Lymphocyte beta-adrenergic receptor modification in bulimia.

beta-Adrenergic receptor binding on circulating lymphocytes was evaluated in young female bulimic patients (n = 12) and age- and sex-matched normal control volunteers (n = 10). Using iodine 125-labeled cyanopindolol, antagonist binding was evaluated (number of receptors [Bmax] and dissociation constant [KD]), and using isoproterenol competition of cyanopindolol binding, the concentration required to inhibit binding by 50% (IC50) for isoproterenol and the agonist affinity measure of KL/KH (ratio of dissociation constants for the low- and high-affinity states of the receptor) were determined. Plasma norepinephrine (NE) level was also measured. There was a trend toward lower plasma NE levels in the bulimic patients. The KL/KH ratio in bulimic patients was significantly greater than that for the normal volunteers, indicating increased receptor coupling. The KL/KH ratio was not significantly correlated with plasma NE level. Neither Bmax nor KD was different between the two groups. These findings suggest that beta-adrenergic receptors in bulimic patients may be more responsive than in normal subjects, without alteration of the traditional measures of receptor responses, a difference that cannot be explained on the basis of plasma NE. These findings provide another line of evidence for altered regulation of the noradrenergic system in bulimic patients during a controlled phase of their illness.

Inhibition of [3H]Gpp(NH)p release from human platelet membranes by GDP.

Previous reports have described hormone-stimulated release of radiolabeled guanine nucleotides from prelabeled membranes. In this paper, we report the inhibition by GDP of both hormone-independent and hormone-specific stimulation of [3H]Gpp(NH)p release from human platelet membranes. This inhibition was shown to be concentration specific and suggests a more complex mechanism of [3H]Gpp(NH)p release than was previously thought. A model is proposed in which guanosin triphosphates cause dissociation of the subunits of the guanine nucleotide binding protein, whereas guanosine diphosphate prevents this dissociation resulting respectively in stimulation or inhibition of the release of bound [3H]Gpp(NH)p.

[Functional changes of components of the adenylate cyclase complex during heterologous desensitization by isoproterenol]. / Funktsional'nye izmeneniia komponentov adenilattsiklaznogo kompleksa v khode geterologicheskoi desensitizatsii k izoproterenolu.

Long-term exposure of pigeon erythrocytes to isoproterenol produces significant changes in the regulatory properties of adenylate cyclase. The enzyme sensitivity to the action of isoproterenol (0.1 mM) and GTP (0.1 mM) decreases by 50% for 15 min, that to guanylyl imidodiphosphate, a non-hydrolyseable analog of ATP, by 20-25% for 40 min. One of possible causes of the loss of the adenylate cyclase sensitivity to isoproterenol is the disturbances in the interaction between the beta-adrenoreceptor and N-protein. In experiments with restoration of the beta-adrenoreceptor and the catalytic component sensitivity to GTP, the properties of the N-protein both in the control and in desensitized preparations of plasma membranes remained unchanged. It was found that the disturbances in the beta-adrenoreceptor--N-protein interactions are due to the changes in the receptor properties. Similar results were obtained in experiments with membranes isolated from the cells to which isoproterenol or cAMP (0.1 mM) + isobutylmethylxanthine (1 mM) were added; the desensitization in this case was probably due to the phosphorylation of the beta-adrenoreceptor. A model of heterologous desensitization of pigeon erythrocyte adenylate cyclase is proposed.

ADP- and epinephrine-elicited release of [3H]guanylylimidodiphosphate from platelet membranes. Implications for receptor-Ni stoichiometry.

Epinephrine-promoted release of [3H]guanylylimidodiphosphate ( [3H]Gpp(NH)p) from human platelet membranes has been used to probe the interactions between alpha 2-adrenergic receptors and Ni, the guanine nucleotide binding protein that couples those receptors to an inhibition of adenylate cyclase activity. We show here that ADP, which also acts through specific platelet receptors to inhibit adenylate cyclase activity, also promotes the release of [3H]Gpp(NH). The amount of [3H]Gpp(NH)-release elicited by epinephrine and by ADP together is equal to the sum of the amounts released by the two agents acting individually. Furthermore the maximal amounts of [3H]Gpp(NH)-release elicited by each of the two agents approximates the numbers of receptors for ADP and epinephrine present in the platelet membranes. These results suggest that the two receptor types interact with distinct portions of the pool of Ni molecules and that each receptor initiates guanine-nucleotide exchange on a single molecule of Ni.

5'-p-Fluorosulfonylbenzoyl guanosine as a probe for the GTP-binding protein in alpha 2-adrenergic receptor-adenylate cyclase systems.

An analog of GTP, 5'-p-fluorosulfonylbenzoyl guanosine (5'-p-FSO2BzGuo), appears to interact irreversibly with guanine nucleotide-binding sites on human platelet membranes. This conclusion is based on the observation that incubation of human platelet membranes with 1.4 mM 5'-p-FSO2BzGuo followed by extensive membrane washing results in a reduction in the density of binding sites for [3H]guanylylimidodiphosphate (Gpp(NH)p), a hydrolysis-resistant analog of GTP. The alpha 2-adrenergic receptor of human platelets is felt to interact with a GTP-binding protein that modulates alpha 2-receptor-agonist interactions and mediates inhibition of adenylate cyclase. The present data suggest that 5'-p-FSO2BzGuo attains saturating, or near saturating, occupancy of this alpha 2-receptor-associated GTP-binding protein, since incubation of human platelet membranes with 5'-p-FSO2BzGuo mimics the effects of optimal concentrations of Gpp(NH)p (0.1 mM) in reducing alpha 2-receptor affinity for agonist agents: 5'-p-FSO2BzGuo increases the EC50 for epinephrine competition for [3H]yohimbine antagonist binding to alpha 2-receptors from 0.15 to 1.5 microM and promotes a time- and concentration-dependent decrease in high affinity [3H]epinephrine agonist binding. The persistent effects of 5'-p-FSO2BzGuo on alpha 2-receptor-agonist interactions following extensive washing of the platelet membranes suggest that 5'-p-FSO2BzGuo modification of the alpha 2-receptor-associated GTP-binding protein is irreversible. Taken together, the above findings suggest that 5'-p-FSO2BzGuo may be the appropriate reagent to prepare in a radiolabeled form to affinity label the GTP-binding proteins in human platelet membranes and compare the properties of alpha 2-adrenergic receptor-associated GTP-binding protein(s) with those of the presumably distinct GTP-binding protein that mediates stimulation of adenylate cyclase.