Selective 5-HT1A Receptor Antagonist WAY Research Articles

Enhanced prefrontal serotonin 5-HT(1A) currents in a mouse model of Williams-Beuren syndrome with low innate anxiety.

Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder caused by the hemizygous deletion of 28 genes on chromosome 7, including the general transcription factor GTF2IRD1. Mice either hemizygously (Gtf2ird1 +/−) or homozygously (Gtf2ird1 −/−) deleted for this transcription factor exhibit low innate anxiety, low aggression and increased social interaction, a phenotype that shares similarities to the high sociability and disinhibition seen in individuals with WBS. Here, we investigated the inhibitory effects of serotonin (5-HT) on the major output neurons of the prefrontal cortex in Gtf2ird1 −/− mice and their wildtype (WT) siblings. Prefrontal 5-HT receptors are known to modulate anxiety-like behaviors, and the Gtf2ird1 −/− mice have altered 5-HT metabolism in prefrontal cortex. Using whole cell recording from layer V neurons in acute brain slices of prefrontal cortex, we found that 5-HT elicited significantly larger inhibitory, outward currents in Gtf2ird1 −/− mice than in WT controls. In both genotypes, these currents were resistant to action potential blockade with TTX and were suppressed by the selective 5-HT1A receptor antagonist WAY-100635, suggesting that they are mediated directly by 5-HT1A receptors on the recorded neurons. Control experiments suggest a degree of layer and receptor specificity in this enhancement since 5-HT1A receptor-mediated responses in layer II/III pyramidal neurons were unchanged as were responses mediated by two other inhibitory receptors in layer V pyramidal neurons. Furthermore, we demonstrate GTF2IRD1 protein expression by neurons in layer V of the prefrontal cortex. Our finding that 5-HT1A-mediated responses are selectively enhanced in layer V pyramidal neurons of Gtf2ird1 −/− mice gives insight into the cellular mechanisms that underlie reduced innate anxiety and increased sociability in these mice, and may be relevant to the low social anxiety and disinhibition in patients with WBS and their sensitivity to serotonergic medicines.Electronic supplementary materialThe online version of this article (doi:10.1007/s11689-010-9044-5) contains supplementary material, which is available to authorized users.

Predatory Stress as an Experimental Strategy to Measure Fear and Anxiety-related Behaviors in Non-human Primates

Natural defense-inducing stimuli are being increasingly exploited as a means to investigate the neural mechanisms underlying normal and pathological anxiety, as well as for the screening of new compounds with potential therapeutic use in human anxiety disorders. Such an approach, frequently used in rodents, has recently been employed in the Marmoset Predator Confrontation Test (MPCT). In this method, marmoset monkeys are individually confronted with a taxidermized predator (wild oncilla cat) in a previously habituated maze environment, while several easily discernable fear/anxiety-related behaviors are measured. Confrontation with the cat stimulus significantly altered ongoing behaviors, each habituating distinctively during repeated exposures; e.g. complete rapid habituation (alarm call), complete slow habituation (exploration, vigilance) or only partial habituation (proximity avoidance). Pharmacological validating studies with diazepam and buspirone induced a significant dose-dependent reversal of the fear-induced proximic avoidance and scratching/scent-marking behaviors, while exploration (smell/lick the maze, leg stand) was found to increase. The neuropeptide substance P and the selective 5-HT1A receptor antagonist WAY100635 resulted in a similar anxiolytic-like profile. The response pattern observed was not influenced by social isolation, handling/manual restraint, novel environment exposure or habituation to the stimulus or its location. Persistent defensive behavior and response pattern to diazepam was observed when naive versus MPCT-experienced marmosets were tested following a recent predatory stress. Taken together, the results indicate that the MPCT is a valuable experimental procedure to measure fear and anxiety-related behaviors in nonhuman primates.

Activation of 5-HT1Areceptors in medullary raphé disrupts sleep and decreases shivering during cooling in the conscious piglet

Activation of 5-HT1A receptors in the medullary raphé decreases sympathetically mediated brown adipose tissue (BAT) thermogenesis and peripheral vasoconstriction when previously activated with leptin, LPS, prostaglandins, or cooling. It is not known whether shivering is also modulated by medullary raphé 5-HT1A receptors. We previously showed in conscious piglets that activation of 5-HT1A receptors with (+/-)-8-hydroxy-2-(dipropylamino)-tetralin (8-OH-DPAT) in the paragigantocellularis lateralis (PGCL), a medullary region lateral to the raphé that contains substantial numbers of 5-HT neurons, eliminates rapid eye movement (REM) sleep and decreases shivering in a cold environment, but does not attenuate peripheral vasoconstriction. Hoffman JM, Brown JW, Sirlin EA, Benoit AM, Gill WH, Harris MB, Darnall RA. Am J Physiol Regul Integr Comp Physiol 293: R518-R527, 2007. We hypothesized that, during cooling, activation of 5-HT1A receptors in the medullary raphé would also eliminate REM sleep and, in contrast to activation of 5-HT1A receptors in the PGCL, would attenuate both shivering and peripheral vasoconstriction. In a continuously cool environment, dialysis of 8-OH-DPAT into the medullary raphé resulted in alternating brief periods of non-REM sleep and wakefulness and eliminated REM sleep, as observed when 8-OH-DPAT is dialyzed into the PGCL. Moreover, both shivering and peripheral vasoconstriction were significantly attenuated after 8-OH-DPAT dialysis into the medullary raphé. The effects of 8-OH-DPAT were prevented after dialysis of the selective 5-HT1A receptor antagonist WAY-100635. We conclude that, during cooling, exogenous activation of 5-HT1A receptors in the medullary raphé decreases both shivering and peripheral vasoconstriction. Our data are consistent with the hypothesis that neurons expressing 5-HT1A receptors in the medullary raphé facilitate spinal motor circuits involved in shivering, as well as sympathetic stimulation of other thermoregulatory effector mechanisms.

The selective 5-HT1A receptor antagonist WAY-100635 inhibits neuronal activity of the ventromedial prefrontal cortex in a rodent model of Parkinson’s disease

The ventral part of the medial prefrontal cortex (mPFC) plays an important role in initiation and control of voluntary movement, mood and cognition. However, after the degeneration of the nigrostriatal pathway, the neuronal activity of the ventral mPFC and the role of serotonin(1A) (5-hydroxytryptamine, 5-HT(1A)) receptors in the firing of the neurons are still unknown. The present study is to investigate the change of neuronal activity in the ventral mPFC and the effect of systemic administration of the selective 5-HT(1A) receptor antagonist WAY-100635 on the activity of the neurons in normal and 6-hydroxydopamine (6-OHDA)-lesioned rats. Single unit responses were recorded extracellularly with glass microelectrodes from ventral mPFC neurons in normal rats and 6-OHDA unilaterally lesioned rats in vivo. 6-OHDA lesion of the substantia nigra pars compacta (SNc) significantly increased the firing rate with no change in the firing pattern of neurons of the ventral mPFC in rats. Systemic administration of WAY-100635 (0.1 mg/kg, i.v.) did not change the mean firing rate and firing pattern of ventral mPFC neurons in normal rats. In contrast, WAY-100635 significantly decreased the mean firing rate of the neurons in rats with 6-OHDA lesion of the SNc. These data suggest that the degeneration of the nigrostriatal pathway results in an increase of neuronal activity of ventral mPFC and dysfunction of 5-HT(1A) receptor.

Antipsychotic-like effect by combined treatment with citalopram and WAY 100635: involvement of the 5-HT2C receptor

Catalepsy occurs following high dopamine (DA) D2 blockade by typical antipsychotic drugs (APDs). We showed that a combination of a high dose of citalopram, a selective serotonin reuptake inhibitor (SSRI) and the selective 5-HT1A receptor antagonist WAY 100635 produces significant catalepsy in rats, similar to APDs. Here, we investigated the potential antipsychotic activity of lower doses of citalopram+WAY 100635, using the conditioned avoidance response (CAR) test. Cataleptogenic liability of the combination was evaluated with the catalepsy test. Citalopram and WAY 100635 in combination, but not when given alone, produced a significant antipsychotic action in CAR without significant catalepsy, similar to the effect of a low dose of the typical APD haloperidol. Pretreatment with a selective 5-HT2C receptor antagonist, SB 242084, completely prevented the citalopram/WAY 100635-induced suppression of CAR indicating an involvement of the 5-HT2C receptor. In summary, treatment with an SSRI/5-HT1A antagonist combination might prove beneficial in psychiatric disorders with psychotic/depressive symptoms.

Postsynaptic 5‐HT1B receptors modulate electroshock‐induced generalised seizures in rats

1. Although an important regulatory role for serotonin (5-HT) in seizure activation and propagation is well established, relatively little is known of the function of specific 5-HT receptor subtypes on seizure modulation. 2. The aim of the present study was to investigate the role of 5-HT(1A, 1B and 1D) receptors in modulating generalised seizures in the rat maximal electroshock seizure threshold (MEST) test. 3. The mixed 5-HT receptor agonists SKF 99101 (5-20 mg kg(-1) i.p.) and RU 24969 (1-5 mg kg(-1) i.p.), 0.5 h pretest, both produced marked dose-related increases in seizure threshold. These agents share high affinity for 5-HT(1A, 1B and 1D) receptors. 4. Antiseizure effects induced by submaximal doses of these agonists were maintained following p-chlorophenylalanine (150 mg kg(-1) i.p. x 3 days)-induced 5-HT depletion. 5. The anticonvulsant action of both SKF 99101 (15 mg kg(-1) i.p.) and RU 24969 (2.5 mg kg(-1) i.p.) was dose-dependently abolished by the selective 5-HT1B receptor antagonist SB-224289 (0.1-3 mg kg(-1) p.o., 3 h pretest) but was unaffected by the selective 5-HT1A receptor antagonist WAY 100635 (0.01-0.3 mg kg(-1) s.c., 1 h pretest). This indicates that 5-HT1B receptors are primarily involved in mediating the anticonvulsant properties of these agents. 6. In addition, the ability of the 5-HT(1B/1D) receptor antagonist GR 127935 (0.3-3 mg kg(-1) s.c., 60 min pretest) to dose-dependently inhibit SKF 99101-induced elevation of seizure threshold also suggests possible downstream involvement of 5-HT1D receptors in the action of this agonist, although confirmation awaits the identification of a selective 5-HT1D receptor antagonist. 7. Overall, these data demonstrate that stimulation of postsynaptic 5-HT1B receptors inhibits electroshock-induced seizure spread in rats.

Activation of 5-HT1A receptors in raphe pallidus inhibits leptin-evoked increases in brown adipose tissue thermogenesis

To elucidate the central neural pathways contributing to the thermogenic component of the autonomic response to intravenous administration of leptin, experiments were conducted in urethane-chloralose-anesthetized, ventilated rats to address 1) the role of neurons in the rostral ventromedial medulla, including raphe pallidus (RPa), in the leptin-evoked stimulation of brown adipose tissue (BAT) sympathetic nerve activity (SNA); and 2) the potential thermolytic effect of 5-hydroxytryptamine(1A) (5-HT(1A)) receptors on RPa neurons that influence BAT thermogenesis. Leptin (1 mg/kg) administration increased BAT SNA by 1,219% of control, BAT temperature by 2.8 degrees C, expired CO(2) by 1.8%, heart rate by 90 beats/min, and mean arterial pressure by 12 mmHg. Microinjection of the 5-HT(1A) receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) into RPa resulted in a prompt and sustained reversal of the leptin-evoked stimulation of BAT SNA, BAT thermogenesis, and heart rate, with these variables returning to their pre-leptin control levels. Subsequent microinjection of the selective 5-HT(1A) receptor antagonist WAY-100635 into RPa reversed the BAT thermolytic effects of 8-OH-DPAT, returning BAT SNA and BAT temperature to the elevated levels after leptin. In conclusion, activation of neurons in RPa, possibly BAT sympathetic premotor neurons, is essential for the increases in BAT SNA, BAT thermogenesis, and heart rate stimulated by intravenous administration of leptin. Neurons in RPa express 5-HT(1A) receptors whose activation leads to reversal of the BAT thermogenic and the cardiovascular responses to intravenous leptin, possibly through hyperpolarization of local sympathetic premotor neurons. These results contribute to our understanding of central neural substrates for the augmented energy expenditure stimulated by leptin.

Effects of chronic paroxetine treatment on dialysate serotonin in 5-HT1B receptor knockout mice.

The role of serotonin (5-HT)1B receptors in the mechanism of action of selective serotonin re-uptake inhibitors (SSRI) was studied by using intracerebral in vivo microdialysis in conscious, freely moving wild-type and 5-HT1B receptor knockout (KO 5-HT1B) mice in order to compare the effects of chronic administration of paroxetine via osmotic minipumps (1 mg per kg per day for 14 days) on extracellular 5-HT levels ([5-HT]ext) in the medial prefrontal cortex and ventral hippocampus. Basal [5-HT]ext values in the medial prefrontal cortex and ventral hippocampus, approximately 20 h after removing the minipump, were not altered by chronic paroxetine treatment in both genotypes. On day 15, in the ventral hippocampus, an acute paroxetine challenge (1 mg/kg i.p.) induced a larger increase in [5-HT]ext in saline-pretreated mutant than in wild-type mice. This difference between the two genotypes in the effect of the paroxetine challenge persisted following chronic paroxetine treatment. Conversely, in the medial prefrontal cortex, the paroxetine challenge increased [5-HT]ext similarly in saline-pretreated mice of both genotypes. Such a challenge produced a further increase in cortical [5-HT]ext compared with that in saline-pretreated groups of both genotypes, but no differences were found between genotypes following chronic treatment. To avoid the interaction with raphe 5-HT1A autoreceptors, 1 micro m paroxetine was perfused locally through the dialysis probe implanted in the ventral hippocampus; similar increases in hippocampal [5-HT]ext were found in acutely or chronically treated wild-type mice. Systemic administration of the mixed 5-HT1B/1D receptor antagonist GR 127935 (4 mg/kg) in chronically treated wild-type mice potentiated the effect of a paroxetine challenge dose on [5-HT]ext in the ventral hippocampus, whereas systemic administration of the selective 5-HT1A receptor antagonist WAY 100635 did not. By using the zero net flux method of quantitative microdialysis in the medial prefrontal cortex and ventral hippocampus of wild-type and KO 5-HT1B mice, we found that basal [5-HT]ext and the extraction fraction of 5-HT were similar in the medial prefrontal cortex and ventral hippocampus of both genotypes, suggesting that no compensatory response to the constitutive deletion of the 5-HT1B receptor involving changes in 5-HT uptake capacity occurred in vivo. As steady-state brain concentrations of paroxetine at day 14 were similar in both genotypes, it is unlikely that differences in the effects of a paroxetine challenge on hippocampal [5-HT]ext are due to alterations of the drug's pharmacokinetic properties in mutants. These data suggest that there are differences between the ventral hippocampus and medial prefrontal cortex in activation of terminal 5-HT1B autoreceptors and their role in regulating dialysate 5-HT levels. These presynaptic receptors retain their capacity to limit 5-HT release mainly in the ventral hippocampus following chronic paroxetine treatment in mice.

Effect of chronic fluoxetine and WAY-100635 treatment on serotonergic neurotransmission in the frontal cortex.

Clinical augmentation strategies have shown that some improvement in antidepressant efficacy can be achieved by combining the beta-adrenergic/serotonin (5-HT)1A/1B receptor antagonist (+/-)pindolol with a selective serotonin reuptake inhibitor (SSRI). This has lead to the hypothesis that a combination of a 5-HT1A receptor antagonist with an SSRI will lead to a faster onset of antidepressant action. Although there is a significant accumulation of acute preclinical data supporting this rationale, until recently, there have been no investigations examining the chronic effects of combining an SSRI with a 5-HT1A receptor antagonist. Here, we determined the chronic effects of fluoxetine (10 mg/kg o.d.), administered in combination with the selective 5-HT1A receptor antagonist WAY-100635 (1 mg/kg b.i.d.), on serotonergic neurotransmission in the frontal cortex using in-vivo microdialysis. Following chronic administration of fluoxetine +/- WAY-100635, functional changes in serotonergic neurotransmission, as well as 5-HT1A autoreceptors, were assessed by administering fluoxetine or (+/-) 8-hydroxy-2-(di-n-propylamino)tetralin [(+/-) 8-OH-DPAT] 24 h after the last chronic dose. Chronic administration of WAY-100635 alone produced no detectable change in the functional status of the 5-HT1A receptor. However, fluoxetine alone produced a time-dependent adaptation in serotonergic transmission such that fluoxetine (acutely administered on day 15) was able to produce a two-fold increase in extracellular 5-HT levels but the decrease in response to 8-OH-DPAT was completely attenuated. These data indicate that the fluoxetine-induced adaptation was mediated by desensitization of the 5-HT1A receptor. WAY-100635 given chronically in combination with fluoxetine blocked the SSRI-induced desensitization of the 5-HT1A receptor. Furthermore, chronic treatment with this combination produced no tolerance in terms of its ability to acutely increase forebrain 5-HT levels. These data suggest that augmentation of an SSRI by combined pharmacotherapy with a 5-HT1A antagonist would be effective upon prolonged exposure.

Effects of the Selective Norepinephrine Reuptake Inhibitor Reboxetine on Norepinephrine and Serotonin Transmission in the Rat Hippocampus

Given that norepinephrine (NE) and serotonin (5-HT) neurons are implicated in the mechanisms of action of antidepressant drugs and both project to the hippocampus, the impact of acute and long-term administration of the selective NE inhibitor reboxetine was assessed on CA3 pyramidal neuron firing in this postsynaptic structure. Cumulative injections of reboxetine (1–4 mg/kg, i.v.) dose-dependently increased the recovery time of the firing of these neurons following iontophoretic applications of NE, but not 5-HT. In rats treated with reboxetine for 2.5 mg/kg/day for 21 days, a robust increase in the recovery time following NE applications was observed, and a small but significant prolongation occurred following 5-HT applications. In controls and reboxetine-treated rats, 1 and 5 Hz stimulations of the afferent 5-HT bundle to the hippocampus, which allows determination of terminal 5-HT1B autoreceptor sensitivity, produced similar frequency-dependent decreases in pyramidal neuron firing in both groups. However, after low and high doses of clonidine (10 and 400 μg/kg, i.v.), which assesses α2-adrenergic auto- and heteroreceptor sensitivity, respectively, only the effect of the high dose of clonidine was attenuated. Interestingly, administration of the selective 5-HT1A receptor antagonist WAY 100,635 induced a 140% increase in basal pyramidal neuron firing in reboxetine as compared to saline-treated rats. This increase in tonic activation of postsynaptic 5-HT1A receptors might be attributable in part to a desensitization of α2-adrenergic heteroreceptors, presumably resulting from sustained NE reuptake inhibition. These results indicate that even a selective NE reuptake inhibitor can modulate 5-HT transmission.

Anxiety-like effects induced by acute fluoxetine, sertraline or m-CPP treatment are reversed by pretreatment with the 5-HT2C receptor antagonist SB-242084 but not the 5-HT1A receptor antagonist WAY-100635

The possible role of 5-HT1A and 5-HT2C receptors in the anxiety induced by fear, acute treatment with SSRI antidepressants or the 5-HT receptor agonist m-CPP were tested in the social interaction anxiety test in male Sprague-Dawley rats. Fluoxetine (2.5-10 mg/kg, i.p.), sertraline (15 mg/kg, i.p.) and m-CPP (0.5-2.0 mg/kg, i.p.) all had an anxiogenic-like profile (decrease in time of total social interaction and increase in self-grooming compared to vehicle) under low-light, familiar arena test conditions. All these effects were reversed by pretreatment with the highly subtype-selective 5-HT2C receptor antagonist, SB-242084 at doses of either 0.05 or 0.2 mg/kg, i.p. In contrast, the selective 5-HT1A receptor antagonist WAY-100635 (0.05 and 0.2 mg/kg, s.c.) failed to reverse SSRI-induced decrease in time of total social interaction, further, it augmented self-grooming response. SB-242084 (0.2 mg/kg) and WAY-100635 (0.05 and 0.2 mg/kg) reversed hypolocomotion caused by the SSRI antidepressants. SB-242084, tested alone against vehicle under high-light, unfamiliar arena test conditions associated with fear, caused significant anxiolysis at 0.2 mg/kg and higher doses. These results suggest that increased anxiety in rodents, and possibly, also in humans (e.g. agitation or jitteriness after SSRIs and panic after m-CPP), caused by acute administration of SSRI antidepressants or m-CPP, are mediated by activation of 5-HT2C receptors. Blockade of 5-HT1A autoreceptors may exacerbate certain acute adverse effects of SSRI antidepressants. Both 5-HT1A and 5-HT2C receptors are involved in the SSRI-induced decrease in locomotor activity. In addition, our studies confirm data that subtype-selective 5-HT2C receptor antagonists have strong anxiolytic actions.

Serotonergic modulation of retinal input to the mouse suprachiasmatic nucleus mediated by 5-HT1B and 5-HT7 receptors.

Serotonin (5-HT) and 5-HT receptor agonists can modify the response of the mammalian suprachiasmatic nucleus (SCN) to light. It remains uncertain which 5-HT receptor subtypes mediate these effects. The effects of 5-HT receptor activation on optic nerve-mediated input to SCN neurons were examined using whole-cell patch-clamp recordings in horizontal slices of ventral hypothalamus from the male mouse. The hypothesis that 5-HT reduces the effect of retinohypothalamic tract (RHT) input to the SCN by acting at 5-HT1B receptors was tested first. As previously described in the hamster, a mixed 5-HT(1A/1B) receptor agonist, 1-[3-(trifluoromethyl)phenyl]-piperazine hydrochloride (TFMPP), reduced the amplitude of glutamatergic excitatory postsynaptic currents (EPSCs) evoked by selectively stimulating the optic nerve of wild-type mice. The agonist was negligibly effective in a 5-HT1B receptor knockout mouse, suggesting minimal contribution of 5-HT1A receptors to the TFMPP-induced reduction in the amplitude of the optic nerve-evoked EPSC. We next tested the hypothesis that 5-HT also reduces RHT input to the SCN via activation of 5-HT7 receptors. The mixed 5-HT(1A/7) receptor agonist, R(+)-8-hydroxy-2-(di-n-propylamino) tetralin hydrobromide (8-OH-DPAT), reduced the evoked EPSC amplitude in both wild-type and 5-HT1B receptor knockout mice. This effect of 8-OH-DPAT was minimally attenuated by the selective 5-HT1A receptor antagonist WAY 100635 but was reversibly and significantly reduced in the presence of ritanserin, a mixed 5-HT(2/7) receptor antagonist. Taken together with the authors' previous ultrastructural studies of 5-HT1B receptors in the mouse SCN, these results indicate that in the mouse, 5-HT reduces RHT input to the SCN by acting at 5-HT1B receptors located on RHT terminals. Moreover, activation of 5-HT7 receptors in the mouse SCN, but not 5-HT1A receptors, also results in a reduction in the amplitude of the optic nerve-evoked EPSC. The findings indicate that 5-HT may modulate RHT glutamatergic input to the SCN through 2 or more 5-HT receptors. The likely mechanism of altered RHT glutamatergic input to SCN neurons is an alteration of photic effects on the SCN circadian oscillator.

Two-lever drug-drug discrimination with the 5-HT1 receptor agonists flesinoxan and eltoprazine.

Previous drug discrimination studies with the 5-HT1 receptor agonists flesinoxan and eltoprazine showed a clear 5-HT1A receptor-mediated effect for flesinoxan and the involvement of both 5-HT1A and 5-HT1B receptors in eltoprazine. However, there was no clear antagonism of eltoprazine's cue, possibly due to the compound nature of the eltoprazine stimulus. In the present experiments, in order to create a specific 5-HT1A vs. 5-HT1B receptor-mediated discrimination, rats were trained to discriminate between flesinoxan and eltoprazine. All rats learned the discrimination readily (mean = 41.3 sessions to criterion). With training doses of 1.0 mg/kg, p.o. flesinoxan and 1.5 mg/kg, p.o. eltoprazine, saline administration resulted in 50% of the responses made on both levers. Substitution tests showed that the flesinoxan stimulus was mediated by the 5- HT1A receptor (8-OH-DPAT, buspirone) and the eltoprazine stimulus probably mediated by the 5-HT1B receptor (anpirtoline, TFMPP, RU-24969). The selective 5-HT1A receptor antagonist WAY-100635 antagonized the flesinoxan cue, and the discriminative stimulus of eltoprazine could be completely antagonized with the 5-HT1B/1D receptor antagonist GR-127935. When the training doses of both flesinoxan and eltoprazine were administered concurrently, complete substitution for eltoprazine was obtained. We conclude that rats can learn to discriminate between two serotonergic drugs with overlapping stimulus properties and that the flesinoxan stimulus is mediated by 5-HT1A receptors and the eltoprazine stimulus, under these particular training conditions, by 5-HT1B receptors.

In vivo actions of the selective 5-HT1A receptor agonist BAY x 3702 on serotonergic cell firing and release.

We investigated the effects of the novel 5-HT1A receptor agonist BAY x 3702 on the serotonergic function in rat brain using single unit recordings in the dorsal raphe nucleus (DR) of anesthetized rats and in vivo microdialysis in freely moving rats. The administration of BAY x 3702 (0.25-4 microg/kg i.v.) suppressed the firing activity of 5-HT neurones. This effect was antagonized by a low dose of the selective 5-HT1A receptor antagonist WAY 100635 (5 microg/kg i.v.). In microdialysis experiments, BAY x 3702 (10-100 microg/ kg s.c.) reduced dose-dependently the 5-HT output in the dorsal and median raphe (MnR) nucleus, dorsal hippocampus (DHPC) and medial prefrontal cortex (mPFC) in a regionally selective manner. Maximal effects were observed in the MnR and mPFC, with reductions to approximately 15% of baseline at a dose of 0.1 mg/kg s.c. The decrease in 5-HT output produced in the DR and DHPC was more moderate, to 45% of baseline at 0.1 mg/kg s.c. BAY x 3702. WAY 100635 (0.3 mg/kg s.c.) completely antagonized the effect of BAY x 3702 (30 microg/kg s.c.). The application of BAY x 3702 in the DR (1-100 microM) reduced the local 5-HT output to 25% of baseline. In rats implanted with two dialysis probes (in DR and mPFC) the application of BAY x 3702 (30 microM) in the DR reduced the 5-HT output in the DR and that in mPFC. These effects were significantly antagonized by the co-perfusion of WAY 100635 (100 microM) in the DR. Overall, these results indicate that the systemic administration of BAY x 3702 reduces the 5-HT release with high potency through the activation of midbrain 5-HT1A receptors.

Rapid visual learning in the rat: effects at the 5-HT1a receptor subtype.

The 5-hydroxytryptamine1a (5-HT1a) receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT; 0.15 mg/kg) impaired rats' rapid visual learning on a computerized maze. This treatment also increased decision time (DT) but the learning impairment was not necessarily a side-effect of slower responding because, in this task, responses made at long DT are more accurate than those at short DT. The selective 5-HT1a receptor antagonist WAY-100635 (0.3 mg/kg) was itself without effect on accuracy, but was effective in reversing effects of 8-OH-DPAT (on both accuracy and DT). Within problems (i.e., over the 40-60 trials of a single discrimination), performance was reduced by treatment with 8-OH-DPAT at all stages of learning. We conclude that this effect is mediated through the 5-HT1a receptor site (rather than through some other serotonergic receptor site or non-specific mechanism) as it was reversible by treatment with WAY-100635. Although it could still arise from behaviourally non-specific effects, the performance deficit finds its best account in terms of the psychological processes necessary to visual learning. Its reversal with WAY-100635 offers support to the hypothesis that 5-HT1a receptor antagonists could improve cognitive function, under conditions of pre-existing impairment due to overactive serotonergic inhibition, as is thought to occur in Alzheimer's disease.

The 5-HT(1A) receptor agonist 8-OH-DPAT reduces rats' accuracy of attentional performance and enhances impulsive responding in a five-choice serial reaction time task: role of presynaptic 5-HT(1A) receptors.

Whilst several studies have investigated the role of serotonergic receptor subtypes in learning and memory, relatively few studies have examined their role in attentional processes. The present study investigated the role of pre- and postsynaptic 5-HT1A receptors on rats' attentional performance in the five-choice serial reaction time task (5-CSRT). Hungry rats were trained in the 5-CSRT task to detect brief (0.5 s) flashes of light presented randomly in one of five locations with a fixed intertrial interval of 5 s paced by the rat. We studied the effects of 8-OH-DPAT, a 5-HT1A receptor agonist, at various subcutaneous (SC) doses (10-100 microg/kg) on measures of rats' discriminative accuracy (the index of attentional functioning) and various behavioural indices of response control and motivation. Manipulations of basic task parameters, intracerebroventricular (ICV) injections of 5,7-dihydroxytryptamine (5,7-DHT) to deplete forebrain 5-HT and treatments with a selective 5-HT1A receptor antagonist WAY 100635 were made in order to determine the behavioural and neural specificity of the effects of 8-OH-DPAT. A dose of 100 microg/kg, but not lower doses, significantly reduced choice accuracy and increased errors of omission, latencies to respond correctly and to collect food reward and premature responses. All these effects were completely blocked by WAY 100635, injected SC 5 min before 8-OH-DPAT at doses from 10-100 microg/kg. WAY 100635 by itself had no effect in the task. Dimming the visual stimuli to one-third of the usual brightness did not modify the effect of 8-OH-DPAT on choice accuracy. Prolonging the stimuli from 0.5 to 1.0 s reversed 8-OH-DPAT's effect on choice accuracy but did not modify the other effects on rats' performance. An ICV injection of 150 microg 5,7-DHT, which depleted forebrain serotonin by 90%, reversed 8-OH-DPAT's effect on choice accuracy but did not modify the effects on errors of omission and latency to make correct responses. Similar effects were found by infusing 1.0 microg/0.5 microl WAY 100635 in the dorsal raphe 5 min before 8-OH-DPAT. 8-OH-DPAT increased the latency to collect the reinforcement; this effect was attenuated by ICV 5,7-DHT and completely antagonized by WAY 100635 in the dorsal raphe. Rats treated with 5,7-DHT or 8-OH-DPAT showed more premature responses and these effects were markedly reduced by the combined treatment. The results suggest that stimulation of presynaptic 5-HT1A receptors is involved in the ability of 8-OH-DPAT to cause attentional dysfunction and enhance impulsivity while slowing of responding and increase in errors of omission mainly depend on stimulation of post-synaptic 5-HT1A receptors.

Pharmacological characterisation of 5-HT receptors positively coupled to adenylyl cyclase in the rat hippocampus.

The pharmacological properties of 5-hydroxytryptamine (5-HT) receptors positively coupled to adenylyl cyclase in the rat hippocampus were investigated using selective agonists and antagonists. 5-HT (0.008-125 microM) stimulated cyclic AMP formation in homogenates of rat hippocampus in a concentration-dependent manner. The maximal increase in cyclic AMP formation occurred at 1 microM (141+/-6%) and the half-maximal effect (EC50) at 50+/-22 nM. Cyclic AMP accumulation induced by 1 microM 5-HT was partly inhibited by the selective 5-HT1A receptor antagonist WAY 100,635 (1 microM), the selective 5-HT4 receptor antagonist SB 203,186 (1 microM), and the 5-HT2A/c/ 5-HT7 receptor antagonist mesulergine (25 microM). WAY 100,635, SB 203,186 and mesulergine inhibited the effect of 5-HT (1 microM) by 47%, 33% and 49%, respectively. The combination of WAY 100,635 (1 microM) with SB 203,186 (1 microM) or mesulergine (25 microM) resulted in stronger inhibition than with each antagonist alone, and the combination of all three antagonists produced almost total blockade (95%) of 5-HT-induced cyclic AMP accumulation. 5-Carboxamidotryptamine (5-CT; 0.008-125 microM), a 5-HT1/5-HT7 receptor agonist, and SDZ 216-454 (0.008-125 microM), a selective 5-HT4 receptor agonist, concentration-dependently stimulated cyclic AMP formation, but the maximal effect of each agonist was smaller than that of 5-HT alone. SDZ 216-454 (5 microM) and 5-CT (5 microM) in combination stimulated cyclic AMP formation in an additive manner. 8-OH-PIPAT and 8-OH-DPAT, two selective 5-HT1A agonists, produced a small but significant increase in cyclic AMP formation at concentrations above 0.04 microM and 10 microM, respectively. These findings suggest that at least three 5-HT receptor subtypes, i.e. 5-HT1A, 5-HT7 and 5-HT4 receptors, are involved in mediating 5-HT-induced cyclic AMP formation in rat hippocampus.

Postsynaptic 5-HT1A receptors control 5-HT release in the rat medial prefrontal cortex.

5-HTt1A receptor agonists reduce the neuronal release of 5-hydroxytryptamine (5-HT) by activation of raphe 5-HT1A autoreceptors. Using in vivo microdialysis in unanesthetized rats, we show that the local application of the selective 5-HT1A receptor agonist 8-OH-DPAT decreased the 5-HT output to approximately 50% of controls in medial prefrontal cortex (mPFC) but not in dorsal hippocampus. The decrease in 5-HT output was counteracted by the concurrent application of the selective 5-HT1A receptor antagonist WAY-100635. This agent also reversed the decrease in 5-HT output elicited by the novel 5-HT1A receptor agonist BAY x 3702 (30 microM) in mPFC and dorsal raphe nucleus. These results indicate that postsynaptic 5-HT1A receptors in mPFC also participate in the control of serotonergic activity.

Venlafaxine: discrepancy between in vivo 5-HT and NE reuptake blockade and affinity for reuptake sites.

Using an in vivo electrophysiological paradigm, venlafaxine and paroxetine displayed similar potency for suppressing the firing activity of dorsal raphe 5-HT neurons (ED50: 233 and 211 microg/kg i.v., respectively), while venlafaxine was three times less potent than desipramine (ED50: 727 and 241 microg/kg i.v., respectively) to suppress the firing activity of locus coeruleus NE neurons. The selective 5-HT1A receptor antagonist WAY 100635 (100 microg/kg, i.v.) reversed the suppressant effect of venlafaxine and paroxetine on the firing activity of 5-HT neurons and the alpha2-adrenoceptor antagonist piperoxane (1 mg/kg, i.v.) reversed those of venlafaxine and desipramine on the firing activity of NE neurons. The ED50 of venlafaxine on the firing activity of 5-HT neurons was not altered (ED50: 264 microg/kg) in noradrenergic-lesioned rats, while the suppressant effect of venlafaxine on the firing activity of NE neurons was greater in serotonergic-lesioned rats (ED50: 285 microg/kg). Taken together, these results suggest that, in vivo, venlafaxine blocks both reuptake processes, its potency to block the 5-HT reuptake process being greater than that for NE. Since the affinities of venlafaxine for the 5-HT and NE reuptake carriers are not in keeping with its potencies for suppressing the firing activity of 5-HT and NE neurons, the suppressant effect of venlafaxine on the firing activity of 5-HT and NE neurons observed in vivo may not be mediated solely by its action on the [3H]cyanoimipramine and [3H]nisoxetine binding sites. In an attempt to unravel the mechanism responsible for this peculiarity, in vitro superfusion experiments were carried out in rat brain slices to assess a putative monoamine releasing property for venlafaxine. (+/-)Fenfluramine and tyramine substantially increased the spontaneous outflow of [3H]5-HT and [3H]NE, respectively, while venlafaxine was devoid of such releasing properties.

Inhibition of hippocampal 5-HT synthesis by fluoxetine and paroxetine: evidence for the involvement of both 5-HT1A and 5-HT1B/D autoreceptors.

Hippocampal serotonin (5-hydroxytryptamine, 5-HT) synthesis, as determined by the accumulation of 5-hydroxytryptophan (5-HTP) following inhibition of L-aromatic amino acid decarboxylase with NSD 1015, was inhibited by systemic administration of the selective serotonin reuptake inhibitors fluoxetine (10 mg/kg i.p.) and paroxetine (3 mg/kg i.p.). Pretreatment of rats with the selective 5-HT1A receptor antagonist WAY 100635 for a period of 7 days using subcutaneously implanted osmotic minipumps (1 mg/kg/day) was sufficient to block the inhibition of 5-HT synthesis following the 5-HT 1A receptor agonist 8-OH-DPAT (0.3 mg/kg s.c.), but failed to inhibit the decrease of hippocampal 5-HT synthesis by fluoxetine (10 mg/kg i.p.) or paroxetine (3 mg/kg i.p.). Similarly, pretreatment of rats with GR 127935 (5 mg/kg i.p.), an antagonist with high affinity for 5-HT1B/D receptors, blocked the reduction of hippocampal 5-HT synthesis following the 5-HT receptor agonist TFMPP (3 mg/kg s.c.) without affecting the reduction of hippocampal 5-HT synthesis by either fluoxetine or paroxetine. In contrast, pretreatment with WAY 100635 (1 mg/kg/day, for 7 days s.c. in osmotic minipumps) in combination with GR 127935 (5 mg/kg i.p.) significantly attenuated the decrease of hippocampal 5-HT synthesis by both fluoxetine and paroxetine. These results indicate that both 5-HT1A and 5-HT1B/1D receptors, which function in the rat as inhibitory somatodendritic and nerve terminal autoreceptors, independently regulate hippocampal 5-HT synthesis and must be simultaneously blocked to prevent the inhibition of 5-HT synthesis by selective serotonin reuptake inhibitors which increase 5-HT availability at both nerve terminals in hippocampus and 5-HT cell bodies in the raphe nuclei.