Showing papers in "Journal of Pharmacology and Experimental Therapeutics in 1995"

Immunological characterization of urinary 8-epi-prostaglandin F2 alpha excretion in man.

Abstract: F2-isoprostanes are prostaglandin (PG) F2-like compounds that are formed in vivo directly by free radical-catalyzed lipid peroxidation. One of the compounds that can be produced in abundance by such mechanism is 8-epi-PGF2 alpha, a potent vasoconstrictor. We have developed an enzyme immunoassay and a radioimmunoassay for measuring urinary concentrations of 8-epi-PGF2 alpha by raising antibodies against this compound. The antisera presented high titers (> 1/300,000) and provided highly sensitive assays (IC50, 8 and 24 pg/ml, for EIA and RIA, respectively); cross-reactivity with other PG was negligible. The interassay reproducibility of EIA was assessed by measuring the same urine stored frozen in aliquots after solid phase extraction and thin-layer chromatography (17%, n = 13). Measurements of urinary 8-epi-PGF2 alpha by immunoassays were validated using different antisera and by comparison with gas chromatography/mass spectrometry. Healthy volunteers excreted 25 +/- 12 ng of 8-epi-PGF2 alpha/mmol creatinine (n = 19), with no circadian variation over three consecutive 8-hr collection periods (n = 10); preliminary results showed that excretion increased as a function of age. Urinary excretion of 8-epi-PGF2 alpha was unchanged by treatment with two nonsteroidal antiinflammatory drugs, Ibuprofen at 1.2 g/day for 4 days (n = 4) or aspirin as a single administration of 1 g (n = 6). In contrast, the urinary excretion of 11-dehydro-thromboxane B2, a platelet cyclooxygenase-derived metabolite was reduced by more than 80% after aspirin administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Serotonin and norepinephrine uptake inhibiting activity of centrally acting analgesics: structural determinants and role in antinociception.

Abstract: Although it is well established that the analgesic effects of morphine are mediated by opioid receptors, previous studies have shown that some opioids additionally inhibit the uptake of serotonin and norepinephrine. The present investigation of a diverse group of opioids revealed that structurally identifiable subgroups inhibited the neuronal reuptake of these monoamines. Phenanthrene opioids with an oxygen bridge between C4 and C5, such as morphine and naloxone (group I), did not block norepinephrine or serotonin uptake, whereas phenanthrene opioids without the oxygen bridge and the C6-OH moiety, such as levorphanol and levomethorphan (group II), did inhibit uptake, as did nonphenanthrene opioids, such as d-propoxyphene and methadone (group III). Affinity at the mu opioid receptor correlated with antinociceptive potency (r = 0.87, P < .05). Although the antinociceptive activity of the "active enantiomers" of group II and III compounds also correlated with their affinity at the mu opioid receptor (r = 0.85, P = .007), additional consideration of serotonin uptake inhibiting activity (but not of norepinephrine uptake inhibiting activity) significantly improved the correlation between antinociceptive potency and the in vitro activity of these compounds (r = 0.915, P = .0017). Additionally, for group II and III (but not group I) compounds, smaller differences between enantiomers in antinociceptive potency than in mu receptor affinity were noted, presumably because of the contribution of uptake inhibition to the antinociceptive activity of group II and III compounds. Evidence also is provided suggesting a broader role for the combination of mu opioid affinity and 5-hydroxytryptamine uptake inhibition in the activity of other antinociceptive agents.

Nebivolol vasodilates human forearm vasculature: evidence for an L-arginine/NO-dependent mechanism.

Abstract: Nebivolol, a beta 1 selective adrenergic receptor antagonist with additional properties, is a racemic mixture of (S,R,R,R)- and (R,S,S,S)-enantiomers. We investigated its effects on human forearm vasculature. Blood flow was measured using venous occlusion plethysmography during brachial artery infusion of drugs. Interaction between nebivolol and the L-arginine/nitric oxide pathway was investigated via comparison with carbachol (an endothelium-dependent agonist) and nitroprusside, and by coinfusion of a competitive inhibitor of nitric oxide synthase, NG-monomethyl L-arginine (LNMMA) +/- L-arginine. Nebivolol (354 micrograms/min) increased blood flow by 91 +/- 18% (mean +/- SEM, n = 8, P < .01) whereas an equimolar dose of atenolol had no significant effect. L-NMMA (1 mg/min) inhibited vasodilation to nebivolol (by 65 +/- 10%) and carbachol (by 49 +/- 8%) to a significantly greater extent than it reduced responses to nitroprusside. Inhibition of nebivolol response by L-NMMA was abolished by L-arginine (62 +/- 11% inhibition by L-NMMA, 15 +/- 17% inhibition by L-NMMA with L-arginine, 10 mg/min, n = 8). Vasodilation caused by the (S,R,R,R)- and (R,S,S,S)-enantiomers was similar. We conclude that nebivolol vasodilates human forearm vasculature via the L-arginine/nitric oxide pathway.

Pharmacology of a selective cyclooxygenase-2 inhibitor, L-745,337: a novel nonsteroidal anti-inflammatory agent with an ulcerogenic sparing effect in rat and nonhuman primate stomach.

Abstract: Recent studies have shown that there are two isoforms of cyclooxygenases. The constitutive form, cyclooxygenase 1 (COX-1), is believed to be involved in the maintenance of physiological functions. A second isoform, cyclooxygenase 2 (COX-2), has been shown to be induced in inflammation. In the present study, the pharmacology of a selective inhibitor of COX-2, L-745,337 (5-methanesulfonamido-6-(2,4-difluorothiophenyl)-1-indano ne), is described. L-745,337 has IC50 values of 23 +/- 8 nM and > 10 microM for the inhibition of prostaglandin E2 production in whole-cell assays for COX-2 and COX-1, respectively. This compound inhibited carrageenan-induced rat paw edema and rat paw hyperalgesia with ID50 values of 2.00 and 0.37 mg/kg, respectively. In an endotoxin-induced pyresis assay in the rat, L-745,337 significantly reversed the pyretic responses (ID50 = 3.75 mg/kg). L-745,337 did not cause visible gastric lesions in rats at up to 30 mg/kg (4 hr after dosing). In a fecal 51chromium (51Cr) excretion assay to detect gastrointestinal integrity in rats and primates, L-745,337 had no effect at doses up to 100 mg/kg (rat) or after chronic dosing at 20 mg/kg per day for 5 days (primates). In contrast, oral administration of indomethacin, diclofenac or flurbiprofen resulted in substantial increase in fecal 51Cr excretion and/or frank gastric ulceration (rats). L-745,337 significantly inhibited the prostaglandin E2 levels in the inflammatory exudates from the rat pleural cavity after injection with carrageenan but did not inhibit prostaglandin E2 levels in the stomach.(ABSTRACT TRUNCATED AT 250 WORDS)

Ultrarapid hydroxylation of debrisoquine in a Swedish population. Analysis of the molecular genetic basis.

Abstract: Hydroxylation of debrisoquine, catalyzed by the cytochrome P450 CYP2D6 exhibits genetic polymorphism, with large inter-individual differences in metabolic capacity. About 7% of Caucasians carry deficient CYP2D6 alleles and lack the CYP2D6 enzyme (poor metabolizers). We have shown in two Swedish families, individuals carrying duplicated or amplified functional CYP2D6L-genes (CYP2D6L2), causing the opposite phenomenon, ultrarapid metabolism of debrisoquine. In the present study, the occurrence of extra copies of CYP2D6L-alleles was studied in relation to debrisoquine metabolic ratio (MR) in 270 Swedish Caucasians including 64 selected subjects with very rapid metabolism (MR < or = 0.2). Thirteen of the 64 subjects carried a duplicated CYP2D6-gene as identified by EcoRI and XbaI restriction fragment length polymorphism and allele-specific polymerase chain reaction-amplification of genomic DNA. A new allele with three active CYP2D6L-genes was identified, characterized by an XbaI 54 kilobase fragment. This indicates a preference of the CYP2D6L-gene to be amplified compared to other CYP2D6 genes. Only one subject with an MR higher than 0.2 carried the duplicated CYP2D6L-allele, also being heterozygous for the defect CYP2D6B-allele. The overall frequency of the duplicated/amplified CYP2D6-allele was about 1%, and was present in 40% of subjects with MRs < or = 0.1. Thus, other variant CYP2D6-genes may exist that cause increased CYP2D6 activity. In conclusion, a haplotype with duplicated or amplified functional CYP2D6 genes predicts, with high accuracy, ultrarapid metabolism of debrisoquine. Genotyping for this CYP2D locus variant might be of value in patients not responding to generally recommended doses of CYP2D6 substrates, to distinguish between high metabolic capacity and noncompliance.

Lipopolysaccharide-induced changes in plasma nitrite and nitrate concentrations in rats and mice: pharmacological evaluation of nitric oxide synthase inhibitors.

Abstract: The benefits of nitric oxide synthase (NOS) inhibitors in the treatment of endotoxemia or sepsis presumably arise from inhibition of the type II (inducible) NOS. However, inasmuch as the effect of these inhibitors on NOS function in vivo is rarely assessed, NOS activity was evaluated in rats and mice by measuring changes in plasma nitrite and nitrate concentrations ([NOx]) after administration of lipopolysaccharide (LPS). In both species, [NOx] peaked at 20 hr, returning to base line by 48 to 72 hr. The ED50 values (dose that elicited a 50% inhibition of the LPS-dependent increase in [NOx] 6 hr after LPS administration) for L-NG-monomethylarginine acetate, L-NG-nitroarginine methyl ester and aminoguanidine (administered 3 hr after LPS) were 34, 21 and 19 mg/kg in the rat and 32, 5 and 4 mg/kg in the mouse. These compounds also decreased the survival of LPS-challenged animals, which in the case of L-NG-nitroarginine methyl ester was reversed by L-arginine. Dexamethasone (which prevents the induction of type II NOS) also inhibited the LPS-dependent increase in [NOx] with ED50 values of 0.05 mg/kg (rat) and 1 mg/kg (mouse), but did not lead to decreased survival. Thus, inhibition of the type I (neuronal) or type III (endothelial) NOS, rather than the type II isoform, may be a possible mechanism for the animal mortality. These models provide a simple and reproducible means for assessing the in vivo inhibition of type II NOS by various compounds.

Ziprasidone (CP-88,059): a new antipsychotic with combined dopamine and serotonin receptor antagonist activity.

Abstract: Ziprasidone (CP-88,059) is a combined 5-HT (serotonin) and dopamine receptor antagonist which exhibits potent effects in preclinical assays predictive of antipsychotic activity. Whereas the compound is a dopamine antagonist in vitro and in vivo, its most potent action is antagonism of 5-HT2A receptors, where its affinity is an order of magnitude greater than that observed for dopamine D2 sites. Laboratory and clinical findings have led to a hypothesis that antagonism of 5-HT2A receptors in the brain limits the undesirable motor side effects associated with dopamine receptor blockade and improves efficacy against the negative symptoms of schizophrenia. Ziprasidone possesses an in vitro 5-HT2A/dopamine D2 receptor affinity ratio higher than any clinically available antipsychotic agent. In vivo, ziprasidone antagonizes 5-HT2A receptor-induced head twitch with 6-fold higher potency than for blockade of d-amphetamine-induced hyperactivity, a measure of central dopamine D2 receptor antagonism. Ziprasidone also has high affinity for the 5-HT1A, 5-HT1D and 5-HT2C receptor subtypes, which may further enhance its therapeutic potential. The prediction of antipsychotic efficacy without severe motor side effects is supported by the relatively weak potency of ziprasidone to produce catalepsy in animals, contrasted with its potent antagonism of conditioned avoidance responding and dopamine agonist-induced locomotor activation and stereotypy. The compound is well tolerated in animals at doses producing effective dopamine antagonism in the brain. Ziprasidone should be a valuable addition to the treatment of psychotic disorders.

Effects of duloxetine, a combined serotonin and norepinephrine reuptake inhibitor, on central neural control of lower urinary tract function in the chloralose-anesthetized female cat.

Abstract: Because all three components of lower urinary tract control (parasympathetic, sympathetic and somatic) are intimately associated with serotonin (5-hydroxytryptamine [5HT])- and norepinephrine (NE)- containing terminals and receptors, in the present study, we examined the effects of increasing extracellular levels of 5HT and NE with duloxetine, a 5HT and NE reuptake inhibitor, on lower urinary tract function under "normal" or nonirritated conditions (transvesical infusion of saline) and in a model of bladder irritation (i.e., transvesical infusion of 0.5% acetic acid) in chloralose-anesthetized cats. Irritation reduced bladder capacity (to 20% of control) and produced insignificant increases in periurethral electromyographic (EMG) activity compared with nonirritated control animals. Duloxetine produced insignificant increases in bladder capacity and sphincter EMG activity when administered under nonirritated bladder conditions. However, this duloxetine "pretreatment" did prevent the typical acetic acid-induced reductions in bladder capacity and unmasked a marked activation of sphincter EMG activity on acetic acid infusion (by 8-fold). Furthermore, when administered initially under irritated bladder conditions, duloxetine produced dose-dependent increases in bladder capacity (by 5-fold) and increased periurethral striated muscle EMG activity (by 8-fold). The effects on bladder activity were due to central mechanisms since bladder contractions evoked by direct electrical stimulation of efferent fibers in the pelvic nerve were not effected by duloxetine. The effects of duloxetine on bladder capacity were antagonized by methiothepin, a non-selective 5HT receptor antagonist, but not by the other 5HT and NE receptor antagonists examined: LY53857, a 5HT2 antagonist; prazosin, an alpha-1-adrenergic receptor antagonist; idazoxan, an alpha-2-adrenergic receptor antagonist; or propranolol, a beta-adrenergic receptor antagonist. The facilitatory effects of duloxetine on periurethral sphincter EMG were significantly antagonized to various degrees by methiothepin, LY53857 and prazosin but not by idazoxan or propranolol. These results indicate that duloxetine, through inhibition of 5HT and NE reuptake, has weak effects under normal conditions. However, under conditions of bladder irritation, duloxetine suppresses bladder activity through 5HT receptor mechanisms and enhances external urethral sphincter activity through 5HT2 and alpha-1-adrenergic mechanisms.

Repeated administration of cocaine or amphetamine alters neuronal responses to glutamate in the mesoaccumbens dopamine system

Abstract: The development of behavioral sensitization during repeated administration of psychomotor stimulants is a well characterized phenomenon which involves alterations in dopaminergic neurotransmission within the mesoaccumbens system. However, recent evidence indicating that both behavioral sensitization and certain of its neuronal correlates can be prevented by excitatory amino acid receptor antagonists suggests an integral role for glutamate systems in sensitization processes. Therefore, we have determined whether repeated psychomotor stimulant administration can alter responsiveness of the mesoaccumbens dopamine (DA) system to glutamate. After five daily injections of either cocaine (15.0 mg/kg) or d-amphetamine (5.0 mg/kg), rats were subjected to in vivo single cell recording to determine the efficacy of iontophoretically administered glutamate in altering the firing of ventral tegmental area DA neurons and nucleus accumbens neurons. Current-response determinations indicated that the responsiveness of ventral tegmental area DA neurons to glutamate was significantly enhanced in d-amphetamine-treated and cocaine-treated rats in that the neurons entered a state of apparent depolarization block at significantly lower iontophoretic currents. In contrast, nucleus accumbens neurons in psychomotor stimulant-treated rats were significantly less sensitive to the rate-enhancing effects of glutamate. Thus, sensitization appears to be associated with alterations in glutamate transmission at both the origin and termination of the mesoaccumbens DA pathway.

Acute administration of buprenorphine in humans: Partial agonist and blockade effects

Abstract: Buprenorphine, a mixed opioid agonist-antagonist, is being investigated as a treatment for opioid dependence. This study compared the acute subjective and physiological effects of sublingual buprenorphine to those of p.o. methadone over a wide range of doses and compared the ability of both drugs to alter the effects of an opioid challenge. Male inpatient volunteers (n = 9) with histories of opioid abuse participated in this double-blind, double-dummy study. Sublingual buprenorphine (0, 0.5, 2, 8, 16 and 32 mg) and p.o. methadone (3.75, 15 and 60 mg) were administered once weekly according to a Latin-square design, and subjects were monitored on a variety of physiological and subjective measures. Twenty-four hours later, subjects were tested with ascending doses of i.m. hydromorphone (0, 1 and 4 mg) given 45 min apart. Buprenorphine and methadone produced typical opioid agonist effects of long duration, including pupillary constriction, respiratory depression and elevations on subject-rated and observer-rated indices of euphoria, sedation and opioid-like symptoms. The buprenorphine dose-effect curves were nonlinear and maximal effects for most physiological and subjective measures were observed between 4 and 8 mg, with no greater effects observed at higher doses. The methadone dose-effect curves were linear across the range of doses tested. High doses of buprenorphine and methadone both attenuated the response to hydromorphone challenge 24 hr later. These data indicate that there is a ceiling on the effects of buprenorphine in humans that may reduce its abuse liability and increase its safety, and indicate that opioid blockade occurs after acute administration of buprenorphine or methadone.

Effects of acute and repeated administration of antidepressant drugs on extracellular levels of 5-hydroxytryptamine measured in vivo.

Abstract: Extracellular levels of serotonin (5-HT) and the regulation of 5-HT release by the 5-HT1A receptor were examined after single and repeated treatment with different types of antidepressant drugs: the selective 5-HT uptake inhibitor fluoxetine, the selective norepinephrine uptake inhibitor desipramine and the 5-HT2A/2C/alpha 2 receptor antagonist mianserin (each at 15.0 mg/kg). Extracellular levels of 5-HT were measured using in vivo microdialysis in the striatum and hippocampus of rats anesthetized with chloral hydrate. Acute administration of fluoxetine transiently elevated the levels of 5-HT in the striatum and hippocampus; desipramine did not change 5-HT levels, and mianserin slightly decreased 5-HT levels in the hippocampus. Rats were administered these antidepressant drugs for either 1 or 14 days and studied 48 hr after the final injection. Repeated treatment with fluoxetine increased base-line levels of 5-HT in the striatum and hippocampus; repeated treatment with desipramine increased base-line 5-HT levels in the striatum only, and repeated treatment with mianserin did not alter base-line 5-HT levels. Repeated fluoxetine treatment attenuated the ability of the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) to decrease 5-HT release in both the striatum and hippocampus. Repeated desipramine treatment did not significantly alter the effects of 8-OH-DPAT on 5-HT release, but there was a hint of a decreased effect in the hippocampus. Repeated mianserin treatment did not significantly alter the effects of 8-OH-DPAT on 5-HT release, but there was a hint of an increased effect in the striatum. The results of the present study suggest that repeated treatment with antidepressant drugs alters extracellular levels of 5-HT and the ability of 5-HT1A receptors to regulate the release of 5-HT in a regionally selective manner. These changes in the regulation of 5-HT release produced by antidepressant drugs may be associated with their therapeutic effects, because they are caused by repeated rather than acute treatment.

Muscarinic acetylcholine receptor subtypes mediating urinary bladder contractility and coupling to GTP binding proteins.

Abstract: Activation of muscarinic acetylcholine receptors is primarily responsible for urinary bladder emptying. Because multiple subtypes of muscarinic receptors exist, we wished to characterize those present in bladder and ultimately to attribute function to those that regulate bladder contractility, neurotransmitter release and perhaps other cholinergic functions in this tissue. Although the m2 and m3 subtypes could be immunoprecipitated after solubilization from human, rat, rabbit and guinea pig bladder membranes, the m1, m4 and m5 subtypes could not. The m2:m3 ratio was 9:1 in rat bladder but was only 3:1 in the other species examined. Immunoprecipitation of the m2 subtype correlated with the relative levels of high-affinity agonist binding sites measured by competition of carbachol for [3H]N-methylscopolamine binding or measured directly using [3H]oxotremorine-M. In the presence of agonist, but not antagonist, GTP binding proteins could be immunoprecipitated in concert with the m2 or m3 receptors using anti-receptor antibodies. These proteins were members of the Gi and Gq/11 subfamilies for both the m2 and the m3 receptor subtypes. In spite of the preponderance of the m2 receptor in all species studied, Schild analysis using somewhat selective antagonists showed that the pharmacologically defined m3 receptor mediated contractility in strips of rat and rabbit bladder. Thus acetylcholine activates bladder smooth muscle via the m3 receptor subtype, and subsequent contractility may be transduced by guanine nucleotide binding proteins such as the Gi and Gq/11 subfamilies.

(1-(2,5-dimethoxy-4 iodophenyl)-2-aminopropane)-induced head-twitches in the rat are mediated by 5-hydroxytryptamine (5-HT) 2A receptors: modulation by novel 5-HT2A/2C antagonists, D1 antagonists and 5-HT1A agonists.

Abstract: In this study, the involvement of serotonergic and dopaminergic receptors in the modulation of the head-twitch (HTW) response to the 5-hydroxytryptamine (5-HT)2A/5-HT2C agonist, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane, was characterized in rats using novel and selective ligands at 5-HT2A, 5-HT2C, D1, D2 and 5-HT1A receptors. HTW were dose-dependently inhibited by the 5-HT2A/2C antagonists, ritanserin, metergoline, mesulergine, mianserin, ICI 169,369 and LY 58,537, by the preferential 5-HT2A antagonist, ketanserin and by the novel, selective 5-HT2A antagonist, SR 46349B. A further selective 5-HT2A antagonist, MDL 100,907, very potently abolished HTW (ED50 = 0.005 mg/kg). The order of relative potency correlated highly with their affinity at 5-HT2A (r = 0.83) but not 5-HT2C receptors (r = 0.06). In addition, the novel, selective 5-HT2C antagonist, SB 200,646A, failed to abolish HTW and the 5-HT2C agonists/5-HT2A antagonists, 1-(3-chlorophenyl)piperazine and 1-(3-trifluoromethylphenyl)piperazine, blocked, rather than elicited, HTW. The D1 antagonists, SCH 23390, NNC 112, NNC 756, SCH 39166 and A 69024, in this order of relative potency that correlated with their affinity at D1 receptors (r = 0.98), blocked HTW. The D2 antagonists, raclopride, eticlopride and haloperidol also blocked HTW. The 5-HT1A agonists, S 14671, S 14506, 8-hydroxy-2-(di-n-propylamino)tetralin, buspirone, ipsapirone and (+)-flesinoxan, abolished HTW. The action of 8-hydroxy-2-(di-n-propylamino)tetralin was blocked by (-)-tertatolol (ID50 = 4.5 mg/kg), a novel 5-HT1A receptor antagonist. Similarly, (-)-tertatolol attenuated the action of S 14506 and abolished that of S 14671, buspirone and ipsapirone. A role of postsynaptic 5-HT1A receptors in the action of 5-HT1A agonists was suggested by the finding that parachlorophenylalanine (3 x 300 mg/kg, i.p.), which depleted cerebral pools of 5-HT, did not modify the activity of ipsapirone. The present data demonstrate that 5-HT2A receptors mediate HTW in rats and that both D1 and D2 receptors as well as (postsynaptic) 5-HT1A receptors play a role in their expression.

7-(4-[4-(2,3-Dichlorophenyl)-1-piperazinyl]butyloxy)-3,4-dihydro-2(1H)-quinolinone (OPC-14597), a new putative antipsychotic drug with both presynaptic dopamine autoreceptor agonistic activity and postsynaptic D2 receptor antagonistic activity.

Abstract: The effects of 7-(4-[4-(2,3-dichlorophenyl)-1-piperazinyl]butyloxy)-3,4-dihydro-2 (1H)- quinolinone (OPC-14597), a derivative of the dopamine (DA) autoreceptor agonist 7-(3-[4-(2,3-dimethylphenyl)piperazinyl]propoxy)-2(1H)-quinolinone (OPC-4392), on DA receptors were biochemically and behaviorally studied and compared with those of OPC-4392. Both OPC-14597 and OPC-4392 inhibited reserpine- and gamma-butyrolactone (GBL)-induced increase in tyrosine hydroxylase activity in the mouse and rat brain. The effects of OPC-14597 were comparable to those of OPC-4392 and were completely antagonized by haloperidol. OPC-14597, unlike apomorphine, did not evoke postsynaptic DA receptor-stimulating behavioral signs such as hyperlocomotion in the reserpinized mice and contralateral rotation in rats with unilateral striatal 6-hydroxydopamine lesions. Both OPC-14597 and OPC-4392 inhibited such apomorphine-induced postsynaptic behavioral changes as stereotypy and hyperlocomotion in mice and rats and rotation in rats with unilateral striatal kainic acid lesions. The anti-apomorphine effects of OPC-14597 were about 30 to 140 times greater than those of OPC-4392 and were observed at doses that inhibit the increases in tyrosine hydroxylase activity. The affinities of OPC-14597 for 3H-spiperone-labeled D2 receptors in the rat frontal cortex, limbic forebrain and striatum were higher than those of OPC-4392. These results suggest that OPC-14597 is a unique antipsychotic drug candidate, being a DA autoreceptor agonist that has a stronger postsynaptic D2 receptor antagonistic activity than that of OPC-4392.

Sensitization to the conditioned rewarding effects of cocaine: pharmacological and temporal characteristics.

Abstract: An unbiased place preference conditioning procedure was used to determine whether the repeated administration of cocaine results in sensitization to its conditioned rewarding effects. Rats received noncontingent injections of saline or cocaine (10 mg/kg i.p.) for 5 days. Place preference conditioning commenced 72 hr later. A minimum of three drug conditioning sessions was necessary for the establishment of cocaine-induced conditioned place preferences (CPP) in saline-pretreated rats. The minimum dose producing this effect was 10.0 mg/kg. In contrast, pre-exposure to cocaine resulted in significant place preferences occurring after only two drug conditioning sessions. Furthermore, CPP was observed in response to doses as low as 5.0 mg/kg. This shift in the cocaine dose-response curve was apparent when conditioning commenced either 3 or 7, but not 14, days after the cessation of cocaine pretreatment. An increased sensitivity to cocaine was also observed in rats which received only two cocaine (25.0 mg/kg) injections before conditioning and in those which had received either d-amphetamine (0.5 mg/kg) or morphine (5.0 mg/kg) for 5 days. Repeated administration of the D1 dopamine (DA) receptor antagonist, SCH-23390 (0.01-0.05 mg/kg), or the D2 antagonist, raclopride (0.1-1.0 mg/kg), for 5 days did not modify cocaine-induced place conditioning. Administration of SCH-23390 (0.05 mg/kg) in combination with cocaine, however, prevented the sensitized response to cocaine. In contrast, raclopride did not influence the sensitized response to cocaine. These data demonstrate that sensitization occurs to the conditioned rewarding effects of cocaine and suggest an involvement of D1 DA receptors in the development of this phenomenon.

Cloning and characterization of a rat H+/peptide cotransporter mediating absorption of beta-lactam antibiotics in the intestine and kidney.

Abstract: A complementary DNA (cDNA) encoding the rat H+/peptide cotransporter (PepT1) was isolated, and the transport characteristics of orally active beta-lactam antibiotics were assessed by measuring uptake into Xenopus oocytes expressing the rat PepT1. The rat PepT1 cDNA encoded a 710-amino acid protein with 77% identity to the rabbit PepT1. The message for rat PepT1 was approximately 2.9 kilobases and was found predominantly in the small intestine, whereas reverse transcription-polymerase chain reaction amplification revealed that the message was expressed both in the small intestine and in the kidney cortex. The 75-kDa protein was identified by translation of in vitro synthesized transcript of rat PepT1 cDNA by use of rabbit reticulocyte lysates and by Western blot analysis with a specific antibody against the rat PepT1. When expressed in Xenopus oocytes, rat PepT1 stimulated the uptake of ceftibuten (anion) and cephradine (zwitterion) in the presence of an inward H+ gradient, and the expressed uptake was inhibited by excess dipeptides. Kinetic analysis revealed that ceftibuten has 14-fold higher affinity for the rat PepT1 than cephradine. These findings suggest that the rat PepT1 mediates H(+)-coupled uphill transport of the oral beta-lactam antibiotics across the brush-border membranes of intestinal and renal proximal tubular cells.

Methamphetamine-induced hyperthermia and dopaminergic neurotoxicity in mice: pharmacological profile of protective and nonprotective agents.

Abstract: Neurotoxic doses of methamphetamine (METH) can cause hyperthermia in experimental animals. Damage sustained to dopaminergic nerve terminals by this stimulant can be reduced by environmental cooling or by pharmacological manipulation which attenuates the hyperthermia. Many pharmacological agents with very diverse actions protect against METH-induced neuropathology. Several of these compounds, as well as drugs which do not protect, were investigated to determine if there was a relationship between protection and METH-induced hyperthermia. Mice received METH with or without concurrent administration of other drugs and core (i.e., colonic) temperature was monitored during treatment. The animals were sacrificed > or = 5 days later and neostriatal tyrosine hydroxylase activity and dopamine were measured. Core temperature was significantly elevated (> or = 2 degrees C) in mice treated with doses of METH which produced > or = 90% losses in striatal dopamine but not in mice less severally affected (only 50% loss of dopamine). Concurrent treatment of mice with METH and pharmacological agents which protected partially or completely from METH-induced toxicity also prevented the hyperthermic response (i.e., dopamine receptor antagonists, fenfluramine, dizocilpine, alpha-methyl-p-tyrosine, phenytoin, aminooxyacetic acid and propranol). These findings are consistent with the hypothesis that the hyperthermia produced by METH contributes to its neuropathology. However, studies with reserpine, a compound which dramatically lowers core temperature, demonstrated that hyperthermia per se is not a requirement for METH-induced neurotoxicity. Although core temperature was elevated in reserpinized mice treated with METH as compared with reserpinized control mice, their temperatures remained significantly lower than in nonreserpinized control mice. However, the hypothermic state produced in the reserpinized mice did not provide protection from METH-induced toxicity. These data demonstrate that hyperthermia per se contributes to but is not solely responsible for the METH-induced neuropathology.

Interindividual variation in expression of P-glycoprotein in normal human liver and secondary hepatic neoplasms.

Abstract: P-glycoprotein (Pgp), a drug transport protein, pumps many drugs out of hepatocytes. To begin to determine how variation in the level of human hepatic Pgp might influence individual differences in drug disposition, we have used Northern blot and immunochemical analysis to determine the variation in Pgp and in the mRNA for Pgp (MDR1) in liver from 41 individuals. These samples were divided into two groups, normal and perineoplastic (normal liver adjacent to secondary hepatic neoplasms). There was large variation in MDR1 mRNA and Pgp protein expression between all human liver samples. The average amount of Pgp was 2.5-fold greater in normal than in perineoplastic liver. Hepatic Pgp expression was associated with gender, with males expressing 2-fold higher amounts of Pgp than females. There was no correlation between expression of MDR1 and cytochrome P4501A1, but there was a trend toward Pgp and cytochrome P4503A proteins being inversely correlated, although it did not reach statistical significance. MDR1 expression was increased in three of four individuals who had previously received chemotherapy. Pgp expression appeared to be regulated developmentally as MDR1 mRNA was undetectable in six fetal livers, but Pgp was present as early as 1 month postnatally. The level of Pgp was then compared between nine paired samples consisting of seven secondary metastatic hepatic neoplasms, one primary heptocellular carcinoma, one hepatic adenoma and their adjacent normal perineoplastic liver. There was no consistent increase or decrease in Pgp expression in secondary hepatic neoplasms compared with paired perineoplastic liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Amphetamine produces sensitized increases in locomotion and extracellular dopamine preferentially in the nucleus accumbens shell of rats administered repeated cocaine.

Abstract: Alterations in dopamine transmission in the nucleus accumbens, which is composed of two anatomically distinct compartments termed the shell and core, contribute to the expression of cocaine-induced behavioral sensitization. To test potential presynaptic components of behavioral sensitization, the behavioral and neurochemical response to amphetamine administration in the accumbens shell and core was measured at early (days 1-3) and late (days 20-22) withdrawal in rats pretreated with systemic cocaine (15 mg/kg x 2 days, 30 mg/kg x 5 days) or saline. Behavioral sensitization was observed at late, but not early withdrawal when amphetamine was microinjected into the nucleus accumbens shell of cocaine-pretreated rats. There were no significant differences between cocaine- and saline-pretreated animals when behavior was monitored after amphetamine injections into the core at either withdrawal period. After both withdrawal periods, the amphetamine-induced increase in extracellular dopamine was potentiated among cocaine-pretreated animals in the shell by the local administration of amphetamine (0.03, 0.3, 3.0 and 30 microM through the dialysis probe). In the core at early withdrawal there was tolerance to the amphetamine-induced increase in extracellular dopamine in the cocaine group, whereas there was no difference between the repeated saline and cocaine groups at late withdrawal. In a second experiment designed to evaluate potential postsynaptic influences, the D1 partial agonist, SFK-38393 (0.01 or 0.1 microgram/side), was microinjected into the nucleus accumbens core or shell regions after behavioral sensitization to cocaine. Although there was a motor-stimulant effect of SKF-38393 at both withdrawal periods, there was no difference between rats pretreated with repeated cocaine or saline. Collectively, these results demonstrate that the augmentation in dopamine transmission in the nucleus accumbens that is associated with behavioral sensitization is more robust in the shell than the core.

Cannabinoids modulate voltage sensitive potassium A-current in hippocampal neurons via a cAMP-dependent process.

Abstract: Previous studies have shown that cannabinoid receptor analogs increase voltage-dependent potassium A-current (IA) in cultured hippocampal cells. Because cannabinoid receptors inhibit adenylate cyclase, the present study explored whether cAMP played a role in mediating this effect on IA. The specific issue of whether cannabinoid receptor modulation of voltage-dependent IA acts via a cAMP-dependent process was investigated. The cAMP analog, 8-bromo-cAMP, as well as the adenylate cyclase stimulant forskolin, produced concentration-dependent shifts in IA that were opposite those produced by cannabinoid receptor ligands. Moreover, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine also produced a marked negative shift in the steady-state voltage dependence of IA and increased the effect of forskolin on IA. As shown in previous studies, the cannabinoid agonist WIN 55,212-2 increased IA via a decrease in steady-state voltage-dependent inactivation of IA. WIN 55,212-2 also reversed the effects of forskolin on IA. The electrophysiological studies were paralleled by direct assays of cAMP in these cells, where cannabinoids inhibited forskolin-stimulated cAMP by 50% in a pertussis toxin-sensitive manner. The results confirmed that pertussis toxin-sensitive cannabinoid receptor-mediated changes in IA were probably the result of inhibition of adenylate cyclase. The findings are discussed in terms of modulation of IA conductance properties via cannabinoid receptor-mediated inhibition of cAMP levels within the cell.

Metabolism and bioactivation of clozapine by human liver in vitro.

Abstract: The metabolism of clozapine by human liver has been investigated in vitro. Irreversible protein-binding and conjunction with model nucleophiles have been used as markers for bioactivation of clozapine, while stable metabolite formation has been assessed using radiometric HPLC. In all nine liver microsomal preparations investigated, clozapine was extensively metabolized to the stable products desmethylclozapine (range 19%-27.2%), N-oxide (1.5-20.5%) and three polar metabolites (0-20.8%), and was bioactivated to a protein-reactive metabolite (0.6-2.1%). The CYP2D6 genotype did not influence the capacity of the livers to form these metabolites. All metabolic pathways were inhibited by ketoconazole, indicating the involvement of the cytochrome P450 enzymes. Isozyme-selective inhibitor studies demonstrated that whereas demethylation was performed by CYP1A2, N-oxidation and chemically reactive metabolite formation were dependent upon multiple forms of P450. The N-oxide was readily reduced back to clozapine in the presence of NADPH, this conversion being inhibited by ascorbic acid. Glutathione (1 mM) decreased covalent binding by 70%. The amount of putative adduct formed in the presence of glutathione (13.4 +/- 0.9%) was much greater than the covalent binding (mean 1.1 +/- 0.2%). The bioactivation of clozapine was, like the N-oxidation of clozapine, a reversible process. In summary, our results indicate clozapine undergoes extensive metabolism by human liver to both stable and chemically reactive metabolites, the formation of which is catalyzed by the cytochrome P450 enzymes. The role of the reactive metabolite, which may be a free radical, in the pathogenesis of clozapine agranulocytosis and hepatotoxicity requires further study.

Reversal of nerve ligation-induced allodynia by spinal alpha-2 adrenoceptor agonists.

Abstract: After nerve injury, a sympathetically dependent allodynia may be observed. Spinal alpha-2 agonists inhibit preganglionic neurons. The nature of the effect of alpha-2 agonists on allodynia induced by L5 and L6 nerve ligation (Chung model) was thus examined. Rats were implanted with spinal intrathecal catheters directed at the upper lumbar (L1-L2) or the lower cervical (C5-C6) spinal levels. After nerve injury, rats displayed a tactile allodynia (mean withdrawal thresholds, oxymetazoline (14) = guanfacine (17) = 5-bromo-6-(2-imidazolin-2-ylamino)quinoxaline (19) = 2-(2,6-diethylphenylamino)-2-imidazoline (21) = clonidine (22) > morphine (> 30) = methoxamine (> 30 micrograms). The effects of clonidine and 2-(2,6-diethylphenylamino)-2-imidazoline were reversed by the intrathecal injection of yohimbine. At equivalent doses, clonidine delivered systemically, i.c.v. by chronic ventricular guides, or at the level of the cervical spinal cord, produced substantially less antiallodynic action. These results jointly suggest that the sites of the antiallodynic action of spinal alpha-2 agonists are located at the level of the spinal preganglionic neurons and correspond to their ability to diminish preganglionic sympathetic outflow. The failure of morphine to exert an antiallodynic action reflects the fact that 1) opiates act presynaptically on small primary afferents and the allodynia is mediated by large afferent input and 2) opiates, unlike alpha-2 agonists do not have an effect on autonomic outflow.

Ursodeoxycholate (UDCA) inhibits the mitochondrial membrane permeability transition induced by glycochenodeoxycholate: a mechanism of UDCA cytoprotection.

Abstract: Ursodeoxycholate (UDCA), a hydrophilic bile salt, ameliorates hepatocellular injury by toxic bile salts and is used to treat cholestatic liver disease. However, the mechanisms of bile salt-mediated hepatocyte necrosis and UDCA cytoprotection remain unclear. Hepatocyte necrosis is thought to be caused by the mitochondrial membrane permeability transition (MMPT). Thus, the aims of our study were to determine if a toxic bile salt, glycochenodeoxycholate (GCDC) induces the MMPT and if so, whether UDCA prevents the bile salt-induced MMPT. The MMPT was assessed in isolated rat liver mitochondria. Cell viability was measured in isolated rat hepatocytes. GCDC induced the MMPT in a dose-dependent manner. The GCDC-induced MMPT was partially blocked by cyclosporin A plus trifluoperazine, known inhibitors of the MMPT. UDCA also inhibited the GCDC-induced MMPT, and partially blocked the MMPT by phenylarsene oxide, an established mediator of the MMPT. UDCA or cyclosporin A plus trifluoperazine protected against loss of hepatocyte viability during treatment with GCDC. In conclusion, GCDC induces a MMPT; a finding providing a physicochemical explanation for the bioenergetic form of cell necrosis caused by toxic bile salts. UDCA cytoprotection may, in part, be due to inhibition of the bile salt-induced MMPT.

Serotonin dysfunction in the nucleus accumbens of rats during withdrawal after unlimited access to intravenous cocaine.

Abstract: To test the hypothesis that cocaine withdrawal is associated with abnormalities in serotonin (5-HT) neurotransmission, 5-HT concentrations in the nucleus accumbens (NAC) of rats were analyzed with microdialysis both during and after 12 h of unlimited-access intravenous cocaine self-administration. For comparison with previous work, dopamine (DA) levels were also monitored. Self-administration produced sustained increases of both 5-HT and DA to approximately 340% of base line. During the first 6 h of withdrawal, dialysate 5-HT concentrations decreased to 41% of base-line levels obtained before self-administration and to 25% of levels in drug-naive control animals. During the same period, dialysate DA decreased to 72% of presession base-line concentrations but did not decline below control levels. Extracellular 5-HT concentrations were subsequently estimated by use of a quantitative microdialysis technique. After 12 h of self-administration extracellular 5-HT levels were significantly lower (0.6 +/- 0.3 nM) than levels in drug-naive animals (2.0 +/- 0.5 nM) or in rats given only limited-access to cocaine (3 h/day; 1.4 +/- 0.2 nM). Additionally, after 12 h of self-administration low concentrations of intra-accumbens 5-HT applied by reverse dialysis significantly elevated DA efflux. This effect was not observed in either control animals or in rats given only limited access to cocaine. These results suggest that deficient 5-HT neurotransmission may be a significant factor in the cocaine withdrawal symptomatology and provide key information regarding nondopaminergic mechanisms involved in cocaine dependence.

The immunosuppressant leflunomide inhibits lymphocyte proliferation by inhibiting pyrimidine biosynthesis.

Abstract: Leflunomide is a novel immunosuppressive compound that is effective in the treatment of animal models of autoimmune disease and human rheumatoid arthritis. The mechanism of action is unknown. Here we show that leflunomide blocked 1) increases in nucleolar size and number, 2) upregulation of the nuclear protein antigens (PCNA and Ki-67), 3) increases in uridine incorporation and total RNA and DNA content, 4) cell cycle progression and 5) proliferation in mitogen-stimulated rat spleen mononuclear cells and human peripheral blood mononuclear cells (HPBMC). Exogenous uridine reversed the leflunomide-dependent inhibition of the normal increase in total RNA and DNA content in mitogen-stimulated HPBMC and rat spleen cells. Uridine reversed the leflunomide-dependent inhibition of cell cycle progression in stimulated rat cell cultures. Either uridine or cytidine, which can be converted to uridine by cytidine deaminase, reversed the antiproliferative effect of leflunomide in HPBMC. Dihydroorotate accumulated in leflunomide-treated human T-lymphoblastoid cells, suggesting that the compound inhibited the fourth enzyme in the pyrimidine biosynthetic pathway, dihydroorotate dehydrogenase. The results support the hypothesis that the in vitro effects of leflunomide on T-lymphocytes are due to inhibition of de novo pyrimidine synthesis.

SNC 80, a selective, nonpeptidic and systemically active opioid delta agonist.

Abstract: The present study has investigated the pharmacology of SNC 80, a nonpeptidic ligand proposed to be a selective delta agonist in vitro and in vivo. SNC 80 was potent in producing inhibition of electrically induced contractions of mouse vas deferens, but not in inhibiting contractions of the guinea pig isolated ileum (IC50 values of 2.73 nM and 5457 nM, respectively). The delta selective antagonist ICI 174,864 (1 microM) and the mu selective antagonist CTAP (1 microM) produced 236- and 1.9-fold increases, respectively, in the SNC 80 IC50 value in the mouse vas deferens. SNC 80 preferentially competed against sites labeled by [3H]naltrindole (delta receptors) rather than against those labeled by [3H]DAMGO (mu receptors) or [3H]U69, 593 kappa receptors) in mouse whole-brain assays. The ratios of the calculated Ki values for SNC 80 at mu/delta and kappa/delta sites were 495- and 248-fold, respectively, which indicates a significant degree of delta selectivity for this compound in radioligand binding assays. SNC 80 produced dose- and time-related antinociception in the mouse warm-water tail-flick test after i.c.v., i.th. and i.p. administration. The calculated A50 values (and 95% C.I.) for SNC 80 administered i.c.v., i.th. and i.p. were 104.9 (63.7-172.7) nmol, 69 (51.8-92.1) nmol and 57 (44.5-73.1) mg/kg, respectively. The i.c.v. administration of SNC 80 also produced dose- and time-related antinociception in the hot-plate test, with a calculated A50 value (and 95% C.I.) of 91.9 (60.3-140.0) nmol.(ABSTRACT TRUNCATED AT 250 WORDS)

Cloning and pharmacological characterization of human alpha-1 adrenergic receptors: sequence corrections and direct comparison with other species homologues.

Abstract: We have cloned cDNAs encoding three human alpha-1 adrenergic receptor (AR) subtypes and characterized pharmacological properties of the expressed receptor protein. A number of significant sequence corrections have been identified and compared with previously published data, at both nucleotide and amino acid levels; the most major differences occur for the human alpha-1a/dAR. Pharmacological characterization was performed simultaneously using six cloned alpha-1AR subtypes (human and rat alpha-1a/d, human and hamster alpha-1b, human and bovine alpha-1c) stably expressed in rat-1 fibroblasts at approximately equal receptor concentrations (1-2 pmol/mg of total protein). In general, human alpha-1AR subtypes have similar pharmacology compared to their rat, hamster and bovine homologs, although a few minor species differences important for alpha-1AR classification are noted. In addition, much lower inactivation (approximately 20%) by the alkylating agent chloroethylclonidine is noted in this study compared to previous reports for both human and bovine alpha-1cAR membrane preparations. All six alpha-1AR subtypes couple to phosphoinositide hydrolysis in a pertussis toxin-insensitive manner, including the cloned human alpha-1a/dAR which had not been expressed previously. In spite of significant sequence differences between human alpha-1ARs and their other species counterparts, previously established ligand selectivity remains fairly comparable. In summary, these data represent the first side-by-side comparison of pharmacological properties between species homologs of alpha-1AR subtypes and should facilitate the development of alpha-1AR subtype selective drugs for clinical use.

Pharmacological studies of the regulation of chronic FOS-related antigen induction by cocaine in the striatum and nucleus accumbens.

Abstract: Previous work has demonstrated that chronic administration of cocaine induces apparently novel Fos-like transcription factors, termed chronic Fras (Fos-related antigens), in the rat striatum and nucleus accumbens. Induction of these proteins is associated with prolonged increases in AP-1 DNA binding activity that parallel the long half-life of the chronic Fras in brain. The goal of the present study was to characterize pharmacologically the regulation of chronic Fra induction by cocaine. Chronic Fra induction was examined with respect to the cocaine dose, time course and administration intervals used. Cocaine was found to induce the chronic Fras over widely differing treatment regimens in the striatum and nucleus accumbens, although clear differences between the two brain regions were observed. In general, maximal induction occurred with moderate treatment conditions, with more or less intensive treatments resulting in lower levels of chronic Fras. The pharmacological mechanisms underlying cocaine induction of the chronic Fras were also investigated. Pretreatment with a D1 receptor antagonist, which did not affect chronic Fra levels by itself, attenuated cocaine induction of the chronic Fras in striatum and nucleus accumbens. In contrast, treatment with a D2 receptor antagonist alone greatly induced chronic Fra levels, with no further increase seen in response to combined treatment with cocaine. Combined treatment with D1 and D2 receptor agonists, or with amphetamine, led to a strong induction of chronic Fras. Similarly, repeated treatment with a specific dopamine transporter inhibitor increased chronic Fra levels, whereas treatment with a specific serotonin or norepinephrine transporter inhibitor failed to produce this effect. These results support an important role for dopaminergic neurotransmission in the induction of chronic Fras by cocaine. Taken together, the results of the present study provide a more complete understanding of the pharmacological properties underlying cocaine regulation of the chronic Fras, which will assist in identifying the functional role played by these proteins in cocaine action.

Cocaine sensitization in periadolescent and adult rats.

Abstract: Periadolescent rats have been reported to be affected differentially by catecholaminergic agents when compared with younger or adult animals. The present study evaluated the behavioral responsivity of periadolescent (34- to 39-day-old) and adult (60- to 70-day-old) Sprague-Dawley rats of both sexes to i.p. cocaine (Coc) administration (0, 10 or 20 mg/kg, once daily for 4 days). All animals received injections of both saline and Coc every day paired with a different context, with one-half of the animals receiving the drug in the home cage (Coc-Home) and the other half in the testing chamber (Coc-Test). Forty-eight hours after the last drug injection, all animals were challenged with 10 mg/kg i.p. of Coc, and their behavior in the test chamber was scored. As expected, acute Coc induced a prominent increase in a number of behaviors, and this response profile was less marked in periadolescent relative to adult animals. In contrast, Coc-Test animals of both ages showed a clear behavioral sensitization relative to the chronic saline group. No evidence of carry-over effects was found in Coc-Home animals. Females were in general more sensitive than males to acute Coc effects. The development of behavioral sensitization to Coc was a function of age-specific alterations in sensitivity to psychostimulants. Periadolescent rats of both sexes showed sensitization to the locomotor activating effects (matrix crossings) of Coc, whereas a consistent sensitization profile for both stereotyped head scanning and focused sniffing activities were found in adults but not in periadolescents. Chronic Coc reduced body weight and food consumption, particularly in adult males, whereas it did not affect periadolescent patterns. No evidence of sensitization to Coc was found in the hormonal parameters considered.

Neurochemical and functional characterization of the preferentially selective dopamine D3 agonist PD 128907.

Abstract: The present study determined the biochemical and pharmacological effects of PD 128907 [R-(+)-trans-3,4,4a,10b-tetrahydro-4-propyl-2H,5H- [1]benzopyrano[4,3-b]-1,4-oxazin-9-ol], a dopamine (DA) receptor agonist that shows a preference for the human D3 receptor. In transfected Chinese hamster ovary cells (CHO K1), PD 128907 displaced [3H]spiperone in a biphasic fashion which fit best to a two-site model, generating Ki values of 20 and 6964 nM for the high- and low-affinity sites for the D2L receptors and 1.43 and 413 nM for the corresponding sites for the D3 receptors. Addition of sodium and the GTP analog Gpp(NH)p to both the D2L and D3 caused a modest reduction in the affinity of the compound suggestive of an agonist type action. In agonist binding ([3H]N-0437), PD 128907 exhibited an 18-fold selectivity for D3 versus D2L, a selectivity similar to that found with antagonist binding to the high-affinity sites. PD 128907 exhibited only weak affinity for D4.2 receptors (Ki = 169 nM). No significant affinity for a variety of other receptors was observed. PD 128907 stimulated cell division (measured by [3H]thymidine uptake) in CHO p-5 cells transfected with either D2L or D3 receptors exhibiting about a 6.3-fold greater potency in activating D3 as compared to D2L receptors. In vivo the compound was active in reducing DA synthesis both in normal and gamma-butyrolactone (GBL) treated rats; in the GBL model, the decrease was greater in the higher D3-expressing mesolimbic region as compared with striatum which has a lower expression of D3 receptors. PD 128907 decreased DA release (as measured by brain microdialysis) both in rat striatum, nucleus accumbens and medial frontal cortex, as well as in monkey putamen. Behaviorally PD 128907 decreased spontaneous locomotor activity (LMA) in rats at low doses, whereas at higher doses stimulatory effects were observed. PD 128907 at high doses reversed the reserpine-induced decrease in LMA and induced stereotypy in combination with the D1 agonist SKF 38393 indicating postsynaptic DA agonist actions. It is unclear which of the subtypes of DA receptors might be mediating the pharmacological effects of PD 128907. However, the present findings indicating that PD 128907 shows a preference for DA D3 over D2L and D4.2 receptors indicates that its action at low doses may be due to interaction with D3 receptors and at higher doses, with both D2 and D3 receptors.