MK212, a 5-hydroxytryptamine 2C receptor agonist, reverses prepulse inhibition deficits in the medial prefrontal cortex and ventral hippocampus.

Prepulse inhibition (PPI) is disrupted in many neuropsychiatric diseases. Molecules such as 5-HT2C receptor agonists alleviate PPI deficits in rodents; however, the precise mechanisms and critical regions of the brain responsible for the reversal effect of these agonists remain inconclusive. The present study aimed to investigate the areas of the brain critical for the reversal effect of 5-HT2C receptor agonists on PPI deficits in mice. The results showed that systemic administration of the 5-HT2C receptor agonist MK212 did not affect normal PPI behavior, but reversed the PPI deficits induced by the N-methyl d-aspartate receptor antagonist MK801 in mice. In addition, the 5-HT2C receptor antagonist SB242084 had no effect on PPI behavior despite MK801 treatment. Moreover, local infusion of MK212 into the medial prefrontal cortex and ventral hippocampus, excluding the nucleus accumbens or ventral tegmental area, rescued the PPI deficits induced by MK801. These data suggest that the medial prefrontal cortex and ventral hippocampus are critical brain areas responsible for the reversal of 5-HT2C agonists on PPI deficits. The results will contribute to our current knowledge on the molecular and neural mechanisms underlying the antipsychotic effects of 5-HT2C receptor agonists, especially the neural circuits modulated by 5-HT2C receptor activity.

A selective serotonin reuptake inhibitor ameliorates obsessive-compulsive disorder-like perseverative behavior by attenuating 5-HT2C receptor signaling in the orbitofrontal cortex.

Perseveration is a characteristic of patients with obsessive-compulsive disorder (OCD). Clinically, neuronal activity in the lateral orbitofrontal cortex (OFC) is increased in OCD patients. Successful treatment with selective serotonin reuptake inhibitors (SSRIs) reduces activity in the lateral OFC of OCD patients, but the precise mechanisms underlying this effect are unclear. Previously, we reported that repeated injection of the dopamine D2 receptor agonist quinpirole (QNP) resulted in OCD-like deficits, including perseveration in a reversal learning task. QNP-treated mice showed hyperactivity in lateral OFC pyramidal neurons. The present study demonstrated that 4-week administration of an SSRI increased the rate of correct choice in a reversal learning task. Using the electrophysiological approach, we revealed that an SSRI decreased the activity of lateral OFC pyramidal neurons in QNP-treated mice by potentiating inhibitory inputs. The 4-week administration of an SSRI inhibited the potentiation of neuronal activity induced by a 5-HT2C receptor agonist. Additionally, both 4-week administration of SSRI and acute application of 5-HT2C receptor antagonist prevented the QNP-induced potentiation of inhibitory inputs to fast-spiking interneurons in the lateral OFC. Administration of a 5-HT2C receptor antagonist to mice for 4 days increased the rate of correct choice in a reversal learning task. Collectively, these results indicate that chronic SSRI ameliorated perseverative behavior in QNP-treated mice by modulating inhibitory inputs in the lateral OFC. Short-term 5-HT2C receptor blockade also ameliorated QNP-induced behavioral and neurological abnormalities by, at least in part, a common mechanism with chronic SSRI.

Barbadin Potentiates Long-Term Effects of Lorcaserin on POMC Neurons and Weight Loss.

Obesity is a serious global health problem because of its increasing prevalence and comorbidities, but its treatments are limited. The serotonin 2C receptor (5-HT2CR), a G-protein-coupled receptor, activates proopiomelanocortin (POMC) neurons in the arcuate nucleus of hypothalamus (ARH) to reduce appetite and weight gain. However, several 5-HT analogs targeting this receptor, e.g., lorcaserin (Lor), suffer from diminished efficacy to reduce weight after prolonged administration. Here, we show that barbadin (Bar), a novel ß-arrestin/ß2-adaptin inhibitor, can prevent 5-HT2CR internalization in cells and potentiate long-term effects of Lor to reduce appetite and body weight in male mice. Mechanistically, we demonstrate that Bar co-treatment can effectively maintain the sensitivity of the 5-HT2CR in POMCARH neurons, despite prolonged Lor exposure, thereby allowing these neurons to be activated through opening the transient receptor potential cation (TRPC) channels. Thus, our results prove the concept that inhibition of 5-HT2CR desensitization can be a valid strategy to improve the long-term weight loss effects of Lor or other 5-HT2CR agonists, and also provide an intellectual framework to develop effective long-term management of weight by targeting 5-HT2CR desensitization.SIGNIFICANCE STATEMENT By demonstrating that the combination of barbadin (Bar) with a G-protein-coupled receptor (GPCR) agonist can provide prolonged weight-lowering benefits in a preclinical setting, our work should call for additional efforts to validate Bar as a safe and effective medicine or to use Bar as a lead compound to develop more suitable compounds for obesity treatment. These results prove the concept that inhibition of serotonin 2C receptor (5-HT2CR) desensitization can be a valid strategy to improve the long-term weight loss effects of lorcaserin (Lor) or other 5-HT2CR agonists. Since GPCRs represent a major category as therapeutic targets for various human diseases and desensitization of GPCRs is a common issue, our work may provide a conceptual framework to enhance effects of a broad range of GPCR medicines.

MicroRNA-34a regulates 5-HT2C expression in dorsal raphe and contributes to the anti-depressant-like effect of fluoxetine.

Selective serotonin reuptake inhibitors (SSRIs) are designed to improve mood by raising extracellular serotonin levels through the blockade of the serotonin transporter. However, they exhibit a slow onset of action, suggesting the involvement of adaptive regulatory mechanisms. We hypothesized that the microRNA-34 family facilitates the therapeutic activity of SSRIs. We show that genetic deletion of these microRNAs in mice impairs the response to chronic, but not acute, fluoxetine treatment, with a specific effect on behavioral constructs that are related to depression, rather than anxiety. Moreover, using a pharmacological strategy, we found that an increased expression of the serotonin 2C (5-HT2C) receptor in the dorsal raphe region of the brain contributes to this phenotype. The onset of the therapeutic efficacy of SSRIs is paralleled by the desensitization of the 5-HT2C receptor in the dorsal raphe, and 5-HT2C is a putative target of microRNA-34. In this study, acute and chronic fluoxetine treatment differentially alters the expression of 5-HT2C and microRNA-34a in the dorsal raphe. Moreover, by in vitro luciferase assay, we demonstrated the repressive regulatory activity of microRNA-34a against 5-HT2C mRNA. Specific blockade of this interaction through local infusion of a target site blocker was sufficient to prevent the behavioral effects of chronic fluoxetine. Our results demonstrate a new miR-34a-mediated regulatory mechanism of 5-HT2C expression in the dorsal raphe and implicate it in eliciting the behavioral responses to chronic fluoxetine treatment.

Role of 5-HT1A and 5-HT2C receptors of the dorsal periaqueductal gray in the anxiety- and panic-modulating effects of antidepressants in rats.

Antidepressant drugs are first-line treatment for panic disorder. Facilitation of 5-HT1A receptor-mediated neurotransmission in the dorsal periaqueductal gray (dPAG), a key panic-associated area, has been implicated in the panicolytic effect of the selective serotonin reuptake inhibitor fluoxetine. However, it is still unknown whether this mechanism accounts for the antipanic effect of other classes of antidepressants drugs (ADs) and whether the 5-HT interaction with 5-HT2C receptors in this midbrain area (which increases anxiety) is implicated in the anxiogenic effect caused by short-term treatment with ADs. The results showed that previous injection of the 5-HT1A receptor antagonist WAY-100635 in the dPAG blocked the panicolytic-like effect caused by chronic systemic administration of the tricyclic AD imipramine in male Wistar rats tested in the elevated T-maze. Neither chronic treatment with imipramine nor fluoxetine changed the expression of 5-HT1A receptors in the dPAG. Treatment with these ADs also failed to significantly change ERK1/2 (extracellular-signal regulated kinase) phosphorylation level in this midbrain area. Blockade of 5-HT2C receptors in the dPAG with the 5-HT2C receptor antagonist SB-242084 did not change the anxiogenic effect caused by a single acute injection of fluoxetine or imipramine in the Vogel conflict test. These results reinforce the view that the facilitation of 5-HT1A receptor-mediated neurotransmission in the dPAG is a common mechanism involved in the panicolytic effect caused by chronic administration of ADs. On the other hand, the anxiogenic effect observed after short-term treatment with these drugs does not depend on 5-HT2C receptors located in the dPAG.

Early ethanol pre-exposure alters breathing patterns by disruptions in the central respiratory network and serotonergic balance in neonate rats.

Early ethanol exposure alters neonatal breathing plasticity. Respiratory EtOH's effects are attributed to central respiratory network disruptions, particularly in the medullary serotonin (5HT) system. In this study we evaluated the effects of neonatal pre-exposure to low/moderate doses upon breathing rates, activation patterns of brainstem's nuclei and expression of 5HT 2A and 2C receptors. At PD9, breathing frequencies, tidal volumes and apneas were examined in pups pre-exposed to vehicle or ethanol (2.0 g/kg) at PDs 3, 5 and 7. This developmental stage is equivalent to the 3rd human gestational trimester, characterized by increased levels of synaptogenesis. Pups were tested under sobriety or under the state of ethanol intoxication and when subjected to normoxia or hypoxia. Number of c-Fos and 5HT immunolabelled cells and relative mRNA expression of 5HT 2A and 2C receptors were quantified in the brainstem. Under normoxia, ethanol pre-exposed pups exhibited breathing depressions and a high number of apneas. An opposite phenomenon was found in ethanol pre-treated pups tested under hypoxia where an exacerbated hypoxic ventilatory response (HVR) was observed. The breathing depression was associated with an increase in the neural activation levels of the raphe obscurus (ROb) and a high mRNA expression of the 5HT 2A receptor in the brainstem while desactivation of the ROb and high activation levels in the solitary tract nucleus and area postrema were associated to the exacerbated HVR. In summary, early ethanol experience induces respiratory disruptions indicative of sensitization processes. Neuroadaptive changes in central respiratory areas under consideration appear to be strongly associated with changes in their respiratory plasticity.

Investigating the role of 5-HT2A and 5-HT2C receptor activation in the effects of psilocybin, DOI, and citalopram on marble burying in mice.

Psychedelic drugs acting as 5-hydroxyptryptamine 2A receptor (5-HT2AR) agonists have shown promise as viable treatments of psychiatric disorders, including obsessive-compulsive disorder. The marble burying test is a test of compulsive-like behavior in mice, and psychedelics acting as 5-HT2AR agonists can reduce digging in this test. We assessed the 5-HT2R contribution to the mechanisms of two 5-HT2A agonists on digging behavior in female NMRI mice, using citalopram as a reference compound. While the 5-HT2AR antagonist M100907 blocked the effect of DOI and the 5-HT2CR antagonist SB242084 blocked the effect of citalopram, neither antagonist blocked the effect of psilocybin. This study confirms 5-HT2AR agonism as a mechanism for reduced compulsive-like digging in the MB test and suggests that 5-HT2A and 5-HT2CRs can work in parallel on this type of behavior. Our results with psilocybin suggest that a 5-HT2R-independent mechanism also contributes to the effect of psilocybin on repetitive digging behavior.

Contribution of serotonin receptor subtypes to hallucinogenic activity of 25I-NBOMe and to its effect on neurotransmission.

BACKGROUND: 4-Iodo-2,5-dimethoxy-N-(2-methoxybenzyl)phenethylamine (25I-NBOMe) is a potent serotonin (5-HT) receptor agonist with hallucinogenic properties. The aim of our research was to examine the role of the 5-HT2A, 5-HT2C and 5-HT1A serotonin receptor subtypes in 25I-NBOMe hallucinogenic activity and its effect on dopamine (DA), 5-HT and glutamate release in the rat frontal cortex. METHODS: Hallucinogenic activity was investigated using the wet dog shake (WDS) test. The release of DA, 5-HT and glutamate in the rat frontal cortex was studied using a microdialysis in freely moving rats. Neurotransmitter levels were analyzed by HPLC with electrochemical detection. The selective antagonists of the 5-HT2A, 5-HT2C and 5-HT1A serotonin receptor subtypes: M100907, SB242084 and WAY100635, respectively were applied through a microdialysis probe. RESULTS: The WDS response to 25I-NBOMe (1 and 3 mg/kg) was significantly reduced by local administration of M100907 and SB242084 (100 nM). The 25I-NBOMe-induced increase in glutamate, DA and 5-HT release was inhibited by M100907 and SB242084. WAY100635 had no effect on 25I-NBOMe-induced WDS and glutamate release, while it decreased DA and 5-HT release from cortical neuronal terminals. CONCLUSION: The obtained results suggest that 5-HT2A and 5-HT2C receptors play a role in 25I-NBOMe-induced hallucinogenic activity and in glutamate, DA and 5-HT release in the rat frontal cortex as their respective antagonists attenuated the effect of this hallucinogen. The disinhibition of GABA cells by the 5-HT1A receptor antagonist seems to underlie the mechanism of decreased DA and 5-HT release from neuronal terminals in the frontal cortex.

Acute EPA-induced learning and memory impairment in mice is prevented by DHA.

Eicosapentaenoic acid (EPA), an omega-3 fatty acid, has been widely used to prevent cardiovascular disease (CVD) and treat brain diseases alone or in combination with docosahexaenoic acid (DHA). However, the impact of EPA and DHA supplementation on normal cognitive function and the molecular targets of EPA and DHA are still unknown. We show that acute administration of EPA impairs learning and memory and hippocampal LTP in adult and prepubescent mice. Similar deficits are duplicated by endogenously elevating EPA in the hippocampus in the transgenic fat-1 mouse. Furthermore, the damaging effects of EPA are mediated through enhancing GABAergic transmission via the 5-HT6R. Interestingly, DHA can prevent EPA-induced impairments at a ratio of EPA to DHA similar to that in marine fish oil via the 5-HT2CR. We conclude that EPA exhibits an unexpected detrimental impact on cognitive functions, suggesting that caution must be exercised in omega-3 fatty acid supplementation and the combination of EPA and DHA at a natural ratio is critical for learning and memory and synaptic plasticity.

A Novel Synthetic Interfering Peptide Tat-3L4F Attenuates Olanzapine-Induced Weight Gain Through Disrupting Crosstalk Between Serotonin Receptor 2C and Protein Phosphatase and Tensin Homolog in Rats.

BACKGROUND: Accompanied with profound efficacy, atypical antipsychotics (AAPs) contribute to metabolic adverse effects with few effective strategies to attenuate. Serotonin 5-HT2C receptor (HTR2C) plays a critical role in hyperphagia and weight gain induced by AAPs, and expression of phosphatase tensin homolog (PTEN) in the hypothalamus also affects feeding behavior and weight change. Moreover, PTEN has a physical crosstalk between PTEN and a region in the third intracellular loop (3L4F) of the HTR2C. Tat-3L4F has the property to disrupt crosstalk between PTEN and HTR2C. This is the first study to our knowledge to investigate the effect of Tat-3L4F on olanzapine-induced metabolic abnormalities and PTEN/ phosphatidylinositol 3-kinase/protein kinase B expression in the hypothalamus in rats. METHODS: The effects of Tat-3L4F were investigated through measuring body weight, food intake, and blood glucose. In addition, PTEN/phosphatidylinositol 3-kinase/protein kinase B level in the hypothalamus was detected by immunofluorescence assay and western blot. Metabolites in the liver tissue were detected by liquid chromatography-mass spectrometry and analyzed by multivariate analyses and pairwise comparison. RESULTS: Our results showed that hyperphagia and weight gain were evident in the olanzapine alone-fed rats but was attenuated after Tat-3L4F treatment. In addition, oral glucose tolerance test indicated blood glucose at 120 minutes was higher in the olanzapine alone-treated group than in groups treated with vehicle and olanzapine + Tat-3L4F (10 µmol kg-1 per day). Furthermore, compared with olanzapine alone treatment, treatment with Tat-3L4F (10 µmol kg-1 per day) significantly inhibited PTEN expression in the hypothalamus. The olanzapine alone-treated group had the highest bile acid level, followed by the olanzapine with Tat-3L4F (1 µmol kg-1) group, olanzapine with Tat-3L4F (10 µmol kg-1) group, and vehicle group. CONCLUSIONS: Our present results reveal that Tat-3L4F is a potential pharmacological strategy for suppressing hyperphagia and weight gain induced by olanzapine, which acts through disrupting crosstalk between HTR2C and PTEN as a result of PTEN downregulation in the hypothalamus.

Identification of Novel 1-O-Substituted Aporphine Analogues as Potent 5-HT2C Receptor Agonists.

The 5-HT2C receptor has emerged as a promising target in the treatment of a variety of central nervous system disorders. We have first identified aporphines as a new class of 5-HT2C receptor agonists. Structure-activity relationship results indicate that the aporphine core may be required for 5-HT2C receptor activity, and substitutions at its C1 position are important for 5-HT2C receptor activity. Our efforts to optimize our hit 15781 lead to the identification of the highly potent and selective 5-HT2C agonist 18b (MQ02-439) with an EC50 value of 104 nM and weak antagonism at the 5-HT2A and 5-HT2B receptors. The findings may serve as good starting points for the development of more potent and selective 5-HT2C agonists as valuable pharmacological tools or potential drug candidates.

Fluoxetine improves behavioural deficits induced by chronic alcohol treatment by alleviating RNA editing of 5-HT2C receptors.

The alcoholism and major depressive disorder are common comorbidity, with alcohol-induced depressive symptoms being eased by selective serotonin re-uptake inhibitors (SSRIs), although the mechanisms underlying pathology and therapy are poorly understood. Chronic alcohol consumption affects the activity of serotonin 2C receptors (5-HT2CR) by regulating adenosine deaminases acting on RNA (ADARs) in neurons. Astrogliopathic changes contribute to alcohol addiction, while decreased release of ATP from astrocytes can trigger depressive-like behaviours in mice. In this study, we discovered that chronic alcohol treatment increased editing of RNA of 5-HT2CR via up-regulating the expression of ADAR2, consequently reducing the release of ATP from astrocytes induced by 5-HT2CR agonist, MK212. Moreover, SSRI antidepressant fluoxetine decreased the expression of ADAR2 through the transactivation of EGFR/PI3K/AKT/cFos signalling pathway. The increased release of astroglial ATP by MK212 which was suppressed by chronic alcohol consumption, and reduction in ADAR2 activity eliminated the RNA editing of 5-HT2CR increased by alcohol in vitro and recovered the release of ATP from astrocytes induced by MK212. Meanwhile, fluoxetine improved the behavioural and motor symptoms induced by alcohol addiction and decreased the alcohol intake. Our study suggests that the astrocytic 5-HT2CR contribute to alcohol addiction; fluoxetine thus can be used to alleviate depression, treat alcohol addiction and improve motor coordination.

Investigating serotonergic contributions to cognitive effort allocation, attention, and impulsive action in female rats.

BACKGROUND: Individuals must frequently evaluate whether it is worth allocating cognitive effort for desired outcomes. Motivational deficits are a common feature of psychiatric illness such as major depression. Selective serotonin reuptake inhibitors are commonly used to treat this disorder, yet some data suggest these compounds are ineffective at treating amotivation, and may even exacerbate it. AIMS: Here we used the rodent Cognitive Effort Task (rCET) to assess serotonergic (5-hydroxytryptamine, 5-HT) contributions to decision-making with cognitive effort costs. METHODS: The rCET is a modified version of the 5-choice serial reaction time task, a well-validated test of visuospatial attention and impulse control. At the start of each rCET trial, rats chose one of two levers, which set the difficulty of an attentional challenge, namely the localization of a visual stimulus illuminated for 0.2 or 1 s on hard versus easy trials. Successful completion of hard trials was rewarded with double the sugar pellets. Twenty-four female Long-Evans rats were trained on the rCET and systemically administered the 5-HT1A agonist 8-OH-DPAT, the 5-HT2A antagonist M100907, the 5-HT2C agonist Ro-60-0175, as well as the 5-HT2C antagonist SB 242, 084. RESULTS: 5-HT2A antagonism dose-dependently reduced premature responding, while 5-HT2C antagonism had the opposite effect. 8-OH-DPAT impaired accuracy of target detection at higher doses, while Ro-60-0175 dose-dependently improved accuracy on difficult trials. However, none of the drugs affected the rats' choice of the harder option. CONCLUSION: When considered with existing work evaluating decision-making with physical effort costs, it appears that serotonergic signalling plays a minor role in guiding effort allocation.

Involvement of 5-HT2A, 5-HT2B and 5-HT2C receptors in mediating the ventrolateral orbital cortex-induced antiallodynia in a rat model of neuropathic pain.

The present study examined the roles of 5-HT2A, 5-HT2B and 5-HT2C receptor subtypes in mediating the ventrolateral orbital cortex (VLO)-induced antiallodynia in a rat model of neuropathic pain induced by spared nerve injury (SNI). Change of mechanical paw withdrawal threshold (PWT) was measured using von-Frey filaments. Microinjection of preferential or selective 5-HT2A/C, 5-HT2B and 5-HT2C receptor agonists, (±)-1-(2,5-Dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI), α-methyl-5-(2-thienylmethoxy)-1H-Indole-3-ethanamine hydrochloride (BW723C86) and 1-(3-Chlorophenyl)-piperazine hydrochloride (m-CPP) into the VLO significantly depressed allodynia induced by SNI, and the inhibitory effect of DOI was blocked or attenuated by selective 5-HT2A/C receptor antagonists ketanserin (+)-tartrate salt (ketanserin) and 5-HT2A receptor antagonist R-(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidinemethanol (M100907); the effects of BW723C86 and m-CPP were antagonized by 5-HT2B receptor antagonists N-(1-Methyl-1H-5-indolyl)-N'-(3-methyl-5-isothiazolyl)urea (SB204741) and 5-HT2C receptor antagonist RS102221 hydrochloride hydrate (RS-102221), respectively. These results suggest that 5-HT2A, 5-HT2B, 5-HT2C receptor subtypes are involved in mediating the VLO-induced antiallodynia in the neuropathic pain state.

Serotonin 2C receptors in the basolateral amygdala mediate the anxiogenic effect caused by serotonergic activation of the dorsal raphe dorsomedial subnucleus.

BACKGROUND: Stimulation of serotonergic neurons within the dorsal raphe dorsomedial subnucleus facilitates inhibitory avoidance acquisition in the elevated T-maze. It has been hypothesized that such anxiogenic effect is due to serotonin release in the basolateral nucleus of the amygdala, where facilitation of serotonin 2C receptor-mediated neurotransmission increases anxiety. Besides the dorsal raphe dorsomedial subnucleus, the dorsal raphe caudal subnucleus is recruited by anxiogenic stimulus/situations. However, the behavioral consequences of pharmacological manipulation of this subnucleus are still unknown. AIMS: Investigate whether blockade of serotonin 2C receptors in the basolateral nucleus of the amygdala counteracts the anxiogenic effect caused by the stimulation of dorsal raphe dorsomedial subnucleus serotonergic neurons. Evaluate the effects caused by the excitatory amino acid kainic acid or serotonin 1A receptor-modulating drugs in the dorsal raphe caudal subnucleus. METHODS: Male Wistar rats were tested in the elevated T-maze and light-dark transition tests after intra-basolateral nucleus of the amygdala injection of the serotonin 2C receptor antagonist SB-242084 (6-chloro-2,3-dihydro-5-methyl-N-[6-[(2-methyl-3-pyridinyl)oxy]-3-pyridinyl]-1H-indole-1-carboxyamide dihydrochloride) followed by intra-dorsal raphe dorsomedial subnucleus administration of the serotonin 1A receptor antagonist WAY-100635 (N-[2-[4-2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinil-cyclohexanecarboxamide maleate). In the dorsal raphe caudal subnucleus, animals were injected with kainic acid, WAY-100635 or the serotonin 1A receptor agonist 8-OH-DPAT ((±)-8-hydroxy-2-(di-n-propylamino) tetralin hydrobromide) and tested in the elevated T-maze. RESULTS: SB-242084 in the basolateral nucleus of the amygdala blocked the anxiogenic effect caused by the injection of WAY-100635 in the dorsal raphe dorsomedial subnucleus. Kainic acid in the dorsal raphe caudal subnucleus increased anxiety, but also impaired escape expression in the elevated T-maze. Neither WAY-100635 nor 8-OH-DPAT in the dorsal raphe caudal subnucleus affected rat's behavior in the elevated T-maze. CONCLUSION: Serotonin 2C receptors in the basolateral nucleus of the amygdala mediate the anxiogenic effect caused by the stimulation of serotonergic neurons in the dorsal raphe dorsomedial subnucleus. The dorsal raphe caudal subnucleus regulates anxiety- and panic-like behaviors, presumably by a serotonin 1A receptor-independent mechanism.

Inverse agonists of the 5-HT2A receptor reduce nicotine withdrawal signs in rats.

Previous work has shown that chronic nicotine administration causes adaptive changes in 5-HT2A receptor expression. Based on this relationship, it was hypothesized that inactivating 5-HT2A receptors with the inverse agonists pimavanserin and volinanserin (MDL100907), would reduce the symptoms of nicotine withdrawal syndrome. Sprague-Dawley rats were rendered nicotine-dependent by subcutaneous infusion of nicotine bitartrate, 9 mg/kg/day for seven days. The infusions were then terminated, and 22 h later, rats were observed under "blind" conditions for somatically expressed behavioral nicotine withdrawal signs. One hour before observations, the nicotine dependent rats were injected i.p. with saline alone, or either 0.3 or 1.0 mg/kg pimavanserin in saline. Total withdrawal signs were reduced in a dose-dependent manner. A one-way ANOVA (total withdrawal signs as a function of dose) was highly significant, as was the descending linear trend of withdrawal signs as a function of dose. The 1.0 mg/kg dose reduced withdrawal signs nearly to the level exhibited by a comparison group of non-dependent rats injected with saline. A second experiment was conducted in a similar manner, which showed that volinanserin at 1.0 mg/kg but not 0.25 mg/kg also reduced nicotine withdrawal signs to nearly the level of non-dependent rats. In vitro experiments demonstrated that both pimavanserin and volinanserin potently antagonize 5-HT2A receptors, with approximately 25-fold, and 300-fold selectivity over 5-HT2C receptors, respectively. The results suggest that the 5-HT2A receptor contributes to mediating nicotine withdrawal syndrome, and thus represents a potential target for interventions to aid smoking cessation.

Adaptation in 5-HT2 receptors-CaMKII signaling in lateral habenula underlies increased nociceptive-sensitivity in ethanol-withdrawn rats.

Alcoholics often experience hyperalgesia, especially during abstinence, yet the underlying cellular and molecular bases are unclear. Recent evidence suggests that 5-HT type 2 receptors (5-HT2Rs) at glutamatergic synapses on lateral habenula (LHb) neurons may play a critical role. We, therefore, measured paw withdrawal responses to thermal and mechanical stimuli, and alcohol intake in a rat model of intermittent drinking paradigm, as well as spontaneous glutamatergic transmission (sEPSCs), and firing of LHb neurons in brain slices. Here, we report that nociceptive sensitivity was higher in rats at 24 h withdrawal from chronic alcohol consumption than that of alcohol-naive counterparts. The basal frequency of sEPSCs and firings was higher in slices of withdrawn rats than that of Naïve rats, and 5-HT2R antagonists attenuated the enhancement. Also, an acute ethanol-induced increase of sEPSCs and firings was smaller in withdrawal than in Naïve rats; it was attenuated by 5-HT2R antagonists but mimicked by 5-HT2R agonists. Importantly, intra-LHb infusion of 5-HT2R agonists increased nociceptive sensitivity in Naïve rats, while antagonists or 5-HT reuptake blocker decreased nociceptive sensitivity and alcohol intake in withdrawn rats. Additionally, KN-62, a CaMKII inhibitor, attenuated the enhancement of EPSCs and firing induced by acute alcohol and by 5-HT2R agonist. Furthermore, intra-LHb KN-62 reduced nociceptive sensitivity and alcohol intake. Quantitative real-time PCR assay detected mRNA of 5-HT2A and 2C in the LHb. Thus adaptation in 5-HT2R-CaMKII signaling pathway contributes to the hyper-glutamatergic state, the hyperactivity of LHb neurons as well as the higher nociceptive sensitivity in rats withdrawn from chronic alcohol consumption.

Dual-mode dopamine increases mediated by 5-HT1B and 5-HT2C receptors inhibition, inducing impulsive behavior in trained rats.

Patients with eating disorders exhibit problems with appetitive impulse control. Interactions between dopamine and serotonin (5-HT) neuron in this setting are poorly characterized. Here we examined 5-HT receptor-mediated changes in extracellular dopamine during impulsive appetitive behavior in rats. Rats were trained to perform a cued lever-press (LP) task for a food reward such that they stopped experiencing associated dopamine increases. Trained rats were administered the mixed 5-HT1B/2C-receptor antagonist metergoline, the 5-HT2A/2C-receptor antagonist ketanserin, and p-chlorophenylalanine (PCPA). We measured dopamine changes in the ventral striatum using voltammetry and examined the number of premature LPs, reaction time (RT), and reward acquisition rate (RAR). Compared with controls, metergoline increased premature LPs and shortened RT significantly; ketanserin decreased premature LPs and lengthened RT significantly; and PCPA decreased premature LPs, lengthened RT, and decreased RAR significantly. Following metergoline administration, rats exhibited a fast phasic dopamine increase for 0.25-0.75 s after a correct LP, but only during LP for an incorrect LP. No dopamine increases were detected with ketanserin or PCPA, or in controls. After LP task completion, metergoline also caused dopamine to increase slowly and remain elevated; in contrast, ketanserin caused dopamine to increase slowly and decrease rapidly. No slow dopamine increase occurred with PCPA. Inhibition of 5-HT1B- and 5-HT2C-receptors apparently induced dual modes of extracellular dopamine increase: fast phasic, and slow long-lasting. These increases may be associated with the suppression of acquired prediction learning and retention of high motivation for reward, leading to impulsive excessive premature LPs.

Blockade of Serotonin 2C Receptors with SB-242084 Moderates Reduced Locomotor Activity and Rearing by Cannabinoid 1 Receptor Antagonist AM-251.

The endocannabinoid and serotonin (5-HT) systems have key roles in the regulation of several physiological functions such as motor activity and food intake but also in the development of psychiatric disorders. Here we tested the hypothesis, whether blockade of serotonin 2C (5-HT2C) receptors prevents the reduced locomotor activity and other behavioral effects caused by a cannabinoid 1 (CB1) receptor antagonist. As a pretreatment, we administered SB-242084 (1 mg/kg, ip.), a 5-HT2C receptor antagonist or vehicle (VEH) followed by the treatment with AM-251 (5 or 10 mg/kg, ip.), a CB1 receptor antagonist or VEH. The effects of the two drugs alone or in co-administration were investigated in social interaction (SI) and elevated plus maze (EPM) tests in male Wistar rats. Our results show that AM-251 decreased the time spent with rearing in the SI test and decreased locomotor activity in EPM test. In contrast, SB-242084 produced increased locomotor activity in SI test and evoked anxiolytic-like effect in both SI and EPM tests. When applied the drugs in combination, these behavioral effects of AM-251 were moderated by SB-242084. Based on these findings, we conclude that certain unwanted behavioral effects of CB1 receptor antagonists could be prevented by pretreatment with 5-HT2C receptor antagonists.

Identification of dual role of piperazine-linked phenyl cyclopropyl methanone as positive allosteric modulator of 5-HT2C and negative allosteric modulator of 5-HT2B receptors.

Allosteric modulators of G-protein-coupled receptors have lately gained significant traction in drug discovery. Recent studies have shown that allosteric modulation of serotonin 2C receptor (5-HT2C) as a viable strategy for the treatment of various central nervous system (CNS) disorders. Considering the critical role of 5-HT2C in the modulation of appetite, a selective positive allosteric modulator (PAM) of 5-HT2C offers a new opportunity for anti-obesity therapeutic development. In this study, phenyl cyclopropyl-linked N-heterocycles were synthesized and evaluated at 5-HT2C for agonist and PAM activity. Our study shows that imidazole linked phenyl cyclopropyl methanones has PAM activity on both 5-HT2C and serotonin 2B receptor (5-HT2B). Interestingly, piperazine linked phenyl cyclopropyl methanones (58) was active as PAM of 5-HT2C (increased the Emax of 5-HT to 139%), and as negative allosteric modulator (NAM) of 5-HT2B (decreases EC50 of 5-HT 10 times without affecting Emax). Similar effect of compound 58 was observed with synthetic orthosteric agonist lorcaserin on 5-HT2B. Molecular docking study revealed that all active compounds were binding to the predicted allosteric site on 5-HT2C and shared a common interacting residues. Finally, compound 58 suppressed food intake in Sprague Dawley (SD) rats similar to lorcaserin after i.c.v. administration. Therefore, these results suggest that piperazine moiety is essential for dual activity (PAM & NAM) of compounds 58, and supports the hypothesis of 5-HT2C PAM for the treatment of obesity similar to the full agonist.