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Biomedical subjects

E Gerlach

Publications and source records attributed to E Gerlach.

At least 55 records · Page 3Linked to original sources
PubMed1989

Inhibition of platelet aggregation following chronic in vivo treatment of rats with nicotine: prevention by simultaneous application of propranolol.

Platelet aggregability is known to be enhanced and platelet-survival time shortened in smokers when compared with nonsmokers. Up to now it is unknown which of the substances in tobacco smoke are responsible for these effects. To evaluate a possible role of nicotine, rats were chronically treated with the alkaloid (10 mg/kg/day), continuously released from subcutaneously implanted osmotic minipumps. Surprisingly, after 8 weeks, platelet sensitivity toward the aggregating stimulus adenosine 5'-diphosphate (ADP) was markedly reduced. The mean ADP concentration required to induce half the maximum rate of aggregation (EC50) was 0.88 mumol/L in nicotine-treated animals, as compared with 0.67 mumol/L in controls (p less than 0.002). Platelet aggregability remained normal when the rats were treated simultaneously with nicotine and the beta blocker propranolol (3.5 mg/kg/day); for these animals, the mean EC50 for ADP was 0.73 mumol/L. These results are suggestive of a catecholamine-mediated action of nicotine. However, neither the basal levels of cAMP in platelet-rich plasma, nor the cAMP levels attained after stimulation of platelet adenylate cyclase with prostaglandin E1 (PGE1), were affected by 8 weeks of treatment with nicotine or nicotine plus propranolol. No effect on platelet aggregation was observed when the rats were treated with nicotine for only 2 weeks, or when nicotine or nicotine plus cotinine were added to platelet-rich plasma in vitro in concentrations equal to those attained in vivo after 8 weeks. Thus, prolonged application of nicotine in vivo caused an inhibition of ADP-induced rat platelet aggregation presumably mediated by beta-catecholaminergic stimulation of platelets.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1989

Purine metabolism in cultured aortic and coronary endothelial cells.

Purine salvage pathways in cultured endothelial cells of macrovascular (pig aorta) and microvascular (guinea pig coronary system) origin were investigated by measuring the incorporation of radioactive purine bases (adenine or hypoxanthine) or nucleosides (adenosine or inosine) into purine nucleotides. These precursors were used at initial extracellular concentrations of 0.1, 5, and 500 microM. In both types of endothelial cells, purine nucleotide synthesis occurred with all four substrates. Aortic endothelial cells salvaged adenine best among purines and nucleosides when applied at 0.1 microM. At 5 and 500 microM, adenosine was the best precursor. In contrast, microvascular endothelial cells from the coronary system used adenosine most efficiently at all concentrations studied. The synthetic capacity of salvage pathways was greater than that of the de novo pathway. As measured using radioactive formate or glycine, de novo synthesis of purine nucleotides was barely detectable in aortic endothelial cells, whereas it readily occurred in coronary endothelial cells. Purine de novo synthesis in coronary endothelial cells was inhibited by physiological concentrations of purine bases and nucleosides, and by ribose or isoproterenol. The isoproterenol-induced inhibition was prevented by the beta-adrenergic receptor antagonist propranolol. The end product of purine catabolism in aortic endothelial cells was found to be hypoxanthine, whereas coronary endothelial cells degraded hypoxanthine further to xanthine and uric acid, a reaction catalyzed by the enzyme xanthine dehydrogenase.

Animals↗
PubMed1988

Endothelium-mediated coronary dilatation by adenosine does not depend on endothelial adenylate cyclase activation: studies in isolated guinea pig hearts.

Adenosine, applied to the coronary system of guinea pigs at up to 10(-6) M, elicits dilatation solely via an endothelium-mediated process. We investigated the role of coronary A2 receptors in this dilation, since the coronary endothelium possesses adenosine A2-receptors with a stimulatory action on the adenylate cyclase. In situ, A2 receptor stimulation can be assessed by prelabeling the coronary endothelial adenine nucleotide pool with 3H-adenosine and subsequently determining the rate of release of radiolabeled cAMP induced by A2 agonists. Thus, perfusion of isolated hearts with 5'-N-ethylcarboxamidoadenosine (NECA) dose-dependently increased coronary flow and the release of 3H-cAMP from the endothelium. In the presence of 50 microM 2',5'-dideoxyadenosine (ddA), a P-site agonist which inhibits the catalytic activity of adenylate cyclase, coronary flow increases induced by both adenosine and NECA were unaffected. In contrast, ddA reduced the release of labeled cAMP in response to NECA by about 60%. In cultured endothelial cells, ddA likewise inhibited cAMP accumulation due to NECA by about 70%. Moreover, ddA antagonized the adenylate cyclase mediated flow response due to the PGI2 analogue, iloprost, as well as the positive chronotropic and inotropic actions of isoproterenol. The dissociation elicited by ddA between the coronary flow response and the release of cAMP strongly indicates that the endothelial A2 receptors which are linked to adenylate cyclase are not causally involved in endothelium-dependent coronary dilatation induced by adenosine.

Adenosine↗
PubMed1988

Blood platelet function after chronic treatment of rats and guinea pigs with nicotine.

To establish whether long-term application of nicotine can at all alter thrombocyte function, the alkaloid was administered to rats and guinea pigs for up to 8 weeks, using subcutaneously implanted ALZET minipumps. These two species were examined, since rat platelets are known to possess both alpha 2- and beta 2-adrenoceptors, whereas guinea pig platelets have practically none of either type. Platelet activity was assessed ex vivo by determining rates of ADP-induced aggregation and by applying a new in vitro technique giving a measure of primary hemostasis. Nicotine in doses of 2 mg/kg/day (rats) and 10 mg/kg/day (guinea pigs), continuously applied over 8 weeks, yielded alkaloid plasma levels in the respective species in the same range as found for smokers. This chronic in vivo pretreatment had no discernible effect on platelet function. However, platelets from rats having received the higher dose of 10 mg nicotine/kg/day for 8 weeks required a significantly higher concentration of ADP to induce the half-maximal rate of aggregation. Concomitantly, the in vitro determined bleeding time doubled. The nicotine-induced decrease in platelet sensitivity towards ADP could be prevented by simultaneously treating the rats with the beta-blocker propranolol. Adrenaline applied chronically to rats for 8 weeks stimulated platelet response instead of mimicking nicotine effects. The unexpected inhibition of rat platelet function at higher alkaloid-dosage thus seems to be an indirect nicotine action, mediated via platelet beta-adrenoceptors. Since chronic treatment with adrenaline did not exert such an effect, the action of nicotine may, rather, be attributed to increases in endogenous noradrenaline. These results could explain varying responses of human blood platelets in smokers.

Animals↗
PubMed1986

Increasing platelet aggregability after venepuncture is platelet, not plasma derived.

The time course of ADP induced aggregation of human platelets was determined in aliquots of stored platelet rich plasma 3.5, 10, 30 and 100 minutes after venepuncture. The maximal rate of aggregation was found to increase throughout this entire period, even though pH (7.4), CO2 (7 volume per cent) and temperature (35 degrees C) of the samples were kept constant. The mean acceleration (+/- SEM) between 3.5 and 100 minutes was 41.7 +/- 6.9 per cent (n = 67) at an ADP-concentration of 1 mumol/l and 18.3 +/- 6.2 per cent (n = 23) at 2 mumol/l ADP. The effect did not result from changes of any platelet regulatory factors putatively present alone in the plasma. Acceleration of aggregability was only found when the platelets themselves underwent storage, but not when freshly prepared plasma was given to prestored platelets. The change in aggregability was not diminished after inhibition of platelet cyclooxygenase by oral administration of acetylsalicylic acid.

Adenosine Diphosphate↗
PubMed1985

[Recent knowledge about the metabolic regulation of coronary circulation, with a contribution on adenine nucleotide metabolism of coronary endothelial cells].

The adenosine hypothesis in its original form neglects other myocardial tissues besides the predominant cardiomyocyte compartment. We could, however, demonstrate that the coronary endothelium comprises a metabolically very active adenosine and adenine nucleotide compartment of the heart, and functions as an impermeable metabolic barrier for interstitially or intravascularly accumulating adenosine if the vasoactive nucleoside is present at concentrations less than 10(-6) M. As a consequence, the vasodilatory action of intracoronarily applied adenosine cannot result from a direct action on the smooth muscle cells of the arterioles, but must be mediated by the endothelium. Since high molecular weight derivatives of adenosine, which are clearly confined to the coronary system, can also induce a very prompt coronary flow increase when applied intravascularly, smooth muscle relaxation must be triggered by an extracellular adenosine receptor at the luminal surface of the endothelium. According to preliminary pharmacological studies, this receptor belongs to the A2-type and thus stimulates the endothelial adenylate cyclase system. On the basis of our findings it is evident that with respect to the vasodilating effect of adenosine one has to distinguish between its action from the interstitial space directly via the putative receptor at the surface of the arteriolar smooth muscle cells, and its action from the intravascular space via the newly detected endothelial A2-receptor. It is a matter of further experimentation to determine to what extent both receptor populations actually participate in the metabolic regulation of coronary flow under physiological and pathophysiological conditions.

Adenine Nucleotides↗
PubMed1984

Acute effects of nicotine on hemodynamic and metabolic parameters of isolated, perfused hearts of guinea pigs and rats.

Nicotine infused in concentrations greater than 10(-6) M exerted a dose-dependent negative inotropic effect on isolated, perfused hearts of guinea pigs (Langendorff and working heart preparations). This effect became manifest after an initial positive inotropic and positive chronotropic response had subsided. Furthermore, the stimulation of cardiac contractility by norepinephrine (concentration less than 10(-7) M) was attenuated by nicotine in the guinea pig heart, but not in isolated, perfused hearts of Sprague-Dawley rats. Nicotine inhibited the positive inotropic effect of catecholamines according to the order norepinephrine greater than epinephrine greater than isoproterenol. In no case did nicotine have a marked negative chronotropic action. Changes in coronary flow, oxygen consumption, and the release of lactate and adenosine were in keeping with the state of myocardial activity in the presence of nicotine. The drug did not significantly alter myocardial uptake or release of norepinephrine during expression of its negative inotropic action. Prazosin, phentolamine, cocaine, and hexamethonium suppressed both the nicotine-induced transient stimulatory response and the negative inotropic effect in norepinephrine-stimulated hearts. In the presence of theophylline and at high perfusate calcium levels (7.5 mEqu/l) the negative inotropic effect was alleviated. The cardiodepressive action of nicotine may possibly evolve from a weak blockade of beta 1-adrenoceptors. Direct modifications of myocardial Ca2+-transport could not be observed.

Animals↗
PubMed1984

Compartmentation and hemodynamic effects of nicotine in isolated, perfused guinea pig hearts.

Tachyphylaxis with respect to the positive inotropic and positive chronotropic effects of nicotine in isolated perfused hearts of guinea pigs was found to persist for 3 and 10 min, respectively, after discontinuation of intracoronary infusion of nicotine (5 X 10(-5) M). The longer duration of chronotropic tachyphylaxis correlated with a slower release of nicotine from the tissue of the right atrium in comparison to the washout from the ventricular myocardium. A similar disparity in washout between atrial and ventricular tissue was found for inulin. Thus, greater perfusion of the ventricular tissue, not a difference in affinity for nicotine, seems to account for the more rapid recovery of the inotropic response to nicotine. The apparent volume of distribution of nicotine in both atrial and ventricular myocardium was about 2 ml/g wet weight; uptake and release of nicotine was rapid. Kinetic analyses of coronary venous washout yielded four components, indicating that nicotine is distributed in at least four (kinetically defined) compartments within the ventricular myocardium. Washout from the largest tissue compartment was controlled by the rate of perfusion of the vascular space, as demonstrated by comparison with washout of erythrocytes, inulin, sucrose, and urea from the heart. Hexamethonium reduced the nicotine content of this fast-emptying compartment from 85-57 nmol/g (prelabeling concentration of nicotine 5 X 10(-5) M), giving an upper estimate for the number of specific nicotine binding sites.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1981

Different sites of adenosine formation in the heart.

In an attempt to further define the site of myocardial adenosine formation, isolated guinea pig hearts were perfused with potent inhibitors of 5'-nucleotidase [alpha, beta-methylene adenosine 5'-diphosphate (AOPCP)] and of nucleoside transport [4-nitrobenzyl thioinosine (NBMPR)]. AOPCP (50 microM) inhibited the activity of cardiac ecto-5'-nucleotidase by 85% but did not influence the release of adenosine, inosine, and hypoxanthine formed at an accelerated rate by the heart during hypoxic perfusion (30% O2). In contrast, NBMPR (5 microM) diminished the hypoxia-induced release of adenosine and its degradatives and greatly potentiated the increase of myocardial tissue levels of respective purine compounds. Studies carried out with 5'-deoxyadenosine, an adenosine derivative that is not metabolized, indicate NBMPR to inhibit both uptake and release of adenosine in the isolated heart and in human erythrocytes. Cell fractionation studies on guinea pig ventricular muscle revealed that 5'-nucleotidase, though mainly associated with the membrane fraction, is also found in the cardiac cytosol (200,000-g supernatant), exhibiting a different substrate specificity. Furthermore, S-adenosylhomocysteine hydrolase as well as adenosine kinase and adenosine deaminase proved to be exclusively present in the cytosolic fraction. Our findings suggest that in the hypoxic heart a) ecto-5'-nucleotidase most likely is not involved in the formation of adenosine, b) release of adenosine from the heart requires adenosine to be transported across the sarcolemma membrane, and c) adenosine is predominantly formed intracellularly, a process involving cytosolic 5'-nucleotidase and/or S-adenosylhomocysteine hydrolase.

5'-Nucleotidase↗
PubMed1981

Inhibitory action of adenosine on histamine- and dopamine-stimulated cardiac contractility and adenylate cyclase in guinea pigs.

Infusion of adenosine into the coronary arteries of isolated guinea pig hearts produced a dose-dependent inhibition of cardiac contractile force development elicited by bolus injections of histamine (7.5 X 10(-9) mol) or dopamine (1.5 X 10(-8) mol). Threshold concentration of adenosine was 10(-7) M and maximal inhibition (90%) was obtained at 3 X 10(-5) M. Adenosine in the effective concentration range did not alter Ca2+-induced increases in contractile force. The rise in tissue levels of cAMP induced by equieffective doses of histamine (7.5 X 10(-9) mol) and dopamine (1.5 X 10(-8) mol) was inhibited by adenosine (3.5 X 10(-5) M) by about 60%. In a particular membrane preparation of guinea pig ventricles the adenylate cyclase activity stimulated by the histamine (10(-5) M) and dopamine (10(-4) M) was inhibited in a dose-dependent manner by adenosine. This effect could be reversed by theophylline (5 X 10(-5) M) in a competitive manner. The hormone-insensitive adenylate cyclase of a Lubrol-PX solubilized membrane preparation stimulated by NaF or 5'-guanylylimido-diphosphate (GppNHp) was also inhibited by adenosine (40% and 90% inhibition at 10(-5) M and 10(-4) M, respectively). Adenosine did not influence the Km value of the adenylate cyclase for ATP, but markedly lowered Vmax of the enzyme. From additional studies with purine-substituted (N6-methyl-adenosine, N6-phenylisopropyl-adenosine) and ribose-substituted (2'-deoxy-adenosine and arabino-furanosyl-adenine) adenosine analogues, we conclude that adenosine may inhibit the inotropic responses to hormones as well as the adenylate cyclase activity by specifically interacting with at least two different sites associated with the adenylate cyclase.

Adenosine↗
PubMed1981

Isolation, identification, and continuous culture of coronary endothelial cells from guinea pig hearts.

Viable and homogeneous endothelial cells were obtained from isolated guinea pig hearts by application of a special perfusion technique of the coronary system with an isotonic collagenase-trypsin solution and subsequent purification of the dissociated cells by Percoll density gradient centrifugation. The coronary endothelial cells were grown in tissue culture for periods up to 7 months. Serial passage proved to be possible. During logarithmic growth, generation time was found to be 18 h; it could be reduced to 16 h by addition of thrombin to the culture medium. Light, phase contrast and scanning electron microscopy as well as autoradiography revealed that cultured coronary endothelial cells grew as strict monolayers of closely apposed, polygonal large cells. By scanning electron microscopy, it could be demonstrated that the morphology of the cultured cells changes characteristically during attachment of the cells to their substratum. The changes observed were very similar to those of proliferating endothelial cells of isolated coronary vessels kept in organ culture. According to transmission electron microscopy studies, cultured coronary endothelial cells proved to contain only an extremely small number of Weibel-Palade bodies. Nucleoside phosphorylase (EC 2.4.2.5.) and 5'-nucleotidase (EC 3.1.3.5.) were identified in freshly isolated as well as in cultured endothelial cells. Their specific and total activities proved to be much higher than in myocardial tissue, thus indicating a prominent role of nucleotide metabolism in the coronary endothelium.

Animals↗
PubMed1980

Inhibition by (aminooxy)acetate of the malate-aspartate cycle in the isolated working guinea pig heart.

The quantitative importance of the malate-aspartate cycle and the sn-glycerol 3-phosphate cycle, with respect to the flux of cytosolic reducing equivalents across the mitochondrial membrane, was studied in isolated perfused guinea pig hearts. The heart preparations performed pressure-volume work and metabolized glucose, lactate, pyruvate or 3-hydroxybutyrate. With glucose or lactate as the substrate, (aminooxy)acetate, an inhibitor of the malate-aspartate cycle, caused left ventricular failure, manifested by reduced aortic pressure and cardiac output, in association with a decrease in myocardial oxygen consumption and a depletion of high energy phosphate stores; lactate and particularly sn glycerol 3-phosphate accumulated in the myocardium. Moreover, lactate release rates increased more than 10-fold in presence of glucose and (aminooxy)acetate. Pretreatment of the animals with high doses of triiodothyronine did not prevent the hemodynamic and metabolic alterations caused by (aminooxy)acetate. In contrast, (aminooxy)acetate did not affect performance and energy-yielding metabolism when hearts metabolized pyruvate or 3-hydroxybutyrate as the substrate. From the findings it is concluded that the malate-aspartate cycle preponderates over the sn-glycerol 3-phosphate cycle in the working guinea pig heart, even when sn-glycerol 3-phosphate accumulates.

Acetates↗
PubMed1980

Early metabolic alterations during the development of experimentally induced cardiac hypertrophy.

The main biochemical features of protein synthesis in the myocardium are described as well as several metabolic alterations that occur during the very early stages in three models of cardiac hypertrophy resulting from pressure overload in vivo and in vitro, application of a high dose of isoprenaline and repeated daily administration of triiodothyronine. Furthermore, both mechanical and metabolic factors possibly participating in the development of cardiac hypertrophy are discussed and critically evaluated.

Adenosine Triphosphate↗
PubMed1979

Purine metabolism in cultured coronary endothelial cells.

Endothelial cells from coronary vessels of guinea pig hearts were isolated, cultivated and morphologically characterized. --Cells from confluent cultures contained adenine nucleotides and their dephosphorylated degradatives in exceptionally high amounts. --Adenine nucleotide levels were only slightly influenced by the growth state of the cultures and remained stable during incubation for three days in purine-free medium. In contrast, brief incubation of endothelial cells under anoxic conditions resulted in a substantial breakdown of adenine nucleotides associated with an enhanced formation and release of adenosine. --Measurements of specific activities of enzymes involved in adenine nucleotide synthesis and degradation lend additional support to the view that a very active adenine nucleotide metabolism is a typical feature of cultured coronary endothelial cells.

Adenine Nucleotides↗