Receptors In Rat Striatum Research Articles

Striatal dopamine receptor sensitivity after subchronic fencamfamine in the rat

Dopamine (DA) receptor sensitivity was assessed in the rat striatal system following subchronic treatment with fencamfamine or saline for 7 days (10 mg/kg i.p.). Seventy-two hours after the last injection the stereotyped behaviour and general activity induced by apomorphine or saline were evaluated. Apomorphine (2.0 mg/kg s.c.) induced a decrease of the stereotypic response when fencamfamine-pretreated animals were compared to saline-treated ones while apomorphine (0.02 mg/kg s.c.) failed to alter the general activity of animals, treated with fencamfamine or not. In biochemical experiments subchronic fencamfamine did not alter the effects of apomorphine (0.02 mg/kg s.c.) in reducing homovanillic acid (HVA) and dihydroxyphenylacetic acid (DOPAC) striatal levels, when compared to saline. In addition we observed a slight but significant reduction in the total dopamine receptor content in the striatum labelled by [ 3H]spiroperidol. These findings indicate that subchronic fencamfamine treatment leads to a desensitization of postsynaptic DA receptors in rat striatum.

Effects of pertussis toxin on D 2-dopamine receptor in rat striatum: Evidence for coupling of Ni regulatory protein with D 2-receptor

It is well established that a pertussis toxin, islet activating protein (IAP), interacts directly with the Ni regulatory protein involved in the receptor-adenylate cyclase system. In this study we investigated the effect of the toxin on the dopaminergic function of the central nervous system in conjunction with the adenylate cyclase system. Direct bilateral microinjection of the toxin into rat striatum reduced the stereotyped behavior induced by apomorphine. This inhibitory effect was observed even 40 h after administration of the toxin, whereas the inhibition by haloperidol disappeared within 15 h. Toxin administration did not influence either the Bmax or affinity of specific binding of [3H]spiroperidol to the striatal membrane. However, it increased the IC50 value of apomorphine for the specific binding of [3H]spiroperidol. GTP (10(-4) M) had little effect on the apparent affinity of apomorphine to the [3H]spiroperidol binding sites in IAP-treated membrane, though an effect was clearly observed in untreated membrane. [3H]Spiroperidol binding sites were increased 34 +/- 8% (n = 4) on chronic treatment of haloperidol for 2 weeks. On the contrary, on long-term administration of IAP the ligand binding sites were decreased by 25 +/- 10% (n = 4). These results indicate that pertussis toxin can interact with the Ni protein coupled with striatal D2-dopamine receptor. Inhibition of the coupling between Ni protein and the D2-dopamine receptor attenuated the manifestation of rat stereotyped behavior. Furthermore, chronic administration of dopamine antagonist resulted in the up-regulation of the D2-dopamine receptor, while long-term inhibition of coupling between the D2-dopamine receptor and Ni regulatory protein reduced the amount of receptors.

Binding of a novel dopaminergic agonist radioligand [ 3H]-fenoldopam 1 (SKF 82526) to D-1 receptors in rat striatum

Fenoldopam (SKF 82526), a dopamine agonist which exhibits D-1 receptor subtype selectivity, was evaluated as a radioligand for this receptor subtype. In saturation studies in rat striatal membrane preparations, [ 3H]-fenoldopam appeared to label a single binding site with a K D of 2.3 ± 0.1 nM and a B max of 590 ± 40 fmoles/mg protein. In competition binding experiments, binding was shown to be stereoselective, and rank ordering of affinities of dopaminergic and non-dopaminergic compounds closely correlated with potencies of these compounds in stimulating or inhibiting dopamine-sensitive adenylate cyclase (D-1) and in binding to D-1 sites labelled with the antagonist [ 3H]-cis-flupenthixol. The most potent competitors were the recently identified D-1 selective antagonists, SCH 23390 and SKF R-83566. [ 3H]-Fenoldopam was also used to assess agonist/D-1 receptor interactions. The results suggest that [ 3H]-fenoldopam is a useful and selective agonist radioligand for the D-1 receptor.

P2B0222) - Effect of the oxotremorine-induced down-regulation on biochemical parameters involved in protein synthesis system of muscarinic acetylcholine receptor in rat striatum.

P2B0222) - Effect of the oxotremorine-induced down-regulation on biochemical parameters involved in protein synthesis system of muscarinic acetylcholine receptor in rat striatum.

Effects of alloxan-induced diabetes on dopaminergic receptors in rat striatum and anterior pituitary.

The binding of [3H]-spiroperidol after 4 weeks of hyperglycemia was determined in the rat striatum and anterior pituitary. Alloxan-induced diabetes increased the number of dopaminergic binding sites in the striatum but not in the anterior pituitary. The interaction of metoclopramide with striatal dopaminergic receptors was slightly modified, while that of dopamine, bromocriptine and haloperidol was unaffected. These results suggest that chronic hyperglycemia exerts selective effects on nigrostriatal dopaminergic system in the rat.

Distinct target size of dopamine D-1 and D-2 receptors in rat striatum

Frozen rat striatal tissue was exposed to 10 MeV electrons from a linear accelerator. Based on the theory of target size analysis, the molecular weights of dopamine D-1 receptors (labelled by 3H-piflutixol) and dopamine D-2 receptors (labelled by 3H-spiroperidol) were 79,500 daltons and 136,700 daltons, respectively. The size of the dopamine-stimulated adenylate cyclase was 202,000 daltons. The estimated molecular sizes were deduced by reference to proteins with known molecular weights which were irradiated in parallel. The results showed that the molecular entities for 3H-piflutixol binding and 3H-spiroperidol binding were not identical. The present results do not allow conclusions as to whether D-1 and D-2 receptors are two distinct proteins in the membrane, or whether the receptors are located on the same protein. In the latter case the binding of 3H-spiroperidol needs the presence of a second molecule.

3H]5?-N-ethylcarboxamide adenosine binds to both Ra and Ri adenosine receptors in rat striatum

Adenosine analogs such as 5'-N- ethylcarboxamide adenosine and N6-cyclohexyladenosine stimulate or inhibit adenosine cyclase activity in preparations of rat striatum depending on the assay conditions. N6-cyclohexyladenosine inhibits but does not stimulate adenosine cyclase activity in preparations of hippocampus. These findings suggest that the striatum contains both Ra (stimulatory) and Ri (inhibitory) adenosine receptors while the hippocampus contains only Ri receptors. We have previously shown that [3H]N6-cyclohexyladenosine binds to Ri receptors in rat hippocampus ( Yeung and Green 1983). Comparisons of the characteristics of [3H]5'-N- ethylcarboxamide adenosine and [3H]N6-cyclohexyladenosine binding to hippocampus show that [3H]5'-N- ethylcarboxamide adenosine also binds to Ri receptors with high affinity. [3H]5'-N- ethylcarboxamide adenosine binds to Ri receptors in the striatum and to a second site that is present in striatum but not hippocampus. High affinity binding of both ligands to Ri receptors can be blocked by treatments with N-ethylmaleimide that do not markedly affect [3H]5'-N- ethylcarboxamide adenosine binding to the second site in the striatum. The pharmacological characteristics of the second site indicate that it is the Ra adenosine receptor.

3]5?-N-ethylcarboxamide adenosine binds to both Ra and Ri adenosine receptors in rat striatum

3]5?-N-ethylcarboxamide adenosine binds to both Ra and Ri adenosine receptors in rat striatum

Differential affinity of dopaminergic agonists and antagonists for D 1 and D 2 dopamine receptors in rat striatum

Dopamine (DA) receptors have been classified into two types, D1 and D2, on the basis of their pharmacological specifities and localization. We now report that lesions of striatal dopaminergic neurons by kainic acid (KA) injection or cerebral neocortex (CN) ablation, differentially affect the potency of DA agonists and antagonists in displacing 3H-spiroperidol specific binding. In particular, agonists show a preferential affinity in displacing 3H-spiroperidol from those receptors situated on intrastriatal neurons, following CN lesion. On the other hand, antagonists show a higher displacing activity for those receptors mainly located on cortical terminals following KA lesion. These data support the hypothesis of the existence of different types of DA receptors with a differential anatomical location.

Inhibition of dopamine-activated adenylate cyclase and dopamine binding by opiate receptors in rat striatum.

Low-affinity (micromolar) 3H-dopamine binding was measured under conditions which permitted dopamine activation and opiate inhibition of adenylate cyclase in rat striatal membranes. Opiate drugs and peptides inhibited the dopamine binding in the presence of both GTP and Gpp(NH)p. Opiate inhibition of adenylate cyclase was, however, observed only in the presence of GTP. It is suggested that the dopamine D1 receptor in striatum may be modulated by the opiate delta receptor through a shared guanine nucleotide binding subunit.

3H]dopamine binds to D-1 and D-2 receptors in rat striatum

Although saturation studies suggest that [ 3H]dopamine binds to a homogeneous class of stereospecific sites in rat striatal membranes, pharmacologic analysis reveals two distinct sites, one with high and another with low antagonist affinities. The affinity of antagonists for the first site is correlated with their affinity for [ 3H]butyrophenone binding sites; affinity for the second site is correlated with inhibitory potency against the dopamine-stimulated adenylate cyclase. These result suggest that [ 3H]dopamine binds to D-1 and D-2 receptors.

Selective changes in the density of ß1-adrenergic receptors in rat striatum following chronic drug treatment and adrenalectomy

The corpus striatum has a high density of ß-adrenergic receptors though it appears to contain low levels of ß-hydroxylated catecholamines. In an attempt to determine whether these receptors normally receive an endogenous input, the densities of ß 1 and ß 2-adrenergic receptors in rat caudate have been measured following adrenalectomy and after various pharmacological manipulations. Chronic administration of either pargyline, an inhibitor of mononamine oxidase activity, or desmethylimipramine, an inhibitor of norepinephrine uptake, resulted in a 20–25% decrease in the density of ß 1-adrenergic receptors while either adrenalectomy or the chronic adminitsration of the non-selective ß-adrenergic receptor antagonist propranol resulted in small but significant increases in the density of ß 1-receptors. These treatments did not lead to significant changes in the density of ß 2-receptors. It thus appears that the density of ß 1-receptors in the caudate is normally affected by changing levels of endogenous catecholamines.

Selective labelling of dopamine (D2) receptors in rat striatum by [3H]domperidone but not by [3H]spiperone

Specific binding of [3H]spiperone and [3H]domperidone, displaceable by 1 microM d-butaclamol, was examined in rat striatal membranes. Initial saturation and displacement experiments indicated that [3H]spiperone bound to more sites than [3H]domperidone and that, whilst all displacing drugs were more potent against [3H]domperidone, this difference in potency was greatest for dopamine agonists and specific antagonists and least for 5HT-related drugs. Sulpiride displaced [3H]spiperone biphasically, and was used at a concentration of 50 microM to examine two classes of [3H]spiperone binding: site 1 displaceable by sulpiride, and site 2 displaceable by butaclamol but not by sulpiride. Site 1 had twice the capacity of site 2 and ten times the affinity for [3H]spiperone. Dopaminergic drugs displaced preferentially from site 1, whilst 5HT-related drugs were more potent against site 2. GTP reduced the potency of dopamine, noradrenaline and, to a lesser extent, 5HT at site 1, but had no effect at site 2. [3H]Domperidone sites had the same capacity as [3H]spiperone site 1, and dopamine, noradrenaline and 5HT, in the absence or presence of GTP, and sulpiride had essentially identical affinities for [3H]domperidone sites and [3H]spiperone site 1. It is concluded that [3H]domperidone and [3H]spiperone label an identical population of dopamine (D2) receptors, whilst [3H]spiperone also labels a substantial number of non-dopamine sites, at least some of which are 5HT-related. [3H]spiperone also labels a substantial number of non-dopamine sites, at least some of which are 5HT-related. [3H]Domperidone is the better radioligand for dopamine receptors.

Effects of proteolytic enzymes and monovalent ions demonstrate proteasesensitive and protease-insensitive stereospecific binding sites on dopaminergic receptors in rat striatum

Rat striatal membranes from different subcellular fractions were treated with various proteolytic and other enzymes and the binding of a dopamine agonist ([ 3H]NPA) and of an antagonist ([ 3H]haloperidol) was assayed in several conditions. In membranes of striatal microsomal and mitochondrial fractions, stereospecific binding of both [ 3H]NPA and [ 3H]haloperidol assayed in a monovalent ion-poor buffer was potently and rapidly inhibited by trypsin and certain related proteases. The enzymes did not affect the binding of the ligands when assayed in a buffer containing monovalent ions (⩾40 mM NaCl or KCl or a physiological mixture of electrolytes). The inhibition, seen in the monovalent ion-poor buffer, was dependent on the enzyme concentration. The endoproteases (trypsin, α-chymotrypsin, papain, ficin) showed nanomolar IC 50-values for inhibition of both [ 3H]NPA and [ 3H]haloperidol binding. The inhibition occurred very rapidly at 0° and was different from the slow proteolytic inactivation seen by prolonged incubation at 37°. It was demonstrated that monovalent ions did not themselves interfere with the interaction between the proteases and the membranes. The observations provide evidence for two different types of stereospecific dopaminergic binding sites which are differentially exposed for ligand binding depending on the concentration of monovalent ions. These sites are protease-sensitive sites, labelled in monovalent ion-poor media and protease-insensitive sites, labelled in media with higher concentrations of monovalent ions. Both types of binding sites bind dopamine agonists and dopamine antagonists with high affinity, but some differences were noted in the binding properties and the drug binding selectivity of the sites. It is argued that both sites form part of the same dopamine receptor macromolecular complex. The findings corroborate the hypothesis that dopamine receptors are composed of different sub-unit binding sites, but these are not distinct agonist and antagonist specific sites. The mechanism by which the protease-sensitive sites are rapidly inactivated by particular proteases, is probably a complexation between the enzymes and certain essential peptide moieties of the receptor sites involved.

Multiple postsynaptic dopamine receptors and behavioral manifestation

Previously we demonstrated the existence of non- and positive-cooperative dopamine receptors in rat striatum in [ 3H] apomorphine binding experiments. Non-cooperative sites were sensitive to the inhibition of sulpiride while cooperative sites were not. In the present study results dealing with the involvement of those two types of postsynaptic dopamine receptors in different behavioral manifestations is shown employing lisuride hydrogen maleate (LHM). LHM elicited contralateral turnings in 6-hydroxydopamine lesioned rats unilaterally in the striatum whereas it caused ipsilateral turnings in kainic acid lesioned rats as was observed following the administration of apomorphine. Furthermore, the effect of LHM on rotating behavior was abolished by the pretreatment with sulpiride. On the other hand, LHM inhibited apomorphine induced stereotyped behavior whereas sulpiride failed to block it. These results suggested the dual action of LHM on multiple postsynaptic dopamine receptors. The results also indicated that non-cooperative postsynaptic dopamine receptors are involved in rotating behavior while cooperative receptors participate in the elicitation of stereotypy.

Kainic acid lesions dissociate [ 3H]spiperone and [ 3H]cis-flupenthixol binding sites in rat striatum

The effects of striatal kainic acid lesions on [ 3H]cis-flupenthixol ([ 3H]FPT) and [ 3H]spiperone ( 3H-SPIP) binding to dopamine (DA) receptors in control and lesioned striata were examined. Significant reductions in both binding parameters were observed, [ 3H]FPT binding was depleted to a greater extent than [ 3H]SPIP binding. Reductions in both [ 3H]FPT and [ 3H]SPIP binding were significantly correlated with reductions in glutamic acid decarboxylase activity. After complete loss of GAD activity, 30–40% of [ 3H]SPIP binding sites remained in the lesioned striata, whereas all [ 3H]FPT binding sites were destroyed. The results are discussed in relation to the presence of distinct types of DA receptors in rat striatum, and ligand binding to these receptors.

Optimal conditions for [3H]apomorphine binding and anomalous equilibrium binding of [3H]apomorphine and [3H]spiperone to rat striatal membranes: involvement of surface phenomena versus multiple binding sites.

I. Binding of [3H] apomorphine to dopaminergic receptors in rat striatum was most reproducible and clearly detectable when incubations were run at 25 degrees C in Tris-HCl buffer, pH 7.5, containing 1 mM-EDTA and 0.01% ascorbic acid, using a washed total-membrane fraction. The receptor binding was stereospecifically inhibited by (+)-butaclamol, and dopamine agonists and antagonists showed high binding affinity for these sites. Unlabelled apomorphine inhibited an additional nonstereospecific binding site, which was unrelated to dopamine receptors. EDTA in the incubation mixture considerably lowered nonstereospecific [3H]apomorphine binding, apparently by preventing the complexation of the catechol moiety with metal ions which were demonstrated in membrane preparations. Stereospecific [3H]apomorphine binding was not detectable in the frontal cortex, whereas in the absence of EDTA much saturable nonstereospecific binding occurred. II. Kinetic patterns of stereospecific [3H]spiperone and [3H]apomorphine binding to rat striatal membranes and the inhibition patterns of a dopamine antagonist and an agonist were evaluated at different temperatures in high-ionic-strength Tris buffer with salts added and low-ionic-strength Tris buffer with EDTA. Apparent KD values of spiperone decreased with decreasing tissue concentrations. KD values of both spiperone and apomorphine were little influenced by temperature changes. Scatchard plots of the stereospecific binding changed from linear to curved; the amount of nonstereospecific binding of the 3H ligands varied considerably, but in opposite directions for spiperone and apomorphine in the different buffers. In various assay conditions, interactions between agonists, and between antagonists, appeared fully competitive, but agonist-antagonist interactions were of mixed type. The anomalous binding patterns are interpreted in terms of surface phenomena occurring upon reactions of a ligand with complex physicochemical properties and nonsolubilized sites on membranes suspended in a buffered aqueous solution. It is concluded that anomalous binding patterns are not necessarily an indication of binding to multiple sites or involvement of distinct receptors for high-affinity agonist and antagonist binding.

Persistence of ß-adrenergic receptors in rat striatum following kainic acid administration

Persistence of ß-adrenergic receptors in rat striatum following kainic acid administration

Domperidone, a specific in vitro dopamine antagonist, devoid of in vivo central dopaminergic activity

When tested in different in vitro binding assays and in dopamine-sensitive adenylate cyclase, domperidone was found to be a very potent and specifie dopamine antagonist. Accordingly, [ 3H]ldomperidone binding sites were exclusively detected in homogenates of brain dopaminergic regions. Despite its powerful dopamine antagonism, domperidone did not act centrally in a similar way to neuroleptic drugs. Ex vivo and in vivo experiments indicated that domperidone is unable to reach dopamine receptors in rat striatum. The very low amount of labelling and atypical distribution found in rat brain after administration of [ 3H]ldomperidone is compatible with a lack of penetration into brain structures. Finally, domperidone did not elicit marked HVA increase as is found with classical neuroleptic drugs and metoclopramide. From the present experiments, one may conclude that domperidone, though a very potent and specific dopamine antagonist, cannot readily cross the blood brain barrier, which thus prevents the occurrence of central effects.

Dopamine receptors localised on cerebral cortical afferents to rat corpus striatum.

STRIATAL dopamine receptors, monitored by dopamine-sensitive adenylate cyclase activity1,2 or binding of 3H-haloperidol3–5 apparently represent distinct entities because of differences in drug sensitivity and the pattern of their ontogenetic development6,7. Dopamine also elicits both excitatory and inhibitory effects on striatal neurones8,9. We have examined the effects of selective degeneration of striatal intrinsic neurones with the neurotoxin. kainic acid10–12, and elimination of cortico-striate afferents by cortical ablation13 on the dopamine receptors in rat striatum. We present evidence that a substantial portion of striatal 3H-haloperidol receptor sites are localised to axons of cerebral cortical afferents whereas dopamine-sensitive adenylate cyclase is confined to neurones intrinsic to the striatum.