Search PubMed⌕ Search

Biomedical subjects

P Chidiac

Publications and source records attributed to P Chidiac.

13 recordsLinked to original sources
PubMed1995

Serotonergic antagonists differentially inhibit spontaneous activity and decrease ligand binding capacity of the rat 5-hydroxytryptamine type 2C receptor in Sf9 cells.

The activities of serotonergic antagonists as inverse agonists at the rat 5-hydroxytryptamine (5-HT)2C serotonin receptor were compared with their potencies in promoting receptor "down-regulation," after expression of the recombinant receptor in the baculovirus/Sf9 insect cell system. Baculovirus expression yielded high levels of 5-HT2C receptors (up to 10(6) receptors/cell), which were functionally coupled to polyphosphoinositide turnover in Sf9 cells through a pertussis toxin-insensitive pathway. The expressed receptor exhibited spontaneous activation of inositol phosphate production, which was inhibited in a dose-dependent manner by serotonergic antagonists, consistent with inverse agonist activity. The potencies of antagonists as inverse agonists correlated with their respective binding affinities determined in competition binding studies with membrane preparations. The maximal inhibition of spontaneous activity ranged from 32% inhibition for mianserin to no effect for spiroxatrine, indicating that antagonists differ in their intrinsic inverse efficacies. Antagonist treatment of intact Sf9 cells or membranes containing the 5-HT2C receptor, followed by washout of residual drug, resulted in a decrease (up to 90%) in the number of binding sites for [3H]mesulergine and [3H]5-HT, with no change in the affinity for [3H]mesulergine. The decrease in binding was irreversible, was not due to the presence of residual antagonist, and was not observed after treatment with agonists. This effect of antagonists in membranes was dose dependent, but the rank order of potency was clearly different from that for inverse agonist activity, indicating that the two effects reflect distinct actions of antagonists at the 5-HT2C receptor. The relative abilities of antagonists to produce loss of binding showed a good correlation with their reported abilities to down-regulate 5-HT2 receptors in vivo after chronic treatment, suggesting that these actions reflect the same underlying process.

Animals↗
PubMed1994

Beta-adrenergic receptor desensitization in the early stage of hereditary cardiomyopathy in hamsters.

The beta-adrenergic receptor (beta AR)/adenylyl cyclase signalling pathway was examined in cardiac membranes from cardiomyopathic Syrian hamsters. Three stages were examined during the progression of this hereditary cardiomyopathy (30 days old, prenecrotic phase; 60 days old, necrotic phase; and 120 days old, compensatory phase). Isoproterenol-stimulated adenylyl cyclase activity was decreased by 32 +/- 16% in 30-day-old cardiomyopathic hamsters, compared with age-matched controls. This was not accompanied by any change in the fluoride- or forskolin-stimulated activities, suggesting that the decrease reflects a perturbation of the receptor-mediated stimulation. Neither the density nor the subcellular distribution of the beta AR, as assessed by [125I]iodocyanopindolol binding assays, was affected in these animals. However, the agonist binding properties of the beta AR were significantly affected. Indeed, the effect of guanyl nucleotides on isoproterenol binding was decreased in 30-day-old cardiomyopathic hamsters. Given that guanyl nucleotide sensitivity is correlated with the ability of the beta AR to productively interact with Gs protein, these results suggest that the decreased beta-adrenergic-stimulated adenylyl cyclase activity results from a functional uncoupling of the beta AR with no change in receptor density. The desensitization of the beta-adrenergic-stimulated adenylyl cyclase was transient, since no change in isoproterenol-stimulated adenylyl cyclase was detected in 60- and 120-day-old hamsters, compared with age-matched controls. Similarly, the receptor number and distribution were not affected at those ages.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclases↗
PubMed1994

Inverse agonist activity of beta-adrenergic antagonists.

Agonist-independent properties of the human beta 2-adrenergic receptor (beta 2AR) were studied using the baculovirus expression system in Sf9 cells. In the absence of agonist but in the presence of GTP, membranes from cells expressing the beta 2AR exhibited higher levels of cAMP production than did membranes from uninfected cells or from cells infected with wild-type baculovirus. The increase in cAMP production was proportional to the number of beta 2AR expressed, up to 40 pmol/mg of membrane protein, and it could be inhibited in a dose-dependent manner by beta AR antagonists. The increase and its reversal both were independent of the possible presence of contaminating catecholamines in the culture medium and thus appear to reflect spontaneous beta 2AR activity and direct antagonist-receptor interactions, respectively. The maximal level of inhibition varied among the beta AR ligands tested, to yield the following rank order of "inverse efficacy"; timolol > or = propranolol > alprenolol > or = pindolol > labetalol > dichloroisoproterenol. The same rank order was observed using membranes prepared from Chinese hamster fibroblasts expressing beta 2AR. The effect of timolol was partly blocked by labetalol and dichloroisoproterenol, in an apparently competitive manner. The intracellular cAMP content of Sf9 cells cultured in serum-free medium was also increased by the expression of beta 2AR, and that increase was reversed by timolol and propranolol, consistent with observations in membrane preparations. The properties revealed by the expression of the beta 2AR in Sf9 cells suggest two agonist-independent traits of G protein-linked receptors, i.e., 1) that unliganded receptors are able to activate G proteins both in membrane preparations and in whole cells and 2) that antagonists may mediate their effects not only by preventing the binding of agonists but also by decreasing the propensity of the receptor to assume an active state.

Adrenergic beta-Agonists↗
PubMed1992

Effects of adenyl nucleotides and carbachol on cooperative interactions among G proteins.

Muscarinic agonists and adenyl nucleotides are noncompetitive modulators of sites labeled by [35S]GTP gamma S in washed cardiac membranes from Syrian golden hamsters. Specific binding of the radioligand and its inhibition by either GTP gamma S or GDP reveals three states of affinity for guanyl nucleotides. In the absence of adenyl nucleotide, carbachol promotes an apparent interconversion of sites from higher to lower affinity for GDP; the effect recalls that of guanyl nucleotides on the binding of agonists to muscarinic receptors. In the presence of 0.1 mM ATP gamma S, the binding of [35S]GTP gamma S is increased at concentrations up to about 50 nM and decreased at higher concentrations. At a radioligand concentration of 160 pM, binding exhibits a bell-shaped dependence on the concentration of both ATP gamma S and AMP-PNP; with ADP and ATP, there is a second increase in bound [35S]GTP gamma S at the highest concentrations of adenyl nucleotide. ATP gamma S and AMP-PNP also modulate the effect of GDP, which itself emerges as a cooperative process: that is, binding of the radioligand in the presence of AMP-PNP exhibits a bell-shaped dependence on the concentration of GDP; moreover, the GDP-dependent increase in bound [35S]GTP gamma S is enhanced by carbachol. The interactions among GDP, GTP gamma S, and carbachol can be rationalized quantitatively in terms of a cooperative model involving two sites tentatively identified as G proteins. Both GTP gamma S and GDP exhibit negative homotropic cooperativity; carbachol enhances the homotropic cooperativity of GDP and induces or enhances positive heterotropic cooperativity between GDP and [35S]GTP gamma S. An analogous mechanism may underlie the guanyl nucleotide-dependent binding of agonists to muscarinic receptors. The data suggest that the binding properties of G proteins and their associated receptors reflect cooperative effects within heterooligomeric arrays; agonist-induced changes in cooperativity may facilitate the exchange of GTP for bound GDP and thereby constitute the mechanism of G protein activation in vivo.

Adenine Nucleotides↗
PubMed1991

Inefficient muscarinic transduction in cardiomyopathic Syrian hamsters.

Regulation of cyclic AMP (cAMP) production and muscarinic binding were studied in highly washed left ventricular membranes from spontaneously cardiomyopathic Syrian hamsters (TO strain). Basal production of cAMP was decreased relative to that in random-bred (RB) controls, with proportionally similar decreases in stimulated production elicited by 7 beta-deacetyl-7 beta-(gamma-N-methylpiperazino)-butyryl forskolin and by the beta-adrenergic agonist isoproterenol. GTP-stimulated production of cAMP was inhibited fully by the muscarinic agonist carbachol in tissue from controls, but only partially in tissue from TO hamsters. Total muscarinic binding, as revealed by N-[3H]methylscopolamine, was similar in the two strains. Competition between carbachol and the radioligand revealed at least three classes of sites for the agonist, the apparent affinities of which were insensitive to the disease. Upon the addition of guanylyl imidodiphosphate (GMP-PNP, 0.1 mM), there was a disease-dependent redistribution such that the sites appeared to be predominantly of low affinity for the agonist in RB tissue and predominantly of medium affinity in TO tissue. The potency of GMP-PNP in mediating the change in carbachol binding apparently was unaffected by the disease. The loss of muscarinic regulation of cAMP production in TO left ventricular tissue appears to reflect a disease-related change in the coupling of muscarinic receptors to inhibitory GTP-binding proteins.

Adenylyl Cyclases↗
PubMed1987

Lack of induction of nuclear aberrations by captan in mouse duodenum.

The fungicide, captan, is known to induce point mutations in vitro. In extremely high doses, technical grade captan leads to duodenal tumors in mice. In short-term in vivo assays for genotoxicity, equivocal results have been obtained with captan. In this study, the genotoxicity of captan was studied in vivo in the proximal small intestine of the mouse, the site of its oncogenicity. Orally administered captan up to 4,000 mg/kg body weight failed to induce a response in the small intestine nuclear aberration assay in a wide range of doses under a variety of experimental conditions, including pretreatment of animals with L-buthionine-S, R-sulfoximine (an inhibitor of glutathione biosynthesis). 1,2,3,6-Tetrahydrophthalimide and bis (trichloromethyl)disulfide, two compounds that have been identified as impurities in technical grade captan, also failed to produce positive results in this assay.

Animals↗
PubMed1986

An in vivo assay for small intestine genotoxicity.

The induction of nuclear aberrations (NA) (apoptotic bodies and micronuclei) in duodenal crypts in a dose-dependent manner was associated with administration of agents known to induce tumours in the small intestine. These included X-irradiation, N-methyl-N-nitrosourea (MNU), benzo[a]pyrene (B[a]P), and 1,2-dimethylhydrazine (DMH), which were found to induce NA in cells in the proliferative region of crypts 24 h after they were given to mice. Methylurea (MU) and benzo[e]pyrene (B[e]P), which are non-carcinogenic structural analogues of MNU and B[a]P, respectively, did not induce NA under similar conditions. Based on these results, the ability of an agent to induce NA in the small intestine appears to reflect of its oncogenic potential in that organ.

Alkylating Agents↗
PubMed1978

Radionuclide quantitation of left-to-right cardiac shunts in children.

In order to evaluate the clinical usefulness of radionuclide methods measuring the magnitude of left-to-right cardiac shunts, a comparison has been made of the results obtained using these procedures with those estimated by cardiac catheterization. Seventy children in whom the presence or absence of a shunt had been established, were studied. Two methods for radionuclide shunt quantitation, the pulmonary to systemic blood flow ratio (Qp/Qs) of Maltz and Treves and their modified C2/C1, were evaluated. The former was found to be superior and showed a high correlation of shunt size with the catheter results. With attention to the performance of the investigation and in the data analysis, good discrimination can be achieved between normal patients and those with shunts. Because of the atraumatic nature of the investigation, it is concluded that the procedure is of value in the diagnosis of a cardiac murmur, the assessment of postoperative patients and the follow-up of patients known to have a small shunt.

Adolescent↗