A genetic approach to access serotonin neurons forin vivoandin vitrostudies

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.Cre recombinase ͉ pet-1 ͉ transgenic ͉ yellow fluorescent protein ͉ monoamine S erotonin (5HT) is a transmitter of broad relevance to nervous system development and function (1-4). Serotonergic pathways innervate most cytoarchitectonic structures of the CNS, and accordingly they have been implicated in the modulation of circuitry involved in nearly all behaviors and physiological processes (3, 5-7). Additionally, 5HT neurotransmission is modulated through abundant afferent information arising from, for example, other monoaminergic systems (8, 9) and from orexinergic, glutamatergic, and GABAergic pathways (10-12). The remarkably expansive neuromodulatory influence of 5HT is the product of a complex transcriptional cascade that generates 5HT-synthesizing neurons in the ventral hindbrain (13-18). 5HT also figures prominently in mental health disorders as a number of lines of evidence provide strong support for the hypothesis that altered serotonergic signaling contributes to neurological and psychiatric pathogenesis (19)(20)(21)(22). Despite considerable progress in understanding the importance of 5HT neurotransmission, however, the mechanisms governing 5HT neuron development and the precise physiological roles of 5HT in the modulation of CNS circuitry are not yet clear.Studies of 5HT neurons are hind...

Gα i/o -coupled receptor signaling restricts pancreatic β-cell expansion

“…First, we expressed a well-studied Gi-GPCR, serotonin receptor HTR1A, in developing islet cells under the control of the ePet1 enhancer from the Fev gene ( Fig. S1A) (18,19). The transgenic progeny of one ePet1-Htr1a transgenic founder displayed marked hyperglycemia (Fig.…”

Loss of MeCP2 in aminergic neurons causes cell-autonomous defects in neurotransmitter synthesis and specific behavioral abnormalities

“…The TH-Cre transgenic mouse line expresses Cre in all TH expressing regions of the brain, including the substantia nigra/ ventral tegmental area, locus ceruleus, and medullary norepinephrine neurons, as well as in the peripheral nervous system and the adrenal medulla (19). In contrast, the PET1-Cre transgenic mouse line expresses Cre driven by the PET1 enhancer element in the dorsal and medial raphe nuclei (20). The recombination efficiency of each Cre line was tested by immunofluorescence, showing that Mecp2 was effectively deleted from the specified brain regions (SI Text and Fig.…”

The Presynaptic Component of the Serotonergic System is Required for Clozapine's Efficacy

“…These hypotheses predict that deletion of the presynaptic component of the serotonin neuronal system would not affect clozapine's therapeutic actions. To test these hypotheses, we took advantage of the observation that the ETS-domain transcription factor pet1 is essential for normal expression of the serotonergic phenotype in neurons, and that its deletion greatly diminishes expression of presynaptic serotonergic markers (Hendricks et al, 2003;Jensen et al, 2008;Scott et al, 2005).…”

“…Slices from the pet1 À/À mice were also processed to detect expression of the lacZ marker of 5-HT precursor neurons as previously described (Scott et al, 2005), but using chicken anti-b-galactosidase (1 : 1000, Abcam) and FITC-conjugated goat anti-chicken (1 : 250, Abcam) antibodies.…”