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Biomedical subjects

P De Neef

Publications and source records attributed to P De Neef.

At least 91 records · Page 5Linked to original sources
PubMed1984

Effects of verapamil on the binding properties of rat heart muscarinic receptors: evidence for an allosteric site.

The calcium channel antagonist verapamil is known to inhibit competitively antagonist binding to rat heart muscarinic receptors. The present data suggest that this drug recognized two binding sites on the muscarinic receptors: 1) an allosteric site modulating the tracer dissociation rates and 2) the muscarinic drug binding site. The affinity of verapamil for the allosteric site and the efficacy of its effect on muscarinic ligand dissociation rates depended on the ligand studied.

Allosteric Site↗
PubMed1984

Multiple effects of short-chain alcohols on binding to rat heart muscarinic receptors.

Short-chain alcohols inhibited the equilibrium binding of agonists and antagonists to rat heart muscarinic receptors. Methanol, ethanol, propan-2-ol and propan-1-ol, when used at low concentrations, behaved as pseudo-competitive antagonists. Their rank order of potency paralleled their relative partition coefficients, suggesting that this inhibition was simply due to the interaction of the alcohols with a hydrophobic part of the receptor or with membrane lipids. The four alcohols increased the dissociation rate constant of [3H]oxotremorine M from the high-affinity agonist receptors and decreased the stability of this receptor state. These effects might reflect increased membrane fluidity and/or decreased hydrophobic interactions (see below). By contrast, the effects of alcohols on the association and dissociation rates of N-[3H]- methscopolamine (an antagonist) were not correlated to their relative octanol/water partition coefficient (a measure of their affinity for biophases ). Alcohols, at the relatively high concentrations necessary for increased membrane 'fluidity', are known to affect the relative stability of various protein conformations. We believe that the effects of alcohols on antagonist binding to rat heart muscarinic receptors reflected changes in the activation energy of association and dissociation reactions, the inhibition of equilibrium binding being mainly due to decreased 'hydrophobic interactions'.

1-Propanol↗
PubMed1984

A comparison between muscarinic receptor occupancy, adenylate cyclase inhibition, and inotropic response in human heart.

Binding to muscarinic receptors was compared with adenylate cyclase inhibition in membranes derived from human heart auricles, and with inhibition of the contraction of auricular muscle fibers. In the absence of GTP, agonists recognized two classes of receptors both of which bound antagonists with the same affinity. In the presence of GTP, both classes of receptors for agonists were converted into a single low affinity state. Carbachol and oxotremorine inhibited adenylate cyclase activity by 43%, pilocarpine being less efficient (-28%). The 3 agonists exerted similar inhibitory effects on the inotropic response, in 7 out of 9 preparations of electrically- and norepinephrine-stimulated fibers. Dose-effect curves suggested that spareness (or an amplification mechanism) was implicated in the occupancy of low affinity binding sites by carbachol and oxotremorine (but not by the partial agonist pilocarpine) and the resulting inhibition of both adenylate cyclase activity and contractile force.

Adenylyl Cyclase Inhibitors↗
PubMed1984

Secretin-induced changes in rate, contractility and adenylate cyclase activity in rat heart atria.

Secretin stimulated adenylate cyclase activity in crude membrane preparations from right and left rat atria, when tested in the presence of the potentiating activator forskolin. Its maximal effect on adenylate cyclase activity was greater in the right atrium than in the left atrium but the peptide efficiency was lower, in both atria, than that of D,L-isoproterenol. Secretin stimulated the rate of contraction of the spontaneously beating right atrium, but less efficiently than D,L-isoproterenol. Although this positive chronotropic action of secretin was inhibited by D,L-propranolol, it was probably not mediated by the release of endogenous catecholamines as: a) the inhibitory effects of D- and L-propranolol were similar and b) the efficiency of secretin on the in vitro beating of right atrium was the same in control and reserpinized rats. Secretin stimulated the force of contraction of spontaneously beating right atrium, even in the presence of propranolol, i.e. when the chronotropic effect of secretin was abolished. The hormone exerted also a positive inotropic effect on the electrically stimulated rat atrium. This effect was blocked neither by tetrodotoxin nor by propranolol. When comparing the dose-effect curves of secretin and D,L-isoproterenol on adenylate cyclase activation on the one hand, and on the stimulation of rate and contractility on the other hand, it is tempting to suggest that cyclic AMP might be involved in the modulation by secretin of the mechanical properties of rat atria.

Adenylyl Cyclases↗
PubMed1984

The in vitro chronotropic and inotropic effects of vasoactive intestinal peptide (VIP) on the atria and ventricular papillary muscle from Cynomolgus monkey heart.

The in vitro chronotropic and inotropic effects of vasoactive intestinal peptide (VIP) and of isoproterenol, two agents known to stimulate cardiac adenylate cyclase were compared on the heart from Cynomolgus monkey using the spontaneously beating right atrium, the electrically stimulated left atrium, and the electrically-stimulated ventricular papillary muscle. VIP increased concentration-dependently the rate of beating of the right atrium as well as the contractility of both atria but its efficiency was lower than that of D,L-isoproterenol. VIP also stimulated concentration-dependently, and this time as efficiently as D,L-isoproterenol, the contractility of papillary muscle. These VIP effects were unaltered by the neuronal blocker tetrodotoxin. In addition, the moderate inhibition exerted by the beta-adrenergic blocker D,L-propranolol on VIP effects argued against the implication of beta-adrenergic receptors in VIP effects. These results indicate that VIP exerts a direct stimulatory influence on the rate and contractility of Cynomolgus monkey heart.

Animals↗
PubMed1983

The adenylate cyclase activity in heart membranes from normotensive and spontaneously hypertensive rats, after chemical sympathectomy, suggests the presence of presynaptic secretin receptors.

Normotensive (WKY) and spontaneously hypertensive (SHR) male adult rats were sacrificed 2 and 3 weeks after 6-hydroxydopamine treatment. Untreated WKY and SHR rats served as controls. In rat heart membranes from WKY rats, 6-hydroxydopamine treatment increased guanosine 5'-O-(2-3-imido)-triphosphate (Gpp(NH)p)-, NaF-, D,L-isoproterenol- and glucagon-stimulated adenylate cyclase activities by 18-38% while secretin stimulation was unaffected. In heart membranes from SHR rats, Gpp(NH)p, NaF, D,L-isoproterenol, or glucagon stimulation of the enzyme was similarly increased by 14-38% whilst the low secretin responsiveness which is characteristic of these animals decreased even further (by 24-47%). These results are consistent with: (1) an up regulation of postsynaptic beta-adrenergic receptors coupled to adenylate cyclase after degeneration of adrenergic nerves, and (2) a differential response of secretin receptors coupled to adenylate cyclase in the two strains of rats: there was no change in WKY rats and a decreased response in SHR rats. The possible presence and contribution of presynaptic secretin cardiac receptors is considered.

Adenylyl Cyclases↗
PubMed1983

Increased number of high-affinity muscarinic receptors in rat heart after phenylhydrazine treatment.

The effect of phenylhydrazine-induced heart hypertrophy on muscarinic cholinergic binding sites was studied in heart membranes by determining total (with [3H]NMS) and high-affinity (with [3H]Oxo-M) binding sites and estimating low-affinity binding sites by difference. Phenylhydrazine induced an increased concentration of high-affinity binding sites without affecting the concentration of low-affinity binding sites. The KD values for the ligands were unaffected.

Animals↗
PubMed1983

Topographical distribution of the secretin- and VIP-stimulated adenylate cyclase system in the heart of five animal species.

Adenylate cyclase stimulation by secretin and VIP was compared to the effect of glucagon, D,L-isoproterenol, Gpp[[NH]p, and NaF in atria and ventricles from rat, guinea pig, rabbit, dog and Cynomolgus monkey. In rat ventricular membranes, secretin was a better stimulant than VIP and was as active as D,L-isoproterenol. In rat auricular membranes both peptides were inactive. In guinea pig and rabbit heart membranes (ventricular and auricular) VIP and secretin were inactive. In dog and monkey atria, VIP stimulation of adenylate cyclase was comparable to that of D,L-isoproterenol, secretin being inactive. In dog ventricles, VIP was less efficient than D,L-isoproterenol, secretin being inactive. In monkey ventricles, by contrast, VIP was slightly more efficient than D,L-isoproterenol, secretin having a small effect only in left ventricles. The present results established a clear difference between animal species with respect to the efficacy of the peptides of the secretin/VIP family: the presence of "secretin-preferring" receptors in rat heart contrasted with the presence of "VIP-preferring" receptors in dog and monkey heart. Our results in dog and monkey hearts suggest that VIP might be a candidate for a physiological control of heart function.

Adenylyl Cyclases↗
PubMed1983

The cardiac inotropic response to secretin is lower in genetically obese (fa/fa) than in lean (fa/?) Zucker rats.

The efficacy and potency of secretin in rat heart was documented by comparing secretin and D,L-isoproterenol effects on: 1. the inotropic response of papillary muscles electrically stimulated in vitro, and 2. adenylate cyclase activity in a crude membrane preparation from heart ventricle. In male adult lean (FA/?) Zucker rats, the extent of the positive inotropic response and the stimulatory effect on adenylate cyclase were similar in magnitude for secretin and D,L-isoproterenol, the two corresponding EC50 values for inotropic effects being 5- to 10-fold lower than the EC50 values for adenylate cyclase stimulation. In male adult obese (fa/fa) Zucker rats, the stimulatory effects exerted by secretin on the inotropic response and adenylate cyclase activity were reduced by 73% and 70%, respectively, while the extent of similar D,L-isoproterenol responses were reduced, respectively, by 29% and 25% only. With D,L-isoproterenol, the EC50 value for the inotropic effect was 5-fold lower than the EC50 value for adenylate cyclase activation whereas both EC50 values were comparable when secretin was used as a stimulant.

Adenylyl Cyclases↗
PubMed1983

Thermodependence of basal and stimulated cardiac adenylate cyclase activity in normotensive and spontaneously hypertensive rats.

The cardiac adenylate cyclase activity from normotensive and spontaneously hypertensive (SHR) rats was studied as a function of temperature between 17 degrees and 37 degrees C. Arrhenius plots of adenylate cyclase activity displayed a break around 31 degrees C when tested under basal conditions or in the presence of GTP but were linearized after activation with p[NH]ppG, NaF, secretin, glucagon or isoproterenol. The energy of activation of adenylate cyclase activity in the presence of GTP (9.5 +/- 0.5 kcal/mol) was significantly lower than in the presence of p[NH]ppG (17.7 +/- 0.8 kcal/mol). A hormone was without effect on the energy of activation observed with either GTP or p[NH]ppG but the simultaneous presence of hormone and nucleotide increased markedly the activity of the enzyme. The energies of activation were analyzed in terms of variation of enthalpy and entropy and discussed in relation with the process of activation and coupling of the guanine nucleotide regulatory protein. These thermodynamic characteristics were similar in cardiac membranes from normotensive and spontaneously hypertensive rats, suggesting that the impairment of hormone-stimulated adenylate cyclase activity observed in the heart membranes of hypertensive rats was not a consequence of a defect in the activation process of the enzyme.

Adenylyl Cyclases↗
PubMed1983

The human heart beta-adrenergic receptors. I. Heterogeneity of the binding sites: presence of 50% beta 1- and 50% beta 2-adrenergic receptors.

Beta-adrenergic receptors were characterized in a particulate fraction of human auricles obtained from patients operated upon for coronary insufficiency or valvular disease. [125I] Hydroxybenzylpindolol binding was evaluated in terms of kinetics; KD and Bmax values; and inhibition of binding in the presence of 10 microM GTP and of increasing concentrations of four nonselective agonists giving a Hill coefficient of 1 (isoproterenol, salbutamol, fenoterol, and epinephrine), of two nonselective antagonists giving a Hill coefficient of 1 (pindolol and propranolol), and of a series of selective drugs giving a Hill coefficient of 0.60-0.72 that included three beta 1-selective antagonists (practolol, metoprolol, and atenolol) and two beta 2- selective agonists (procaterol and zinterol). KD values for all drugs were compatible with the coexistence in membranes from human auricles of beta 1- and beta 2-adrenergic receptors, the relative proportions of receptors of each subclass being approximately the same.

Adrenergic beta-Agonists↗
PubMed1982

Early decrease in secretin-, glucagon-, and isoproterenol-stimulated cardiac adenylate cyclase activity in rats treated with isoproterenol.

Male Wistar albino rats received three times daily for one to five days 0.25 to 5.0 mg/kg D,L-isoproterenol intraperitoneally. D.L-Isoproterenol injections provoked a time dependent- and dose-related cardiac hypertrophy. With moderate hypertrophy, a selective decrease in secretin-stimulated adenylate cyclase activity occurred. When heart hypertrophy was more pronounced, greater losses in secretin-, as well as in D,L-isoproterenol-, glucagon-, guanine nucleotide-, and fluoride-stimulated enzyme activity developed. Hormone stimulations of adenylate cyclase were more severely, curtailed (60 to 65%) than guanine nucleotide or fluoride stimulations (40 to 45%). The accompanying loss in beta-receptors (35%) was proportionately lower than the loss of D,L-isoproterenol sensitivity of adenylate cyclase. This complex pattern of adenylate cyclase desensitization in heart membranes from animals chronically treated with D,L-isoproterenol is reminiscent of that observed in heart membranes from spontaneously hypertensive rats.

Adenylyl Cyclases↗
PubMed1982

Effects of PHI on vasoactive intestinal peptide receptors and adenylate cyclase activity in lung membranes. A comparison in man, rat, mouse and guinea pig.

The presence of receptors, recognized by vasoactive intestinal peptide (VIP) as well as by PHI (a peptide with N-terminal histidine and C-terminal isoleucine amide), was documented in lung membranes from rat, mouse, guinea pig and man by the ability of these receptors, once occupied, to stimulate adenylate cyclase. In lung membranes from rat, mouse and guinea pig, the capacity of VIP, PHI and secretin to stimulate the enzyme and the potency of the same peptides to compete with 125I-VIP for binding to VIP receptors were similar, the affinity decreasing in the order: VIP greater than PHI greater than secretin. In addition, dose-effect curves were compatible with the coexistence of high-affinity and low-affinity VIP receptors, in the four animal species considered. If PHI was able to recognize all VIP receptors it could not, however, discriminate the subclasses of VIP receptors.

Adenylyl Cyclases↗
PubMed1982

Characterization of secretin and vasoactive intestinal peptide receptors in rat pancreatic plasma membranes using the native peptides, secretin-(7-27) and five secretin analogues.

A comparison has been made of the ability of vasoactive intestinal peptide (VIP), secretin, secretin analogues, and secretin-(7-27) to stimulate adenylate cyclase in rat pancreatic plasma membranes. A parallel study of the capacity of peptides of the VIP-secretin family to compete with 125I-VIP for binding to the same plasma membranes was conducted. This allowed a classification of VIP-secretin receptors into three subtypes: (1) VIP-preferring receptors; (2) high-affinity secretin receptors, and (3) low-affinity secretin receptors. The properties of secretin at high-affinity secretin receptors were likely to reflect a contribution of membranes from centroacinar and duct cells.

Adenylyl Cyclases↗
PubMed1981

Presence of vasoactive intestinal peptide receptors coupled to adenylate cyclase in rat lung membranes.

(1) The binding of 125I-labelled vasoactive intestinal peptide (VIP) to a particulate fraction from rat lung was rapid, temperature dependent, saturable and specific. This process was also reversible and 125I-labelled VIP dissociation was accelerated by guanine triphosphate nucleotides. The curves describing the inhibition of tracer binding by peptides of the VIP-secretin family suggested the presence of at least two classes of VIP receptor: a "high-affinity' type with decreasing affinity for VIP in the order: VIP = [Val5]secretin greater than [Ala4, Val5]secretin; and a "low-affinity type' with decreasing affinity for VIP in the order: VIP greater than [Val5]secretin greater than [Ala4, Val5]secretin = secretin greater than [Ala4]secretin. (2) VIP and related peptides stimulated the adenylate cyclase activity of the same lung membrane preparation more efficiently than beta-adrenergic agonists and prostaglandins E1 and E2. The dose-effect curves of stimulation of adenylate cyclase by VIP and parent peptides were also compatible with the existence of two classes of VIP receptor, the relative peptide potencies being identical with their ability to compete with 125I-labelled VIP for binding.

Adenylyl Cyclases↗
PubMed1981

Binding of vasoactive intestinal peptide and its stimulation of adenylate cyclase through two classes of receptors in rat liver membranes. Effects of 12 secretin analogues and 2 secretin fragments.

1. Vasoactive intestinal peptide (VIP) receptors were identified in crude rat hepatic membranes by 125I-labelled VIP binding and by the ability of VIP to stimulate adenylate cyclase activity. The specificity of these receptors was evaluated by the capacity of secretin, synthetic secretin analogues, and secretin fragments to inhibit 125I-labelled VIP binding and to stimulate adenylate cyclase. 2. The results were compatible with the existence of two classes of VIP binding sites that could be distinguished according to their affinity for VIP and their specificity. High-affinity sites were more specific for VIP as secretin was 175 times less potent than VIP for recognition of these sites while being only 33 times less potent than VIP for recognition of low-affinity sites. 3. Secretin analogues, monosubstituted in position 2, 3, 4 or 6 were less potent than secretin for adenylate cyclase stimulation as well as for the recognition of the two classes of receptors. [Val5]secretin was more potent than secretin and appeared definitely more VIP-like than secretin; [Ala4, Val5] and [D-Ala4,Val5]secretin were equipotent to secretin. 4. The fragment secretin (7-27) was unable to recognize VIP receptors and to stimulate adenylate cyclase. The substituted fragment [Gln9,Asn15]secretin (5-27) recognized these receptors with weak potency but could not activate the enzyme.

Adenylyl Cyclases↗
PubMed1981

Comparative effects of alpha-methyldopa, propranolol and hydralazine therapy on cardiac adenylate cyclase activity in normal and spontaneously hypertensive rats.

Normotensive (WKY) and spontaneously hypertensive (SHR) male rats were treated orally, one week after weaning and for 9 weeks, with alpha-methyldopa (100 mg/kg per day), propranolol (30 mg/kg per day) or hydralazine (10 mg/kg per day). Untreated WKY and SHR rats served as controls. The development of hypertension in SHR rats were attenuated by treatment but none of the drugs was able to restore the impairment in isoproterenol, secretin and glucagon responsiveness of cardiac adenylate cyclase activity which is characteristic of these animals. In heart membranes from both WKY and SHR rats, alpha-methyldopa treatment increased the number of beta-adrenoceptors by 20-32% and the maximal response of adenylate cyclase activity to isoproterenol and glucagon by 20-34%. By contrast, the beta-blocker propranolol was ineffective on these parameters. The results obtained are consistent with the hypothesis that the change in adenylate cyclase seen in SHR rats is genetic in origin and is not a consequence of hypertension.

Adenylyl Cyclases↗