At present the most investigated 5-HT receptor that has been shown to play a role in the control of micturition is the 5-HT(1A) receptor followed by 5-HT(7), 5-HT(2) and 5-HT(3) receptors. Most experiments focus on the control these receptors have on the parasympathetic outflow to the bladder and the somatic outflow to the external urethral sphincter (EUS) in the rat. Furthermore, 5-HT(1A) and 5-HT(7) receptors have been identified as having an excitatory physiological role in the control of bladder function. 5-HT(1A) receptors act, at least in the rat, at both a spinal (probably a heteroreceptor) and supraspinal (probably an autoreceptor) level, while 5-HT(7) receptors only act at a supraspinal level. Additionally, in the rat, 5-HT administered at a spinal or supraspinal site has an excitatory action, although earlier experiments have shown that activating 5-HT-containing brain areas causes inhibition of the bladder. Recent experiments have also indicated that blockade of the 5-HT(1A) receptor pathway shows rapid tolerance. However, no data exist for the development of tolerance for the 5-HT(7) receptor pathway. Neither receptor seems to play a role in the control of the urethra. Regarding 5-HT(2) receptors, activation of this receptor subtype inhibits micturition, and this inhibitory action may occur at a spinal, supraspinal or both levels. Although no physiological role for 5-HT(2C) receptors can yet be identified, 5-HT(2C) receptors have been implicated in the proposed supraspinal tonically active 5-HT(1A) autoreceptor (negative feedback) pathway. This proposition reconciles the data that central 5-HT-containing pathways are inhibitory to micturition, while 5-HT(1A) receptors, although inhibitory to adenylyl cyclase, have an excitatory function. This is because activation of 5-HT(1A) autoreceptors reduces the release of 5-HT thus reducing the activation of the 5-HT(2C) receptors, which are inhibitory in the control of micturition (disinhibition). Furthermore, 5-HT(2A) receptors in the rat and 5-HT(2C) receptors in the guinea pig cause activation of the EUS. In this respect, 5-ht(5A) receptors have also been identified in Onuf's nucleus, the site of somatic motoneurones controlling this sphincter. In the cat there is very little evidence to indicate that 5-HT receptors are involved in micturition except under pathological conditions in which activation of 5-HT(1A) receptors causes inhibition of micturition. Interestingly, under such conditions 5-HT(1A) receptors cause excitation of the EUS. Nevertheless, spinal <em>5HT</em>(3) receptors have been implicated in the physiological control of micturition in the cat, but not yet in the rat. Overall, the data support the view that 5-HT receptors are important in the control of micturition. However, many more studies are required to fully understand these roles and why there are such species differences.

The facilitatory effect of serotonin (<em>5HT</em>) on hypoglossal (XII) motoneurons is likely to be reduced during rapid eye movement (REM) sleep, when the activity of the brainstem serotonergic system reaches its nadir. Therefore, we assessed the hypothesis that application of exogenous <em>5HT</em> will attenuate the REM sleep-like suppression of XII motoneurons produced in decerebrate cats by pontine microinjections of a cholinergic agonist, carbachol. Microinjections of <em>5HT</em> or 5-carboxamidotryptamine into the XII nucleus increased XII nerve activity to 182 +/- 53% (standard deviation; SD) of control. Subsequent pontine microinjections of carbachol reduced XII nerve activity by 55 +/- 21% of the pre-<em>5HT</em> level (n = 12). Microinjections of methysergide (a <em>5HT</em> antagonist) into the XII nucleus reduced XII nerve activity to 54 +/- 17% of the pre-methysergide control (n = 6). Pontine carbachol injections after methysergide further reduced XII nerve activity by 49 +/- 20% of the pre-methysergide level. Treatments with both agonists and the antagonist attenuated the carbachol-induced decrease when compared to two previous studies using the same model: 1) In experiments with no injections of serotonergic agents, pontine carbachol injections decreased XII nerve activity by 90 +/- 6% of control. 2) After enhancement of XII nerve activity by inhibitory amino acid antagonists (to 135 +/- 60%), the subsequent carbachol-induced decrease was even larger, 112 +/- 62% of control. We propose that serotonergic excitation can significantly attenuate the REM sleep-like suppression of XII nerve activity, and that this is achieved, in part, by substituting for the decreased endogenous <em>5HT</em> in the XII nucleus. The study also demonstrates that other, non-serotonergic, mechanisms also contribute to the carbachol-induced suppression.

Stimulation in the nucleus raphe magnus (NRM) inhibits transmission of nociceptive information within the spinal cord through activation of bulbospinal pathways. This study used microdialysis in combination with high pressure liquid chromatography to measure the release of serotonin (<em>5HT</em>) and several amino acids, including glutamate, aspartate and glycine, from the lumbar dorsal horn during electrical stimulation within the NRM in the alpha-chloralose anesthetized cat. Observed release of putative neurotransmitters was correlated with inhibition of nociceptive projection neurons recorded from sites within 800 microns rostral or caudal to the dialysis fiber. NRM stimulus parameters considered to preferentially activate myelinated fibers caused inhibition of nociceptive evoked activity, and increased the release of excitatory amino acids and glycine within the spinal cord, with no detectable release of <em>5HT</em>. When pulse widths were lengthened and unmyelinated fibers were also activated, increases in <em>5HT</em> in the spinal dialysate were observed as well. Strychnine administered through the dialysis fiber (0.02-1 mM) antagonized NRM-induced inhibition when <em>5HT</em> release was not detected. Inhibition produced by stimulation that increased <em>5HT</em> concentrations was relatively strychnine resistant. These results point to a raphe-spinal inhibitory pathway that is not dependent on <em>5HT</em>, the activation of which results in the spinal release of glycine.

There is evidence that repeated electroconvulsive shocks (ECS) in rats potentiate dopamine (DA)-, serotonin (<em>5HT</em>)- and alpha-noradrenergic (alpha-NA)-mediated drug-induced behaviour and reduce opiate-induced behaviours. These studies suggest changes at the level of the receptor or beyond. However, high affinity in vitro 3H-ligand binding studies in brain membranes from ECS-treated control rats failed to demonstrate generalized ECS-induced changes in <em>5HT</em>, DA, alpha-NA or opiate receptor binding. Binding of the beta-receptor ligand dihydroalprenolol (3H-DHA) was significantly reduced in ECS-treated rat brain membranes. This may be secondary to effects on NA neurones since ECS-induced reduction of 3H-DHA binding did not occur in animals with 6-hydroxydopamine-induced depletion of cortical noradrenaline. In conjunction with other studies, the results suggest that electroconvulsive therapy may have a noradrenergic mechanism of action.

This study examined the cellular actions of the anti-migraine drug sumatriptan, on neurons in the substantia gelatinosa of the spinal trigeminal nucleus pars caudalis. Sumatriptan inhibited the miniature EPSC (mEPSC) rate in a dose dependent fashion, with an EC(50) of 250 nM. Sumatriptan (3 microM) inhibited the mEPSC rate by 36%, without altering the mEPSC amplitude. This effect was partially reversed by the <em>5HT</em>(1D) specific antagonist BRL15572 (10 microM). In contrast, the <em>5HT</em>(1B) agonist CP93129 (10 microm) did not alter the mEPSC rate. Furthermore, sumatriptan (3 microM) decreased the amplitude of electrically evoked EPSCs (eEPSC) by 40%. After incubating the slices in ketanserin (an antagonist which shows selectivity for <em>5HT</em>(1D) over <em>5HT</em>(1B) receptors) sumatriptan had little effect on eEPSC amplitude. In control conditions paired stimuli resulted in paired pulse depression (PPD; the ratio eEPSC(2)/eEPSC(1)=0.7+/-0.01), whilst in the presence of sumatriptan the PPD was blocked (ratio eEPSC(2)/eEPSC(1)=0.9+/-0.1). Sumatriptan produced no post-synaptic membrane current and had no significant effect on membrane conductance over a range of membrane potentials (-60 to -130 mV). RT-PCR experiments revealed the presence of mRNA for both <em>5HT</em>(1D) and <em>5HT</em>(1B) receptor subtypes in the trigeminal ganglia and subnucleus caudalis. These data suggest that sumatriptan acts pre-synaptically on trigeminal primary afferent central terminals to reduce the probability of release of glutamate, and that this action is mediated through <em>5HT</em>(1D) receptors.

Serotonin (5-HT) can either increase or decrease anxiety-like behaviour in animals, actions that depend upon neuroanatomical site of action and 5-HT receptor subtype. Although systemic studies with 5-HT(2) receptor agonists and antagonists suggest a facilitatory role for this receptor subtype in anxiety, somewhat inconsistent results have been obtained when such compounds have been directly applied to limbic targets such as the hippocampus and amygdala. The present study investigated the effects of the 5-HT(2B/2C) receptor agonist mCPP bilaterally microinjected into the dorsal hippocampus (DH: 0, 0.3, 1.0 or 3.0nmol/0.2microl), the ventral hippocampus (VH: 0, 0.3, 1.0 or 3.0nmol/0.2microl) or the amygdaloid complex (0, 0.15, 0.5, 1.0 or 3.0nmol/0.1microl) in mice exposed to the elevated plus-maze (EPM). Test sessions were videotaped and subsequently scored for conventional indices of anxiety (percentage of open arm entries and percentage of open arm time) and locomotor activity (closed arm entries). Results showed that mCPP microinfusions into the DH or VH failed to affect any behavioural measure in the EPM. However, when injected into the amygdaloid complex, the dose of 1.0nmol of this <em>5HT</em>(2B/2C) receptor agonist increased behavioural indices of anxiety without significantly altering general activity levels. This anxiogenic-like effect of mCPP was selectively and completely blocked by local injection of a behaviourally-inactive dose of SDZ SER-082 (10nmol/0.1microl), a preferential 5-HT(2C) receptor antagonist. These data suggest that <em>5HT</em>(2C) receptors located within the amygdaloid complex (but not the dorsal or ventral hippocampus) play a facilitatory role in plus-maze anxiety in mice.

To understand the mechanism of allosteric coupling between the ligand-binding domain and the ion channel of the Cys-loop ligand-gated ion channels (LGICs), we fused the soluble acetylcholine-binding protein (AChBP), which lacks an ion channel, to either the cationic serotonin type-3A ion channel (<em>5HT</em>(3A)) or the anionic glycine ion channel. Both linear chimeras expressed in HEK-293 cells display high affinity for the nicotinic agonist epibatidine (K(D) = 0.2-0.5 nM), but are not targeted to the cell surface. Only after substituting a ring of three loops located at the putative membrane side of the AChBP three-dimensional structure by the homologous residues of <em>5HT</em>(3A), the resulting chimera AChBP(ring)/<em>5HT</em>(3A) (i) still displayed on intact cells an apparent high affinity for epibatidine, yet with a fourfold decrease (K(D) = 2.1 nM), (ii) displayed a high proportion of low affinity sites (11 +/- 7 microM) for the resting state stabilizing competitive antagonist alpha-bungarotoxin and (iii) was successfully targeted to the cell surface, as seen by immunofluorescence labelling. The AChBP(ring)/<em>5HT</em>(3A) chimera forms a pentameric structure, as revealed by sucrose gradient sedimentation. However, no whole-cell patch-clamp currents were detectable. Interestingly, binding assays with membrane fragments prepared from cells expressing AChBP(ring)/<em>5HT</em>(3A) showed a decrease in the apparent affinity for the agonists nicotine and epibatidine (5-fold), concomitant with an increase in the proportion of high-affinity sites (48 +/- 1 nM) for alpha-bungarotoxin. These results indicate that fusion of AChBP to an ion channel forms a pentameric receptor exposed to the cell surface and able to convert between discrete allosteric states, but stabilized in a high affinity state for epibatidine that likely corresponds to a desensitized form of LGICs. These artificial chimeras might offer a useful system to investigate signal transduction in LGICs.

The steady state concentration of dopamine (DA), norepinephrine (NE), and serotonin (<em>5HT</em>) was determined and turnover estimated in several brain regions of young (3-4 months) and old (21 months) male Wistar rats. An estimate of DA and NE turnover was obtained by determining their depletion rates after treatment with alpha-methylpara-tyrosine, a tyrosine hydroxylase inhibitor. Serotonin turnover was estimated by determining its rate of increase after monoamine oxidase inhibition with pargyline. In old males, medial basal hypothalamic (MBH) DA concentration and depletion rate were significantly lower than in young males. DA concentration of the remaining hypothalamus also was lower in old than in young males, but depletion rates were not different. DA concentration and depletion rate in the olfactory tubercle were the same in both age groups. The steady state concentration of NE in the MBH and remaining hypothalamus, and the hypothalamic NE depletion rate, were significantly lower in old than in young animals. In both brain and hypothalamus, steady state concentrations of <em>5HT</em> were the same in young and old rats, but by 30 min after monoamine oxidase inhibition with pargyline, hypothalamic, but not brain, <em>5HT</em> increased more in old than in young males. This may indicate a greater turnover of <em>5HT</em> in the hypothalamus of old than of young males. In non-drug treated (control) old male rats, serum LH and FSH were lower, serum prolactin was higher and serum TSH was the same in young male rats. These data suggest that a decrease in catecholamine and an increase in serotonin metabolism occur in the hypothalamus of old male rats. These changes may be related to the decrease in release of gonadotropins and increase in release of prolactin observed in these old male rats.

The current study assessed whether various co-administered serotonin (5-HT) receptor antagonists could prevent some of the acute behavioral effects of 3,4-methylenedioxymethamphetamine (MDMA, "Ecstasy") in rats. In the social interaction test, MDMA (5 mg/kg) significantly increased the duration of total social interaction between two conspecifics meeting for the first time. Microanalysis showed that MDMA increased adjacent lying and approach behaviours while reducing anogenital sniffing. MDMA (5 mg/kg) also caused elements of the serotonin syndrome including low body posture and piloerection. In the emergence test, MDMA significantly increased hide time and emergence latency indicating increased anxiety-like behavior. Pretreatment with the <em>5HT</em> 1A receptor antagonist, WAY 100635 (1 mg/kg), prevented MDMA-induced increases in social interaction and markers of the serotonin syndrome while the 5-HT 1B receptor antagonist GR 55562 (1 mg/kg) and 5-HT 2A receptor antagonist ketanserin (1 mg/kg) were ineffective. The 5-HT 2B/2C receptor antagonist, SB 206553 (2 mg/kg), prevented MDMA-induced prosocial effects but caused pronounced thigmotaxis (hyperactivity at the periphery of the testing chamber). The anxiogenic effect of MDMA on the emergence test was not prevented by pretreatment with any of the 5-HT receptor antagonists tested. These results indicate that prosocial effect of MDMA may involve 5-HT 1A and possibly 5-HT 2B/2C receptors. In contrast, MDMA-induced generalised anxiety, as measured by the emergence test, seems unlikely to involve the 5-HT 1A, 5-HT 1B or 5-HT 2A, 5-HT 2B or 5-HT 2C receptors.

A new phenylpiperidine derivative, FG4963, and several tricyclic antidepressants were compared in various in vitro and in vivo tests for central <em>5HT</em>- and NA-uptake inhibition. FG4963 was found to be a <em>5HT</em>-pump blocker with activity similar to that of chlorimipramine. FG4963 inhibited NA-uptake less than the most potent tricyclic thymoleptics. In contrast to imipramine derivatives FG4963 was a weak inhibitor of peripheral NA-uptake. FG4963 seems to produce central <em>5HT</em>-potentiation without affecting organ functions regulated by the autonomic nervous system as much as tricyclic antidepressants.

The ability of subanesthetic doses of N-methyl-D-aspartate (NMDA) antagonists to induce positive, negative, and cognitive schizophrenia-like symptoms suggests that reduced NMDA receptor function may contribute to the pathophysiology of schizophrenia. An increasing body of evidence indicates that antipsychotic drugs, especially those with "atypical" properties, can antagonize the effects of NMDA antagonists in a variety of experimental paradigms. We demonstrated previously that clozapine, the prototype of atypical antipsychotics, but not haloperidol, the typical antipsychotic, blocked ketamine-induced alterations in brain metabolism. In this study, effects of clozapine were compared with two of the newer atypical antipsychotic drugs, risperidone and olanzapine, on ketamine-induced alterations in regional [(14)C]2-deoxyglucose (2-DG) uptake. A subanesthetic dose of ketamine (25 mg/kg) induced robust increases in 2-DG uptake in limbic cortical regions, hippocampal formation, nucleus accumbens, and basolateral amygdala. Pretreatment of rats with risperidone (0.3 mg/kg) before ketamine administration did not alter the effects of ketamine. These data suggest that novel pharmacological properties may contribute to the effects of clozapine in this model, in addition to the well characterized actions at D(2) and <em>5HT</em>(2A) receptors. In contrast to the results with risperidone, olanzapine blocked ketamine-induced increases in 2-DG uptake. However, a higher dose of olanzapine (10 mg/kg) was required to completely block the effects of ketamine than would be expected if D(2) and <em>5HT</em>(2) receptor blocking properties of the drug were solely responsible for its action. The results suggest that the ketamine challenge 2-DG paradigm may be a useful model to identify antipsychotic drugs with atypical characteristics and to explore mechanisms of atypical antipsychotic action.

Both serotonin and NMDA signaling in prefrontal cortex (PFC) are implicated in mental disorders, including depression and anxiety. To understand their potential contributions to PFC neuronal excitability, we examined the effect of co-activation of 5-HT and NMDA receptors on action potential firing elicited by depolarizing current injection in PFC pyramidal neurons. In the presence of NMDA, a low concentration of the 5-HT(1A) agonist 8-OH-DPAT substantially reduced the number of spikes, and a low concentration of the 5-HT(2A/C) agonist alpha-Me-<em>5HT</em> significantly enhanced it, while both agonists were ineffective when applied alone. The 8-OH-DPAT effect on firing was mediated by inhibition of protein kinase A (PKA), whereas the alpha-Me-<em>5HT</em> effect was mediated by activation of protein kinase C (PKC). Moreover, the extracellular signal-regulated kinase (ERK), a signaling molecule downstream of PKA and PKC, was involved in both 5-HT(1A) and 5-HT(2A/C) modulation of neuronal excitability. Biochemical evidence showed that 5-HT(1A) decreased, whereas 5-HT(2A/C) increased the activation of ERK in an NMDA-dependent manner. In animals exposed to acute stress, the enhancing effect of 5-HT(2A/C) on firing was lost, while the decreasing effect of 5-HT(1A) on firing was intact. Concomitantly, the effect of 5-HT(2A/C), but not 5-HT(1A), on ERK activation was abolished in stressed animals. Taken together, our results demonstrate that distinct 5-HT receptor subtypes, by interacting with NMDA receptors, differentially regulate PFC neuronal firing, and the complex effects of 5-HT receptors on excitability are selectively altered under stressful conditions, which are often associated with mental disorders.

There is a need for rigorous positron emission tomography (PET) and endocrine methods to address inconsistencies in the literature regarding age, sex, and reproductive hormone effects on central serotonin (<em>5HT</em>) 1A and 2A receptor binding potential (BP). Healthy subjects (n=71), aged 20-80 years, underwent <em>5HT</em>1A and 2A receptor imaging using consecutive 90-min PET acquisitions with [(11)C]WAY100635 and [(18)F]altanserin. Logan graphical analysis was used to derive BP using atrophy-corrected distribution volume (V(T)) in prefrontal, mesiotemporal, occipital cortices, and raphe nucleus (<em>5HT</em>1A only). We used multivariate linear regression modeling to examine BP relationships with age, age(2), sex, and hormone concentrations, with post hoc regional significance set at p<0.008. There were small postsynaptic <em>5HT</em>1A receptor BP increases with age and estradiol concentration in women (p=0.004-0.005) and a tendency for small <em>5HT</em>1A receptor BP declines with age and free androgen index in men (p=0.05-0.06). Raphe <em>5HT</em>1A receptor BP decreased 4.5% per decade of age (p=0.05), primarily in men. There was a trend for 15% receptor reductions in prefrontal cortical regions in women relative to men (post hoc p=0.03-0.10). The significant decline in <em>5HT</em>2A receptor BP relative to age (8% per decade; p<0.001) was not related to sex or hormone concentrations. In conclusion, endocrine standardization minimized confounding introduced by endogenous hormonal fluctuations and reproductive stage and permitted us to detect small effects of sex, age, and endogenous sex steroid exposures upon <em>5HT</em>1A binding. Reduced prefrontal cortical <em>5HT</em>1A receptor BP in women vs men, but increased <em>5HT</em>1A receptor BP with aging in women, may partially explain the increased susceptibility to affective disorders in women during their reproductive years that is mitigated in later life. <em>5HT</em>1A receptor decreases with age in men might contribute to the known increased risk for suicide in men over age 75 years. Low hormone concentrations in adults <50 years of age may be associated with more extreme <em>5HT</em>1A receptor BP values, but remains to be studied further. The <em>5HT</em>2A receptor declines with age were not related to sex or hormone concentrations in this sample. Additional study in clinical populations is needed to further examine the affective role of sex-hormone-serotonin receptor relationships.

Frontal cerebral cortex brain samples from schizophrenics and controls have been assayed for binding associated with muscarinic cholinergic, serotonin (<em>5HT</em>), gamma-aminobutyric acid (GABA), and beta-adrenergic receptors as well as for the activity of the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD). Binding levels of tritium-LSD, presumably associated with postsynaptic <em>5HT</em> receptors, were reduced 40% to 50% in samples from schizophrenics in three independent studies, whereas no other consistent alteration was observed in levels of binding associated with other receptors or in the activity of GAD. This change in receptor binding levels does not seem to be attributable to postmortem changes, to influence of drugs received by the patients, or to demographic features of the patient populations.

OBJECTIVE

Elevated serum serotonin is associated with carcinoid heart disease, the hallmark of which is valvular thickening. Yet, the mechanistic role of serotonin in carcinoid heart disease is poorly understood. We postulated that serotonin has a direct mitogenic effect on cardiac valvular subendocardial cells, and that this effect is mediated by serotonin receptors.

METHODS

The dose-dependent proliferative effects of serotonin (10(-8) to 10(-4)M) on cultured porcine aortic valve cells via a [3H]thymidine assay were determined in vitro. Serotonin receptor antagonist studies in culture were also performed using methiotepin, a <em>5HT</em>1b antagonist, and ketanserin, a <em>5HT</em>2 receptor antagonist, to determine the mechanism of serotonin action. The ex-vivo proliferation level in human carcinoid (n = 26) and normal valves (n = 10) was compared using proliferating cell nuclear antigen (PCNA) staining, a marker for proliferation. Identification and localization of specific <em>5HT</em> receptor was assessed by immunostaining for serotonin receptors in the valves.

RESULTS

Serotonin increased valvular proliferation in vitro in a dose-dependent manner (10-fold increase) (p <0.001), and this mitogenic effect was inhibited by methiotepin but not ketanserin. In human carcinoid heart valves the level of proliferation was 35-fold higher than in normal human valves (p <0.001). <em>5HT</em>1b receptors were found only in the carcinoid valves, and not in the normal valves.

CONCLUSIONS

Serotonin is a powerful mitogen for valvular subendocardial cells. The mitogenic effect is at least partly mediated via <em>5HT</em>1b receptors. Subendothelial cell proliferation is significantly elevated in human carcinoid valves in vivo. The data suggest a mechanism whereby serotonin may contribute to valvular proliferation in carcinoid heart disease.

RNA editing is a post-transcriptional modification in which adenosine residues are converted to inosine (adenosine-to-inosine editing). Commonly used methodologies to quantify RNA editing levels involve either direct sequencing or pyrosequencing of individual cDNA clones. The limitations of these methods lead to a small number of clones characterized in comparison to the number of mRNA molecules in the original sample, thereby producing significant sampling errors and potentially erroneous conclusions. We have developed an improved method for quantifying RNA editing patterns that increases sequence analysis to an average of more than 800,000 individual cDNAs per sample, substantially increasing accuracy and sensitivity. Our method is based on the serotonin 2C receptor (5-hydroxytryptamine(2C); <em>5HT</em>(2C)) transcript, an RNA editing substrate in which up to five adenosines are modified. Using a high-throughput multiplexed transcript analysis, we were able to quantify accurately the expression of twenty <em>5HT</em>(2C) isoforms, each representing at least 0.25% of the total <em>5HT</em>(2C) transcripts. Furthermore, this approach allowed the detection of previously unobserved changes in <em>5HT</em>(2C) editing in RNA samples isolated from different inbred mouse strains and dissected brain regions, as well as editing differences in alternatively spliced <em>5HT</em>(2C) variants. This approach provides a novel and efficient strategy for large-scale analyses of RNA editing and may prove to be a valuable tool for uncovering new information regarding editing patterns in specific disease states and in response to pharmacological and physiological perturbation, further elucidating the impact of <em>5HT</em>(2C) RNA editing on central nervous system function.

Nicotinic acetylcholine receptors (nAChRs) mediate fast synaptic transmission in the insect brain and are target sites for neonicotinoid insecticides. Seven nAChR subunits (four alpha-type and three beta-type) have been cloned previously from Drosophila melanogaster, the model insect system and characterized by heterologous expression. Recently, three further putative nAChR alpha subunits (Dalpha5, Dalpha6 and Dalpha7) with sequence similarity to the vertebrate alpha7 subunit have been identified from Drosophila genome sequence data but there have been no reports, as yet, of their characterization by heterologous expression. In the present study, we report the first isolation of a full-length Dalpha7 cDNA and the independent molecular cloning of Dalpha6. Binding of nicotinic radioligands was not detected to full-length Dalpha6 or Dalpha7 subunits when expressed alone or when or co-expressed with other nAChR subunits in Drosophila or mammalian cell lines, but specific cell-surface binding of [(125)I]alpha-bungarotoxin (K(d) = 0.68 +/- 0.22 nm) and [(3)H]methyllycaconitine (K(d) = 0.27 +/- 0.06 nm) was detected after expression of a subunit chimera containing the ligand-binding domains of Dalpha6 fused to the C-terminal domain of the 5-hydroxytryptamine receptor <em>5HT</em>(3A). Although cell-surface binding was not detected with a Dalpha7/<em>5HT</em>(3Alpha) chimera expressed alone, co-expression of the two subunit chimeras resulted in significantly enhanced levels of nicotinic radioligand binding (with no change in affinity). This is the first evidence for the formation of a nAChR binding site by heterologously expressed Drosophila nAChR subunits in the absence of a co-expressed vertebrate nAChR subunit. In addition to the formation of homomeric nAChR complexes, evidence has been obtained from both radioligand binding and co-immunoprecipitation studies for the co-assembly of Dalpha6 and Dalpha7 into heteromeric cell surface complexes.

A-to-I RNA editing is a post-transcriptional modification of single nucleotides in RNA by adenosine deamination, which thereby diversifies the gene products encoded in the genome. Thousands of potential RNA editing sites have been identified by recent studies (e.g. see Li et al, Science 2009); however, only a handful of these sites have been independently confirmed. Here, we systematically and quantitatively examined 109 putative coding region A-to-I RNA editing sites in three sets of normal human brain samples by ultra-high-throughput sequencing (uHTS). Forty of 109 putative sites, including 25 previously confirmed sites, were validated as truly edited in our brain samples, suggesting an overestimation of A-to-I RNA editing in these putative sites by Li et al (2009). To evaluate RNA editing in human disease, we analyzed 29 of the confirmed sites in subjects with major depressive disorder and schizophrenia using uHTS. In striking contrast to many prior studies, we did not find significant alterations in the frequency of RNA editing at any of the editing sites in samples from these patients, including within the <em>5HT</em>(2C) serotonin receptor (HTR2C). Our results indicate that uHTS is a fast, quantitative and high-throughput method to assess RNA editing in human physiology and disease and that many prior studies of RNA editing may overestimate both the extent and disease-related variability of RNA editing at the sites we examined in the human brain.

Glutamatergic hypofunction occurs in Alzheimer's disease (AD). MK801, a noncompetitive blocker of glutamate N-methyl-D-aspartate receptors, was used to disrupt the cognitive performance of rats trained on a delayed nonmatching to sample radial maze task. Drugs which act by blocking serotonin (5-HT) receptors were evaluated for their ability to reduce the cognitive impairment produced by MK801. Specifically, WAY-100635, a selective 5-HT1A receptor antagonist, buspirone, a 5-HT1A partial agonist, ritanserin, a 5-HT2 antagonist, and ondansetron, a 5-HT3 antagonist, were assessed. In addition, the muscarinic agonist arecoline was evaluated for its potential cognitive benefit in this model. It was found that WAY-100635 significantly reduced the cognitive impairment induced by MK801. Treatment with single doses of ritanserin, ondansetron, or arecoline in combination with MK801 did not result in a cognitive impairment, indicating that these drugs attenuated the MK801 impairment. The combination of buspirone and MK801 resulted in an inability of the animals to complete the task. These results suggest that interactions between 5-HT and glutamate may mediate the beneficial effects of reducing cognitive impairment and that 5-HT antagonists, especially selective 5-HT1A antagonists, may be useful in treating AD. Further, it is indicated that the MK801 model of cognitive impairment may add to the armamentarium of tools available to predict treatment efficacy in AD.

The identification of a second 5-HT(3) (5-HT(3B)) subunit provides an explanation for 5-HT(3) receptor heterogeneity. We investigated whether introduction of recombinant 5-HT(3B) subunits would alter the functional properties of mouse neuroblastoma 5-HT(3) receptors. RT-PCR analysis revealed that NB41A3 cells contain mRNAs encoding 5-HT(3A) and 5-HT(3B) subunits. 5-HT increased intracellular Ca(2+) concentration ([Ca(2+)](i)) and caused the concentration-dependent activation of inward currents recorded at -60 mV. Both actions of 5-HT were antagonized by ondansetron. The 5-HT concentration-response relationship of NB41A3 cells was indistinguishable from that of the related NG108-15 cell line. The selective 5-HT(3)-receptor agonist mCPBG also elevated [Ca(2+)](i) and activated inward currents. 2-M-<em>5HT</em> was less efficacious than 5-HT as an activator of 5-HT(3) receptors in NB41A3 cells and did not significantly increase [Ca(2+)](i). The 5-HT induced increase in [Ca(2+)](i) did not involve caffeine- or thapsigargin-sensitive intracellular Ca(2+) stores. The introduction of the 5-HT(3B) subunit by transient transfection of NB41A3 cells caused 5-HT to become less potent as an activator of 5-HT(3) receptors and altered the kinetics of 5-HT activated currents so that they resembled currents mediated by 5-HT(3AB) receptors. The 5-HT(3B) subunit also abolished the 5-HT induced [Ca(2+)](i) increase seen in untransfected NB41A3 cells. These data are consistent with the hypothesis that NB41A3 cells predominantly express homomeric 5-HT(3A) receptors that become heteromeric 5-HT(3AB) receptors upon introduction of the recombinant 5-HT(3B) subunit.

Paroxetine is a new antidepressant drug with potent serotonin (<em>5HT</em>) uptake inhibitory properties. In this double-blind comparative study, the antidepressant effect of paroxetine and amitriptyline has been compared in 44 patients with depressive illnesses of an endogenous nature. Each drug was given for 6 weeks. The 17-item Hamilton Depression Scale was used to measure the antidepressant effect. Reported events were assessed applying a 22-item check list. Non-parametric statistical analyses were applied in the evaluation of treatment outcome for the 30 patients who completed the study. The results showed no significant differences in overall antidepressant efficacy between paroxetine and amitriptyline and that paroxetine displayed significantly fewer instances of dry mouth and orthostatic dizziness than amitriptyline. No obvious relationship was demonstrated between the plasma levels of the drugs and their clinical effects.

OBJECTIVE

The available literatures show that 5-HT(1A) receptors are widely distributed throughout the basal ganglia, and their activation facilitate dopamine release. Neuroleptic drugs such as haloperidol induce Parkinson-like syndrome through blocking brain D(2) receptors. This study aimed to investigate effect of buspirone, a partial agonist of <em>5HT</em>(1A) receptor, on motor dysfunctions induced by haloperidol and involvement of <em>5HT</em>(1A) receptors in this regard.

METHODS

Study was performed on the male mice weighing 25-30 g. Animals were divided randomly to groups of 10 animals. Motor dysfunction was induced by intraperitoneal (i.p.) injection of haloperidol (1 mg/kg). Catalepsy was assayed by bar-test method 5, 60, 120 and 180 minutes after drug administration and motor imbalance was studied by rotarod test.

CONCLUSIONS

Results showed that buspirone (20 mg/kg, i.p.) decreased significantly haloperidol-induced catalepsy and balance disorder in a dose dependent manner. Furthermore, 8-OH-DPAT (10 mg/kg, i.p.), as an agonist of 5-HT(1A) receptor, decreased haloperidol-induced catalepsy and balance disorder. The effect of buspirone (20 mg/kg, i.p.) on haloperidol-induced motor disorders was abolished by NAN-190 (10 mg/kg, i.p.), as a 5-HT(1A) receptor antagonist. From the results it may be concluded that buspirone improves haloperidol-induced catalepsy and balance disorder through activation of 5-HT(1A) receptors.

BACKGROUND

Previous studies have suggested involvement of <em>5HT</em>(1) receptors in the control of gastric tone.

OBJECTIVE

To study the effect of buspirone, a <em>5HT</em>(1A) agonist, on gastric sensorimotor function in healthy volunteers.

METHODS

Ten healthy volunteers (six males and four females, ages 20-29 years) participated in a barostat study evaluating the influence of single oral doses of buspirone (5, 10, 20, 30 and 40 mg) on tone and sensitivity of the proximal stomach. In addition, the effect of placebo or the three lowest doses of buspirone on gastric emptying was assessed using a solid and liquid gastric emptying breath test.

RESULTS

Compared to preadministration volumes, buspirone increased proximal stomach volumes in a dose-dependent manner, with significant fundic relaxation after 30 and 40 mg doses (intra-balloon volume increases of respectively 258 +/- 80 mL and 273 +/- 49 mL, P < or = 0.05). Pressure thresholds during gastric distention were not altered, but corresponding intraballoon volumes were significantly increased after 30 and 40 mg doses (respective discomfort volumes 596 +/- 73 vs. 791 +/- 87 mL and 630 +/- 73 vs.741 +/- 60 mL, both P < 0.05). Buspirone significantly slowed solid and liquid gastric emptying at the 20-mg dose.

CONCLUSIONS

Buspirone dose-dependently relaxes the proximal stomach in the fasting state and decreases the gastric emptying rate in healthy volunteers.