A study on the localization of [H]sulpiride binding sites in rat striatal membranes

The specific binding of [³H]sulpiride to purified rat striatal synaptic membranes has been examined. This binding component was unchanged in rats with 6-hydroxydopamine-induced lesions but was significantly reduced (36%) in 24 hr kainate-lesioned, and (53%) in rats with cortical ablations (28 days postoperative). These results suggest that specific [³H]sulpiride binding sites are present on both slriatal interneurones and on the cortico-striatal pathway. This conclusion is inconsistent with the present classification of D₁ and D₂ receptors.     

Characterization of [3H]zetidoline binding to rat striatal membranes

The binding of [3H]zetidoline, a novel neuroleptic agent, to rat brain striatal membranes was investigated in-vitro. The optimal binding conditions for [3H]zetidoline differed from those for [3H]spiperone in pH, temperature and time. [3H]Zetidoline has high affinity for striatal dopamine receptors. Its binding is saturable, stereo-specific, has a low non-specific component and is reversible and tissue specific. The Scatchard analysis gave a biphasic curve, indicating that [3H]zetidoline interacts with more than one population of receptor sites (B'max = 67 fmol mg-1 protein, K'd = 0.11 nM; B"max = 500 fmol mg-1 protein, K'd = 2.49 nM). Kinetic analysis of rates of association and dissociation yielded a Kd value in agreement with that measured at equilibrium. Inhibition studies indicated that only dopamine and dopaminergic agents are able to displace [3H]zetidoline from its binding sites, and in a different rank order from that for displacement of [3H]spiperone. (-)-Sulpiride was especially effective in inhibiting [3H]zetidoline specific binding. Furthermore, like that of [3H]benzamides, [3H]zetidoline binding appears to be highly Na+-dependent and Li+ only partially substitutes Na+.     

The binding of [3H]N-(chloroethyl)-norapomorphine to rat striatal membranes

On the stretch-induced discharge activity of the isolated crayfish sensory neuron flurazepam (less than or equal to 3 X 10(-4) M) and Ro 15-1788 (less than or equal to 10(-3) M) produced reversible concentration-dependent excitation, but oxazepam only produced depression (less than or equal to 5 X 10(-4) M). Similar divergent effects on the membrane properties were observed. Oxazepam increased the threshold to firing without changing resting potential, membrane resistance or the GABA-mediated IPSP. In contrast flurazepam and Ro 15-1788 produced a concentration-dependent decrease in threshold. Flurazepam did not alter membrane resistance or resting potential but facilitated GABA transmission. Ro 15-1788 had the opposite effect on the GABA synapse, and also depolarized the resting potential but did not alter membrane resistance. The change in spike threshold appeared to be an important component in producing discharge excitation or depression. These results not only demonstrate the capability of the sensory neuron to discriminate between structures of benzodiazepines, but also that these agents can produce divergent effects on synaptic and non-synaptic properties of a single neuron.     

Rapid in vitro modulation of [3H]hemicholinium-3 binding sites in rat striatal slices

The effects of depolarizing concentrations of potassium chloride on the modulation of [3H]hemicholinium-3 binding sites and high affinity choline uptake were examined in vitro. When rat striatal slices were incubated in Krebs buffer for 20 min, [3H]hemicholinium-3 binding sites diminished to 60% of binding measured in fresh un-incubated tissue, and remained stable for 60 min. Upon addition of Krebs buffer containing 40 mM KCl, the number of binding sites increased during a 20 min period, and remained stable for 40 min. Changes in [3H]hemicholinium-3 binding sites closely paralleled changes in high affinity choline uptake. Scatchard analysis revealed that changes in binding result from alterations in the number of binding sites (Bmax), and not in the affinity (KD). These results suggest that neuronal depolarization rapidly alters the velocity of choline transport into cholinergic neurons by increasing the number of available carriers.     

Multiple binding sites of [3H]clonidine on hepatic plasma membranes

The binding of [3H]clonidine on mouse liver plasma membrane was a rapid, saturable and reversible process. It was characterized by two types of population: high affinity receptors with KD of 6.76 +/- 1.02 nM and Bmax of 106.15 +/- 24.05 fmol/mg protein, and low affinity receptors with KD of 63.66 +/- 12.85 nM and Bmax of 818.06 +/- 128.49 fmol/mg protein. Displacement of [3H]clonidine from its binding sites by various ligands indicated that alpha 1--as well as alpha 2--adrenoceptors were involved in the high affinity system. The respective participation of these two types of receptors was discussed.     

Paraxanthine displaces the binding of [3H]SCH 23390 from rat striatal membranes

We present evidence showing that paraxanthine (1,7-dimethylxanthine), the main metabolite of caffeine in man, displaces the binding of [3H]SCH 23390, a radioligand which selectively labels dopamine D-1 receptors when used at low concentrations, from striatal membranes of the rat. The displacement was competitive and indicated the existence of two affinity states (Hill coefficient = 0.49; K(high) = 0.15 microM; K(low) = 95.9 microM, %R(high) = 32.4). When the stable GTP analog Gpp(NH)p was included, the displacement curve indicated the presence of only the low-affinity state (Hill coefficient = 1.16; Ki = 72.1 microM). However, paraxanthine did not displace the specific binding of [3H]spiperone. After injection of 30 mg/kg s.c. of caffeine, a maximum of 10 microM of paraxanthine was found in striatal homogenates, which could be sufficient to occupy dopamine D-1 receptors. Our results suggest that a dopaminergic action of paraxanthine could be involved in the behavioural stimulation produced by caffeine.     

3H]DTG and [3H](+)-3-PPP label pharmacologically distinct sigma binding sites in guinea pig brain membranes.

The interaction of various compounds with sigma binding sites was examined in membranes prepared from whole guinea pig brain. Whereas [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine labeled a single population of binding sites exhibiting a Kd of 43 nM, [3H]1,3-di-o-tolylguanidine bound to two sites having Kds of 35 and 212 nM, and to a greater maximum number of sites than [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine. Haloperidol, 1,3-di-o-tolylguanidine, BMY 14802, and (-)-pentazocine each displayed nearly equal affinity for binding sites labeled by [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine and [3H]1,3-di-o-tolylguanidine, whereas (+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine was 3 times more potent in inhibiting [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine than [3H]1,3-di-o-tolylguanidine binding. In contrast, (+)-SKF 10,047, (+)-cyclazocine and (+)-pentazocine exhibited more than 9-fold higher affinity for [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine than [3H]1,3-di-o-tolylguanidine binding sites. Dextromethorphan was 15-fold more potent against [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine than [3H]1,3-di-o-tolylguanidine, inhibited [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine binding in a biphasic manner, and inhibited [3H]haloperidol and [3H](+)-SKF 10,047 binding with potencies similar to those obtained against [3H]1,3-di-o-tolylguanidine and [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine, respectively. Phenytoin increased [3H](+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine and [3H](+)-SKF 10,047 binding, but did not enhance [3H]1,3-di-o-tolylguanidine or [3H]haloperidol binding. However, the potency of dextromethorphan to inhibit [3H]1,3-di-o-tolylguanidine binding was increased in the presence of phenytoin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Binding of [3H]ADTN to rat striatal membranes

The conformationally restricted dopamine analogue ADTN binds in a specific saturable manner to rat striatal membranes. Analysis of the data suggests a single binding site. Binding of [3H]ADTN is displaced by a wide range of dopamine agonists and antagonists (both typical and atypical). The potency of ADTN derivatives to displace [3H]ADTN correlates well with their activity as agonists in other tests both in vivo and in vitro.     

Chemical heterogeneity of [3H]imipramine binding sites on human platelet membranes

Functional groups essential for high- and low-affinity [3H]imipramine (IMI) binding were determined by the method of chemical modification. The high-affinity recognition sites contained cysteine and lysine amino acid residues, but not aspartic or glutamic acid residues. The low-affinity recognition sites contained only cysteine residues. Moreover, probably only part of these sites contained these residues. The arginine, tyrosine and histidine residues are not likely to be functionally important for the [3H]IMI binding process. Analysis of the structure-function interaction of drug molecules reveals that, for all substances with high displacement ability, there is a conformation in which they can react with high-affinity IMI recognition sites. Data obtained allowed us to construct a tentative structure model of the high-affinity recognition IMI binding site.     

3H]GBR-12935 binding sites in human striatal membranes: binding characteristics and changes in parkinsonians and schizophrenics

The binding of the diphenyl-substituted piperazine derivative, [3H]GBR-12935, a selective dopamine uptake inhibitor, to the post-mortem human putamen was studied. Inhibition curves by dopamine uptake inhibitors suggested the existence of two populations of [3H]GBR-12935 binding sites: one is potently inhibited by mazindol and/or nomifensine, and the second binding site is benztropine- and/or GBR 12909-sensitive. In the human putamen, [3H]GBR-12935 labeled both these two distinct binding sites. The [3H]GBR-12935 binding displaced by mazindol was enriched in the mouse and rat striatum, but not in the cultured mouse neuroblastoma cell N1E-115. The mazindol-sensitive [3H]GBR-12935 binding site increased in the presence of sodium and markedly decreased in the putamen from parkinsonians (45% of controls). On the other hand, the [3H]GBR-12935 binding displaced by benztropine showed no sodium-dependent increase and was not decreased in the putamen from parkinsonians. In the putamen from schizophrenics, the [3H]GBR-12935 binding did not significantly change in the density, while that displaced by mazindol tended to increase. It is concluded that in the human putamen, [3H]GBR-12935 binds to two distinct sites. One site is partially sodium-dependent and appears to be associated with a high-affinity dopamine uptake system on dopaminergic nerve terminals. The second binding site shows no sodium-dependency and may be associated with a nondopaminergic and/or extraneuronal DA uptake system.     

Localisation of [3H]clonidine binding to membranes from guinea pig renal tubules

The selective radioligand [3H]clonidine has been used to localise alhpa 2 adrenoceptors in guinea pig kidney. Chemical sympathectomy with 6-hydroxydopamine produced no significant change in the number of sites labeled by [3H]clonidine indicating that the majority of binding sites were not located on sympathetic nerve terminals. Binding was enhanced in membranes prepared from renal tubules and considerably reduced in preparations from glomeruli. Subcellular fractions of renal cortex revealed that binding was to plasma membranes and that the greatest binding capacity was present in the fraction rich in basal lateral membranes. It is concluded that the major concentration of renal alpha 2 adrenoceptors are present on renal tubules and that they may be localised to a particular pole of the renal tubule cell.     

Estradiol-induced increase of specific [3H]ketanserin binding sites on rat uterine membranes

[3H]Ketanserin, a specific serotonin (5-HT) antagonist, was used to investigate whether 5-HT receptors increased in the uterine membranes of ovariectomized rats on administration of 17 beta-estradiol-3-benzoate (estradiol) and also to investigate the characteristics of specific [3H]ketanserin binding to the uterine membranes from estradiol-treated ovariectomized rat. Administration of estradiol significantly increased the amount of [3H]ketanserin specifically bound at equilibrium but did not change the apparent affinity of specific [3H]ketanserin binding. The specific [3H]ketanserin binding to estradiol-treated ovariectomized preparations was rapid and reversible. The Scatchard plots of the saturation curves of specific [3H]ketanserin binding to untreated and estradiol-treated ovariectomized preparations were convex. The apparent Ki values of various serotonergic agents deduced from displacements by these compounds of specific [3H]ketanserin binding to estradiol-treated ovariectomized preparations were two to four orders of magnitude smaller than those of adrenergic, dopaminergic and histaminergic agents. These results suggest that [3H]ketanserin binds mainly to 5-HT receptors in the uterine membranes of estradiol-treated ovariectomized rats.     

(+)-[3H]SKF 10,047, (+)-[3H]ethylketocyclazocine, mu, kappa, delta and phencyclidine binding sites in guinea pig brain membranes

Binding of the opiates (+)-[3H]SKF 10,047 [N-allylnormetazocine; (+)-[3H]SKF] and (+)-[3H]ethylketocyclazocine [(+)-[3H]EKC] were compared to mu, kappa and delta and phencyclidine (PCP) receptor binding in guinea pig brain membranes. (+)-[3H]SKF and (+)-[3H]EKC binding were not blocked by naloxone, and had different drug selectivity compared to mu, kappa and delta binding sites. The number of binding sites, drug selectivity and region distribution in brain were similar for (+)-[3H]SKF and (+)-[3H]EKC. Sigma opiates that are associated with psychotomimetic activities, such as pentazocine, cyclazocine, SKF 10,047 and bremazocine, were potent inhibitors of (+)-[3H]SKF and (+)-[3H]EKC binding. Haloperidol was the most potent inhibitor of (+)-[3H]SKF binding. Haloperidol and sigma opiates demonstrated biphasic displacement of [3H]PCP binding, suggesting that [3H]PCP labelled two sites. PCP had a similar affinity for both (+)-[3H]SKF and [3H]PCP binding sites in the presence of 100 mM NaCl. The highest concentrations of (+)-[3H]SKF and (+)-[3H]EKC bindings sites were in the hypothalamus, anterior pituitary, midbrain, pons and medulla.