Biomedical subjects

S W O'Connor

Publications and source records attributed to S W O'Connor.

8 recordsLinked to original sources

PubMed1986

Temperature and isoproterenol modulation of beta-adrenergic receptor characteristics.

Since agonists and temperature affect receptor affinity, and since these factors may influence the actual determination of receptor affinity, we assessed the in vitro effects of temperature and isoproterenol on the high and low affinity states of beta-adrenergic receptors in rat membrane preparations. There was a temperature-dependent decrease in beta-adrenergic receptor agonist affinity which was further promoted by the presence of isoproterenol. The decrease in receptor agonist affinity was reflected by a decrease in the number of receptors in the high affinity state. These data suggest that receptor desensitization may occur in membrane preparations in vitro and that the present methodology used to assess agonist affinity in vitro may itself alter the very properties being measured.

Animals↗

PubMed1985

Cholesterol modulation of beta-adrenergic receptor characteristics.

Cholesterol, a major structural component of plasma membranes, has a profound influence on cell surface receptor characteristics and on adenylate cyclase activity. beta-Adrenergic receptor number, adenylate cyclase activity, and receptor-cyclase coupling were assessed in rat lung membranes following preincubation with cholesteryl hemisuccinate. beta-Adrenergic receptor number increased by 50% without a change in antagonist affinity. However, beta-adrenergic receptor affinity for isoproterenol increased 2-fold as a result of an increase in the affinity of the isoproterenol high-affinity binding site. The increase in agonist affinity did not potentiate hormone-stimulated adenylate cyclase activity, which decreased 3-fold following cholesterol incorporation. However, the ratio of isoproterenol to GTP-stimulated activity was unchanged with cholesterol. Stimulation distal to the receptor by GTP, NaF, GppNHp, Mn2+ and forskolin also demonstrated 50-80% reduced enzyme activity following cholesterol incorporation. These data suggest that membrane cholesterol incorporation decreases catalytic unit activity without affecting transduction of the hormone signal.

Adenylyl Cyclases↗

PubMed1985

Characterization of the beta-adrenergic receptor of the rat peritoneal macrophage.

The beta-adrenergic receptor was characterized on BCG-activated rat peritoneal macrophage membranes by radio-ligand binding studies. Saturable binding with [125I]iodocyanopindolol (125I-ICYP) was demonstrated. With Scatchard analysis, rat macrophages demonstrate approximately 1000 receptors per cell with a Kd of 5 X 10(-11) M for 125I-ICYP. Competition curves with (-) and (+) propranolol at concentrations below 10(-6) M confirmed stereospecificity. The potency of various ligands to compete for 125I-ICYP binding sites followed the order: propranolol greater than isoproterenol greater than epinephrine greater than norepinephrine with apparent Kd of 2.0 X 10(-9), 3.9 X 10(-7), 1.0 X 10(-5), and 2.5 X 10(-5) M, respectively. Isoproterenol-stimulated adenylate cyclase activity was two-fold above basal activity. The potential physiologic significance of a beta-adrenergic receptor on rat peritoneal macrophages was suggested by a dose-dependent decrease in phagocytosis of soluble, model immune complexes (aggregated gamma-globulin) by macrophages incubated with metaproterenol. We conclude that the rat macrophage has a beta-adrenergic receptor and that catecholamines may thereby modulate macrophage function.

1-Methyl-3-isobutylxanthine↗

PubMed1984

The effect of age and cholesterol on the rat lung beta-adrenergic system.

To assess the influence of membrane lipid composition on beta-adrenergic receptor number and adenylate cyclase activity in aging, we investigated the effect of cholesteryl hemisuccinate on these parameters in lung membranes of 3-, 12-, and 24-month-old CDF (F-344) rats. When cholesteryl hemisuccinate (0.5 mg/ml) was incubated with lung membranes, beta-adrenergic receptor density was increased by 70%. This effect was the same for each age group studied and indicated that the density of both basal and CHS-sensitive receptors is unaltered in rat lung with age. Forskolin, NaF, p[NH]ppG, and isoproteronol-stimulated adenylate cyclase activity is 30% lower in lung membranes from aged rats. Since enzyme activity is affected by the lipid environment and membrane composition often changes with age, we assessed adenylate cyclase activity following cholesteryl hemisuccinate incorporation. There was up to a 75% decrease in adenylate cyclase activity following cholesteryl hemisuccinate incorporation in lung membranes in each of the three age groups. In untreated membranes, there was no significant difference in cholesterol or lipid phosphate content with age. These data suggest that cholesterol content does not account for alterations in senescent rat lung adenylate-cyclase activity.

Adenylyl Cyclases↗

PubMed1984

Accessibility of circulating immunoglobulin G to the extravascular compartment of solid rat tumors.

We have measured the rate of influx (kin) of normal rat immunoglobulin G (IgG) from the blood into the fluid surrounding the cells of three syngeneic rat fibrosarcomas as well as rat skin, muscle, lung, and kidney. Also measured was the rate of efflux (kout) of IgG from the tumor and tissue back into the circulation. The value of kin ranged from 0.11 to 9.0% of the blood value (activity/ml blood) transferred/hr/g for the nonmalignant tissues and from 6.2 to 7.9% of the blood value transferred/hr/g for the three tumors. Dividing kin by the plasma volume of the tissue gave a measure of the permeability of the vascular bed of that tissue. This ratio was rather constant for the normal tissues studied; however, it was at least an order of magnitude larger in all three tumors, indicating that the vasculature of the tumors was very permeable to IgG. The interstitial fluid volume (IF) of the tumor and tissue was calculated. The IF of the three tumors contained approximately 0.5 ml of fluid per g, while the IF of normal tissues had values that ranged from 0.14 to 0.34 ml of fluid per g. Knowledge of the IF, kin, and kout allowed a calculation of the concentration of IgG in the fluid surrounding the cells of tumors and tissues. The concentration of IgG in the IF of the tumors was found to be 50% of the plasma concentration; this was larger than the concentration of IgG in the IF of normal tissues, where the values ranged from 9 to 28% of the plasma concentration. A model for Ab localization onto solid tumors was developed. The model was used to discuss the mechanism of localization as well as the physiological limits of drug- or isotope-coupled Ab localization.

Animals↗

PubMed1983

The thermal lability of adenylate cyclase: mechanisms of stabilization.

The thermal inactivation of adenylate cyclase was investigated in human lymphocytes and in the N-protein deficient cyc- S49 mouse lymphoma cell line. The enzyme is rapidly inactivated at 37C with a t1/2 of 5.5 and 4.5 min respectively in human and cyc- membranes. Thermal inactivation is prevented by at least two mechanisms. The first mechanism involves ATP which stabilizes adenylate cyclase in a concentration dependent manner similar to the Km of ATP for cAMP formation. However, the inhibition of inactivation does not require Mg++ while the enzyme catalysis of ATP to cAMP does. The second mechanism involves substances which activate the enzyme. The human lymphocyte enzyme is equally stabilized by either NaF, GppNHp, or forskolin. In contrast, the cyc- enzyme is fully stabilized by forskolin but only partially stabilized by NaF. When human erythrocyte N-protein extract is added to cyc- membranes, NaF fully stabilizes the enzyme. These data suggest that an activated N-protein is instrumental in stabilizing adenylate cyclase and that there is some N-protein component in cyc- membranes through which NaF may be exerting its stabilizing action.

Adenosine Triphosphate↗

PubMed1981

Age-associated decrease of adenylate cyclase activity in rat myocardium.

Myocardial inotropic and chronotropic responses to beta-adrenergic agonists are diminished with aging. Since myocardial beta-adrenergic receptors are unaltered with age, we tested the hypothesis that this decreased responsiveness is related to a defect in the adenylate cyclase system. Isoproterenol-stimulated adenylate cyclase activity was assessed in myocardial membranes from Fischer 344 rats of 3, 12, and 24 months of age. Basal, as well as F-, GTP-, and hormone-stimulated adenylate cyclase activity was decreased by 20-30% with age. The Km of the enzyme for ATP was not found to be statistically different for any age group studied. These data support the hypothesis that the diminished responsiveness seen in senescence is a result of an alteration in either the catalytic subunit or the coupling protein (N) of the adenylate cyclase complex.

Adenosine Triphosphate↗

PubMedDate not supplied

Age-associated decrease in the catalytic unit activity of rat myocardial adenylate cyclase.

Two techniques were employed to define the site of diminished adenylate cyclase activity observed in age-rat myocardium. We previously reported no change in beta-adrenergic receptor number or affinity but decreased NaF- and 5'-guanylylimidodiphosphate (GppNHp)-stimulated adenylate cyclase activity with age in the rat myocardium. These data suggest a defect in either N-protein or the catalytic unit component of adenylate cyclase. N-protein activity was assessed by the ability of myocardial membrane extracts from 3-, 12, and 24-month-old Fischer 344 rats to complement the N-protein deficient cyc- S49 mouse lymphoma cell line. Catalytic unit activity was assessed by the ability of forskolin to stimulate adenylate cyclase in myocardial membranes from young and old rats. The results demonstrate that both N-protein activity and catalytic unit activity are diminished with age in the rat myocardium. However, since N-protein is present in excess relative to catalytic unit, the data are consistent with the hypothesis that the loss of catalytic unit activity accounts for the loss of overall enzyme activity. The effect is tissue-specific, since erythrocyte membranes do not show the same age-dependent loss of adenylate cyclase activity.

Adenylyl Cyclases↗