BGC20-1531, a novel, potent and selective prostanoid EP receptor antagonist: a putative new treatment for migraine headache.

BACKGROUND AND PURPOSE: Prostanoid EP(4) receptor antagonists may have therapeutic utility in the treatment of migraine since EP(4) receptors have been shown to be involved in prostaglandin (PG)E(2)-induced cerebral vascular dilatation, which may be an important contributor to migraine pain. This study reports the pharmacological characterization of BGC20-1531, a novel EP(4) receptor antagonist. EXPERIMENTAL APPROACH: BGC20-1531 was characterized in radioligand binding and in vitro functional assays employing recombinant and native EP(4) receptors. Changes in canine carotid haemodynamics were used to assess the pharmacodynamic profile of BGC20-1531 in vivo. KEY RESULTS: BGC20-1531 exhibited high affinity at recombinant human EP(4) receptors expressed in cell lines (pK(B) 7.6) and native EP(4) receptors in human cerebral and meningeal artery (pK(B) 7.6-7.8) but showed no appreciable affinity at a wide range of other receptors (including other prostanoid receptors), channels, transporters and enzymes (pKi < 5). BGC20-1531 competitively antagonized PGE(2)-induced vasodilatation of human middle cerebral (pK(B) 7.8) and meningeal (pK(B) 7.6) arteries in vitro, but had no effect on responses induced by PGE(2) on coronary, pulmonary or renal arteries in vitro. BGC20-1531 (1-10 mg.kg(-1) i.v.) caused a dose-dependent antagonism of the PGE(2)-induced increase in canine carotid blood flow in vivo. CONCLUSIONS AND IMPLICATIONS: BGC20-1531 is a potent and selective antagonist at EP(4) receptors in vitro and in vivo, with the potential to alleviate the symptoms of migraine that result from cerebral vasodilatation. BGC20-1531 is currently in clinical development for the treatment of migraine headache.

Parasite loss and inhibited development of Teladorsagia circumcincta in relation to the kinetics of the local IgA response in sheep.

Groups of yearling sheep, which had been trickle infected with Teladorsagia circumcincta for 8 weeks and then drenched, were challenged with 50 000 T. circumcincta larvae together with groups of worm-free controls. Fewer parasites and a greater proportion of early fourth stage larvae were recovered from previously infected sheep compared to controls. Worm loss and arrested development were evident by 5 days after challenge whereas growth retardation of developing worms was observed by day 10. In the previously infected sheep a secondary IgA response was observed in the efferent gastric lymph from 5 days post-infection. Western blot analysis showed the lymph IgA to be predominantly dimeric and nonsecretory in nature and that the somatic antigens recognized were predominantly in the 100-250 kDa range. The concentration of IgA in lymph was always higher than in blood and in the previously infected sheep increased fivefold 8 days post-challenge in contrast to blood where IgA levels were unchanged. The timing of the response suggested that it occurred too late to have been the cause of worm loss or arrested development, though it may have retarded the growth of developing parasites.

Plasma level-dependent effects of methylphenidate on task-related functional magnetic resonance imaging signal changes.

RATIONALE: Methylphenidate (MPH) is a dopamine and noradrenaline enhancing drug used to treat attentional deficits. Understanding of its cognition-enhancing effects and the neurobiological mechanisms involved, especially in elderly people, is currently incomplete. OBJECTIVES: The aim of this study was to investigate the relationship between MPH plasma levels and brain activation during visuospatial attention and movement preparation. METHODS: Twelve healthy elderly volunteers were scanned twice using functional magnetic resonance imaging (fMRI) after oral administration of MPH 20 mg or placebo in a within-subject design. The cognitive paradigm was a four-choice reaction time task presented at two levels of difficulty (with and without spatial cue). Plasma MPH levels were measured at six time points between 30 and 205 min after dosing. FMRI data were analysed using a linear model to estimate physiological response to the task and nonparametric permutation tests for inference. RESULTS: Lateral premotor and medial posterior parietal cortical activation was increased by MPH, on average, over both levels of task difficulty. There was considerable intersubject variability in the pharmacokinetics of MPH. Greater area under the plasma concentration-time curve was positively correlated with strength of activation in motor and premotor cortex, temporoparietal cortex and caudate nucleus during the difficult version of the task. CONCLUSION: This is the first pharmacokinetic/pharmacodynamic study to find an association between plasma levels of MPH and its modulatory effects on brain activation measured using fMRI. The results suggest that catecholaminergic mechanisms may be important in brain adaptivity to task difficulty and in task-specific recruitment of spatial attention systems.

Dopaminergic drug effects on physiological connectivity in a human cortico-striato-thalamic system.

Cortico-striato-thalamic (CST) systems are anatomical substrates for many motor and executive functions and are implicated in diverse neuropsychiatric disorders. Electrophysiological studies in rats, monkeys and patients with Parkinson's disease have shown that power and coherence of low frequency oscillations in CST systems can be profoundly modulated by dopaminergic drugs. We combined functional MRI with correlational and path analyses to investigate functional and effective connectivity, respectively, of a prefronto-striato-thalamic system activated by object location learning in healthy elderly human subjects (n = 23; mean age = 72 years). Participants were scanned in a repeated measures, randomized, placebo-controlled design to measure modulation of physiological connectivity between CST regions following treatment with drugs which served both to decrease (sulpiride) and increase (methylphenidate) dopaminergic transmission, as well as non-dopaminergic treatments (diazepam and scopolamine) to examine non-specific effects. Functional connectivity of caudate nucleus was modulated specifically by dopaminergic drugs, with opposing effects of sulpiride and methylphenidate. The more salient effect of sulpiride was to increase functional connectivity between caudate and both thalamus and ventral midbrain. A path diagram based on prior knowledge of unidirectional anatomical projections between CST components was fitted satisfactorily to the observed inter-regional covariance matrix. The effect of sulpiride was defined more specifically in the context of this model as increased strength of effective connection from ventral midbrain to caudate nucleus. In short, we have demonstrated enhanced functional and effective connectivity of human caudate nucleus following sulpiride treatment, which is compatible both with the anatomy of ascending dopaminergic projections and with electrophysiological studies indicating abnormal coherent oscillations of CST neurons in parkinsonian states.

Novel (4-piperazin-1-ylquinolin-6-yl) arylsulfonamides with high affinity and selectivity for the 5-HT(6) receptor.

The discovery of (4-piperazin-1-ylquinolin-6-yl) arylsulfonamides and their binding affinities for a selection of 5-HT and dopamine subreceptors is described. Many compounds show high affinity (pK(i)>8) for the 5-HT(6) receptor and >100-fold selectivity against a range of other receptors. Structure-activity relationships of these compounds are discussed.

Effects of centrally administered orexin-B and orexin-A: a role for orexin-1 receptors in orexin-B-induced hyperactivity.

RATIONALE: Orexin-A and orexin-B are hypothalamic neuropeptides derived from a 130-amino acid precursor, prepro-orexin, and are potent agonists at both the orexin-1 (OX1) and orexin-2 (OX2) receptors. Orexin-A has been ascribed a number of in vivo functions in the rat after intracerebroventricular (ICV) administration, including hyperphagia, neuroendocrine modulation and a role in the regulation of sleep-wake function. The in vivo role of orexin-B is not as clear. OBJECTIVES: To investigate the behavioural, endocrine and neurochemical effects of orexin-B in in-vivo tests. In a number of experiments, these effects were compared with those of orexin-A. METHODS: Experiments were carried out in male, Sprague-Dawley rats with a guide cannula directed towards the lateral ventricle. The effects of orexin-B (ICV) upon grooming behaviour were compared with those of orexin-A. The effects of orexin-B upon the motor activity response to both novel and familiar environments were assessed in an automated activity monitor. Orexin-B was tested upon startle reactivity and body temperature. Further, plasma hormones and [DOPAC+ HVA]/[DA] and [5-HIAA]/[5-HT] ratios in six brain areas were measured 40 min post-orexin-B or orexin-A. RESULTS: The clearest behavioural response to orexin-B was increased motor activity in both novel and familiar environments. Orexin-B-induced hyperactivity was blocked by an OX1 receptor antagonist, SB-334867-A, implicating OX1 receptors in this behavioural response. In common with orexin-A, orexin-B reduced plasma prolactin and failed to influence startle reactivity. However, in contrast with orexin-A, orexin-B increased head grooming but failed to cause a robust whole body grooming response or increase plasma corticosterone levels. Further, orexin-B, but not orexin-A, increased plasma TSH and increased hypothalamic and striatal [5-HIAA]/[5-HT] ratios. CONCLUSIONS: The present study has demonstrated a number of behavioural, neuroendocrine and neurochemical effects of orexin-B that distinguish it from orexin-A. Further, we have demonstrated a role for OX1 receptors in the actions of orexin-B upon motor activity.

Phenyl benzenesulfonamides are novel and selective 5-HT6 antagonists: identification of N-(2,5-dibromo-3-fluorophenyl)-4-methoxy-3-piperazin-1-ylbenzenesulfonamide (SB-357134).

Substituted N-phenyl-4-methoxy-3-piperazin-1-ylbenzenesulfonamides and conformationally restricted analogues have been identified as high affinity and selective 5-HT6 antagonists. Compounds from this series had a range of pharmacokinetic profiles in rat and in general there was a correlation between clearance and CNS penetration. Based on its overall biological profile 2 (SB-357134) was selected for further pre-clinical evaluation.

The effect of SB-236057-A, a selective 5-HT1B receptor inverse agonist, on in vivo extracellular 5-HT levels in the freely-moving guinea-pig.

5-HT1B autoreceptors are involved in the control of extracellular 5-HT levels from both the terminal and cell body regions of serotonergic neurones. In this study we report on the effect of a selective and potent 5-HT1B receptor inverse agonist, SB-236057-A (1'-ethyl-5-(2'-methyl-4'-(5-methyl- 1,3,4-oxadiazolyl-2-yl)biphenyl-4-carbonyl)-2,3,6,7-tetrahydros piro [furo[2,3-f]indole-3,4' -piperidine] hydrochloride), on extracellular 5-HT levels in the cortex and dentate gyrus of the freely-moving guinea-pig, using the technique of in vivo microdialysis. SB-236057-A had ca. 23% bioavailability following oral drug administration. In vivo hypothermia pharmacodynamic assays demonstrated it was brain penetrant with a duration of action in excess of 18 h. SB-236057-A (0.75 mg/kg p.o.) increased extracellular 5-HT levels in the dentate gyrus to a maximum of 167+/-7% of basal but had no effect in the frontal cortex. However, a small increase in cortical 5-HT levels (117+11% of basal) was evident at 2.5 mg/kg p.o. In addition, SB-236057-A (0.75 mg/kg and 2.5 mg/kg p.o.) antagonised the sumatriptan-induced inhibition of extracellular 5-HT levels in the guinea-pig frontal cortex. These differences were attributed to MRN-innervated regions (e.g. dentate gyrus) being more responsive to 5-HT1B receptor-mediated negative feedback than DRN-innervated regions (e.g. frontal cortex). In the dentate gyrus, the increase in 5-HT release induced by SB-236057-A (0.75 mg/kg p.o.) was comparable to that after 14 days of paroxetine (10 mg/kg p.o.) administration, reaching a maximum of 183+/-13% of basal. These data suggest that acute 5-HT1B receptor blockade, by virtue of increased 5-HT release in the dentate gyrus, may provide a rapidly acting antidepressant.

Pharmacological actions of a novel, high-affinity, and selective human dopamine D(3) receptor antagonist, SB-277011-A.

SB-277011-A (trans-N-[4-[2-(6-cyano-1,2,3, 4-tetrahydroisoquinolin-2-yl)ethyl]cyclohexyl]-4-quinolininecarboxamide), is a brain-penetrant, high-affinity, and selective dopamine D(3) receptor antagonist. Radioligand-binding experiments in Chinese hamster ovary (CHO) cells transfected with human dopamine D(3) or D(2 long) (hD(3), hD(2)) receptors showed SB-277011-A to have high affinity for the hD(3) receptor (pK(i) = 7.95) with 100-fold selectivity over the hD(2) receptor and over 66 other receptors, enzymes, and ion channels. Similar radioligand-binding data for SB-277011-A were obtained from CHO cells transfected with rat dopamine D(3) or D(2). In the microphysiometer functional assay, SB-277011-A antagonized quinpirole-induced increases in acidification in CHO cells overexpressing the hD(3) receptor (pK(b) = 8.3) and was 80-fold selective over hD(2) receptors. Central nervous system penetration studies showed that SB-277011-A readily entered the brain. In in vivo microdialysis studies, SB-277011-A (2. 8 mg/kg p.o.) reversed the quinelorane-induced reduction of dopamine efflux in the nucleus accumbens but not striatum, a regional selectivity consistent with the distribution of the dopamine D(3) receptor in rat brain. SB-277011-A (2-42.3 mg/kg p.o.) did not affect spontaneous locomotion, or stimulant-induced hyperlocomotion. SB-277011-A (4.1-42.2 mg/kg p.o.) did not reverse prepulse inhibition deficits in apomorphine- or quinpirole-treated rats, but did significantly reverse the prepulse inhibition deficit in isolation-reared rats at a dose of 3 mg/kg p.o. SB-277011-A (2.5-78. 8 mg/kg p.o.) was noncataleptogenic and did not raise plasma prolactin levels. Thus, dopamine D(3) receptor blockade produces few of the behavioral effects characteristic of nonselective dopamine receptor antagonists. The effect of SB-277011-A on isolation-induced prepulse inhibition deficit suggests that blockade of dopamine D(3) receptors may benefit the treatment of schizophrenia.

Characterization of [(125)I]-SB-258585 binding to human recombinant and native 5-HT(6) receptors in rat, pig and human brain tissue.

SB-258585 (4-Iodo-N-[4-methoxy-3-(4-methyl-piperazin-1-yl)-phenyl]-benzen esulphonamide) is a high affinity ligand at 5-HT(6) receptors. It displays over 100 fold selectivity for the 5-HT(6) receptor over all other 5-HT receptors tested so far. SB-258585 has been radiolabelled, to high specific activity, for its characterization as a 5-HT(6) receptor selective radioligand. [(125)I]-SB-258585 bound, with high affinity, to a single population of receptors in a cell line expressing human recombinant 5-HT(6) receptors. Kinetic and saturation binding experiments gave pK(D) values of 9.01+/-0.09 and 9.09+/-0.02, respectively. In membranes derived from rat or pig striatum and human caudate putamen, [(125)I]-SB-258585 labelled a single site with high levels (>60%) of specific binding. Saturation analysis revealed pK(D) values of 8.56+/-0.07 for rat, 8.60+/-0.10 for pig and 8.90+/-0.02 for human. B(max) values for the tissues ranged from 173+/-23 and 181+/-25 fmol mg(-1) protein in rat and pig striatum, respectively, to 215+/-41 fmol mg(-1) protein in human caudate putamen. The pK(i) rank order of potency for a number of compounds, determined in competition binding assays with [(125)I]-SB-258585, at human caudate putamen membranes was: SB-271046>SB-258585>SB-214111>methiothepin>clozapine>5-Me-OT>5-HT>Ro 04-6790>mianserin>ritanserin=amitriptyline>5-CT>mesulergine. Similar profiles were obtained from pig and rat striatal membranes and recombinant 5-HT(6) receptors; data from the latter correlated well with [(3)H]-LSD binding. Thus, [(125)I]-SB-258585 is a high affinity, selective radioligand which can be used to label both recombinant and native 5-HT(6) receptors and will facilitate further characterization of this receptor subtype in animal and human tissues.

Characterization of SB-271046: a potent, selective and orally active 5-HT(6) receptor antagonist.

SB-271046, potently displaced [(3)H]-LSD and [(125)I]-SB-258585 from human 5-HT(6) receptors recombinantly expressed in HeLa cells in vitro (pK(i) 8.92 and 9.09 respectively). SB-271046 also displaced [(125)I]-SB-258585 from human caudate putamen and rat and pig striatum membranes (pK(i) 8.81, 9.02 and 8.55 respectively). SB-271046 was over 200 fold selective for the 5-HT(6) receptor vs. 55 other receptors, binding sites and ion channels. In functional studies on human 5-HT(6) receptors SB-271046 competitively antagonized 5-HT-induced stimulation of adenylyl cyclase activity with a pA(2) of 8.71. SB-271046 produced an increase in seizure threshold over a wide-dose range in the rat maximal electroshock seizure threshold (MEST) test, with a minimum effective dose of < or =0.1 mg kg(-1) p.o. and maximum effect at 4 h post-dose. The level of anticonvulsant activity achieved correlated well with the blood concentrations of SB-271046 (EC(50) of 0.16 microM) and brain concentrations of 0.01-0.04 microM at C(max). These data, together with the observed anticonvulsant activity of other selective 5-HT(6) receptor antagonists, SB-258510 (10 mg kg(-1), 2-6 h pre-test) and Ro 04-6790 (1-30 mg kg(-1), 1 h pre-test), in the rat MEST test, suggest that the anticonvulsant properties of SB-271046 are likely to be mediated by 5-HT(6) receptors. Overall, these studies demonstrate that SB-271046 is a potent and selective 5-HT(6) receptor antagonist and is orally active in the rat MEST test. SB-271046 represents a valuable tool for evaluating the in vivo central function of 5-HT(6) receptors.

Design and synthesis of trans-N-[4-[2-(6-cyano-1,2,3, 4-tetrahydroisoquinolin-2-yl)ethyl]cyclohexyl]-4-quinolinecarboxamide (SB-277011): A potent and selective dopamine D(3) receptor antagonist with high oral bioavailability and CNS penetration in the rat.

A selective dopamine D(3) receptor antagonist offers the potential for an effective antipsychotic therapy, free of the serious side effects of currently available drugs. Using clearance and brain penetration studies as a screen, a series of 1,2,3, 4-tetrahydroisoquinolines, exemplified by 13, was identified with high D(3) affinity and selectivity against the D(2) receptor. Following examination of molecular models, the flexible butyl linker present in 13 was replaced by a more conformationally constrained cyclohexylethyl linker, leading to compounds with improved oral bioavailability and selectivity over other receptors. Subsequent optimization of this new series to improve the cytochrome P450 inhibitory profile and CNS penetration gave trans-N-[4-[2-(6-cyano-1, 2,3, 4-tetrahydroisoquinolin-2-yl)ethyl]cyclohexyl]-4-quinolinecarbo xamide (24, SB-277011). This compound is a potent and selective dopamine D(3) receptor antagonist with high oral bioavailability and brain penetration in the rat and represents an excellent new chemical tool for the investigation of the role of the dopamine D(3) receptor in the CNS.

Enhancement of 5-HT1B and 5-HT1D receptor antagonist effects on extracellular 5-HT levels in the guinea-pig brain following concurrent 5-HT1A or 5-HT re-uptake site blockade.

The effects of selective serotonin re-uptake inhibitor (SSRI), paroxetine, and 5-HT1A, 5-HT1B and 5-HT1B/1D receptor antagonists on in vivo extracellular 5-HT levels in the guinea-pig frontal cortex and dorsal hippocampus were investigated using the technique of microdialysis. The aim of the study was to further investigate the autoreceptor roles of the 5-HT1A, 5-HT1B and 5-HT1D receptors in the median vs dorsal raphe nuclei. In the frontal cortex, 5-HT1A (WAY 100635, 1 mg/kg i.p.) or 5-HT1B (SB-224289, 4 mg/kg i.p.) receptor antagonists had no effect on extracellular levels of 5-HT, whilst the mixed 5-HT1B/1D receptor antagonist (GR 127935, 0.3 mg/kg i.p) produced a significant decrease in extracellular 5-HT levels. Paroxetine (10 microM) significantly increased extracellular 5-HT levels when perfused locally into the cortex. Administration of SB-224289, followed 120 min later by WAY 100635, had no effect on extracellular 5-HT levels. In contrast, sequential administration of either WAY 100635 and GR 127935, or SB-224289 and paroxetine significantly increased extracellular 5-HT levels. In the dorsal hippocampus, whilst 5-HT1A receptor antagonism elicited by administration of WAY 100635 had no effect, both 5-HT1B and mixed 5-HT1B/1D receptor blockade significantly increased extracellular 5-HT levels. Administration of SB-224289 followed 120 min later with WAY 100635, or WAY 100635 followed 30 min later with GR 127935, potentiated the effect of the three compounds alone, significantly increasing extracellular 5-HT levels. These data demonstrate that to simultaneously increase extracellular 5-HT in both frontal cortex and dorsal hippocampus of the guinea-pig brain concurrent 5-HTA1A, 5-HT1B and 5-HT1D receptor blockade is required. Whereas in the dorsal hippocampus, 5-HT1B receptor blockade is sufficient to elicit an increase in extracellular 5-HT levels.

Orexin A activates locus coeruleus cell firing and increases arousal in the rat.

The localization of orexin neuropeptides in the lateral hypothalamus has focused interest on their role in ingestion. The orexigenic neurones in the lateral hypothalamus, however, project widely in the brain, and thus the physiological role of orexins is likely to be complex. Here we describe an investigation of the action of orexin A in modulating the arousal state of rats by using a combination of tissue localization and electrophysiological and behavioral techniques. We show that the brain region receiving the densest innervation from orexinergic nerves is the locus coeruleus, a key modulator of attentional state, where application of orexin A increases cell firing of intrinsic noradrenergic neurones. Orexin A increases arousal and locomotor activity and modulates neuroendocrine function. The data suggest that orexin A plays an important role in orchestrating the sleep-wake cycle.

Development of a protocol for the automated analysis of amino acids in brain tissue samples and microdialysates.

An automated precolumn derivatisation method has been developed for the measurement of fourteen amino acids in brain tissue and microdialysate samples. The method involves labelling amino acids with naphthalene-2,3-dicarboxaldehyde (NDA) in the presence of cyanide (CN-). The resulting highly stable N-substituted 1-cyanobenz[f]isoindole (CBI) derivatives were separated using a binary gradient elution profile and detected fluorometrically. The order of elution of the derivatised amino acids was confirmed by using liquid chromatography with fluorescence and mass spectrometric detection in tandem. Linear calibration plots were obtained for all amino acids in the range studied (0.2-12.5 microM). The limit of detection for CBI derivatives of amino acids was in the range 5-20 fmol (S/N=2) using a 5 microl injection volume. The method has been used for the measurement of amino acids in microdialysates from rat brain and tissue homogenates from different regions of mouse brain.

Differential effects of 5-HT1B/1D receptor antagonists in dorsal and median raphe innervated brain regions.

The effect of SB-224289 (2,3,6.7-tetrahydro-1'-methyl-5-¿2'-methyl-4'-[(5-methyl-1,2,4-oxadiazol e-3-yl)biphenyl-4-yl]carbonyl¿Furo[2,3-F]-indole-3-spiro-4'-piperidine oxalate) (4 mg/kg i.p., 5-HT1B receptor antagonist), GR 127935 (N-[4-methoxy-3-(4-methyl-1-piperizinyl)phenyl]-2'-methyl-4'-(5-me thyl-1,2,4-oxadiazole-3-yl)[1,1'-biphenyl]-carboxamide) (0.3 mg/kg i.p., 5-HT1B/1D receptor antagonist), and paroxetine (10 mg/kg p.o.) were investigated on extracellular 5-hydroxytryptamine (5-HT) levels in the frontal cortex, striatum and dentate gyrus of the freely moving guinea-pig with microdialysis. In the frontal cortex and striatum (dorsal raphe innervated areas), GR 127935 evoked a significant decrease in extracellular 5-HT, reaching minima of 41+/-12% and 32+/-6% of basal, respectively. This decrease may be explained by antagonism of inhibitory 5-HT1B/1D receptors on raphe cell bodies, leading to a local increase in 5-HT, which, in turn, stimulated 5-HT1A receptors to decrease cell firing, and hence 5-HT release from terminals. In contrast, SB-224289 had no effect on 5-HT levels in either region. In the dentate gyrus (median raphe innervated area), GR 127935 and SB-224289 significantly increased extracellular 5-HT, reaching maxima of 146+/-11% and 151+/-19% of basal, respectively. The ability of both compounds to increase 5-HT levels in the dentate gyrus suggests a lack of 5-HT1B/1D receptors in the median raphe nucleus. Paroxetine produced a small but non-significant increase in extracellular 5-HT in the frontal cortex, and a small decrease in the dentate gyrus. The lack of effect of paroxetine in terminal areas may be due to the limiting effects of cell body 5-HT autoreceptors. In summary, the above data demonstrate that 5-HT1B/1D receptor antagonists increase 5-HT levels in the dentate gyrus, implying that acute administration of 5-HT1B/1D receptor antagonists will achieve a similar effect to chronic selective serotonin re-uptake inhibitor treatment in median raphe innervated areas. This, in turn, suggests that such compounds may be efficacious in the treatment of depression.

The selective 5-HT1B receptor inverse agonist 1'-methyl-5-[[2'-methyl-4'-(5-methyl-1,2, 4-oxadiazol-3-yl)biphenyl-4-yl]carbonyl]-2,3,6,7-tetrahydro- spiro[furo[2,3-f]indole-3,4'-piperidine] (SB-224289) potently blocks terminal 5-HT autoreceptor function both in vitro and in vivo.

5-HT1 receptors are members of the G-protein-coupled receptor superfamily and are negatively linked to adenylyl cyclase activity. The human 5-HT1B and 5-HT1D receptors (previously known as 5-HT1Dbeta and 5-HT1Dalpha, respectively), although encoded by two distinct genes, are structurally very similar. Pharmacologically, these two receptors have been differentiated using nonselective chemical tools such as ketanserin and ritanserin, but the absence of truly selective agents has meant that the precise function of the 5-HT1B and 5-HT1D receptors has not been defined. In this paper we describe how, using computational chemistry models as a guide, the nonselective 5-HT1B/5-HT1D receptor antagonist 4 was structurally modified to produce the selective 5-HT1B receptor inverse agonist 5, 1'-methyl-5-[[2'-methyl-4'-(5-methyl-1,2, 4-oxadiazol-3-yl)biphenyl-4-yl]carbonyl]-2,3,6, 7-tetrahydrospiro[furo[2,3-f]indole-3,4'-piperidine] (SB-224289). This compound is a potent antagonist of terminal 5-HT autoreceptor function both in vitro and in vivo.

Evidence to suggest that agonist modulation of hyperlocomotion is via post-synaptic dopamine D2 or D3 receptors.

It has been suggested that a sub-population of dopamine D3 receptors is located pre-synaptically and these serve as autoreceptors in dopamine projection areas such as the nucleus accumbens/ventral striatum. To study further the physiological role and synaptic location of the dopamine D3 receptor, we have investigated the in vivo effect of the D3/D2 receptor agonist quinelorane on amphetamine-induced hyperactivity and extracellular dopamine release from the nucleus accumbens of the conscious rat. Amphetamine increased dopamine release to 202 +/- 34% of pre-injection control values, but quinelorane at 2.5 micrograms/kg, a dose which effectively blocked amphetamine-induced hyperlocomotion, had no significant effect on amphetamine-induced dopamine release. These data suggest that hyperlocomotion is mediated via post-synaptic rather than pre-synaptic dopamine receptors. Since quinelorane has significant affinity for the dopamine D3 receptor, these effects may be via post-synaptic D3 receptors; however, D2 receptor effects cannot be disregarded. In summary, these data indicate that the quinelorane effect on amphetamine-stimulated hyperlocomotion is not mediated via D3 or D2 autoreceptors, but rather a population of receptors located post-synaptically, which appear to mediate the inhibition of rat locomotor activity.