Concentration Of High-affinity Receptors Research Articles

Serotoninergic agonist and antagonist are modulators of the α2-adrenoceptor activity in rat brain cortex membranes

The influence of activation and inhibition of serotonin receptors by serotonin (5HT) and miancerin on binding of specific nonselective α2-antagonist [3H]RX821002 in rat cerebral cortex membranes was studied. It was shown that the ligand-receptor interaction for α2-adrenoceptors corresponded to the model suggesting the presence of one pool of receptors and binding of two ligand molecules to the receptor. The parameters of the [3H]RX821002 binding to α2-adrenoceptors were as follows: Kd = 1.57 ± 0.276 nM, Bmax = 7.24 ± 1.63 fmol/mg protein, n = 2. In the case of activation of 5HT-receptors by serotonin, the character of ligand binding was different: two pools of receptors were detected with the parameters Kd1 = 0.82 ± 0.06; Kd2 = 2.65 ± 0.22 nM; Bm1 = 1.65 ± 0.23; Bm2 = 4.20 ± 0.11 fmol/mg protein; n = 2. The affinity of high-affinity receptors increased twofold and the affininty of low-affinity receptors decreased by 69% as compared to control values. The concentration of high-affinity receptors decreased 4.4-fold, and of low-affinity, 1.7-fold. The value of maximal reaction (Bmax) decreased by 20%. In the case of miancerin-induced inhibition of 5HT-receptors the character of ligand binding also changed; two pools of receptors were detected with the following parameters: Kd1 = 0.48 ± 0.09; Kd2 = 3.79 ± 0.71 nM; B1 = 0.63 ± 0.17; B2 = 4.75 ± 0.21 fmol/mg protein; n = 2. The affinity of high-affinity receptors pool increased by 70% and that of low-affinity receptors decreased by 76% as compared to control values. The concentration of active high-affinity and low-affinity α2-adrenoceptors decreased by 70% and 141%, respectively. The total amount of the receptors (Bmax) decreased by 26%. The data suggest that α2-adrenoceptors in rat cerebral cortex exist as dimers. Modulatory effects of serotonin and miancerin on specific antagonist binding to α2-adrenoceptors may be accomplished by altering the character and binding parameters of the nonselective α2-antagonist [3H]RX821002.

Localization of the Na +-sugar cotransport system in a kidney epithelial cell line (LLC PK 1)

Studies of the localization of the Na +-dependent sugar transport in monolayers of LLC PK 1 cells show that the uptake of a methyl α- d-glucoside, a nonmetabolizable sugar which shares the glucose-galactose transport system, occurs mainly from the apical side of the monolayer. Kinetics of [ 3H]phlorizin binding to monolayers of LLC PK 1 cells were also measured. These studies demonstrate the presence of two distinct classes of receptor sites. The class comprising high affinity binding sites had a dissociation constant ( K d ) of 1.2 μM and a concentration of high affinity receptors of 0.30 μmol binding sites per g DNA. The other class involving low affinity sites had a K d of 240 μM with the number of binding sites equal to 12 μmol/g DNA. Phlorizin binding at high affinity binding sites is a Na +-dependent process. Binding at the low affinity sites on the contrary is Na +-independent. The mode of action of Na + on the high affinity binding sites was to increase the dissociation constant without modifying the number of binding sites. The Na + dependence and the matching of K d for high affinity binding sites with the K i of phlorizin for the inhibition of methyl α- d-glucoside strongly suggest that the high affinity phlorizin binding site is, or is part of the methyl α- d-glucoside transport system. Binding studies from either side of the monolayer also show that the binding of phlorizin at the Na + dependent high affinity binding sites occurs mainly from the apical rather than the basolateral side. The specific location of the Na +-dependent sugar transport system in the apical membrane of LLC PK 1 cells is, therefore, another expression of the functional polarization of epithelial cells that is retained under tissue culture condition. In addition, since this sugar transport almost disappears after the cells are brought into suspension, it can be used as a marker to study the development of the apical membrane in this cell line.

Increased affinity and number of insulin receptors in the fetus

Insulin has been implicated as a growth factor in fetal life. Clinical and experimental studies have shown that fetal hyperinsulinism leads to macrosomia. Studies in adult humans have demonstrated that changes in insulin binding to mononuclear cells mirror changes in other tissues. In an attempt to clarify the role of insulin in the fetus, we have studied insulin receptors on mononuclear leukocytes in cord blood from 12 normal newborns. Eight healthy young adults served as controls. The average specific binding in the absence of unlabelled insulin was 24.3 ± 3.5% and 4.7 ± 0.9% in newborns and adults respectively. This increase in binding is caused by an increase in number of receptor sites per cell, as well as by an increase in receptor affinity. The newborns had an average of 44600 receptor sites per cell compared with 7100 for the adult controls. An average affinity constant Ke was 5.9 × 108M−1 for newborns, and 2.9 × 108M−1 for the adults. This finding of markedly high concentrations of high affinity receptors for insulin on fetal cells may reflect the importance of insulin in intrauterine growth and development.