3H-spiperone Binding Sites Research Articles

Specific binding of 3h-spiperone to peripheral blood cells: Relevance for the interpretation of binding studies in psychiatric disorders

1. 1. There is an ongoing discussion regarding the elevated binding of 3H-spiperone to lymphocytes of schizophrenic patients. Several authors described an atypical binding pattern with a saturable high-affinity and a nonsaturable binding site both displaceable by (+)-butaclamol. Recent findings are still controversial (Fartacek et al., 1997; Wodarz et al., 1996), possibly due to methodological differences. The authors investigated 3H-spiperone binding to different peripheral blood cells including B- and T-lymphoblastoids. 2. 2.B-lymphocytes (K d = 0.081 nM; B max = 0.46 × 10 −15 mol/10 6 cells) and macrophages (K d = 0.11 nM, B max = 2.44 × 10 −15 mol/10 6 cells) are characterized by a minor but saturable binding of 3H-spiperone in a concentration range between 0.5 and 1 nM. Above 1 nM, only nonsaturable binding was measurable. Interestingly, Epstein-Barr virus (EBV) transformed lymphoblastoids (K d = 0.13 nM) have nearly the same affinity for 3H-spiperone as native B-cells, but an increased number of binding sites ( B max = 1.76 × 10 −15 mol/ 10 6 cells). 3. 3.Membranes from B-lymphoblastoids displayed a saturable binding in a concentration between 0 and 1.8 nM of 3H-spiperone (K d = 0.5 nM and B max = 1.72 × 10 −15 mol/mg protein>). Extraction with 1 % digitonin resulted in a similar binding characteristic (K d = 0.17 nM and B max = 1.97 × 10 −15 mol/mg protein). 4. 4.T-cells, granulocytes and MOLT-3-cells did not show a saturable binding even not at high concentrations of 3H-spiperone. 5. 5.The pharmacological profile of the high-affinity 3H-spiperone binding site is clearly different from the dopamine D 2 and D 4, serotonin 5-HT 2 histamine H 1 and noradrenergic alpha, and alpha 2 receptor, respectively. 6. 6.In summary, the results suggest that spiperone binding studies to enriched lymphocytes of psychiatric patients should be interpreted cautiously. Variable amounts of leucocytes might result in a higher proportion of nonsaturable, butaclamol displaceable spiperone binding with relevance for the calculation of K d and B max of the saturable high-affinity site. Interestingly, homogenous B-lymphoblastoid cell lines have the same binding characteristics as native B-225 lymphocytes and therefore should be recommended for characterization of 3H-spiperone binding sites.

Age-related changes in dopamine receptor densities in the brain of the honey bee, Apis mellifera

Changes in the levels of binding of 3H-SCH-23390, a vertebrate D1 dopamine receptor ligand, and 3H-spiperone, a vertebrate D2 dopamine receptor ligand were investigated in the brain of the worker honey bee during metamorphic adult development and during the lifetime of the adult bee. Age-related fluctuations in binding levels were markedly different for these two ligands. 3H-SCH-23390 and 3H-spiperone binding sites were present at low levels during metamorphic adult development. After adult emergence, however, 3H-SCH-23390 binding levels, in contrast to those of 3H-spiperone, increased significantly. Within the first 48 h of adult life 3H-SCH-23390 binding reached a level not significantly different from that detected in forager bees. No significant fluctuations in the levels of 3H-spiperone binding were observed during the adult lifetime of the bee. Measurements of dopamine levels in the brains of pupal and adult bees revealed no direct correlation between fluctuations in endogenous amine levels and the amount of binding of either 3H-SCH-23390 or 3H-spiperone. These results provide evidence for subtype-specific patterns of expression of dopamine receptors in the insect brain and show that D1- and D2-like receptors are expressed not only in the adult CNS, but also in the developing brain of the bee.

3H-spiperone binding sites in post-mortem brains from schizophrenic patients: Relationship to neuroleptic drug treatment, abnormal movements, and positive symptoms

In post-mortem putamen samples from 27 schizophrenics and 27 controls D2 receptors were measured by Scatchard analysis using 3H-spiperone as a ligand. Maximum number of binding sites (Bmax) and apparent dissociation constant (KD) were significantly increased only in patients in whom neuroleptic medication had been given within a three-month period before death. When the neuroleptic medication had been withdrawn at least 3 month before death, there was a slight, but not significant, reduction in Bmax values and unchanged KD values. Withdrawal of neuroleptic drugs was followed by a normalization of the KD values within 2 weeks and a slower reduction of Bmax values. There were 6 schizophrenic patients with mainly positive schizophrenic symptoms and 17 patients with mainly negative symptoms; positive schizophrenic symptoms were not related to higher Bmax values. There was no difference in 3H-spiperone binding between patients with and without movement disorders (tardive dyskinesia or extrapyramidal symptoms).

Catecholamine receptor binding in rat kidney: Effect of aging

Binding to several receptors was compared in kidneys from 3- and 24-month-old rats. In crude membrane preparations of aged rat kidneys, the number of beta 2-adrenergic receptors was significantly reduced but the number of total beta-adrenoceptors was unchanged. The high-affinity alpha 1-adrenoceptor component was significantly reduced in old rats, whereas the low-affinity component was unchanged. The number of alpha 2-adrenoceptors showed a non-significant decrease. 3H-spiperone binding sites were similar in young and old rats. For each receptor binding the KD values were the same in young and old animals. The D1 dopamine receptor was significantly reduced in old rats. In our experiments, age-related changes of specific binding sites in the kidney were selective for some receptors studied and did not seem to be due to general aging-induced membrane modifications. Moreover, the renal and central receptors were sensitive to aging differently in the same animal model.

Enhancement of Prolactin-Secreting Effect Produced by Repeated Administration of Haloperidol in the Avoidance Situation and Dopamine Neurons in Rats

Male Wistar strain rats subcutaneously administered haloperidol at the dose of 0.035 mg/kg at 3-4 days interval, 10 times, in the avoidance situation. Enhancement of the prolactin-secreting effect of haloperidol was observed when it was given in the home cages at the 10th day after termination of the repeated administration. In these animals, 3H-spiperone binding sites in the pituitary significantly increased, while the DOPAC/DA ratio in the hypothalamus significantly decreased. The enhancement phenomenon may be produced by a decrease in the activity of dopamine neurons in the hypothalamus.

Tamoxifen counteracts estradiol induced effects on striatal and hypophyseal dopamine receptors

We investigated the ability of Tamoxifen (TAM), an antiestrogen drug, to counteract the modifications induced by estrogens on dopamine (DA) receptors on striatum and on adenohypophysis of ovex female rats. Subacute treatment with 17β-estradiol (E 2) at both low (0.1 μg/kg) and high (20 μg/kg) doses confirmed its ability to increase the number of striatal 3H-Spiperone ( 3H-SPI) binding sites in a dose dependent manner. By contrast in the pituitary, only high doses of estrogen were effective in reducing the number of DA receptors. We treated ovex female rats for 15 days with TAM alone or associated with E 2, to see if these estrogenic effects could be suppressed by an antiestrogenic drug. TAM did not affect the number of striatal DA receptors, but significantly increased the adenohypophyseal DA binding sites, without varying their affinity. No changes were observed in pituitary and striatal DA receptor density, even when TAM was injected in association with estradiol. In conclusion: TAM is able to counteract the effects estrogens have on DA receptors. However there is some evidence that it could influence the pituitary DA systems independently of its antiestrogenic activity.

Modulation of activity of the striatal dopaminergic system during the hibernation cycle

To evaluate how the activity of a well-established neurotransmitter pathway is modulated by a behavioral state, 3H-spiperone binding sites and dopamine (DA) and DA metabolite concentrations were measured in the striata of ground squirrels in 5 phases of the hibernation cycle. Whereas levels of striatal DA and its deaminated metabolite DOPAC did not change significantly, the concentrations of the O-methylated-deaminated metabolite, homovanillic acid (HVA), decreased in all phases of hibernation relative to euthermia. Striatal 3H-spiperone binding sites declined across the hibernation cycle in parallel with the reduction of HVA concentration; receptor binding affinity was unchanged by arousal state. In conjunction with previously reported findings, these results are consistent with the hypothesis that hibernation is associated with a down-regulation of the postsynaptic D2 receptors secondary to increased extracellular DA concentration and reduced DA degradation.

Differential recovery rates of rat D2 dopamine receptors as a function of aging and chronic reserpine treatment following irreversible modification: a key to receptor regulatory mechanisms

The density of D2 dopamine receptors identified by 3H-spiperone binding was significantly lower in the striatum of senescent (28-month-old) than in mature (4-month-old) Fischer 344 rats. The time course of recovery of 3H-spiperone binding to D2 dopamine receptors following irreversible receptor modification by a single injection of N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) was significantly slower in senescent than in mature rats. The kinetics of D2 dopamine receptor repopulation could be adequately described by a model assuming a constant rate of receptor production and a rate of degradation that was dependent on receptor concentration. Both the production rate and degradation rate constant of D2 dopamine receptors were lower in the senescent rats compared to mature rats. The changes in receptor density that occur as a function of development and aging are the result of changes in both the receptor production rate and receptor degradation rate constant, which, while both are reduced, tend to maintain receptor density. That is, receptor density decreased by 26% from 4 to 28 months of age, although the receptor production rate and degradation rate constant decreased by 40-50%. Chronic treatment of rats with reserpine produced a 21% increase in the density of striatal D2 dopamine receptors of mature, but not senescent, rats, accompanied by an increase in the initial recovery rate of 3H-spiperone binding sites following EEDQ treatment. It might have been expected that the reduced formation of agonist-bound receptors would have lead to a decrease in the receptor degradation rate constant.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential effects of proglumide on mesolimbic and nigrostriatal dopamine function.

Cholecystokinin octapeptide (CCK-8) is prevalent as a co-transmitter in the mesolimbic dopamine pathway. The effect of proglumide, a CCK-8 antagonist, on two acute and one chronic behavioral models of dopamine function was tested. First, haloperidol was used to inhibit stereotypies induced by apomorphine in rats. Pre-administration of proglumide significantly shifted the haloperidol dose response curve to the left. Second, rats were injected in the left caudate nucleus with kainic acid. Three weeks later, haloperidol was used to inhibit apomorphine-induced circling. Pre-administration of proglumide had no effect on this haloperidol dose response curve. Third, either proglumide, haloperidol, or combined treatment was administered to rats for 2 weeks. In proglumide-treated animals, a significant increase in 3H-spiperone binding sites in the nucleus accumbens was observed.

Comparison of 3H-spiperone binding in caudate nuclei of rabbits and rats

3H-Spiperone binding and inhibition by competing ligands were studied in caudate nucleus homogenates from Sprague-Dawley rats and New Zealand albino rabbits. 3H-Spiperone binding sites were very similar in the two species and had pharmacological profiles characteristic of D2-dopamine receptors, in accordance with previous reports. However, differences occurred between the species in the potencies with which dopaminergic ergots and other drugs interacted with these binding sites and with regard to the compounds used to define specific binding. These findings suggest that slight differences in relative proportions of subtypesof 3H-spiperone binding sites, or of neuroleptic at these subtypes, may exist between rabbits and rats.

3H-spiperone binding sites in lymphocytes as possible vulnerability marker in schizophrenia

The binding parameters for the dopamine antagonist 3H-spiperone to lymphocytes were investigated in nonmedicated schizophrenics ( N = 43), in medicated schizophrenics ( N = 25), as well as in 38 first- and second-degree relatives of schizophrenics, in a psychiatric control group ( N = 27) and in normal, healthy controls ( N = 40). The density of lymphocyte 3H-spiperone binding sites ( B max) was significantly increased in all acute and chronic schizophrenic patients and in patients undergoing neuroleptic treatment for 2 weeks to several months and remained on a constant level during a drug-free remission period, suggesting that B max is a state independent variable. Elevated binding seems to be associated with schizophrenia; it could not be found in other psychiatric patients. In pedigree and family studies it became obvious that all relatives with one or more previous episodes of schizophrenia had similar increased binding capacity. But even in some well-state relatives elevated spiperone binding could be found, although they never had symptoms of the disease. Among these relatives of schizophrenics, there was an association between increased spiperone binding and the transmission of the disease. Although conclusive data about the precise genetic control obtainable in twin studies are still missing, we would suggest that elevated lymphocyte spiperone binding may be a marker of susceptibility to schizophrenia.

Strain differences in the reverse tolerance to methamphetamine and changes in catecholaminergic neurons in mice.

The effect of methamphetamine (MAP) on ambulatory activity and neurochemical changes in catecholaminergic neurons in the brain were investigated in dd and C57BL/6 strains of male mice. The mice were given repeatedly MAP at 2 mg/kg, s.c., 10 times at a fixed interval of 4 days. Single MAP markedly increased the ambulatory activity in both strains of mice. The ambulation-increasing effect was progressively enhanced without accompanying stereotyped behaviors when the drug was repeatedly given. The dd mice showed higher susceptibility not only to single MAP but also to the enhancing effect of the drug (reverse tolerance) that the C57BL/6 mice. On the other hand, non-treated dd mice exhibited lower maximum densities of 3H-spiperone binding sites in the striatum and 3H-WB4101 binding sites in the cortex and hippocampus than non-treated C57BL/6 mice. In contrast, the dd mice exhibited higher noradrenaline turnover than the C57BL/6 mice in the brain regions examined. The repeated administration of MAP produced decrease in the densities of both 3H-spiperone and 3H-WB4101 binding sites in the corresponding regions with increase in catecholamine turnover in dd mice. However, the similar changes were observed only in 3H-WB4101 binding sites and noradrenaline turnover in C57BL/6 mice. These results suggest that the ambulation-increasing effect of MAP is positively correlated with catecholamine turnover, while it was correlated negatively with the densities of catecholamine binding sites. Furthermore, the enhancing effect of MAP is supposed to have been partially elicited by changes in brain catecholaminergic systems, in particular an increase in catecholamine turnover.(ABSTRACT TRUNCATED AT 250 WORDS)

Muscarinic, benzodiazepine, gaba, chloride channel and other binding sites in frontal cortex in hepatic coma in man

1. 1. Alterations in several neurotransmitter systems in brain have been implicated in the pathophysiology of hepatic coma (HC). Studies on human autopsy material are few. We investigated H-quinuclidlnylbenzllate (QNB), 3H-spiperone, 3H-imipramine, 3H-PN-200-110, 3naloxone, 3H-Flunitrazepam, 3H-muscimol, 35S-t-butylbicyclophosphothionate and 3H-cyclohexyladenosine binding sites In frontal cortex from seven patients with HC and five controls. 2. 2. The density of 3H-QNB binding sites was significantly decreased and the affinity slightly increased in HC. The functional significance of these selective changes in muscarinic receptor binding sites Is unclear. Further studies evaluating cholinergic function In HC are Indicated. 3. 3. Acute studies in animals point to an Increase In GABA and BZ binding sites in HC. The present results show that the BZ/GABA-receptor-chloride-Ionophore complex is unchanged in HC in man. 4. 4. Serotonergic (5HT-2), adenosine (A-l), imipramlne (5HT uptake sites), opiate (naloxone) and calcium channel antagonist binding sites are unchanged in HC.

Enhancement of ambulation-increasing effect produced by repeated administration of methamphetamine in rats and neurochemical changes in catecholaminergic neurons.

Male rats of the Wistar strain were repeatedly given methamphetamine (MAP) at 0.5 mg/kg, s.c., 10 times at a fixed interval of 4 days in different environmental situations. The effects of MAP administration on ambulatory activity and neurochemical parameters in the brain were investigated. MAP (0.5 mg/kg) markedly increased ambulatory activity. The ambulation-increasing effect of MAP was progressively enhanced without accompanying stereotyped behaviors when the drug was repeatedly given in the activity cage. The once enhanced effect was considered to be irreversible. In addition, the pretreatment with MAP in the activity cage produced a decrease in the maximum density of 3H-spiperone binding sites in the striatum and that of 3H-WB4101 binding sites in the cortex and hippocampus, with decreases in catecholamine and increases in its metabolites. However, the enhancement of the ambulation-increasing effect of MAP and changes in the binding sites or catecholamine metabolism were scarcely observed when the drug was repeatedly given to the rats in the narrow cage where horizontal ambulation of the animals was strongly impaired. These results suggest that the enhancing effect of MAP is closely related to the environmental situations to which the animals have been exposed to under the drug effect. Moreover, it is indicated that there is a correlation between the enhancement of the ambulation-increasing effect of MAP and changes in the neurochemical parameters. The enhancing effect of MAP is supposed to have been partially caused by changes in cerebral catecholaminergic systems, in particular an increase in catecholamine turnover.

Enhancement of Ambulation-Increasing Effect Produced by Repeated Administration of Methamphetamine in Rats and Neurochemical Changes in Catecholaminergic Neurons

Male rats of the Wistar strain were repeatedly given methamphetamine (MAP) at 0.5 mg/kg, s.c., 10 times at a fixed interval of 4 days in different environmental situations. The effects of MAP administration on ambulatory activity and neurochemical parameters in the brain were investigated. MAP (0.5 mg/kg) markedly increased ambulatory activity. The ambulation-increasing effect of MAP was progressively enhanced without accompanying stereotyped behaviors when the drug was repeatedly given in the activity cage. The once enhanced effect was considered to be irreversible. In addition, the pretreatment with MAP in the activity cage produced a decrease in the maximum density of 3H-spiperone binding sites in the striatum and that of 3H-WB4101 binding sites in the cortex and hippocampus, with decreases in catecholamine and increases in its metabolites. However, the enhancement of the ambulation-increasing effect of MAP and changes in the binding sites or catecholamine metabolism were scarcely observed when the drug was repeatedly given to the rats in the narrow cage where horizontal ambulation of the animals was strongly impaired. These results suggest that the enhancing effect of MAP is closely related to the environmental situations to which the animals have been exposed to under the drug effect. Moreover, it is indicated that there is a correlation between the enhancement of the ambulation-increasing effect of MAP and changes in the neurochemical parameters. The enhancing effect of MAP is supposed to have been partially caused by changes in cerebral catecholaminergic systems, in particular an increase in catecholamine turnover.

Hypophysectomy may non-selectively alter pharmacokinetic parameters to enhance the ability of haloperidol to increase striatal dopamine receptor density in the rat

We have investigated the effect of a range of doses of haloperidol (0.625–5.0 mg/kg/day) or saline, administered for 14 days, followed by a 3 day drug washout period, to sham operated or hypophysectomized rats. Haloperidol increased the number of specific striatal 3H-spiperone binding sites ( B max) in sham-operated animals at doses of 2.5 and 5.0 mg/kg/day, and in hypophysectomized animals at all doses used (0.625–5.0 mg/kg/day). The inhibition of locomotor activity produced by haloperidol was greater in hypophysectomized than sham-operated animals. Plasma and striatal haloperidol levels after equivalent doses were greater in hypophysectomized than in sham-operated animals. We conclude that hypophysectomy may enhance the ability of haloperidol to induce striatal dopamine receptor supersensitivity in the rat, and that this may be due to differences in the pharmacokinetic handling of haloperidol between sham-operated and hypophysectomized animals.

Behavioral and biochemical studies of dopamine receptor sensitivity in differentially housed mice.

Group housed and individually housed mice were compared in (1) the motor activity responses to direct and indirect dopamine (DA) agonists, (2) in vivo presynaptic autoreceptor sensitivity and (3) in vitro binding of 3H-spiperone. Relative to group housed mice, individually housed mice showed an increased motor activity response to amphetamine, 1.25 and 0.625 mg/kg. Using two in vivo measures of presynaptic DA receptor sensitivity, the antagonism of spontaneous locomotor activity and the antagonism of dihydroxyphenylalanine (DOPA) accumulation by apomorphine (APO), individually housed mice showed greater activity counts and higher DOPA accumulations than group housed mice. Levels of tyrosine were significantly greater in individually housed mice. Significant effects of housing were also noted with the motor activity response to APO, 0.075-0.300 mg/kg, following pretreatment with reserpine, an in vivo measure of postsynaptic receptor sensitivity. However, there was no effect of housing on the number or affinity of 3H-spiperone binding sites in the striatum. These results are discussed in terms of the presynaptic activity of catecholaminergic neurons and the postsynaptic receptor sensitivity to APO in individually housed mice.

Specific in vitro and in vivo binding of 3H-raclopride a potent substituted benzamide drug with high affinity for dopamine D-2 receptors in the rat brain

The substituted benzamide drug raclopride, [((-)-( S)-3,5-dichloro- N-((1-ethyl-2-pyrrolidinyl) methyl)-6-methoxy-salicylamide tartrate; FLA 870(-); A40664] was shown to be a potent and selective antagonist of dopamine D-2 receptors by its high affinity for striatal 3H-spiperone binding sites and low potency to block dopamine stimulated adenylate cyclase in vitro. In vitro studies showed that 3H-raclopride binds with a high affinity ( K D = 1.2 nM) and a low proportion of non-specific binding to rat striatal homogenates. The binding of 3H-raclopride is saturable ( B max = 23.5 pmoles/g wet wt) and reversible (dissociation half-time = 30 min) with a regional distribution of the specifically bound drug showing the following rank-order: striatum > nucleus accumbens > olfactory tubercle > septum > hypothalamus > hippocampus > frontal cortex. After in vivo administration, 3H-raclopride accumulates preferentially in dopamine rich brain areas with approximately 10 times higher levels in the striatum than in the cerebellum, when examined 30 min after injection. The in vivo binding of 3H-raclopride was saturable, reversible and showed a low component of non-specific binding. More than 90% of the drug reached the brain in a non-metabolized form as judged by thin-layer chromatography. Pharmacological analysis of 3H-raclopride binding showed that it could be displaced by dopamine agonists and antagonists but not by serotoninergic or noradrenergic drugs. Taken together, the results suggest that 3H-raclopride labels dopamine D-2 receptors with high specificity in the rat brain both in vitro and in vivo, and thus, that it should be a useful tool in studies of central dopamine D-2 receptors.

Failure of chronic haloperidol to affect prolactin secretion due to acute haloperidol administration

Withdrawal from chronic haloperidol exposure was associated to unaltered circulating levels of prolactin (PRL), decreased 3H-spiperone binding sites in the anterior pituitary and increased 3H-spiperone binding sites in the striatum of male rats. Haloperidol (0.1 mg/kg ip) induced similar rises in plasma PRL in haloperidol-or saline-treated rats and the dose of 0.01 mg/kg was ineffective in both groups. These findings illustrate the poor relatedness existing at the pituitary D 2 receptor between biochemical and functional indices.

Changes in 3H-Spiperone, 3H-WB 4101 and 3H-Dihydroalprenolol Bindings to Brain Membranes Produced by Postnatal Pretreatment with Chlorpromazine in Adult Rats

In order to elucidate possible mechanisms of the learning deficit produced by postnatal pretreatment with chlorpromazine (CPZ), changes in catecholamine receptors in the rat brain were investigated. Male neonates of Wistar strain rats were given s.c. 2 mg/kg/day of CPZ for 7 successive days from days 6 to 12 after birth. Effect of the postnatal pretreatment with CPZ on saturation constants for specific bindings of 3H-spiperone, 3H-WB 4101 and 3H-dihydroalprenolol, respectively, in 8 brain regions was investigated at 60 days after birth. Significant decreases in Bmax values of 3H-WB 4101 binding sites in the cortex, thalamus, hypothalamus, mid brain and medulla oblongata/pons and decreases in Kd values of the binding sites in thalamus, hypothalamus and mid brain were observed in CPZ-pretreated rats when compared with corresponding Bmax and Kd values obtained in saline-pretreated rats. Furthermore, significant decreases in both Bmax and Kd values of 3H-DHA binding sites in the thalamus were detected in CPZ-pretreated rats when compared with those obtained in saline-pretreated rats. However, no alterations in 3H-spiperone binding sites in all brain regions were found between CPZ- and saline-pretreated rats. These results suggest that the learning deficit observed in CPZ-pretreated rats may be produced by a functional disorder of catecholaminergic, in particular alpha 1-noradrenergic neurons in the brain.