Desensitization of adenosine and dopamine receptors in rat brain after treatment with adenosine analogs

Maximally tolerated doses of N6-[(R)-1-methyl-2-phenylethyl] adenosine (0.50 nmol/hr/2 wk), 5'-N-ethylcarboxamide adenosine (NECA, 0.04 nmol/hr/2 wk) or deoxycoformycin (5 nmol/hr/1 wk) were administered i.c.v. to rats using mini-osmotic pumps. Adenosine receptor function was subsequently assayed using both ligand binding and adenylate cyclase assays. Binding to A1 receptors was quantitated using [3H]N6-[(R)-1-methyl-2-phenylethyl]adenosine, a selective agonist ligand at A1 receptors. Differences in the binding of this ligand and that of [3H]NECA, which binds to A1 and A2 receptors with similar affinities, were used to quantitate A2 receptors. None of the treatments affected A1 receptor function as assessed by both ligand binding and adenylate cyclase assays. A2 receptor binding and A2 receptor-mediated stimulation of adenylate cyclase were blunted in striatal membranes from NECA- and deoxycoformycin-treated rats but unaffected in striatal membranes from N6-[(R]-1-methyl-2-phenylethyl]adenosine-treated rats. All three pretreatments attenuated D1 dopamine receptor-mediated stimulation of adenylate cyclase in striatal membranes. These results suggest that 1) the A2 adenosine receptor system is susceptible to desensitization and 2) different mechanisms are involved in the NECA- and deoxycoformycin-induced desensitization of A2 adenosine receptor and D1 dopamine receptor systems. It is suggested that the D1 dopamine receptor desensitization is, in fact, due to the tonic stimulation of adenosine A1 receptors.     

Role of calmodulin-dependent phosphorylation in chronic sulpiride-induced striatal dopamine receptor supersensitivity

In the present study we investigated several pharmacological and biochemical parameters in striatal preparations of rats which were treated chronically with a selective D2 receptor antagonist, sulpiride. Chronic sulpiride treatment in rats (50 mg/kg s.c. for 20 days) potentiated stereotyped responses by apomorphine (0.1-2.5 mg/kg). In association with the dopaminergic behavior supersensitivity, we observed a significant increase in the number of specific [3H]spiperone binding sites (D2 receptors) in the striatum without affecting the ligand binding affinity constant. We further observed a marked increase in the sensitivity of the protein kinase to calcium (0.1-0.5 mM) and calmodulin (1 micrograms) in these rats. The D1 receptor functions which are represented by the basal and dopamine-stimulated adenylate cyclase and the cyclic AMP-dependent protein kinase activity were not changed after chronic sulpiride treatment. In vitro, pretreatment of striatal particulates with the Ca++-chelating agent ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid (1.2 mM) to remove endogenous Ca++ and calmodulin or the addition of Ca++ and calmodulin to the striatal particulates did not affect the binding affinities of dopamine agonists and antagonists to the receptors. Therefore, the increased sensitivity of the calmodulin-dependent system seen in chronic sulpiride-treated rats correlates with the increased number of D2 receptors in striatal dopamine receptor supersensitivity.     

Repeated amphetamine pretreatment alters the responsiveness of striatal dopamine-stimulated adenylate cyclase to amphetamine-induced desensitization

The repeated daily administration of moderate doses of amphetamine results in an augmentation of the behavioral response to subsequent amphetamine challenge. One feature of the augmentation is a shift in the type of perseverative behaviors to those generally associated with higher acute doses of the drug. Consistent with these observations, rats pretreated with six daily injections of amphetamine (3 mg/kg) exhibited primarily oral stereotypies to a challenge dose of 2.5 mg/kg of amphetamine, whereas control animals exhibited focused sniffing and repetitive head movements. Previously we found that the acute administration of amphetamine or methylphenidate only at doses which induce oral stereotypies promotes a rapid desensitization of striatal dopamine-stimulated adenylate cyclase. We therefore examined the effects of repeated amphetamine pretreatment on this index of D1 dopamine receptors. The administration of 2.5 mg/kg of amphetamine produced a 2-fold shift to the right in the concentration-response curve for dopamine-stimulated adenylate cyclase in animals pretreated with amphetamine, but not in saline pretreated controls. No effect of the chronic amphetamine pretreatment on dopamine stimulated cyclase in the absence of amphetamine challenge was observed. The binding of [3H]cis-flupenthixol to striatal D1 dopamine receptors was not affected by acute or chronic amphetamine. These results suggest a relationship between stimulant-induced desensitization of striatal D1 dopamine receptors and the induction of oral stereotypies.     

Development of functional dependence on ethanol in dopaminergic systems

Withdrawal of mice from chronic ethanol treatment results in a decreased responsiveness of striatal (but not mesolimbic) dopamine-sensitive adenylate cyclase activity to stimulation by dopamine. This subsensitivity is not apparent at the time of withdrawal from chronic feeding of ethanol, when animals are still intoxicated, but becomes evident as ethanol is eliminated from the animals. Addition of ethanol in vitro to tissue homogenates from ethanol-withdrawn animals, at concentrations similar to those found in brain at the time of withdrawal, normalizes the response of the adenylate cyclase to dopamine. No difference is evident between control and ethanol-withdrawn animals in stimulation of adenylate cyclase by sodium fluoride. The specificity of the response of striatal adenylate cyclase to stimulation by dopamine, as compared to other transmitters, is unaltered by chronic ethanol feeding. Chronic treatment with ethanol and withdrawal also does not affect the specific binding of spiroperidol in either striatal or mesolimbic regions. It is suggested that the decreased response of adenylate cyclase to dopamine in ethanol-withdrawn animals results from decreased efficiency of coupling between dopamine "receptor" sites and catalytic units of adenylate cyclase.     

Castration blocks chronic sulpiride-induced desensitization of striatal D1 receptor-stimulated adenylate cyclase activity in male rats

Gonadal hormones have been shown to modulate adaptive responses of the mesostriatal dopaminergic system to antipsychotic challenge. We examined the role of endogenous gonadal steroids in the regulation of D1 receptor function after chronic treatment with sulpiride, a D2 specific antagonist. Chronic sulpiride treatment induced a desensitization of striatal D1 receptor-simulated adenylate cyclase activity in intact male rats with no change in the number of D1 or D2 receptors. This desensitization of D1-stimulated adenylate cyclase activity was expressed as a decrease in Vmax with no change in the activation constant. Castration of male rats blocked the chronic sulpiride-induced desensitization of D1 receptor function. Castration of male rats also resulted in a decrease in the number of D1 receptors as measured by [3H]SCH23390 binding. Ovariectomy of female rats had no effect on striatal D1 receptor-stimulated adenylate cyclase activity. Preliminary studies showed no effect of chronic sulpiride treatment on D1 receptor function in intact or ovariectomized female rats. We conclude that testicular hormones have a permissive effect on the expression of the chronic sulpiride-induced desensitization of D1 receptor function.     

Effects of chronic SCH23390 treatment on the biochemical and behavioral properties of D1 and D2 dopamine receptors: potentiated behavioral responses to a D2 dopamine agonist after selective D1 dopamine receptor upregulation

Chronic treatment of rats with SCH23390 (0.5 mg/kg/day s.c.), a D1 dopamine receptor antagonist, for 21 days resulted in an increase in D1 dopamine receptors but produced no change in D2 dopamine receptors. During habituation to locomotor activity cages the rats treated chronically with SCH23390 showed significantly higher locomotor activity than controls treated chronically with saline. When injected with the selective D1 dopamine receptor agonist SKF38393 (3 mg/kg), rats treated chronically with SCH23390 showed significantly greater stereotypy and locomotor activity responses. Surprisingly, rats treated chronically with SCH23390 also showed significantly higher locomotor activity and stereotypy responses when treated with the selective D2 dopamine receptor agonist, quinpirole (LY171555) (0.3 mg/kg). These results indicate that a selective increase in D1 receptors may not be necessary, but is sufficient, to lead to an enhanced behavioral response to either selective D1 or D2 dopamine receptor agonists. If, indeed, an enhanced stereotypy and locomotor activity response to dopaminergic agonists in rats after a brief chronic treatment with a neuroleptic drug is predictive of tardive dyskinesia potential in the clinical setting, these results can suggest that SCH23390 may also induce tardive dyskinesia in humans. Adenylate cyclase activity stimulated by guanine nucleotides, forskolin or dopamine was enhanced after chronic treatment with SCH23390. However, dopamine-stimulated adenylate cyclase activity was not potentiated detectably by the increase in receptor number over the more general increase in guanine nucleotide-stimulated cyclic AMP production. Additionally, no change was observed in dopamine competition for [3H]SCH23390 binding, with dopamine's RH/RL ratio remaining unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)     

K+ channel and adenylate cyclase involvement in regulation of Ca2+-evoked release of [3H]dopamine from synaptosomes

In superfused striatal synaptosomes, previously unexposed to Ca2+ during isolation and superfusion, 1.25 mM Ca2+ evokes the release of [3H]dopamine. This Ca2+-evoked release is produced without elevating K+ (4.5 mM) before or after Ca2+ exposure, can be blocked by the Na+ channel antagonist tetrodotoxin, and modulated by dopamine (D2) receptor agonists and antagonists. We now present evidence that functional K+ channels regulate Ca2+-evoked [3H]dopamine release and may be necessary for the dopamine (D2) modulation of this release. The K+ channel blocker tetraethyl ammonium (TEA) could partially prevent D2 agonist (LY-171555) inhibition of Ca2+-evoked release in both olfactory tubercle and striatal synaptosomes. Another K+ channel blocker, 4-aminopyridine, also partially blocked dopamine (D2) agonist inhibition of release. When both 5 mM tetraethyl ammonium and 0.1 mM 4-aminopyridine were employed, by, dopamine (D2) inhibition of Ca2+-evoked [3H]dopamine release was prevented. However, with both K+ channel blockers present, only the initial portion of the release could be blocked by tetrodotoxin. These results are consistent with what might be expected if K+ channels were linked to dopamine (D2) receptors. In additional experiments we found that stimulation of adenylate cyclase by 1 microM forskolin with 0.25 mM 3-isobutyl-1-methylxanthine present potentiated Ca2+-evoked [3H]dopamine release but that this combination did not affect dopamine (D2) inhibition of [3H]dopamine release. Furthermore, although the protein alkylator n-ethylmaleimide could block dopamine (D2) inhibition of release, pertussis toxin, a specific inactivator of the inhibitory protein regulating adenylate cyclase, had little effect on dopamine (D2) inhibition. Therefore, dopamine (D2) inhibition of dopamine release may not be coupled to adenylate cyclase activity.     

Adaptive mechanisms of striatal D1 and D2 dopamine receptors in response to a prolonged reserpine treatment in mice

Mice receiving reserpine (1 mg/kg/day) during 5 days develop behavioral supersensitivity. To study the possible molecular correlates of these adaptive changes we compared binding parameters of D1 and D2 receptors and adenylate cyclase activity in striata from normal and reserpinized mice. Saturation curves using [3H]SCH 23390 showed no changes in maximum binding capacity (Bmax) or Kd of striatal D1 receptors taken from control or 5 days reserpine-treated mice. However, [3H]spiperone saturation curves showed a 31% increase in D2 receptors Bmax with no changes in Kd. Dopamine competition of [3H]SCH 23390 and [3H]spiperone binding in mouse striatum was also performed. Analysis of data by LIGAND showed that dopamine recognizes two subpopulations for D1 and for D2 receptors. The proportion of receptors in the high affinity state (D1high and D2high) were increased in reserpine-treated animals. The addition of 100 microM GTP produced a complete conversion of D1high and D2high receptors into their low-affinity states in striata from control and reserpinized mice. Five days of reserpine treatment increased basal adenylate cyclase activity of mouse striatum in the presence of Mn++ or Mg++ ions. Concentration curves with dopamine, NaF or forskolin revealed shifts to the left and higher maximum responses without changes in EC50 values in striata from reserpinized mice. Thus, a prolonged reserpine treatment produces marked changes in D1 and D2 receptors increasing the proportion of high affinity state subpopulations and the total Bmax of D2 receptors. Also, dopamine function may be enhanced through an increment of the catalytic component of striatal adenylate cyclase.     

Partial dopamine-agonistic and atypical neuroleptic properties of the amino-ergolines SDZ 208-911 and SDZ 208-912

The aminoergolines SDZ 208-911 [N-[(8-alpha)-2,6-dimethylergoline-8-yl]-2,2- dimethylpropanamide] and SDZ 208-912 [N-[8-alpha)-2-chloro-6-methylergoline-8-yl]- 2,2-dimethylpropanamide] exhibit nonclassical, neuroleptic-like properties in rodents. Thus, they are equipotent to haloperidol as inhibitors of apomorphine-induced gnawing behavior and conditioned avoidance responding, but are essentially devoid of cataleptogenic activity. In addition, they show high affinity for central D-2 receptors in vitro and elevate striatal homovanillic acid levels. In contrast to haloperidol, however, SDZ 208-911 and 208-912 strongly inhibit prolactin secretion and induce contralateral circling behavior in 6-hydroxydopamine-lesioned animals. These profiles are consistent with the drugs exhibiting varying degrees of partial agonistic activity at dopamine D-2 receptors, with SDZ 208-911 being considerably more agonistic than SDZ 208-912. Support for this contention stems from the ability of SDZ 208-911 to reduce the elevation of striatal L-dopa formation induced by gamma-butyrolactone, and SDZ 208-912's partial reversal of apomorphine's inhibitory action on gamma-butyrolactone activity. SDZ 208-911's effects are reduced after the partial alkylation of D-2 receptors with N-ethoxy-carbonyl-2-ethoxy-1,2-dihydroquinoline, confirming its partial agonistic properties. SDZ 208-911 and SDZ 208-912 could be effective against both the positive and negative symptoms of schizophrenia, while exhibiting a reduced incidence of dystonic and parkinsonian side-effects. In addition, their clinical testing might throw more light on the central dopaminergic status of schizophrenic subjects.     

Beta adrenergic receptor subtypes in the atria: evidence for close coupling of beta-1 and beta-2 adrenergic receptors to adenylate cyclase

The relationship between occupancy of beta adrenergic receptors and stimulation of adenylate cyclase in dog atrial tissue was examined by studying the binding of [125I]iodopindolol and the activation of adenylate cyclase. Computer-assisted nonlinear regression analysis was used to analyze the inhibition of isoproterenol-stimulated adenylate cyclase activity by beta-1- or beta-2-selective antagonists. The Ki values for each subtype of receptor for the selective antagonists resulting from studies of the inhibition of adenylate cyclase activity were similar to those determined in studies of the inhibition of the binding of [125I]iodopindolol. To compare further the occupancy of beta-1 or beta-2 adrenergic receptors with the activation of adenylate cyclase mediated by each class of receptor, computer modeling of the stimulation of adenylate cyclase by the beta-1-selective agonist norepinephrine was carried out. The EC50 values of norepinephrine for each receptor subtype, as measured in studies of norepinephrine-stimulated adenylate cyclase activity, were similar to the Ki values for the inhibition by norepinephrine of the binding of [125I]iodopindolol to each receptor subtype. The data led to the conclusion that beta-1 adrenergic receptors make up about 70% of the total number of beta adrenergic receptors and mediate 70% of the increase in adenylate cyclase activity produced by isoproterenol. These results suggest that the relationship between occupancy of each class of receptor and activation of adenylate cyclase is linear and that, when agonist-stimulated adenylate cyclase activity is used as a functional response, neither spare beta-1 nor spare beta-2 adrenergic receptors exist in the atrium.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pharmacological differences between the D-2 autoreceptor and the D-1 dopamine receptor in rabbit retina

The effect of dopamine receptor agonists and antagonists was studied on the calcium-dependent release of [3H]dopamine elicited by field stimulation at 3 Hz for a duration of 1 min (20 mA, 2 msec) from the rabbit retina in vitro and on adenylate cyclase activity in homogenates of rabbit retina. The relative order of potency of dopamine receptor agonists to inhibit the stimulation-evoked [3H]dopamine release was pergolide greater than bromocriptine greater than apomorphine greater than LY 141865 greater than N,N-di-n-propyldopamine greater than or equal to dopamine. The relative order of potencies of dopamine receptor antagonists to increase [3H]dopamine release was: S-sulpiride greater than or equal to domperidone greater than or equal to spiroperidol greater than metoclopramide greater than fluphenazine greater than or equal to R-sulpiride. alpha-Flupenthixol (0.01-1 microM) and (+)-butaclamol (0.01-1 microM) did not increase [3H]dopamine overflow when added alone, but they antagonized the concentration-dependent inhibitory effect of apomorphine (0.1-10 microM). These results suggest that the dopamine inhibitory autoreceptor involved in the modulation of dopamine release from the rabbit retina possesses the pharmacological characteristics of a D-2 dopamine receptor. Maximal stimulation by 30 microM dopamine resulted in a 3-fold increase in adenylate cyclase activity with half-maximal stimulation occurring at a concentration of 2.46 microM. Apomorphine and pergolide elicited a partial stimulation of adenylate cyclase activity. However, at low concentrations both compounds were more potent than dopamine. N,N-di-n-Propyl-dopamine was 30 times less potent than dopamine, and bromocriptine was unable to stimulate adenylate cyclase activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

D-1 and D-2 dopamine receptor blockade: interactive effects in vitro and in vivo

Haloperidol, at low concentrations that block D-2 dopamine (DA) receptors but not D-1 DA receptors (less than 10 microM), potentiated the enhancement of adenylate cyclase activity produced by the D-1 agonist SKF 38393. Low concentrations of haloperidol (less than or equal to 5 microM) also potentiated the K+-evoked release of [3H]acetylcholine from superfused striatal tissue slices. Both of these effects of haloperidol were blocked by nanomolar concentrations of SCH 23390, a D-1 receptor antagonist. In addition, SCH 23390 reduced the ability of haloperidol to antagonize the inhibition of [3H]acetylcholine release produced by the DA agonist apomorphine. By itself, SCH 23390 did not alter either basal adenylate cyclase activity or the K+-evoked release of [3H]acetylcholine. These findings suggest that SCH 23390 can attenuate in vitro responses to D-2 receptor blockade. Likewise, in vivo, very low doses (less than 1 microgram/kg) of SCH 23390 reduced the ability of haloperidol to elevate striatal DA metabolite concentrations and plasma prolactin concentrations. Thus, D-1 receptor blockade may attenuate the effects of D-2 DA receptor blockade both in vitro and in vivo.     

Norepinephrine and BRL 37344 stimulate adenylate cyclase by different receptors in rat brown adipose tissue

The beta adrenergic activation of adenylate cyclase was examined in membrane homogenates of rat interscapular brown adipose tissue (IBAT). In control membranes, isoproterenol and norepinephrine (NE) stimulated adenylate cyclase with activation constants of about 20 and 300 nM, respectively. Exposure of rats to 4 degrees C for 3 days increased the maximal stimulation of adenylate cyclase to these agonists but did not alter the respective activation constants. The beta 1-selective antagonist 1-(2-cyanophenoxy)-3-beta-(3-phenylureido)ethylamino-2-pr opa nol blocked isoproterenol stimulation of adenylate cyclase in control and cold-exposed membranes at a concentration 100 times lower than did the beta 2-selective antagonist erythro-dl-1-(7-methylindan-4-yloxy)-3-isopropylaminobuta n-2-ol. These data indicate that typical adrenergic agonists stimulate IBAT adenylate cyclase via beta 1 receptors. (R*,R*)-4-[2-[2 [9 3-chlorophenyl)-2-hydroxyethyl]amino)propyl) phenyl]phenoxyacetic acid (BRL 37344), an atypical agonist with activity at the beta 3 receptor, stimulated adenylate cyclase in control membranes with an activation constant of approximately 700 nM. Membranes of cold-exposed rats exhibited a high affinity response to BRL 37344 similar to that seen in control membranes and, in addition, a low affinity response. BRL 37344 stimulation of adenylate cyclase was unaffected by 1-(2-cyanophenoxy)-3-beta-(3-phenylureido)ethyl-amino-2-prop anol, whereas stimulation by NE or epinephrine was potently blocked.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of D-2 dopamine receptors in a tumor of the rat anterior pituitary gland

Dopamine receptors in the 7315a transplantable rat anterior pituitary tumor were characterized using radioligand binding assays with [3H]spiroperidol ([3H]SPD) and assays of adenylate cyclase activity. Scatchard analysis of the binding of [3H]SPD yielded linear plots and a Kd value of 73 pM. Studies of the inhibition of the binding of [3H]SPD were performed with a series of competing ligands, including the antagonists domperidone, (+)-butaclamol and sulpiride and the agonists dopamine, bromocriptine and N-propylnorapomorphine. Inhibition curves for the antagonists gave Hill coefficients of approximately 1, consistent with the presence of only a single class of binding sites with a high affinity for [3H]SPD. In contrast, the Hill coefficient for dopamine was significantly less than 1. When assays were carried out in the presence of 300 microM GTP, the inhibition curve for dopamine was shifted to the right and the Hill coefficient increased to approximately 1. An effect of GTP on the affinity of a receptor for agonists is consistent with the existence of at least two agonist affinity states. Inhibition of the binding of [3H]SPD by the partial agonist bromocriptine was not affected when assays were carried out in the presence of GTP. The uniform low affinity of the selective serotonin antagonist ketanserin for these sites indicated that the radioligand was not labeling serotonin-2 receptors in this tissue. A good correlation was observed between the Ki values for competing ligands measured in the tumor and in homogenates of rat striatal tissue. Dopamine was shown to inhibit forskolin-stimulated adenylate cyclase activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Assay of dopamine receptors with [alpha-3H]flupenthixol

Two major classes of dopamine receptors, called D-1 receptors and D-2 receptors, have been identified. [alpha-3H]Flupenthixol has been used as a radioligand for the study of D-1 receptors, which are thought to act through stimulation of adenylate cyclase activity. Previous studies in our laboratory have shown that the D-2 receptors in rat caudate labeled by [3H]spiroperidol also have a high affinity for alpha-flupenthixol. The present experiments show that although Scatchard analysis of the binding of [alpha-3H]flupenthixol is consistent with the presence of a homogeneous population of receptors, subpopulations of these sites can be distinguished by their differing affinities for spiroperidol, which has a Kd for D-1 receptors of about 0.3 microM and a Kd for D-2 receptors of approximately 50 pM. The number of D-1 receptors in rat striatum is approximately 4 times the number of D-2 receptors. D-1 receptors can be studied by including 10 nM spiroperidol in assays carried out with [alpha-3H]flupenthixol, thus blocking the binding of [alpha-3H]flupenthixol to D-2 receptors. The affinity of these receptors for dopamine is decreased by GTP, as has been observed in studies of other receptors whose effects are mediated through changes in adenylate cyclase activity. In the presence of spiroperidol, the Hill coefficients determined from dose-response curves of the inhibition of the binding of [alpha-3H]flupenthixol by antagonists or by agonists in the presence of GTP suggest that the binding reaction obeys simple Michaelis-Menten kinetics for a single class of binding sites.     

Favorable effect of catechol-O-methyltransferase inhibition by OR-462 in experimental models of Parkinson's disease

A selective catechol-O-methyltransferase inhibitor, OR-462, was studied for its ability to affect pharmacokinetic properties of L-dopa after the p.o. administration of the inhibitor to rats and mice. When OR-462 was given to rats at the dose range of 0.3 to 30 mg/kg in conjunction with L-dopa and carbidopa, a dose-related and long-lasting (greater than 5 hr) increase in striatal L-dopa and dopamine levels as well as a reduction in 3-O-methyldopa levels were shown. For a 50% reduction of the 3-O-methyldopa levels a dose of 6 mg/kg of OR-462 was needed. The increase in striatal homovanillic acid, an O-methylated metabolite of dopamine which poorly penetrates the blood brain barrier, indicates that O-methylation was not inhibited in the brain. In order to get the same dopamine levels in striatum the L-dopa dose could be lowered to one-fourth when OR-462 was added. The L-dopa-sparing effect of OR-462 given p.o. was also demonstrated in two behavioral parkinsonian models. OR-462 given at doses of 3 to 30 mg/kg in conjunction with L-dopa and carbidopa, dose-dependently potentiated the L-dopa-induced reversal of hypoactivity in reserpinized mice. Likewise, the same doses of OR-462 caused a marked potentiation of L-dopa-induced contralateral turning behavior in rats with unilateral nigrostriatal lesions produced by 6-hydroxydopamine. The data suggest a possible beneficial effect of OR-462 in the therapy of Parkinson's disease.     

Effect of chronic haloperidol treatment on stimulated synaptic overflow of dopamine in the rat striatum.

In vivo voltammetry was used to assess the change in stimulated striatal dopamine overflow in response to various treatments with the dopamine receptor antagonist haloperidol. Dopamine overflow was induced with stimulating electrodes implanted in the medial forebrain bundle of anesthetized rats while dopamine concentrations were monitored with Nafion-coated, carbon-fiber microelectrodes implanted in the striatum. An acute challenge of haloperidol (0.5 mg kg-1, i.p.) given to naive animals caused stimulated overflow to increase at all stimulation frequencies (10-60 Hz), with the greatest change, 5-fold, occurring at 30 Hz. These results have been compared to those obtained in a different group of rats given daily injections of haloperidol (0.5 mg kg-1, s.c.) for 30 consecutive days. On the 30th day, dopamine striatal tissue levels and uptake kinetics were not altered by this treatment, but 3,4-dihydroxyphenylacetic acid tissue levels were elevated almost 2-fold. A challenge dose of haloperidol (0.5 mg kg-1, i.p.) administered to the animals treated with chronic haloperidol did not elicit a change in stimulated dopamine overflow. In two other groups, rats were withdrawn from 30-day haloperidol treatment for 3 days or 14 days before experimentation. Stimulated dopamine overflow concentrations in both groups were not significantly different from naive animals. When the withdrawn animals were given a haloperidol challenge (0.5 mg kg-1, i.p.), 15- and 12-fold increases in overflow for 3-day and 14-day withdrawal groups, respectively, were observed at a stimulation frequency of 30 Hz. Thus, chronic treatment with haloperidol induces long-lasting effects on the capacity of dopamine receptors to modulate dopamine release.     

Pharmacologic profile of a novel potent direct-acting dopamine agonist, (+)-4-propyl-9-hydroxynaphthoxazine [(+)-PHNO]

The (+)-enantiomer of 1,2,3,4a,5,6-hexahydro-9-hydroxy-4-n-propyl-4H-naphth[1,2-b][ 1,4]-oxazine [(+)-PHNO] is demonstrated to be a potent and direct dopamine (DA) agonist in several in vivo and in vitro test procedures. In vitro (+)-PHNO inhibited binding of [3H]apomorphine (IC50 = 23 nM) or [3H]spiperone (IC50 = 55 nM) to rat striatal membranes. Because (+)-PHNO failed to stimulate adenylate cyclase in carp retina, it was classified as a D-2 agonist. ED50 values (shown in parentheses) derived in DA receptor-related in vivo tests were as follows: in mice, (+)-PHNO produced hypothermia (13 micrograms/kg i.p.) and postural asymmetry in the unilaterally caudectomized animal (4 micrograms/kg i.p.). In the rat, (+)-PHNO produced stereotypy (10 micrograms/kg i.p.) and contralateral turning in 6-hydroxydopamine-lesioned animals (5 micrograms/kg i.p.) that lasted 1 to 3 hr. Whereas both of the latter effects were blocked by haloperidol, prior treatment with depletors of endogenous catecholamines, reserpine or alpha-methylparatyrosine failed to reduce (+)-PHNO-induced stereotypy. The naphthoxazine also produced emesis in beagles (0.05 micrograms/kg i.v.) that was blocked by L-646,462, a peripherally selective DA receptor antagonist. (+)-PHNO was well absorbed when given p.o., producing contralateral turning (10 micrograms/kg) with a ratio of p.o. to i.p. ED50 values of 2. This ratio was much lower than those derived for n-propylnorapomorphine (60) and apomorphine (54). At the DA autoreceptor, (+)-PHNO inhibited the accumulation of dOPA in the gamma-butyrolactone-treated rat (11 micrograms/kg i.p.).(ABSTRACT TRUNCATED AT 250 WORDS)     

Coupling of subtypes of the muscarinic receptor to adenylate cyclase in the corpus striatum and heart

The binding properties of a series of muscarinic antagonists were compared with their ability to antagonize muscarinic receptor mediated inhibition of adenylate cyclase activity in homogenates of the corpus striatum and heart of rats. When measured by the competitive inhibition of the binding of the muscarinic antagonist N-[3H]methylscopolamine, the binding properties of selective muscarinic antagonists in the corpus stratum and cerebral cortex were consistent with a model incorporating a minimum of three populations of muscarinic receptors, a high affinity site for pirenzepine (M1), a high affinity site for AF-DX 116 [11] [2-[ (diethylamino)methyl]-1-piperidinyl] acetyl] -5, 11-dihydro-6H-pyrido [2,3-b] 1,4] benzodiazepine-6-one (M2) and a third population (non-Ml, non-M2 sites) displaying low affinity for the latter antagonists. The results of similar experiments on the heart showed that this tissue contained a uniform population of M2 muscarinic receptors. The binding properties of the M2 receptor in cerebral cortex and corpus stratum were also investigated directly in antagonist [3H] AF-DX 116 competition experiments and, although the high affinity AF-DX 116 site in brain (M2) exhibited selectivity for the cardioselective antagonists AF-DX 116 and gallamine, some differences were noted between M2 sites in brain and heart. The muscarinic adenylate cyclase response in the corpus striatum was relatively insensitive to the M2 selective antagonists AF-DX 116 and gallamine as well as the M1 selective antagonist pirenzepine, suggesting that non-M1, non-M2 sites inhibit adenylate cyclase activity in the corpus striatum. In contrast, the effects of muscarinic antagonists on the muscarinic adenylate cyclase response in the heart were consistent with the postulate that M2 receptors inhibit adenylate cyclase activity in this tissue.     

Modification of brain and spinal cord dopamine D1 receptors labeled with [3H]SCH 23390 after morphine withdrawal from tolerant and physically dependent rats

The effect of morphine tolerance-dependence and abstinence on dopamine D1 receptors in brain regions and spinal cord was determined in rats. Male Sprague-Dawley rats were implanted s.c. under light ether anesthesia with 6 morphine pellets, each containing 75 mg of morphine free base. Rats serving as controls were implanted with placebo pellets. This procedure resulted in the development of tolerance to morphine as evidenced by decreased analgesic response to a challenge dose of morphine. Similarly, the development of physical dependence was evidenced by decreased body weight and colonic temperature after morphine pellet removal (withdrawal). Two sets of animals were used for receptor binding studies. In one, the pellets were left intact and in the other, the pellets were removed. Eighteen hours after pellet removal, the rats were sacrificed. [3H]SCH 23390 [( R-(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1-3-benzapine- 7-ol]) bound to brain region and spinal cord membranes of placebo pellet-implanted rats at a single high affinity site. In rats treated chronically with morphine and then withdrawn, the binding of [3H]SCH 23390 to membranes of spinal cord, hypothalamus and striatum was increased but the binding to amygdalar membranes was decreased in comparison with placebo-treated rats. The changes in binding were due to the changes in Bmax values; the Kd values were unaffected. The behavioral responses to a selective dopamine D1 receptor agonist, SKF 38393 [( 1-phenyl-2,3,4,5-tetrahydro (1H)-3-benzapine-7,8-diol hydrochloride]), were also enhanced in morphine-withdrawn rats when compared to placebo controls.(ABSTRACT TRUNCATED AT 250 WORDS)