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A genetic approach to access serotonin neurons for in vivo and in vitro studies.

ABSTRACT: Serotonin (5HT) is a critical modulator of neural circuits that support diverse behaviors and physiological processes, and multiple lines of evidence implicate abnormal serotonergic signaling in psychiatric pathogenesis. The significance of 5HT underscores the importance of elucidating the molecular pathways involved in serotonergic system development, function, and plasticity. However, these mechanisms remain poorly defined, owing largely to the difficulty of accessing 5HT neurons for experimental manipulation. To address this methodological deficiency, we present a transgenic route to selectively alter 5HT neuron gene expression. This approach is based on the ability of a Pet-1 enhancer region to direct reliable 5HT neuron-specific transgene expression in the CNS. Its versatility is illustrated with several transgenic mouse lines, each of which provides a tool for 5HT neuron studies. Two lines allow Cre-mediated recombination at different stages of 5HT neuron development. A third line in which 5HT neurons are marked with yellow fluorescent protein will have numerous applications, including their electrophysiological characterization. To demonstrate this application, we have characterized active and passive membrane properties of midbrain reticular 5HT neurons, which heretofore have not been reported to our knowledge. A fourth line in which Pet-1 loss of function is rescued by expression of a Pet-1 transgene demonstrates biologically relevant levels of transgene expression and offers a route for investigating serotonergic protein structure and function in a behaving animal. These findings establish a straightforward and reliable approach for developing an array of tools for in vivo and in vitro studies of 5HT neurons.

SUBMITTER: Scott MM

PROVIDER: S-EPMC1283423 | biostudies-literature | 2005 Nov

REPOSITORIES: biostudies-literature

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A genetic approach to access serotonin neurons for in vivo and in vitro studies.

Scott Michael M MM Wylie Christi J CJ Lerch Jessica K JK Murphy Roxanne R Lobur Katherine K Herlitze Stefan S Jiang Weihong W Conlon Ron A RA Strowbridge Ben W BW Deneris Evan S ES

Proceedings of the National Academy of Sciences of the United States of America 20051026 45

Serotonin (5HT) is a critical modulator of neural circuits that support diverse behaviors and physiological processes, and multiple lines of evidence implicate abnormal serotonergic signaling in psychiatric pathogenesis. The significance of 5HT underscores the importance of elucidating the molecular pathways involved in serotonergic system development, function, and plasticity. However, these mechanisms remain poorly defined, owing largely to the difficulty of accessing 5HT neurons for experimen ...[more]

PMID: 16251278

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Project description:Serotonin is a neurotransmitter involved in many psychiatric diseases. In humans, a lack of 5-HT2B receptors is associated with serotonin-dependent phenotypes, including impulsivity and suicidality. A lack of 5-HT2B receptors in mice eliminates the effects of molecules that directly target serotonergic neurons including amphetamine derivative serotonin releasers, and selective serotonin reuptake inhibitor antidepressants. In this work, we tested the hypothesis that 5-HT2B receptors directly and positively regulate raphe serotonin neuron activity. By ex vivo electrophysiological recordings, we report that stimulation by the 5-HT2B receptor agonist, BW723C86, increased the firing frequency of serotonin Pet1-positive neurons. Viral overexpression of 5-HT2B receptors in these neurons increased their excitability. Furthermore, in vivo 5-HT2B-receptor stimulation by BW723C86 counteracted 5-HT1A autoreceptor-dependent reduction in firing rate and hypothermic response in wild-type mice. By a conditional genetic ablation that eliminates 5-HT2B receptor expression specifically and exclusively from Pet1-positive serotonin neurons (Htr2b 5-HTKO mice), we demonstrated that behavioral and sensitizing effects of MDMA (3,4-methylenedioxy-methamphetamine), as well as acute behavioral and chronic neurogenic effects of the antidepressant fluoxetine, require 5-HT2B receptor expression in serotonergic neurons. In Htr2b 5-HTKO mice, dorsal raphe serotonin neurons displayed a lower firing frequency compared to control Htr2b lox/lox mice as assessed by in vivo extracellular recordings and a stronger hypothermic effect of 5-HT1A-autoreceptor stimulation was observed. The increase in head-twitch response to DOI (2,5-dimethoxy-4-iodoamphetamine) further confirmed the lower serotonergic tone resulting from the absence of 5-HT2B receptors in serotonin neurons. Together, these observations indicate that the 5-HT2B receptor acts as a direct positive modulator of serotonin Pet1-positive neurons in an opposite way as the known 5-HT1A-negative autoreceptor.

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Project description:BackgroundSchistosomiasis is a socioeconomically devastating parasitic infection afflicting hundreds of millions of people and animals worldwide. It is the most important helminth infection, and its treatment relies solely on the drug praziquantel. Oral H1-antihistamines are available worldwide, and these agents are among the most widely used of all medications in children and adults. Given the importance of the drug repositioning strategy, we evaluated the antischistosomal properties of the H1-antihistamine drugs commonly used in clinical practices.MethodsTwenty-one antihistamine drugs were initially screened against adult schistosomes ex vivo. Subsequently, we investigated the anthelmintic properties of these antihistamines in a murine model of schistosomiasis for both early and chronic S. mansoni infections at oral dosages of 400 mg/kg single dose or 100 mg/kg daily for five consecutive days. We also demonstrated and described the ability of three antihistamines to induce tegumental damage in schistosomes through the use of scanning electron microscopy.ResultsFrom phenotypic screening, we found that desloratadine, rupatadine, promethazine, and cinnarizine kill adult S. mansoni in vitro at low concentrations (5-15 µM). These results were further supported by scanning electron microscopy analysis. In an animal model, rupatadine and cinnarizine revealed moderate worm burden reductions in mice harboring either early or chronic S. mansoni infection. Egg production, a key mechanism for both transmission and pathogenesis, was also markedly inhibited by rupatadine and cinnarizine, and a significant reduction in hepatomegaly and splenomegaly was recorded. Although less effective, desloratadine also revealed significant activity against the adult and juvenile parasites.ConclusionsAlthough the worm burden reductions achieved are all only moderate, comparatively, treatment with any of the three antihistamines is more effective in early infection than praziquantel. On the other hand, the clinical use of H1-antihistamines for the treatment of schistosomiasis is highly unlikely.

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A genetic approach to access serotonin neurons for in vivo and in vitro studies.

Publications

A genetic approach to access serotonin neurons for in vivo and in vitro studies.

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