5-HT4R agonism reduces L-DOPA-induced dyskinesia via striatopallidal neurons in unilaterally 6-OHDA lesioned mice.

Parkinson's disease is caused by a selective vulnerability and cell loss of dopaminergic neurons of the Substantia Nigra pars compacta and, consequently, striatal dopamine depletion. In Parkinson's disease therapy, dopamine loss is counteracted by the administration of L-DOPA, which is initially effective in ameliorating motor symptoms, but over time leads to a burdening side effect of uncontrollable jerky movements, termed L-DOPA-induced dyskinesia. To date, no efficient treatment for dyskinesia exists. The dopaminergic and serotonergic systems are intrinsically linked, and in recent years, a role has been established for pre-synaptic 5-HT1a/b receptors in L-DOPA-induced dyskinesia. We hypothesized that post-synaptic serotonin receptors may have a role and investigated the effect of modulation of 5-HT4 receptor on motor symptoms and L-DOPA-induced dyskinesia in the unilateral 6-OHDA mouse model of Parkinson's disease. Administration of RS 67333, a 5-HT4 receptor partial agonist, reduces L-DOPA-induced dyskinesia without altering L-DOPA's pro-kinetic effect. In the dorsolateral striatum, we find 5-HT4 receptor to be predominantly expressed in D2R-containing medium spiny neurons, and its expression is altered by dopamine depletion and L-DOPA treatment. We further show that 5-HT4 receptor agonism not only reduces L-DOPA-induced dyskinesia, but also enhances the activation of the cAMP-PKA pathway in striatopallidal medium spiny neurons. Taken together, our findings suggest that agonism of the post-synaptic serotonin receptor 5-HT4 may be a novel therapeutic approach to reduce L-DOPA-induced dyskinesia.

The Serotonin 4 Receptor Subtype: A Target of Particular Interest, Especially for Brain Disorders.

In recent years, particular attention has been paid to the serotonin 4 receptor, which is well expressed in the brain, but also peripherally in various organs. The cerebral distribution of this receptor is well conserved across species, with high densities in the basal ganglia, where they are expressed by GABAergic neurons. The 5-HT4 receptor is also present in the cerebral cortex, hippocampus, and amygdala, where they are carried by glutamatergic or cholinergic neurons. Outside the central nervous system, the 5-HT4 receptor is notably expressed in the gastrointestinal tract. The wide distribution of the 5-HT4 receptor undoubtedly contributes to its involvement in a plethora of functions. In addition, the modulation of this receptor influences the release of serotonin, but also the release of other neurotransmitters such as acetylcholine and dopamine. This is a considerable asset, as the modulation of the 5-HT4 receptor can therefore play a direct or indirect beneficial role in various disorders. One of the main advantages of this receptor is that it mediates a much faster antidepressant and anxiolytic action than classical selective serotonin reuptake inhibitors. Another major benefit of the 5-HT4 receptor is that its activation enhances cognitive performance, probably via the release of acetylcholine. The expression of the 5-HT4 receptor is also altered in various eating disorders, and its activation by the 5-HT4 agonist negatively regulates food intake. Additionally, although the cerebral expression of this receptor is modified in certain movement-related disorders, it is still yet to be determined whether this receptor plays a key role in their pathophysiology. Finally, there is no longer any need to demonstrate the value of 5-HT4 receptor agonists in the pharmacological management of gastrointestinal disorders.

Activation of 5-Hydroxytryptamine 4 Receptor Improves Colonic Barrier Function by Triggering Mucin 2 Production in a Mouse Model of Type 1 Diabetes.

Diabetes leads to intestinal barrier dysfunction. 5-Hydroxytryptamine 4 receptor (5-HT4R) is distributed in the colonic mucosa, but little is known about the role of its activation in diabetes-evoked colonic barrier dysfunction. This study investigates whether activation of 5-HT4Rs on goblet cells (GCs) protects the colon from commensal bacterial translocation in diabetic mice. Expression of 5-HT4R detected inside the colonic epithelium by RNAscope in situ hybridization was further observed within the mucin 2 (MUC2)-immunoreactive GCs. In diabetic mice, neither 5-HT4R transcription nor protein levels were altered compared with those in nondiabetic mice. Bacterial translocation was characterized by 16S rRNA RNAscope in situ hybridization and manifested in both crypts and lamina propria of the colon in diabetic mice. Mucin production and MUC2 expression were significantly decreased in diabetic mice. Furthermore, the loss of mitochondrial cristae of GCs and the down-regulation of mitofilin, the core protein maintaining mitochondrial homeostasis, were observed in diabetic mice. Long-term treatment with 5-HT4R agonist in diabetic mice not only prevented bacterial penetration of the whole colonic mucosa but also promoted mucin production and MUC2 expression. Markedly, 5-HT4R agonist also restored the mitochondrial cristae of GCs and up-regulated mitofilin. However, co-administration of 5-HT4R antagonist abolished the effects of 5-HT4R agonist on diabetic mice. These findings indicate that 5-HT4R in colonic mucosa is an effective target for the treatment of diabetes-induced colonic mucous barrier dysfunction.

Stress Hormone Dynamics Are Coupled to Brain Serotonin 4 Receptor Availability in Unmedicated Patients With Major Depressive Disorder: A NeuroPharm Study.

BACKGROUND: A prominent finding in major depressive disorder (MDD) is distorted stress hormone dynamics, which is regulated by serotonergic brain signaling. An interesting feature of the cerebral serotonin system is the serotonin 4 receptor (5-HT4R), which is lower in depressed relative to healthy individuals and also has been highlighted as a promising novel antidepressant target. Here, we test the novel hypothesis that brain 5-HT4R availability in untreated patients with MDD is correlated with cortisol dynamics, indexed by the cortisol awakening response (CAR). Further, we evaluate if CAR changes with antidepressant treatment, including a selective serotonin reuptake inhibitor, and if pretreatment CAR can predict treatment outcome. METHODS: Sixty-six patients (76% women) with a moderate to severe depressive episode underwent positron emission tomography imaging with [11C]SB207145 for quantification of brain 5-HT4R binding using BPND as outcome. Serial home sampling of saliva in the first hour from awakening was performed to assess CAR before and after 8 weeks of antidepressant treatment. Treatment outcome was measured by change in Hamilton Depression Rating Scale 6 items. RESULTS: In the unmedicated depressed state, prefrontal and anterior cingulate cortices 5-HT4R binding was positively associated with CAR. CAR remained unaltered after 8 weeks of antidepressant treatment, and pretreatment CAR did not significantly predict treatment outcome. CONCLUSIONS: Our findings highlight a link between serotonergic disturbances in MDD and cortisol dynamics, which likely is involved in disease and treatment mechanisms. Further, our data support 5-HT4R agonism as a promising precision target in patients with MDD and disturbed stress hormone dynamics.

Activation of the 5-hydroxytryptamine 4 receptor ameliorates tight junction barrier dysfunction in the colon of type 1 diabetic mice.

Hyperglycemia drives dysfunction of the intestinal barrier. 5-Hydroxytryptaine 4 receptor (5-HT 4R) agonists have been considered therapeutics for constipation in clnic. However, the roles of 5-HT 4R activation in mucosa should be fully realized. Here, we investigate the effects of 5-HT 4R activation on diabetes-induced disruption of the tight junction (TJ) barrier in the colon. Not surprisingly, the TJ barrier in diabetic mice with or without 5-HT 4R is tremendously destroyed, as indicated by increased serum fluorescein isothiocyanate (FITC)-dextran and decreased transepithelial electrical resistance (TER). Simultaneously, decreased expressions of TJ proteins are shown in both wild-type (WT) and 5-HT 4R knockout (KO) mice with diabetes. Notably, chronic treatment with intraperitoneal injection of a 5-HT 4R agonist in WT mice with diabetes repairs the TJ barrier and promotes TJ protein expressions, including occludin, claudin-1 and ZO-1, in the colon, whereas a 5-HT 4R agonist does not improve TJ barrier function or TJ protein expressions in 5-HT 4R KO mice with diabetes. Furthermore, stimulation of 5-HT 4R inhibits diabetes-induced upregulation of myosin light chain kinase (MLCK), Rho-associated coiled coil protein kinase 1 (ROCK1), and phosphorylated myosin light chain (p-MLC), which are key molecules that regulate TJ integrity, in the colonic mucosa of WT mice. However, such action induced by a 5-HT 4R agonist is not observed in 5-HT 4R KO mice with diabetes. These findings indicate that 5-HT 4R activation may restore TJ integrity by inhibiting the expressions of MLCK, ROCK1 and p-MLC, improving epithelial barrier function in diabetes.

Cardiac Roles of Serotonin (5-HT) and 5-HT-Receptors in Health and Disease.

Serotonin acts solely via 5-HT4-receptors to control human cardiac contractile function. The effects of serotonin via 5-HT4-receptors lead to positive inotropic and chronotropic effects, as well as arrhythmias, in the human heart. In addition, 5-HT4-receptors may play a role in sepsis, ischaemia, and reperfusion. These presumptive effects of 5-HT4-receptors are the focus of the present review. We also discuss the formation and inactivation of serotonin in the body, namely, in the heart. We identify cardiovascular diseases where serotonin might play a causative or additional role. We address the mechanisms which 5-HT4-receptors can use for cardiac signal transduction and their possible roles in cardiac diseases. We define areas where further research in this regard should be directed in the future, and identify animal models that might be generated to this end. Finally, we discuss in what regard 5-HT4-receptor agonists or antagonists might be useful drugs that could enter clinical practice. Serotonin has been the target of many studies for decades; thus, we found it timely to summarise our current knowledge here.

Ergotamine Stimulates Human 5-HT4-Serotonin Receptors and Human H2-Histamine Receptors in the Heart.

Ergotamine (2'-methyl-5'α-benzyl-12'-hydroxy-3',6',18-trioxoergotaman) is a tryptamine-related alkaloid from the fungus Claviceps purpurea. Ergotamine is used to treat migraine. Ergotamine can bind to and activate several types of 5-HT1-serotonin receptors. Based on the structural formula of ergotamine, we hypothesized that ergotamine might stimulate 5-HT4-serotonin receptors or H2-histamine receptors in the human heart. We observed that ergotamine exerted concentration- and time-dependent positive inotropic effects in isolated left atrial preparations in H2-TG (mouse which exhibits cardiac-specific overexpression of the human H2-histamine receptor). Similarly, ergotamine increased force of contraction in left atrial preparations from 5-HT4-TG (mouse which exhibits cardiac-specific overexpression of the human 5-HT4-serotonin receptor). An amount of 10 µM ergotamine increased the left ventricular force of contraction in isolated retrogradely perfused spontaneously beating heart preparations of both 5-HT4-TG and H2-TG. In the presence of the phosphodiesterase inhibitor cilostamide (1 µM), ergotamine 10 µM exerted positive inotropic effects in isolated electrically stimulated human right atrial preparations, obtained during cardiac surgery, that were attenuated by 10 µM of the H2-histamine receptor antagonist cimetidine, but not by 10 µM of the 5-HT4-serotonin receptor antagonist tropisetron. These data suggest that ergotamine is in principle an agonist at human 5-HT4-serotonin receptors as well at human H2-histamine receptors. Ergotamine acts as an agonist on H2-histamine receptors in the human atrium.

Serotonin receptor 4 in the hippocampus modulates mood and anxiety.

Serotonin receptor 4 (5-HT4R) plays an important role in regulating mood, anxiety, and cognition, and drugs that activate this receptor have fast-acting antidepressant (AD)-like effects in preclinical models. However, 5-HT4R is widely expressed throughout the central nervous system (CNS) and periphery, making it difficult to pinpoint the cell types and circuits underlying its effects. Therefore, we generated a Cre-dependent 5-HT4R knockout mouse line to dissect the function of 5-HT4R in specific brain regions and cell types. We show that the loss of functional 5-HT4R specifically from excitatory neurons of hippocampus led to robust AD-like behavioral responses and an elevation in baseline anxiety. 5-HT4R was necessary to maintain the proper excitability of dentate gyrus (DG) granule cells and cell type-specific molecular profiling revealed a dysregulation of genes necessary for normal neural function and plasticity in cells lacking 5-HT4R. These adaptations were accompanied by an increase in the number of immature neurons in ventral, but not dorsal, dentate gyrus, indicating a broad impact of 5-HT4R loss on the local cellular environment. This study is the first to use conditional genetic targeting to demonstrate a direct role for hippocampal 5-HT4R signaling in modulating mood and anxiety. Our findings also underscore the need for cell type-based approaches to elucidate the complex action of neuromodulatory systems on distinct neural circuits.

Interplay between 5-HT4 Receptors and GABAergic System within CA1 Hippocampal Synaptic Plasticity.

The type 4 serotonin receptor (5-HT4R) is highly involved in cognitive processes such as learning and memory. Behavioral studies have shown a beneficial effect of its activation and conversely reported memory impairments by its blockade. However, how modulation of 5HT4R enables modifications of hippocampal synaptic plasticity remains elusive. To shed light on the mechanisms at work, we investigated the effects of the 5-HT4R agonist RS67333 on long-term potentiation (LTP) within the hippocampal CA1 area. Although high-frequency stimulation-induced LTP remained unaffected by RS67333, the magnitude of LTP induced by theta-burst stimulation was significantly decreased. This effect was blocked by the selective 5-HT4R antagonist RS39604. Further, 5-HT4R-induced decrease in LTP magnitude was fully abolished in the presence of bicuculline, a GABAAR antagonist; hence, demonstrating involvement of GABA neurotransmission. In addition, we showed that the application of a GABABR antagonist, CGP55845, mimicked the effect of 5-HT4R activation, whereas concurrent application of CGP55845 and RS67333 did not elicit an additive inhibition effect on LTP. To conclude, through investigation of theta burst induced functional plasticity, we demonstrated an interplay between 5-HT4R activation and GABAergic neurotransmission within the hippocampal CA1 area.

Epithelial 5-HT4 Receptors as a Target for Treating Constipation and Intestinal Inflammation.

Because of their importance in the regulation of gut functions, several therapeutic targets involving serotonin-related proteins have been developed or repurposed to treat motility disorders, including serotonin transporter inhibitors, tryptophan hydroxylase blockers, 5-HT3 antagonists, and 5-HT4 agonists. This chapter focuses on our discovery of 5-HT4 receptors in the epithelial cells of the colon and our efforts to evaluate the effects of stimulating these receptors. 5-HT4 receptors appear to be expressed by all epithelial cells in the mouse colon, based on expression of a reporter gene driven by the 5-HT4 receptor promoter. Application of 5-HT4 agonists to the mucosal surface causes serotonin release from enterochromaffin cells, mucus secretion from goblet cells, and chloride secretion from enterocytes. Luminal administration of 5-HT4 agonists speeds up colonic motility and suppresses distention-induced nociceptive responses. Luminal administration of 5-HT4 agonists also decreases the development of, and improves recovery from, experimental colitis. Recent studies determined that the prokinetic actions of minimally absorbable 5-HT4 agonists are just as effective as absorbable compounds. Collectively, these findings indicate that targeting epithelial receptors with non-absorbable 5-HT4 agonists could offer a safe and effective strategy for treating constipation and colitis.

Translating the promise of 5HT4 receptor agonists for the treatment of depression.

Animal experimental studies suggest that 5-HT4 receptor activation holds promise as a novel target for the treatment of depression and cognitive impairment. 5-HT4 receptors are post-synaptic receptors that are located in striatal and limbic areas known to be involved in cognition and mood. Consistent with this, 5-HT4 receptor agonists produce rapid antidepressant effects in a number of animal models of depression, and pro-cognitive effects in tasks of learning and memory. These effects are accompanied by molecular changes, such as the increased expression of neuroplasticity-related proteins that are typical of clinically useful antidepressant drugs. Intriguingly, these antidepressant-like effects have a fast onset of their action, raising the possibility that 5-HT4 receptor agonists may be a particularly useful augmentation strategy in the early stages of SSRI treatment. Until recently, the translation of these effects to humans has been challenging. Here, we review the evidence from animal studies that the 5-HT4 receptor is a promising target for the treatment of depression and cognitive disorders, and outline a potential pathway for the efficient and cost-effective translation of these effects into humans and, ultimately, to the clinic.

Gut microbiota regulates maturation of the adult enteric nervous system via enteric serotonin networks.

The enteric nervous system (ENS) is crucial for essential gastrointestinal physiologic functions such as motility, fluid secretion, and blood flow. The gut is colonized by trillions of bacteria that regulate host production of several signaling molecules including serotonin (5-HT) and other hormones and neurotransmitters. Approximately 90% of 5-HT originates from the intestine, and activation of the 5-HT4 receptor in the ENS has been linked to adult neurogenesis and neuroprotection. Here, we tested the hypothesis that the gut microbiota could induce maturation of the adult ENS through release of 5-HT and activation of 5-HT4 receptors. Colonization of germ-free mice with a microbiota from conventionally raised mice modified the neuroanatomy of the ENS and increased intestinal transit rates, which was associated with neuronal and mucosal 5-HT production and the proliferation of enteric neuronal progenitors in the adult intestine. Pharmacological modulation of the 5-HT4 receptor, as well as depletion of endogenous 5-HT, identified a mechanistic link between the gut microbiota and maturation of the adult ENS through the release of 5-HT and activation of the 5-HT4 receptor. Taken together, these findings show that the microbiota modulates the anatomy of the adult ENS in a 5-HT-dependent fashion with concomitant changes in intestinal transit.

Understanding the Molecular Basis of 5-HT4 Receptor Partial Agonists through 3D-QSAR Studies.

Alzheimer's disease (AD) is a neurodegenerative disorder whose prevalence has an incidence in senior citizens. Unfortunately, current pharmacotherapy only offers symptom relief for patients with side effects such as bradycardia, nausea, and vomiting. Therefore, there is a present need to provide other therapeutic alternatives for treatments for these disorders. The 5-HT4 receptor is an attractive therapeutic target since it has a potential role in central and peripheral nervous system disorders such as AD, irritable bowel syndrome, and gastroparesis. Quantitative structure-activity relationship analysis of a series of 62 active compounds in the 5-HT4 receptor was carried out in the present work. The structure-activity relationship was estimated using three-dimensional quantitative structure-activity relationship (3D-QSAR) techniques based on these structures' field molecular (force and Gaussian field). The best force-field QSAR models achieve a value for the coefficient of determination of the training set of R2training = 0.821, and for the test set R2test = 0.667, while for Gaussian-field QSAR the training and the test were R2training = 0.898 and R2test = 0.695, respectively. The obtained results were validated using a coefficient of correlation of the leave-one-out cross-validation of Q2LOO = 0.804 and Q2LOO = 0.886 for force- and Gaussian-field QSAR, respectively. Based on these results, novel 5-HT4 partial agonists with potential biological activity (pEC50 8.209-9.417 for force-field QSAR and 9.111-9.856 for Gaussian-field QSAR) were designed. In addition, for the new analogues, their absorption, distribution, metabolism, excretion, and toxicity properties were also analyzed. The results show that these new derivatives also have reasonable pharmacokinetics and drug-like properties. Our findings suggest novel routes for the design and development of new 5-HT4 partial agonists.

Memory Disorders Related to Hippocampal Function: The Interest of 5-HT4Rs Targeting.

The hippocampus has long been considered as a key structure for memory processes. Multilevel alterations of hippocampal function have been identified as a common denominator of memory impairments in a number of psychiatric and neurodegenerative diseases. For many years, the glutamatergic and cholinergic systems have been the main targets of therapeutic treatments against these symptoms. However, the high rate of drug development failures has left memory impairments on the sideline of current therapeutic strategies. This underscores the urgent need to focus on new therapeutic targets for memory disorders, such as type 4 serotonin receptors (5-HT4Rs). Ever since the discovery of their expression in the hippocampus, 5-HT4Rs have gained growing interest for potential use in the treatment of learning and memory impairments. To date, much of the researched information gathered by scientists from both animal models and humans converge on pro-mnesic and anti-amnesic properties of 5-HT4Rs activation, although the mechanisms at work require more work to be fully understood. This review addresses a fundamental, yet poorly understood set of evidence of the potential of 5-HT4Rs to re-establish or limit hippocampal alterations related to neurological diseases. Most importantly, the potential of 5-HT4Rs is translated by refining hypotheses regarding the benefits of their activation in memory disorders at the hippocampal level.

Expression of serotonin receptor HTR4 in glucagon-like peptide-1-positive enteroendocrine cells of the murine intestine.

Serotonin (5-hydroxytryptamine [5-HT]) synthesized and released in enterochromaffin (EC) cells participates in various functions in the gastrointestinal tract by acting on a diverse range of 5-HT receptors (HTRs) expressed on smooth muscle, enteric neurons, and epithelial cells. We previously observed that genes encoding HTR2A, HTR2B, and HTR4 are expressed in murine intestinal organoids, suggesting the expression of these HTRs in intestinal epithelial cells. The present study investigated the localization of these HTRs in the murine intestine by immunofluorescence staining. HTR2A, HTR2B, and HTR4 localized in individual solitary cells in the epithelium, while HTR2C was observed in the lamina propria. In the epithelium, HTR2A, HTR2B, and HTR4 colocalized with 5-HT, and HTR4 colocalized with glucagon-like peptide 1 (GLP-1) and peptide YY (PYY). Murine intestinal organoids show a colocalization pattern that is similar to in vivo HTR2A and HTR4 with 5-HT, GLP-1, and PYY. Intraperitoneal and intragastric administration of tegaserod, an HTR4 agonist, failed to alter plasma GLP-1 levels in fasted mice. However, intragastric but not intraperitoneal administration of tegaserod reduced dietary lipid-induced increases of plasma GLP-1 levels. This action of tegaserod was inhibited by co-administration of RS39604, an HTR4 antagonist. These results suggest that murine ileal GLP-1/PYY-producing enteroendocrine (EE) cells express HTR4, while 5-HT-producing EC cells express HTR2A, HTR2B, and HTR4. In addition, the observations regarding in vivo GLP-1 secretion suggest that HTR4 signaling in ileal EE cells suppresses dietary lipid-induced GLP-1 secretion. We thus propose that EC and EE cells may interact with each other through paracrine signaling mechanisms.

Oral contraceptives and the serotonin 4 receptor: a molecular brain imaging study in healthy women.

OBJECTIVE: Sex steroid hormones potently shape brain functions, including those critical to maintain mental health such as serotonin signaling. Use of oral contraceptives (OCs) profoundly changes endogenous sex steroid hormone levels and dynamics. Recent register-based studies show that starting an OC is associated with increased risk of developing depression. Here, we investigate whether use of OCs in healthy women is associated with a marker of the serotonin system in terms of serotonin 4 receptor (5-HT4R) brain imaging. METHODS: [11C]SB207145-PET imaging data on 53 healthy women, of whom 16 used OCs, were available from the Cimbi database. We evaluated global effects of OC use on 5-HT4R binding in a latent variable model based on 5-HT4R binding across cortical and subcortical regions. RESULTS: We demonstrate that OC users have 9-12% lower global brain 5-HT4R binding potential compared to non-users. Univariate region-based analyses (pallidostriatum, caudate, hippocampus, amygdala, anterior cingulate cortex, and neocortex) supported the global effect of OC use with the largest difference present in the hippocampus (-12.8% (95% CI [-21.0; -3.9], Pcorrected = 0.03). CONCLUSION: We show that women who use OCs have markedly lower brain 5-HT4R binding relative to non-users, which constitutes a plausible molecular link between OC use and increased risk of depressive episodes. We propose that this reflects a reduced 5-HT4R gene expression, possibly related to a blunted ovarian hormone state among OC users.

Genotoxicity and carcinogenicity risk assessment of prucalopride, a selective 5-hydroxytryptamine 4 receptor agonist.

Prucalopride, a high affinity, selective serotonin type 4 (5-HT4) receptor agonist, was associated with increased neoplasia incidence (in endocrine tissues and liver) in 2-year rodent bioassays, without evidence of a genotoxic mechanism of action. Proposed mechanisms of action involve prolactin and the constitutive androstane receptor (CAR). Epigenetic mechanisms and their relevance to humans are discussed. Data from in vitro and in vivo rodent studies demonstrated that prucalopride-related stimulation of prolactin secretion (via dopamine receptor D2 antagonism at high doses) is a rodent-specific, non-genotoxic mechanism for inducing hyperplasia and neoplasia in prolactin receptor-expressing endocrine tissues. Additional data demonstrated that CAR-mediated liver enzyme induction underlies the observed hepatocellular adenomas and thyroid follicular adenomas in rodents. A 12-month neonatal mouse carcinogenicity study confirmed the lack of a genotoxic mechanism of action. Furthermore, tumors were observed only at very high exposures (200 and 63 fold higher in mice and rats, respectively, than human exposure after a daily therapeutic dose of 2 mg). The studies indicate that non-genotoxic, rodent-specific, epigenetic mechanisms that are considered clinically irrelevant are responsible for the increased incidence of neoplasias associated with very high exposure to prucalopride in rodents, and that prucalopride does not pose a carcinogenic safety risk to humans.

Related GPCRs couple differently to Gs: preassociation between G protein and 5-HT7 serotonin receptor reveals movement of Gαs upon receptor activation.

How GPCRs and G proteins interact is important for their biologic functions and their functions as pharmacologic targets. It is still an open question whether receptors and G proteins are preassembled in a complex or interact only after receptor activation. We compared the propensity of the two Gs-coupled serotonin (5-HT) receptors 5-HT4 and 5-HT7 to associate with G protein prior to agonist activation. Combining receptor-immobilized fluorescence recovery after photobleaching and fluorescence resonance energy transfer methodologies, we observed that 5-HT7 receptors markedly reduced the diffusion of both Gα and Gßγ at the cell surface, which indicated 5-HT7 receptor preassociation with Gs. This is in sharp contrast to the 5-HT4 receptor for which the diffusion of Gαßγ was not modified, and agonist activation brought together the receptor and Gγ, which is consistent with interaction by collision coupling. Agonist activation of 5-HT7 dissociated Gγ from the receptor, whereas Gαs underwent a rapid conformational change with respect to both Gγ and the receptor, followed by a slower dissociation of Gγ from both Gαs and the receptor. Taken together, these data demonstrate a different propensity among receptors to preassociate with G protein in the absence of ligand and reveals a rapid conformational change in Gαs upon activation by the receptor.-Andressen, K. W., Ulsund, A. H., Krobert, K. A., Lohse, M. J., Bünemann, M., Levy, F. O. Related GPCRs couple differently to Gs: preassociation between G protein and 5-HT7 serotonin receptor reveals movement of Gαs upon receptor activation.

CK2 regulates 5-HT4 receptor signaling and modulates depressive-like behavior.

The serotonergic neurotransmitter system has been widely implicated in the pathophysiology of mood-related disorders such as anxiety and major depressive disorder (MDD). The onset of therapeutic efficacy of traditional antidepressants is delayed by several weeks. The 5-HT4 receptor has emerged as a new therapeutic target since agonists of this receptor induce rapid antidepressant-like responses in rodents. Here we show that the 5-HT4 receptor is regulated by CK2, at transcriptional and post-transcriptional levels. We present evidence, in two different CK2α knockout mouse lines, that this regulation is region-specific, with the 5-HT4 receptor upregulated in prefrontal cortex (PFC) but not striatum or hippocampus where CK2α is also ablated. 5-HT4 receptor signaling is enhanced in vitro, as evidenced by enhanced cAMP production or receptor plasma membrane localization in the presence of CK2 inhibitor or shRNA targeting CK2α. In vivo, 5-HT4 receptor signaling is also upregulated since ERK activation is elevated and sensitive to the inverse agonist, GR113808 in the PFC of CK2α KO mice. Behaviorally, KO mice as well as mice with AAV-mediated deletion of CK2α in the PFC show a robust 'anti-depressed-like' phenotype and display an enhanced response to antidepressant treatment when tested in paradigms for mood and anxiety. Importantly, it is sufficient to overexpress the 5-HT4 receptor in the mPFC to generate mice with a similar 'anti-depressed-like' phenotype. Our findings identify the mPFC as the region that mediates the effect of enhanced 5-HT4 receptor activity and CK2 as modulator of 5-HT4 receptor levels in this brain region that regulates mood-related phenotypes.

Migraine is associated with high brain 5-HT levels as indexed by 5-HT4 receptor binding.

INTRODUCTION: Serotonin (5-HT) plays a role in migraine pathophysiology, but whether brain 5-HT is involved in the conversion from episodic to chronic migraine is unknown. Here, we investigated brain 5-HT levels, as indexed by 5-HT4 receptor binding, in chronic migraine patients and evaluated whether these were associated with migraine frequency. METHODS: Sixteen chronic migraine patients underwent a dynamic PET scan after injection of [11C]SB207145, a specific 5-HT4 receptor radioligand. Data from 15 episodic migraine patients and 16 controls were included for comparison. Quantification of 5-HT4 receptor binding was used as a proxy for brain 5-HT levels, since 5-HT4 receptor binding is inversely related to brain 5-HT levels. RESULTS: Chronic migraine patients had 9.1% (95% CI: [-17%; -1.0%]) lower 5-HT4 receptor binding compared to controls ( p = 0.039). There was no difference in 5-HT4 receptor binding between chronic and episodic migraine patients ( p = 0.48) and no association between number of monthly migraine days and 5-HT4 receptor binding (slope estimate 0.003, 95% CI: [-0.004; 0.715], p = 0.39). CONCLUSION: The finding of low 5-HT4 receptor binding suggests that cerebral levels of 5-HT are elevated in chronic migraine patients. This is in line with observations made in patients with episodic migraine. Elevated brain 5-HT levels may thus be an inherent trait of the migraine brain rather than a risk factor for conversion from episodic to chronic migraine.