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B T Liang

Publications and source records attributed to B T Liang.

44 records · Page 3Linked to original sources
PubMed1987

Reconstitution of muscarinic cholinergic inhibition of adenylate cyclase activity in homogenates of embryonic chick hearts by membranes of adult chick hearts.

We have previously demonstrated that during embryonic development of the chick heart between days 2 1/2 and 10 days in ovo, muscarinic cholinergic inhibition of isoproterenol-stimulated adenylate cyclase activity increased 4-fold, and the sensitivity of isoproterenol-stimulated adenylate cyclase activity to inhibition by carbamylcholine increased 26-fold. Although the number of muscarinic receptors remained constant between days 2 1/2 and 10 in ovo, the levels of a 39- and 41-kDa pertussis toxin substrate increased in parallel with the ability of muscarinic agonist to inhibit adenylate cyclase activity (Liang. B.T., Hellmich, M. R., Neer, E. J., and Galper, J. B. (1986) J. Biol. Chem. 261, 9011-9021). These data are consistent with the hypothesis that between days 2 1/2 and 10 in ovo muscarinic receptors were uncoupled from inhibition of adenylate cyclase activity because of limiting levels of pertussis toxin substrates. In the current studies, in order to test this hypothesis homogenates of embryonic chick hearts 3 1/2 days in ovo were reconstituted with membranes from hearts of hatched chicks. In order to rule out reconstitution by factors from hatched chick hearts other than pertussis toxin substrates, muscarinic receptors from hatched chick hearts were inactivated by covalent binding of benzilycholine mustard and adenylate cyclase inactivated by N-ethylmaleimide prior to reconstitution. Reconstitution of benzilylcholine mustard/N-ethylmaleimide treated hatched chick heart membranes with homogenates of embryonic chick hearts 3 1/2 days in ovo resulted in a 2 1/2-fold increase in the ability of carbamylcholine to inhibit adenylate cyclase activity and reconstitution of hatched chick heart membranes with homogenates of hearts 2 1/2 days in ovo resulted in an approximately 10-fold increase in the sensitivity of isoproterenol-stimulated adenylate cyclase activity to inhibition by carbamylcholine. Membranes from hearts of hatched chicks which had been injected with pertussis toxin were incapable of reconstituting muscarinic inhibition of adenylate cyclase activity in homogenates of hearts 3 1/2 days in ovo. These data support the conclusion that early in embryonic development coupling of muscarinic receptors to inhibition of adenylate cyclase activity is limited by the availability of a pertussis toxin substrate.

Adenylate Cyclase Toxin↗
PubMed1986

Development of muscarinic cholinergic inhibition of adenylate cyclase in embryonic chick heart. Its relationship to changes in the inhibitory guanine nucleotide regulatory protein.

Parasympathetic and sympathetic innervation of the embryonic chick heart proceed non-coordinately. beta-Adrenergic agonists mediate an increase in beating rate in embryonic chick heart prior to ingrowth of the vagus nerve (Culver, N. G., and Fishman, D. A. (1977) Am. J. Physiol. 232, R116-R123) while muscarinic agonists exert relatively little effect on beating rate in hearts 2-4 days in ovo (Papanno, A. J. (1979) Pharmacol. Rev. 29, 3-33). Studies of developmental changes in the ability of muscarinic agonists to inhibit adenylate cyclase activity and their relationship to the development of a physiologic response of the embryonic chick heart to muscarinic stimulation have been inconclusive. In the current studies the ability of isoproterenol to stimulate adenylate cyclase activity did not change during development. Maximum stimulation above basal was 760 pmol of cAMP/10 min/mg of proterin with an IC50 of 1.5 X 10(-6) M for isoproterenol in homogenates of hearts 2 1/2, 3 1/2, and 10 days in ovo and 3 days posthatching. However, inhibition of isoproterenol-stimulated adenylate cyclase activity by carbamylcholine increased from 7.6% with a IC50 for carbamylcholine of 16 +/- 5.0 microM at day 2 1/2 in ovo to 29% with an IC50 of 0.4 +/- 0.1 microM at day 10 in ovo and to 43% with a IC50 of 0.6 +/- 0.1 microM at 3 days posthatching. Since previous data had demonstrated the presence of muscarinic receptors as early as 2 1/2 days in ovo (Galper, J. B., Klein, W., and Catterall, W. A. (1977) J. Biol. Chem. 252, 8692-8699), studies of developmental changes in guanine nucleotide-coupling proteins were carried out to determine whether early in development muscarinic receptors were uncoupled from a physiologic response. Studies of pertussis toxin-catalyzed ADP-ribosylation of homogenates of embryonic chick heart with [32P]NAD demonstrated the presence of two ADP-ribosylated proteins at 39,000 and 41,000 kDa, respectively. Both ADP-ribosylation and immunoblotting of homogenates with an antibody to the 39-kDa guanine nucleotide-binding protein in bovine brain demonstrated that the 39-kDa alpha protein increased 1.8-fold between days 2 1/2 and 3 1/2 in ovo and another 1.8-fold from day 3 1/2 to 10 in ovo in parallel with the increase in the extent of muscarinic inhibition of adenylate cyclase activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Diphosphate Ribose↗
PubMed1986

Beta adrenergic receptor subtypes in the atria: evidence for close coupling of beta-1 and beta-2 adrenergic receptors to adenylate cyclase.

The relationship between occupancy of beta adrenergic receptors and stimulation of adenylate cyclase in dog atrial tissue was examined by studying the binding of [125I]iodopindolol and the activation of adenylate cyclase. Computer-assisted nonlinear regression analysis was used to analyze the inhibition of isoproterenol-stimulated adenylate cyclase activity by beta-1- or beta-2-selective antagonists. The Ki values for each subtype of receptor for the selective antagonists resulting from studies of the inhibition of adenylate cyclase activity were similar to those determined in studies of the inhibition of the binding of [125I]iodopindolol. To compare further the occupancy of beta-1 or beta-2 adrenergic receptors with the activation of adenylate cyclase mediated by each class of receptor, computer modeling of the stimulation of adenylate cyclase by the beta-1-selective agonist norepinephrine was carried out. The EC50 values of norepinephrine for each receptor subtype, as measured in studies of norepinephrine-stimulated adenylate cyclase activity, were similar to the Ki values for the inhibition by norepinephrine of the binding of [125I]iodopindolol to each receptor subtype. The data led to the conclusion that beta-1 adrenergic receptors make up about 70% of the total number of beta adrenergic receptors and mediate 70% of the increase in adenylate cyclase activity produced by isoproterenol. These results suggest that the relationship between occupancy of each class of receptor and activation of adenylate cyclase is linear and that, when agonist-stimulated adenylate cyclase activity is used as a functional response, neither spare beta-1 nor spare beta-2 adrenergic receptors exist in the atrium.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclases↗
PubMed1985

Beta 2-adrenergic receptors contribute to catecholamine-stimulated shortening of action potential duration in dog atrial muscle.

beta 1- and beta 2-adrenergic receptors coexist in atria of humans and a variety of other species. The significance of these receptors and the quantitative extent to which beta 2-adrenergic receptors contribute to the electrophysiologic effects of catecholamines acting at beta-adrenergic receptors have not been determined. In the present study, the beta 1-selective antagonist ICI 89,406 and the beta 2-selective antagonist ICI 118,551 were used to determine the relative densities of beta 1- and beta 2-adrenergic receptors on membranes prepared from dog atria and the contribution that each subtype makes to isoproterenol-induced shortening of action potential duration measured at 75% repolarization (APD75). Computer-aided nonlinear regression analysis of the inhibition of the specific binding of [125I]iodopindolol and the inhibition of isoproterenol-stimulated APD75 shortening by these antagonists showed that a two-site model fits the data better than a one-site model (P less than 0.0001). The affinity constants for each selective antagonist determined by inhibition of APD75 shortening were similar to those determined in studies of the inhibition of the specific binding of [125I]iodopindolol to beta 1- and beta 2-adrenergic receptors. beta 2-Adrenergic receptors made up approximately equal to 25% of the total number of beta-adrenergic receptors but mediated approximately equal to 50% of the electrophysiologic effect of isoproterenol. The inhibition of the binding of [125I]iodopindolol and of the APD75 shortening by propranolol, a nonselective antagonist, was best fit by a one-site model. In other experiments, dogs were treated with reserpine and atropine to eliminate complications caused by the presence of endogenous norepinephrine and acetylcholine. Results obtained with atria from these animals were similar to those obtained in studies with control atria. These data suggest that beta 2-adrenergic receptors as well as beta 1-adrenergic receptors are involved in mediating the electrophysiologic effects of catecholamines in dog atrial muscle and that both beta 1- and beta 2-adrenergic receptors are present on atrial muscle cells.

Action Potentials↗
PubMed1982

Monensin inhibits catecholamine synthesis in pheochromocytoma cells.

The carboxylic ionophore monensin inhibits the activity of tyrosine 3-monooxygenase and decreases the rate of catecholamine synthesis in pheochromocytoma cells incubated in vitro. The ionophore inhibits dopa production in intact pheochromocytoma cells, but does not itself inhibit tyrosine 3-monooxygenase and does not produce a stable inactivation of the enzyme as assayed in cell-free extracts of the cells. The inhibition of dopa production by monensin is dependent upon extracellular Na+, but does not require extracellular Ca++. This effect of monensin is more pronounced in the presence of pargyline. In the absence of pargyline, monensin also depletes the cells of norepinephrine and increases the accumulation of the deaminated norepinephrine metabolite, dihydroxyphenylglycol. Finally, monensin increases the release of catecholamines from isolated chromaffin granules. These results are consistent with the hypothesis that monensin causes the release of norepinephrine from chromaffin granules into the cytoplasm of pheochromocytoma cells and that the increase in cytoplasmic norepinephrine inhibits tyrosine 3-monooxygenase activity.

Adrenal Gland Neoplasms↗
PubMed1981

Studies on dihydropteridine reductase activity in pheochromocytoma cells.

The activity of dihydropteridine reductase (DPR) in pheochromocytoma cells has been studied. The activity of this enzyme in crude extracts of pheochromocytoma cells is approximately 50 nmol/min/mg protein. This activity is very much greater than the activity of tyrosine 3-monooxygenase (TH) in these extracts and the rate of conversion of tyrosine to DOPA in intact pheochromocytoma cells. Incubation of the cells with 56 mM-K+ or with cholera toxin has previously been shown to increase the rate of catecholamine synthesis and to cause a stable activation of TH in the cells. These treatments do not produce a stable activation of DPR, as assayed in vitro. Methotrexate inhibits DPR activity in vitro with an I50 of approximately 20 microM, but has no effect on the rate of DOPA formation in intact pheochromocytoma cells. Therefore, DPR does not appear to be the rate-limiting enzyme in the pathway of catecholamine synthesis in pheochromocytoma cells. Moreover, the activities of DPR and of TH are not regulated coordinately in these cells.

Adrenal Gland Neoplasms↗
PubMed1980

Tyrosine 3-monooxygenase regulates catecholamine synthesis in pheochromocytoma cells.

Incubation of pheochromocytoma cells with 56 mM K+ or with cholera toxin increases the conversion of [14C]tyrosine to [14C]catecholamines (Chalfie, M., Settipani, L., and Perlman, R. L. (1979) Mol. Pharmacol. 15, 263-270). We have now measured the tyrosine content and the rate of dihydroxyphenylalanine production in these cells. Incubation with 56 mM K+ or with cholera toxin increases the rate of dihydroxyphenylalanine production but decreases the tyrosine content of the cells. We have also measured the uptake of tyrosine into pheochromocytoma cells. The rate of tyrosine uptake is more than 1 order of magnitude greater than the rate of dihydroxyphenylalanine production. Moreover, tyrosine uptake is not affected by cholera toxin and is decreased by approximately 30% in media that contain 56 mM K+. These results provide direct evidence that tyrosine 3-monooxygenase regulates catecholamine synthesis in pheochromocytoma cells and that incubation with 56 mM K+ or with cholera toxin causes the activation of this enzyme in these cells.

Biological Transport↗