (DOPAC), in striatum. The increases in striatal DOPAC levels produced by xanomeline were antagonized by the relatively selective M1 antagonist trihexyphenidyl, suggesting that xanomeline interacts with M1 heteroreceptors on dopamine nerve terminals. Xanomeline produced small increases in striatal acetylcholine levels and did not antagonize the large increases in acetylcholine produced by the nonselective muscarinic agonist oxotremorine, indicating that xanomeline did not block M2 autoreceptors. Xanomeline inhibited ex vivo binding of muscarinic radioligands to homogenates of brain and the inhibition of ex vivo binding occurred in the same dose range as increases in DOPAC levels. Xanomeline did not appreciably induce salivation or antagonize oxotremorine-induced salivation indicating that xanomeline does not interact with M3 receptors. The effects of xanomeline on ex vivo binding and DOPAC levels lasted for about 3 hr and were evident after oral administration. An analog of xanomeline with similar in vivo effects did not inhibit acetylcholinesterase or choline acetyltransferase and inhibited choline uptake only at concentrations much higher than those required to inhibit binding. These data indicate xanomeline is selective agonist for M1 over M2 and M3 receptors in vivo in rat. It is not known whether xanomeline interacts with m4 or m5 receptors in vivo.

3,4-Dihydroxyphenylacetic Acid↗

PubMed1993

Muscarinic M1 receptor agonist actions of muscarinic receptor agonists in rabbit vas deferens.

In the electrically field-stimulated rabbit vas deferens, muscarinic receptor agonists increase twitch-height by actions at postjunctional M2 receptors and decrease twitch-height by actions at prejunctional M1 receptors. In the present studies, in contrast to previous reports, muscarinic receptor agonists primarily decreased twitch-height, produced minimal increases in twitch-height, and, produced identical responses in both epididymal and prostatic tissue segments, thus permitting a more detailed investigation of the M1 receptor component of action of muscarinic receptor agonists in the rabbit vas deferens. The nonselective muscarinic receptor agonist carbachol produced biphasic effects on twitch-height in the vas deferens: lower concentrations increased twitch-height to only approximately 25-30% over control, whereas higher concentrations inhibited the twitch. The selective M1 receptor antagonist pirenzepine blocked the inhibitory effects of carbachol, and unmasked carbachol-induced increases in twitch-height. Atropine, 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) and AF-DX 116 (11-2[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,11-dihydro- 6H-pyrido[2,3-b][1,4]benzodiazepin-6-one) blocked both the inhibitory and stimulatory effects of carbachol, but atropine and 4-DAMP were more potent in blocking the inhibitory than the stimulatory effects of carbachol, whereas the reverse was true for AF-DX 116. McN-A-343 (4-hydroxy-2-butynyl)trimethylammonium chloride, m-chlorocarbanilate) and 12 other muscarinic receptor agonists from a variety of chemical classes also produced concentration-dependent decreases in twitch-height. The log IC50s of the muscarinic receptor agonists for decreasing twitch-height were highly correlated with their log Kis for inhibiting [3H]pirenzepine (r = 0.96) and [3H]oxotremorine-M (r = 0.85) binding in rat hippocampal membranes. The present results demonstrate that the muscarinic M1 receptor mediating inhibition of twitch-height in the rabbit vas deferens has pharmacologic properties similar to the muscarinic M1 receptor in rat hippocampus.

Animals↗

PubMed1993

Detection of cytokine mRNA in vivo by polymerase chain reaction. Problems and solutions.

The expression of cytokine mRNAs in murine sponge matrix graft infiltrates was studied using the polymerase chain reaction (PCR). In infiltrates of both C57B1/6 > C57B1/6 isografts and DBA/2 > C57B1/6 allografts, a similar pattern of cytokine expression was observed. On days 6, 8, and 10 postimplant, mRNAs encoding IL-4, IL-6, and IFN-gamma were detected. On days 8 and 10, mRNAs encoding IL-1 alpha, TNF, and lymphotoxin/TNF beta were also expressed. Surprisingly, no IL-2 mRNA was observed in isografts or allografts. Two modified PCR techniques were utilized to compare the level of expression of cytokine mRNAs in isografts versus allografts and to detect the expression of IL-2 mRNA in this system. A semiquantitative PCR protocol based on limiting cycles of amplification is described that was used to determine the amount of IL-4 mRNA expressed in isograft and allograft infiltrates relative to beta-actin mRNA. Using this method, no difference was found in the amount of IL-4 mRNA in infiltrates of day 8 isografts and allografts. The validation of this technique by analysis of samples with known relative mRNA levels is presented. A nested PCR protocol is described that provided greatly enhanced sensitivity over standard PCR analysis. Using this technique, IL-2 mRNA was detected in infiltrates from sponge allografts on all days tested, beginning on day 2 postimplant. No IL-2 mRNA was detected in isograft infiltrates or in peripheral blood from allograft-bearing animals. The pattern of cytokine mRNA expression observed in sponge matrix allografts is consistent with the presence of a weak alloreactive response superimposed upon an intense granulomatous process that occurs in both the isografts and allografts. This report demonstrates modified PCR techniques that can be used to resolve experimental problems associated with the analysis of cytokine mRNA expression in vivo.

Animals↗

PubMed1992

Novel functional M1 selective muscarinic agonists. 2. Synthesis and structure-activity relationships of 3-pyrazinyl-1,2,5,6-tetrahydro-1-methylpyridines. Construction of a molecular model for the M1 pharmacophore.

A series of 3-(3-substituted-pyrazinyl)-1,2,5,6-tetrahydro-1-methylpyridines were synthesized and found to have high affinity for central muscarinic receptors. The ability of some of these compounds to inhibit the electrically stimulated twitch of the guinea pig vas deferens indicated that the compounds were M1 agonists. M1 agonist activity was related to the length of the side chain attached to the pyrazine ring, with maximal activity being obtained with the hexyloxy side chain. The (hexyloxy)pyrazine 3f lacked M2 agonist activity as it failed to affect the guinea pig atria and was also relatively devoid of M3 agonist activity as determined by its lack of tremorogenic and sialogogic effects in mice. A comparison of the M1 agonist efficacy of these pyrazines and related 1,2,5-thiadiazoles and 1,2,5-oxadiazoles suggested that M1 efficacy was related to the magnitude of electrostatic potential located over the nitrogens of the respective heterocycles. The heteroatom directly attached to the 3 position of the pyrazine or 1,2,5-thiadiazole heterocycle markedly influenced the M1 efficacy of the compounds by determining the energetically favorably conformers for rotation about the bond connecting the tetrahydropyridyl ring and the heterocycle. A three-dimensional model for the M1-activating pharmacophore was proposed based on computational studies and the model of the muscarinic pharmacophore proposed by Schulman.

Animals↗

PubMed1992

Steric and electronic requirements for muscarinic receptor-stimulated phosphoinositide turnover in the CNS in a series of arecoline bioisosteres.

A series of arecoline derivatives was utilized to assess steric and electronic effects important for activating muscarinic receptors in the CNS. Arecoline derivatives in which the methyl ester moiety was replaced by hexyloxy-1,2,5-oxadiazole (2b), hexyloxythiophene (3b) or hexyloxypyrazine (4b) were compared with the hexyloxy-1,2,5-thiadiazole compound (1b) (Hexyloxy-TZTP), known from previous work to be active as an M1/M3 partial agonist. MNDO calculations showed that the N-S bonds of the alkoxythiadiazole ring were highly polarized with the ability to form H-bonds to the N's. On the other hand, the smaller oxadiazole had lower polarities in the N-O bonds and reduced ability to form H-bonds, the thiophene was of comparable size to the thiadiazole and had large C-S bond polarities without the H-bond capability and the pyrazine had limited ability to form H-bonds. The compounds were compared with respect to their abilities to stimulate phosphoinositide (Pl) turnover in the hippocampus of the rat brain. 1b was more active than 2b-4b for stimulating the Pl turnover response. The data suggest that the ability to form H-bonds is an important factor for the ability of 1 to stimulate M1 muscarinic receptors in the CNS.

Animals↗

PubMed1992

Alloantigenicity of human endothelial cells. 1. Frequency and phenotype of human T helper lymphocytes that can react to allogeneic endothelial cells.

To determine the relative ability of allogeneic endothelial cells to stimulate helper T lymphocytes (HTL), human PBMC or purified T cells were incubated in conventional lymphocyte microcultures or in limiting dilution microcultures with allogeneic human umbilical vein endothelia (HUVE), with cytokine-treated allogeneic HUVE, or with allogeneic peripheral blood monocytes. These cultures were tested for IL-2 production as an index of HTL stimulation. Dose-response studies in conventional lymphocyte cultures indicated that allogeneic monocytes were better than allogeneic HUVE at stimulating IL-2 production. Limiting dilution analyses revealed that untreated HUVE and TNF-treated HUVE stimulated small numbers of HTL (approximately 1 HTL/30,000 PBMC), whereas 5 to 10 times more HTL were stimulated by IFN-gamma-treated HUVE and 10 to 20 times more HTL were stimulated by allogeneic monocytes. Serologic deletion studies revealed that most of the high frequency HTL responding to IFN-gamma-treated HUVE were CD4+, whereas most of the low frequency HTL responding to nontreated HUVE or to TNF-treated HUVE were CD8+. Interestingly, mAb to MHC class I and class II molecules, which significantly impaired HUVE-induced proliferation, caused little interference with HUVE-induced IL-2 production. Finally, polymerase chain reaction analysis demonstrated that untreated allogeneic HUVE cells could stimulate PBMC to produce mRNA for IFN-gamma, as well as for IL-2. These data demonstrate the following hierarchy of allogeneic stimulatory capacity for human HTL: monocytes greater than IFN-gamma-treated HUVE much greater than TNF-treated HUVE = nontreated HUVE. Further, these data suggest that non-activated allogeneic endothelial cells can initiate immune responses by inducing IL-2 and IFN-gamma. Because IFN-gamma can induce MHC class II expression by the endothelial cells, this could recruit large numbers of CD4+ T cells for IL-2 production.

Antigens, Differentiation, T-Lymphocyte↗

PubMed1992

Novel functional M1 selective muscarinic agonists. Synthesis and structure-activity relationships of 3-(1,2,5-thiadiazolyl)-1,2,5,6-tetrahydro-1-methylpyridines .

A series of novel 3-(3-substituted-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro- 1-methylpyridines (substituted-TZTP; 5a-l, 7a-h, 8, 9c-n, 11, 13j) were synthesized and tested for central muscarinic cholinergic receptor affinity by using [3H]-oxotremorine-M (Oxo-M) and [3H]-pirenzepine (Pz) as ligands. The potency and efficacy of the compounds for the pharmacological defined M1 and M2 muscarinic receptors were determined on isolated electrically stimulated rabbit vas deferens and on spontaneously beating isolated guinea pig atria, respectively. Selected compounds were also tested for M3 activity in the isolated guinea pig ileum. The C1-8 alkoxy-TZTP 5a-l analogues all displaced [3H]-Oxo-M and [3H]-Pz with low nanomolar affinity. Depicting chain length against Oxo-M binding and against Pz binding the unbranched C1-8 alkoxy-TZTP (5a-h) derivatives produced U-shaped curves with butoxy- (5d) and (pentyloxy)-TZTP (5e) as the optimum chain length, respectively. This U-shaped curve was also seen in the ability of the compounds 5a-h to inhibit the twitch height in the vas deferens preparation. The (pentyloxy)- (5e) and the (hexyloxy)-TZTP (5f) analogues produced an over 90% inhibition of the twitch height with IC50 values in the low picomolar range. In both the atria and in the ileum preparations 5f had low efficacy and potency. With the (alkylthio)-TZTP (7a-h) analogues the structure-activity relationship was similar to the one observed with the alkoxy (5a-h) analogues, but generally 7a-h had higher receptor affinity and was more potent than the corresponding 5a-h. However, the C3-8 alkyl-TZTP (9c,e,g,h) analogues had 10-100 times lower affinity for the central muscarinic receptors than the corresponding alkoxy and alkylthio derivatives, and their efficacy in the vas deferens preparation was too low to obtain IC50 values. The unsubstituted TZTP (11) compound was a potent but nonselective muscarinic agonist. The two 3-(3-butoxy/(hexyloxy)-1,2,5-oxadiazol-4-yl)-1,2,5,6-tetrahy dro-1- methylpyridines (butoxy/hexyloxy)-OZTP; 19a/b) were also synthesized and tested. Both 19a and 19b had much lower affinity for the central muscarinic receptors than 5d and 5f, and the efficacy of 19a,b was too low to give IC50 values in the vas deferens preparation. Therefore, the C5-6 (alkyloxy)/(alkylthio)-TZTP's represent a unique series of potent functional M1 selective muscarinic agonists.

Animals↗

PubMed1992

Potent in vivo but not in vitro inhibition of monoamine oxidase by 3-chloro-alpha-phenylpyrazinemethanol.

3-Chloro-alpha-phenylpyrazinemethanol (3-CPM) inhibited monoamine oxidase (MAO) types A and B in vivo in mouse brain, heart and liver. The inhibition was dose-dependent at doses of 0.3-32 mg/kg i.p. and occurred within 1 h after the compound was injected. 3-CPM was a very weak inhibitor of mouse brain mitochondrial MAO activity in vitro, even when preincubated with the enzyme; MAO-A was inhibited only about 50% at a high concentration of 3-CPM (1 mM), and MAO-B was inhibited even less. After a 10 mg/kg i.p. dose of 3-CPM in mice, both MAO-A and MAO-B were inhibited at day 1, but activity had largely recovered within a few days in brain, liver and heart. 3-CPM at doses of 1, 3, 10 and 32 mg/kg i.p. caused dose-dependent antagonism of the depletion of striatal dopamine and of cortical norepinephrine by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. 3-CPM is therefore a potent inhibitor of MAO-A and of MAO-B in mice in vivo despite its weak effect on the enzyme in vitro. A metabolite of the drug may be involved in the in vivo effects.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗