Role of Tyr(356(7.43)) and Ser(190(4.57)) in antagonist binding in the rat beta1-adrenergic receptor.

Site-directed mutagenesis and photoaffinity labeling experiments suggest the existence of at least two distinct binding orientations for aryloxypropanolamine competitive antagonists in the beta-adrenergic receptor (beta-AR), one where the aryloxy moiety is located near transmembrane alpha-helix 7 (tm 7) and another where it is near tm 5. To explore a hydrophobic pocket involving tms 1, 2, 3, and 7 for potential aryloxy interaction sites, we selected Tyr(356(7.43)) and Trp(134(3.

Agonist binding and activation of the rat beta(1)-adrenergic receptor: role of Trp(134(3.28)), Ser(190(4.57)) and Tyr(356(7.43)).

We investigated the role of Trp(134(3.28)), Ser(190(4.57)) and Tyr(356(7.

Site-directed mutagenesis of the rat beta1-adrenoceptor. Involvement of Tyr356 (7.43) in (+/-)cyanopindolol but not (+/-)[125Iodo]cyanopindolol binding.

To determine the role played by Tyr(356 (7.43)) in the rat beta(1)-adrenoceptor in binding the antagonists (+/-)cyanopindolol (4-[3-(t-butylamino]-3-(2'-cyano-indoloxy)-2-propanolol) and its iodinated analogue (+/-)[(125)Iodo]cyanopindolol (1-(t-butylamino]-3-(2'-cyano-3'-iodo-indoloxy)-2-propanolol), Tyr(356 (7.43)) was mutated to either Phe or Ala and binding affinities determined for wild type and mutant rat beta(1)-adrenoceptors.

Synthesis, beta-adrenoceptor pharmacology and toxicology of S-(-)-1-(4-(2-ethoxyethoxy)phenoxy)-2-hydroxy-3-(2-(3,4-dimethoxyphenyl)ethylamino)propane hydrochloride, a short acting beta(1)-specific antagonist.

The synthesis of S-(-)-1-(4-(2-ethoxyethoxy)phenoxy)-2-hydroxy-3-(2-(3,4-dimethoxyphenyl)ethylamino)propane hydrochloride (D140S.HCl 6), a novel short acting beta(1)-specific adrenoceptor antagonist, has been described. The antagonist potency for D140S.

CoMFA analysis of the human beta(1)-adrenoceptor binding affinity of a series of phenoxypropanolamines.

A series of 36 phenoxypropanolamines was examined to determine the structure--activity relationships of beta-adrenoceptor (beta-AR) antagonists for the human beta(1)-AR. The binding affinities of all the compounds were determined for human beta(1)-ARs expressed in Chinese hamster ovary cells and the antagonist potency for rat atrial beta(1)-ARs was determined for 32 of these compounds for comparative purposes. The compounds, based upon a phenoxypropanolamine core structure with various meta-, ortho-, para- and amine-substituents, displayed binding affinities (pK(i)) for the human beta(1)-AR ranging from 5.

Role of beta-adrenergic receptor subtypes in lipolysis.

In vitro lipolysis stimulated by low (-)-isoprenaline concentrations (< or =30 nM) in epididymal white adipocytes from Sprague-Dawley rats was inhibited at least 60-80% by the specific beta1-antagonists LK 204-545 and CGP 20712A (1 microM), suggesting that at these low (10 nM) concentrations of (-)-isoprenaline lipolysis was primarily (80%) but not solely mediated via beta1-adrenergic receptors. Low concentrations (100 nM) of (-)-noradrenaline and formoterol also confirmed a role for beta1-adrenergic receptors in mediating lipolysis at low concentrations of these agonists. At higher agonist concentrations, beta3-adrenergic receptors were fully activated and were the dominant beta-adrenergic receptor subtype mediating the maximum lipolytic response, and the maximum response was not affected by the beta1-antagonists, demonstrating that the beta3-receptor is capable of inducing maximum lipolysis on its own.

beta(1)- and beta(2)-Adrenoceptor antagonist activity of a series of para-substituted N-isopropylphenoxypropanolamines.

To further explore the structure-activity relationships of beta-adrenoceptor (beta-AR) antagonists, a series of 25 para-substituted N-isopropylphenoxy-propanolamines were synthesised, nine of which are new compounds. All have been examined for their ability to antagonise beta(1)-ARs in rat atria and beta(2)-ARs in rat trachea. Substitution in the para-position of the phenyl ring is thought to confer beta(3)-specificity and the selectivity of these compounds for the beta(1)-AR ranges from 1.

LK 204-545, a highly selective beta1-adrenoceptor antagonist at human beta-adrenoceptors.

LK 204-545 ((+/-)-1-(2-(3-(2-cyano-4-(2-cyclopropyl-methoxy-ethoxy)phenoxy)-2-hydro xy-propyl-amino)-ethyl)-3-(4-hydrxy-phenyl) urea), an antagonist that possesses high beta1-/beta2-selectivity in the rat, and a range of cardio-selective and non-selective beta-adrenoceptor antagonists were examined to compare their radioligand binding affinities for human beta1-, beta2- and beta3-adrenoceptors transfected into CHO cells. LK 204-545 and CGP 20712A displayed the highest beta1-/beta2- (approximately 1800 and approximately 650, respectively) and beta1-/beta3-selectivity (approximately 17000 and approximately 2200, respectively) at human beta-adrenoceptors with LK 204-545 being approximately 2.75-fold more beta1-/beta2-selective and approximately 8-fold beta1-/beta3-selective than CGP 20712A.

Pharmacology and subcellular distribution of [3H]rilmenidine binding sites in rat brain.

We have previously reported that in rat brain membranes, [3H]rilmenidine, in addition to labelling alpha2-adrenoceptors and the I2B-subtype of imidazoline receptor binding site (I2B-RBS), may label an additional I-RBS population, distinct from previously classified I1-RBS and I2-RBS. In this study, using crude or fractionated rat brain membranes we examined the possible association of [3H]rilmenidine-labelled I-RBS with the A- and B-isoforms of monoamine oxidase (MAO) by studying the inhibition of [3H]rilmenidine binding by a number of MAO inhibitors; and comparing the maximal binding density (Bmax) and subcellular distribution of [3H]rilmenidine binding sites with that of MAO-A and MAO-B catalytic sites labelled by [3H]RO41-1049 and [3H]RO19-6327 and 12-RBS labelled by [3H]2-BFI. Inhibition of [3H]rilmenidine binding by all MAO inhibitors tested produced very shallow curves (slope 0.

[3H]Rilmenidine-labelled imidazoline-receptor binding sites co-localize with [3H]2-(benzofuranyl)-2-imidazoline-labelled imidazoline-receptor binding sites and monoamine oxidase-B in rabbit, but not rat, kidney.

The distribution and relative densities of imidazoline-receptor binding sites (I-RBS) and monoamine oxidase (MAO)-A and -B enzyme(s) in rat and rabbit kidney were compared autoradiographically using fixed nanomolar concentrations of [3H]rilmenidine and [3H]2-(benzofuranyl)-2-imidazoline ([3H]2-BFI) to label I-RBS, and [3H]RO41-1049 and [3H]RO19-6327 to label MAO-A and -B isoenzymes, respectively. In rat kidney, high densities of I-RBS labelled by [3H]rilmenidine were observed in the cortex and outer stripe (120-280 fmol/mg tissue), in contrast to low I-RBS densities labelled by [3H]2-BFI (<4 fmol/mg). A relatively high density of [3H]RO41-1049 binding to MAO-A enzyme was present in all regions of the rat kidney (160-210 fmol/mg) compared with a low density of [3H]RO19-6327 binding to MAO-B (< 25 fmol/mg).

The effect of levcromakalim (BRL 38227) on bladder function in patients with high spinal cord lesions.

The effects of the potassium channel opener levcromakalim (BRL 38227) 7.5 micrograms kg-1 were examined on urodynamic variables and blood pressure during inflow and voiding cystometry in six high spinal cord lesion patients. Levcromakalim administration significantly increased the duration of bladder contraction (197 +/- 128 s to 267 +/- 167 s, P < 0.

A simple and reliable method for construction of parallel multibarrel microelectrodes.

A modification to the method of construction of parallel or "piggy-back" electrodes for extracellular single-unit recording combined with iontophoresis is described that facilitates the alignment of the two components of the array. The method involves the use of an orthogonal viewing device (a pair of mirrors mounted symmetrically at 45 degrees to the horizontal), which produces a pair of virtual images of the electrode components that can be viewed with a microscope. A slight displacement between the electrodes is easily detected as an uneven separation between the electrode images.

Validation of a solid-phase extraction high-performance liquid chromatographic assay for doxazosin.

The determination of doxazosin by high-performance liquid chromatography with fluorescence detection is described. Propanolol was used as the internal standard. Plasma samples were treated with methanol to precipitate the proteins.

Diurnal blood pressure variation in quadriplegic chronic spinal cord injury patients.

1. Measurement of blood pressure and heart rate over a 24 h period was performed in 10 quadriplegic spinal cord injury patients and 10 immobilized, neurologically intact orthopaedic subjects by using the Spacelabs 90207 automated ambulatory monitoring system. 2.

Differential forskolin activation of rat heart and lung adenylate cyclase. Dependence on membrane-protein interactions.

We have investigated whether the greater ability of forskolin to activate adenylate cyclase (EC 4.6.1.

Forskolin-mediated activation of cardiac, liver and lung adenylate cyclase in the rat. Relation to [3H]forskolin binding sites.

We investigated whether differences in binding sites for [3H]forskolin could account for the low potency of forskolin on adenylate cyclase (EC 4.6.1.

Role of auto-inhibitory feed-back in cardiac sympathetic transmission assessed by simultaneous measurements of changes in 3H-efflux and atrial rate in guinea-pig atrium.

Guinea-pig right atria were labelled with [3H]-noradrenaline or [3H]-dopamine before superfusion in a flow-cell. Choice of label did not significantly alter either the relationship between 3H-efflux and number of electrical field pulses or the inhomogeneity of labelling. The relationship between 3H-efflux and frequency of 4 field pulses (0.

Role of autoinhibitory feedback in cardiac sympathetic transmission.

The relationship between two indices of transmitter release measured simultaneously and the frequency of 4 field pulses (0.125-2 Hz) were obtained from superfused guinea pig right atria after labelling with 3H-noradrenaline. The relationships between 3H-efflux or rate responses and frequency were hyperbolic.

A simple method for the assay of urinary metanephrines using high performance liquid chromatography with fluorescence detection.

A method is described which extends a high performance liquid chromatographic assay for urinary catecholamines to the assay of urinary metanephrines. The amines are separated from urine after acid hydrolysis of conjugates by ion-exchange chromatography, and then further purified by solvent extraction. The final extracts are suitable for direct HPLC assay using the endogenous fluorescence of the amines for detection.