Effects of chronic nicotine treatment on nicotinic receptors in the rabbit urinary bladder

Nicotine induced a phasic contraction in the rabbit urinary bladder. The response was abolished by hexamethonium and partially reduced by atropine and capsaicin. Simultaneous atropine and capsaicin treatment did not abolish the contraction. These findings suggest that the response to nicotine is due to acetylcholine, tachykinins, and unknown mediator release. In contrast, nicotine-induced contraction diminished following the chronic nicotine treatment without a change of its pharmacological properties. These results suggest the possibility that chronic nicotine treatment causes a decrease in nicotinic receptor numbers. Therefore, the binding properties of (-)-[3H]nicotine on rabbit urinary detrusor muscle membrane fractions were studied to evaluate the effects of chronic nicotine treatment on nicotinic receptors. Specific (-)-[3H]nicotine binding reached saturation and Scatchard plots were curvilinear, suggesting the existence of two different affinity sites for (-)-[3H]nicotine. Dissociation constants (KD) and maximum binding sites (Bmax) were KD1 = 4.91 +/- 1.88 nM, Bmax1 = 2.42 +/- 0.22 fmol/mg protein and KD2 = 263 +/- 56 nM, Bmax2 = 25.0 +/- 4.3 fmol/mg protein. In urinary bladder membrane fractions from chronic nicotine-treated rabbits, KD and Bmax values were KD1 = 3.96 +/- 0.38 nM, Bmax1 = 1.07 +/- 0.25 fmol/mg protein and KD2 = 249 +/- 12 nM, Bmax2 = 10.8 +/- 1.5 fmol/mg protein. Dissociation constants for both sites following chronic nicotine treatment did not change but maximum binding site numbers for both sites significantly decreased (p less than 0.05). These results suggest that the decrease in contractile response evoked by nicotine after chronic nicotine treatment in rabbit urinary bladder is due to a decrease in numbers of nicotinic receptors.     

Studies on the properties of muscarinic cholinergic receptor sites in bovine pineal gland

1. Using the tritiated muscarinic receptor antagonist, quinuclidinyl benzilate ([3H]QNB) as a ligand, muscarinic cholinergic receptors have been identified and characterized in the pineal glands of cow and swamp buffalo. 2. At 25 degrees C, the specific binding reached equilibrium within 60 min and remained constant for an additional two hours. Furthermore, the specific binding was saturable, reversible and tissue dependent in nature. 3. The kinetic analyses of muscarinic cholinergic receptor sites revealed KD values of 0.423 +/- 0.01 nM and 0.218 +/- 0.01 nM, and Bmax values of 69.75 +/- 20.91 fmol/mg protein and 74.19 +/- 32.73 fmol/mg protein for the cow's- and the swamp buffalo's pineal glands, respectively. 4. The presence of muscarinic cholinergic receptor sites originating from cholinergic innervation of the pineal gland is suggested.     

Preponderance of alpha 2- over beta 1-adrenergic receptor sites in human fat cells is not predictive of the lipolytic effect of physiological catecholamines

Adrenergic control of human fat cell lipolysis is mediated by two kinds of receptor sites that are simultaneously stimulated by physiological amines. To establish a correlation between the binding characteristics of the receptor and biological functions, the ability of physiological amines to stimulate or inhibit isolated fat cell lipolysis in vitro was compared to the beta- and alpha 2-adrenoceptor properties of the same fat cell batch. The beta-selective antagonist (-)[3H]dihydroalprenolol ([3H]DHA) and the alpha 2-selective antagonists [3H]yohimbine ([3H]YOH) and [3H]rauwolscine ([3H]RAU) were used to identify and characterize the two receptor sites. Binding of each ligand was rapid, saturable, and specific. The results demonstrate 1) the weaker lipolytic effect of epinephrine compared with norepinephrine. This can be explained by the equipotency of the amines at the beta 1-sites and the higher affinity of epinephrine for alpha 2-adrenergic receptors. 2) The preponderance of alpha 2-adrenergic receptor sites labeled by [3H]YOH (Bmax, 586 +/- 95 fmol/mg protein; KD, 2.7 +/- 0.2 nM) or [3H]RAU (Bmax, 580 +/- 100 fmol/mg protein; KD, 3.7 +/- 0.1 nM). These two ligands can be successfully used to label alpha 2-adrenergic receptor sites. 3) The beta 1-adrenergic receptor population labeled by [3H]DHA(Bmax, 234 +/- 37 fmol/mg protein; KD, 1.8 +/- 0.4 nM), although a third as numerous as the alpha 2-adrenergic population, is responsible for the lipolytic effect of physiological amines and is weakly counteracted by simultaneous alpha 2-adrenergic receptor stimulation under our experimental conditions. It is concluded that, in human fat cells, the characterization of beta 1- and alpha 2-adrenergic receptors by saturation studies or kinetic analysis to determine affinity (KD) and maximal number of binding sites (Bmax) is not sufficient for an accurate characterization of the functional adrenergic receptors involved in the observed biological effect.     

Identification of α2-Adrenergic Receptors in Chicken Pineal Gland Using [3H]Rauwolscine

The norepinephrine-induced inhibition of avian pineal N-acetyltransferase activity appears to be mediated by alpha 2-adrenergic receptors. In this study, alpha 2-adrenergic receptors in the chicken pineal gland were directly identified by radioligand binding. Membrane preparations of pineal glands from chickens from 1 to 6 weeks of age were examined using [3H]rauwolscine, a selective alpha 2-adrenergic receptor antagonist, to characterize the binding sites. The results indicate no ontological change in either the affinity (KD) or density of receptor binding sites (Bmax) during the time span examined. The binding was saturable and of high affinity with a mean KD of 0.27 +/- 0.01 nM and a mean Bmax of 242 +/- 12 fmol/mg protein. Further characterization of these binding sites indicated that the alpha 2-adrenergic receptor is of the alpha 2A subtype, since prazosin and ARC-239 bound with low affinities and oxymetazoline bound with high affinity.     

Magnesium-Dependent Enhancement of Endogenous Agonist Binding to A1 Adenosine Receptors: A Complicating Factor in Quantitative Autoradiography

Quantitative autoradiography was used to investigate the effects of Mg2+ on agonist and antagonist binding to A1 receptors in rat striatum. A1 receptors were labelled with the selective agonist N6-[3H]cyclohexyladenosine ([3H]CHA) or the selective antagonist 1,3-[3H]dipropyl-8-cyclopentylxanthine ([3H]DPCPX). Mg2+ had no significant effect on equilibrium binding constants for [3H]CHA [control: KD (95% confidence interval) of 0.34 (0.15-0.80) nM and Bmax of 267 +/- 8 fmol/mg of gray matter; with 10 mM Mg2+: KD of 0.8 (0.13-4.9) nM and Bmax of 313 +/- 8.9 fmol/mg of gray matter] or [3H]DPCPX [control: KD of 0.54 (0.30-0.99) nM and Bmax of 256 +/- 2.3 fmol/mg of gray matter; with 10 mM Mg2+: KD of 1.54 (0.2-11.0) nM and Bmax of 269 +/- 35.7 fmol/mg of gray matter]. In contrast, Mg2+ slowed the apparent association rate for both ligands; this was observed as a shift from a one-component to a two-component model for [3H]DPCPX. Mg2+ also affected the dissociation rates of both ligands; for [3H]CHA, dissociation in the presence of Mg2+ was not detected. Mg2+ produced a concentration-dependent inhibition of [3H]CHA binding only prior to equilibrium. HPLC was performed on untreated sections, sections preincubated with adenosine deaminase (ADA), and sections preincubated with ADA and incubated with ADA in the absence or presence of Mg2+. Adenosine was found in measurable quantities under all conditions, and the concentration was not influenced by Mg2+ or by the inclusion of GTP in the preincubation medium. From these data, we conclude the following: (a) adenosine is present and may be produced continuously in brain sections; (b) ADA is not capable of completely eliminating the produced adenosine; (c) Mg2+ apparently does not influence adenosine production or elimination; (d) A1 receptor-guanine nucleotide binding protein coupling is maximal in this preparation; and (e) Mg2+ decreases the dissociation rate of bound endogenous adenosine from A1 receptors, thus limiting the access of [3H]CHA and [3H]DPCPX to the receptors. Thus, enhancement of endogenous adenosine binding to A1 receptors by Mg2+ is a complicating factor in receptor autoradiography and may be so in other preparations as well.     

Effect of male accessory glands autoaggression on androgenic cytosolic and nuclear receptors of rat prostate.

The effect of immunization against male accessory gland (MAG) homogenates over androgenic cytosolic and nuclear receptors of rat prostate was studied. In the MAG-immunized rats the Bmax of cytosolic receptors was significantly increased (120.3 +/- 44.3 vs 47.7 +/- 24.9 fmol/mg protein, p less than 0.01, mean +/- SD). In contrast, the Bmax of nuclear receptors in the MAG-immunized rats showed no significant difference as regarded controls (kidney immunized rats) when expressed as fmol/100 micrograms DNA (196.1 +/- 84.8 vs 148.3 +/- 88.9) but it show to slight differences (p less than 0.1) when data were reported as percent of weight of tissue (2,189 +/- 918.6 vs 1,303 +/- 611.2 fmol/g wet issue). Results (mean +/- SD) on binding affinity of cytosolic receptors showed no significant differences in MAG-immunized rats as compared with controls (Kd: 1.98 +/- 0.66 vs 1.92 +/- 0.20 nM). Likewise, only a slight difference between both groups was attained for Kds of nuclear receptors (2.34 +/- 0.28 vs 1.80 +/- 0.62 nM, p less than 0.2). On the other hand, 5 alpha 1-dihydrotestosterone (DHT) values obtained in prostate homogenates were significantly decreased in MAG-immunized rats as compared with controls (17.4 +/- 2.0 vs 7.1 +/- 0.9 ng/g tissue, mean +/- SD, p less than 0.01). However, testosterone (T) levels in gland tissue showed no significant differences between both groups (2.4 +/- 0.5 vs 2.6 +/- 0.3 ng/g tissue) with an increase in the T: DHT ratio from 0.14 to 0.37.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of Opioid Receptors in Cultured Neurons

The appearance of mu-, delta-, and kappa-opioid receptors was examined in primary cultures of embryonic rat brain. Membranes prepared from striatal, hippocampal, and hypothalamic neurons grown in dissociated cell culture each exhibited high-affinity opioid binding sites as determined by equilibrium binding of the universal opioid ligand (-)-[3H]bremazocine. The highest density of binding sites (per mg of protein) was found in membranes prepared from cultured striatal neurons (Bmax = 210 +/- 40 fmol/mg protein); this density is approximately two-thirds that of adult striatal membranes. By contrast, membranes of cultured cerebellar neurons and cultured astrocytes were devoid of opioid binding sites. The opioid receptor types expressed in cultured striatal neurons were characterized by equilibrium binding of highly selective radioligands. Scatchard analysis of binding of the mu-specific ligand [3H]D-Ala2,N-Me-Phe4,Gly-ol5-enkephalin to embryonic striatal cell membranes revealed an apparent single class of sites with an affinity (KD) of 0.4 +/- 0.1 nM and a density (Bmax) of 160 +/- 20 fmol/mg of protein. Specific binding of (-)-[3H]bremazocine under conditions in which mu- and delta-receptor binding was suppressed (kappa-receptor labeling conditions) occurred to an apparent single class of sites (KD = 2 +/- 1 nM; Bmax = 40 +/- 15 fmol/mg of protein). There was no detectable binding of the selective delta-ligand [3H]D-Pen2,D-Pen5-enkephalin. Thus, cultured striatal neurons expressed mu- and kappa-receptor sites at densities comparable to those found in vivo for embryonic rat brain, but not delta-receptors.     

Benzodiazepine receptors on human blood platelets

Binding studies conducted on membrane preparation from human platelets using (3H) Ro5-4864 and (3H) diazepam showed specific and saturable binding. Scatchard analysis revealed a single class of binding sites with KD = 10.8 +/- 0.9 nM and Bmax = 775 +/- 105 fmol/mg protein for (3H) Ro5-4864 and KD = 10.5 +/- 1.1 nM and Bmax = 133 +/- 19 fmol/mg for (3H) diazepam. We were unable to detect any GABA binding site on crude membrane preparation, nor did GABA enhance the binding of (3H) Ro5-4864 or (3H) diazepam. This suggests that benzodiazepine receptors are uncoupled to GABA system on human platelets. Ro15-1788, a specific antagonist for "central type" benzodiazepine (BDZ) binding sites was inactive in displacing (3H) Ro5-4864 from membrane receptors, while PK 11195 (a specific ligand for the "peripheral type" receptor) was the most potent of the drugs tested in inhibiting (3H) Ro5-4864 binding. These results indicate that human blood platelets bear "peripheral-type" BDZ receptor. Moreover, we could not detect any (3H) propyl beta carboline specific binding on platelet membranes. Results on benzodiazepine receptors on human circulating lymphocytes are also reported and similarity in pharmacological properties with platelet benzodiazepine receptors is suggested.     

Correlation of changes in cardiac calcium channels with hemodynamics in Syrian hamster cardiomyopathy and heart failure

We compared hemodynamics with [3H]nitrendipine (calcium channel) binding to cardiac membranes from Bio 14.6 cardiomyopathic Syrian hamsters at 4 and 10 months with their F1B controls. A 50% increase in the number (Bmax) of nitrendipine binding sites (calcium channels) was seen only in the 4 month old myopathic vs controls (Bmax = 468 +/- 11 vs 309 +/- 10 fmol/mg prot with no change in affinity (KD) (KD = .65 +/- .12 vs .75 +/- .14 nM), while no differences in Bmax or KD were seen at 10 months (Bmax = 375 +/- 9 vs 362 +/- 7 fmol/mg prot/KD = .82 +/- .18 vs .89 +/- .17 nM) myopathic vs control respectively. Hemodynamic studies revealed no significant differences in cardiac output, cardiac index, stroke volume, heart rate, mean arterial pressure, peripheral resistance, body weight, heart weight at 4 months, but a significant decrease in peripheral resistance (1120 +/- 360 vs 2080 +/- 240) increase in body weight (118 +/- 2 vs 94 +/- 2 grams) and heart weight (97 +/- 5 vs 78 +/- 2 gms/100 gms body weight) in 10 month myopathic vs control animals. We conclude that the onset of cardiomyopathy at 4 months is associated with a selective increase in calcium channel binding sites and heart failure at 10 months is associated with a relative decrease in these sites.     

Presynaptic Cholinergic Mechanisms in the Rat Cerebellum: Evidence for Nicotinic, but Not Muscarinic Autoreceptors

The present study shows that N-[3H]methylcarbamylcholine ([3H]MCC) binds to a single population of high-affinity/low-density (KD = 5.0 nM; Bmax = 8.2 fmol/mg of protein) nicotinic binding sites in the rat cerebellum. Also, there exists a single class of high-affinity binding sites (KD = 4.8 nM; Bmax = 24.2 fmol/mg of protein) in the cerebellum for the M1 specific muscarinic ligand [3H]pirenzepine. In contrast, the M2 ligand, [3H]AF-DX 116, appears to bind to two classes of binding sites, i.e., a high-affinity (KD = 3 nM)/low-capacity (Bmax = 11.7 fmol/mg of protein) class, and a second class of lower affinity (KD = 28.4 nM) and higher capacity (Bmax = 36.3 fmol/mg of protein) sites. The putative M3 selective ligand [3H]4-diphenylacetoxy-N-methylpiperidine also binds to two distinct classes of binding sites in cerebellar homogenates, one of high affinity (KD = 0.5 nM)/low capacity (Bmax = 19.5 fmol/mg of protein) and one of low affinity (KD = 57.5 nM)/high capacity (Bmax = 140.6 fmol/mg of protein). In experiments which tested the effects of cholinergic drugs on acetylcholine release from cerebellar brain slices, the nicotinic agonist MCC enhanced spontaneous acetylcholine release in a concentration-dependent manner, and the maximal increase in acetylcholine release (59.0-68.0%) occurred at 10(-7) M. The effect of MCC to increase acetylcholine release was Ca2+-dependent and tetrodotoxin-insensitive, suggesting an action on cholinergic terminals. Also, the MCC-induced increase in acetylcholine release was effectively antagonized by dihydro-beta-erythroidine, d-tubocurarine, and kappa-bungarotoxin, but was insensitive to either atropine or alpha-bungarotoxin.(ABSTRACT TRUNCATED AT 250 WORDS)     

D2 Dopamine Receptors on Bovine Chromaffin Cell Membranes: Identification and Characterization by [3H]N-Methylspiperone Binding

Although dopamine-containing cells are known to be present in sympathetic ganglia, the site of action and the role of dopamine in ganglion function remain obscure. In the present work, we evaluated the interaction of dopamine receptor ligands with particulate membrane fractions from bovine chromaffin cells and adrenal medullary homogenates using the D2 dopamine receptor radioligand [3H]N-methylspiperone ([3H]NMSP). Scatchard analysis of [3H]NMSP saturation experiments revealed a Bmax of 24.1 +/- 1.6 fmol/mg of protein and a KD of 0.23 +/- 0.03 nM in the particulate fraction from adrenal medulla homogenates and a Bmax of 26.5 +/- 2.7 fmol/mg of membrane protein and a KD of 0.25 +/- 0.02 nM in the particulate fraction prepared from isolated adrenal chromaffin cells. There were approximately 1,000 receptors/cell. There were no detectable levels of specific [3H]NMSP binding in the particulates prepared from adrenal cortical or capsular homogenates. Competition studies with the nonradioactive D2 receptor antagonists spiperone, chlorpromazine, and (-)-sulpiride revealed KI values of 0.28, 21, and 196 nM, respectively. The (+) isomer of butaclamol displayed a 604-fold higher affinity than the (-) isomer. Competition studies with the dopamine receptor agonists dopamine and apomorphine revealed affinities of 3,960 and 417 nM, respectively. A correlation coefficient of 0.96 was obtained in studies comparing the potencies of drugs in inhibiting specific [3H]NMSP binding in bovine adrenal medullary homogenates and in inhibiting specific [3H]NMSP binding to brain D2 dopamine receptors. In summary, radiolabeling studies using [3H]NMSP have revealed the presence of D2 dopamine receptors on bovine adrenal chromaffin cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

A characterization of beta-adrenergic receptors on cellular and perigranular membranes of rat peritoneal mast cells

Beta-adrenergic receptors were characterized by measuring the specific binding of 3H-dihydroalprenolol (DHA) on intact isolated rat peritoneal mast cells (RPMC) and on perigranular membranes derived from purified RPMC granules. The specific binding of 3H-DHA reaches an equilibrium within 30 min at 5 degrees C and is linear with cell number. Scatchard analysis reveals two populations of binding sites on intact cells: with KD = 10.6 +/- 2.6 and 129 +/- 4.7 nM and Bmax of 186 +/- 38 and 1200 +/- 415 fmol/10(6) cells, respectively. Each cell contains 120 X 10(3) high-affinity binding sites and 720 X 10(3) low-affinity binding sites. There appears to be neither alpha-adrenergic nor muscarinic cholinergic receptors on the RPMC. Specific binding of 3H-DHA also occurred to isolated granules with perigranular membranes. The binding was saturable with a single population of binding sites with an affinity (KD) of 7.0 +/- 0.45 nM. Maximum binding (Bmax) was calculated at 56.6 +/- 1.9 fmol/10(9) granules. Subfractionation of granule components demonstrated that the specific binding sites appear to be localized exclusively on the perigranular membrane.     

Calcium mediates the interconversion between two states of the liver inositol 1,4,5-trisphosphate receptor

D-myo-Inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) regulates intracellular Ca2+ by mobilizing Ca2+ from a non-mitochondrial store. We have investigated the effects of Ca2+ on the binding of [32P]Ins (1,4,5)P3 to permeabilized rat hepatocytes and a liver plasma membrane-enriched fraction. Increasing the free Ca2+ concentration in the medium from 0.1 nM to 0.7 microM increased the capacity of a high affinity binding component (KD = 2-3 nM) in permeabilized cells by a factor of 10. If the membrane fraction was preincubated at 37 degrees C before binding was measured at 4 degrees C, all of the Ins(1,4,5)P3 receptors were transformed to a low affinity state (KD = 65 +/- 12 nM, Bmax = 3.1 +/- 0.1 fmol/mg, n = 4). When 0.7 microM of Ca2+ was added, the receptors were totally transformed to a high affinity state (KD = 2.8 +/- 0.4 nM, Bmax = 2.7 +/- 0.4 fmol/mg, n = 4). The EC50 of the Ca2(+)-induced interconversion of the Ins(1,4,5)P3 receptor was 140 nM. This Ca2(+)-induced transformation of the Ins(1,4,5)P3 receptor from a low affinity to a high affinity state was associated with an inhibition of the Ins(1,4,5)P3-induced Ca2+ release in permeabilized hepatocytes. These data suggest that the Ins(1,4,5)P3-dependent hormones, by increasing the intracellular Ca2+ concentration, induce a reversible transformation of the receptor from its low affinity state, coupled to the Ca2+ release, to a desensitized high affinity state. Transformation of the receptor may play a role in the oscillatory release of Ca2+ observed in single isolated hepatocytes.     

Identification of B2 Bradykinin Binding Sites on Cultured Cortical Astrocytes

Bradykinin was found to bind to specific high-affinity sites in cultured cortical astrocytes from rat brain, and this binding appeared to be specific for the B2 bradykinin receptor subtype. Nonlinear regression analysis of saturation experiments using a computer programme revealed a single KD of 16.6 +/- 2.6 nM and a Bmax of 352.2 +/- 30.7 fmol/mg of protein. These results indicate that astrocytes possess bradykinin receptors and that these are predominantly of the B2 subtype.     

Solubilization of the N-Methyl-d-Aspartate Receptor Channel Complex from Rat and Porcine Brain

The N-methyl-D-aspartate (NMDA) receptor complex as defined by the binding of [3H]MK-801 has been solubilized from membranes prepared from both rat and porcine brain using the anionic detergent deoxycholate (DOC). Of the detergents tested DOC extracted the most receptors (21% for rat, 34% for pig), and the soluble complex, stabilized by the presence of MK-801, could be stored for up to 1 week at 4 degrees C with less than 25% loss in activity. Receptor preparations from both species exhibited [3H]MK-801 binding properties in solution very similar to those observed in membranes (Bmax = 485 +/- 67 fmol/mg of protein, KD = 11.5 +/- 2.9 nM in rat; Bmax = 728 +/- 108 fmol/mg of protein, KD = 7.1 +/- 1.6 nM in pig, n = 3). The pharmacological profile of the solubilized [3H]MK-801 binding site was virtually identical to that observed in membranes. The rank order of potency of: MK-801 greater than (-)-MK-801 = thienylcyclohexylpiperidine greater than dexoxadrol greater than SKF 10,047 greater than ketamine, for inhibition of [3H]MK-801 binding, was observed in all preparations. The receptor complex in solution exhibited many of the characteristic modulations observed in membranes.(ABSTRACT TRUNCATED AT 250 WORDS)     

(+)-[3H]MK-801 Binding Sites in Postmortem Human Brain

The pharmacological specificity and the regional distribution of the N-methyl-D-aspartate receptor-associated 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) binding sites in human postmortem brain tissue were determined by binding studies using (+)-[3H]MK-801. Scatchard analysis revealed a high-affinity (KD = 0.9 +/- 0.2 nM, Bmax = 499 +/- 33 fmol/mg of protein) and a low-affinity (KD = 3.6 +/- 0.9 nM, Bmax = 194 +/- 44 fmol/mg of protein) binding site. The high-affinity site showed a different regional distribution of receptor density (cortex greater than hippocampus greater than striatum) compared to the low-affinity binding site (cerebellum greater than brainstem). The rank order pharmacological specificity and stereoselectivity of the high-(cortex) and low-(cerebellar) affinity binding sites were identical. However, all compounds tested showed greater potency at the high-affinity site in cortex. The results indicate that (+)-[3H]MK-801 binding in human postmortem brain tissue shows pharmacological and regional specificity.     

Adrenergic receptors in the liver parenchyma in children with chronic hepatitis]

In the present study adrenergic receptors have been investigated in liver parenchyma, obtained at the resection of extrahepatic portal hypertension children without parenchymal affection (control group, n-7) and the resection of children in parenchymal affection (group of chronic hepatitis children, n-6). It has been shown, that the binding of beta-adrenergic radioligand 3H-dihydroalprenolol (3H-DHA) in liver parenchyma membranes of both control and chronic hepatitis groups was saturable and showed high affinity. The Scatchard analysis of the binding data indicated that the binding site was characterized by Kd and Bmax of 1.2 +/- 0.5 nM, 261.2 +/- 50 fmol/mg, respectively, for the control group; and 0.9 +/- 0.15 nM, 68.5 +/- 18.8 fmol/mg, respectively, for the group of chronic hepatitis patients; (mean+SEM). The binding of alpha 1-adrenergic antagonist 3H-prazosin (3H-PRZ) in liver parenchyma was also saturable and showed high affinity. The binding site is characterized by Kd = 0.6 +/- 0.12 nM, Bmax = 92.8 +/- 8.0 fmol/mg, for the control group; and Kd = 0.8 +/- 0.15 nM, Bmax = 195.0 +/- 22.0 fmol/mg, for the group of chronic hepatitis. It has been found that the number of binding sites of 3H-DHA significantly decreased and the number of binding sites of 3H-PRZ did not change in chronic hepatitis liver parenchyma in comparison with the control group. The results obtained suggest the important role of beta-adrenergic receptors in the pathogenesis of chronic hepatitis and in liver regeneration in children.     

Binding of the hydrophilic beta-adrenergic antagonist [3H]CGP-12177 to cardiac tissue slices: characterization and ontogenetic studies in dogs

A new technique was developed to characterize the binding of a hydrophilic beta-adrenergic antagonist, [3H]CGP-12177, to 1-mm thick slices of canine cardiac tissue. This technique was used to quantify the density (Bmax) and the affinity (Kd) of these receptors in the right ventricular conus (RVC) and the left ventricle (LV) at day 1 to 6 weeks of age, and in the adult. Binding was found to be reversible, saturable, stereospecific, of high affinity, and thermolabile. There was an increase in the density of beta-adrenergic receptors between day 1 (Bmax = 2.2 +/- 0.3 fmol/mg tissue in RVC and 2.9 +/- 0.8 fmol/mg tissue in the LV) and 2 weeks of age postnatally, after which it remained constant until 6 weeks of age (Bmax = 7.5 +/- 0.4 and 6.8 +/- 0.9 fmol/mg tissue in RVC and LV, respectively); however, by 6 weeks of age it had not reached adult levels (10.3 +/- 1.0 fmol/mg tissue). The affinity of these receptors did not change between early neonatal life (Kd = 1.3 +/- 0.4 nM) and adulthood (Kd = 1.4 +/- 0.2 nM). The density of beta-adrenergic receptors in the RVC was similar to that in the LV. This new method of quantifying beta-adrenergic receptors in cardiac tissue is simple and fast, and requires minimal tissue handling. It proved to be useful in studying the development of cardiac beta-adrenergic receptors with age.     

Receptors for Arg8-vasopressin, angiotensin II, and atrial natriuretic peptide in the mesenteric vasculature of pregnant rats.

Blood pressure and sensitivity of blood vessels to vasoconstrictors are decreased in term-pregnant rats (20-21 days). To determine if changes in receptors for vasoactive peptides could account for these observations, receptor kinetics were measured for Arg8-vasopressin (AVP), angiotensin II (Ang II), and atrial natriuretic peptide (ANP) in the mesenteric vascular bed of the rat throughout pregnancy. Receptors for AVP were statistically similar in the five groups of animals (nonpregnant; pregnant 9, 15, and 21 days; and postpartum). The dissociation constant (KD) for [3H]AVP varied from 0.41 to 0.52 nmol/L (NS), while receptor density (Bmax) varied from 310 +/- 110 to 455 +/- 135 fmol/mg protein for six experimental measurements. Similar observations were made for Ang II receptors where KD of 125I-labelled Sar1, Ile8-Ang II was between 0.60 and 0.97 nmol/L and Bmax between 215 +/- 30 and 250 +/- 40 fmol/mg protein in the different groups. 125I-labelled ANP (101-126) receptors were markedly modified in terms of number of sites. Bmax was significantly increased during pregnancy (9 days, 429 +/- 86; 15 days, 541 +/- 54; 20 days, 438 +/- 72) and decreased in the postpartum period (133 +/- 21) by comparison with the nonpregnant group (245 +/- 35 fmol/mg protein), while KD was similar in the different experimental groups (57 to 82 pmol/L). Despite these increases in receptor density, the vasorelaxant effects of ANP was only increased at 9 days of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)     

3H]mepyramine binding to histamine H1 receptors in bovine retina

The promethazine-sensitive [3H]mepyramine binding was used to determine the presence of histamine H1 receptors in membranes from bovine retina. Specific mepyramine binding to retinal membranes was reversible, saturable and of high affinity. The apparent dissociation constant (KD = 2.2 +/- 0.4 nM) and the density of binding sites (Bmax = 60.9 +/- 5.1 fmol/mg protein), obtained in equilibrium studies, were similar to those found in bovine brain cortex. Binding was stereospecific and the inhibitory potencies of H1 and H2 antagonists indicated that [3H] mepyramine binding sites in the retina have characteristics of H1 receptors.