Serotonin uptake and release mechanisms in developing cultures of rat embryonic raphe neurons: age- and region-specific differences

Lautenschlager, Marion; Höltje, Markus; Jagow, B. von; Veh, Ruediger W.; Harms, Christoph; Bergk, A.; Dirnagl, Ulrich; Ahnert‐Hilger, Gudrun; Hörtnagl, Heide

doi:10.1016/s0306-4522(00)00222-0

Cited by 23 publications

(17 citation statements)

References 29 publications

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“…Primary Raphé Nuclei Cultures and Double Fluorescence Deconvolution Microscopy-Rat embryos (E15 Ϯ 1 day) were obtained from pregnant Sprague-Dawley females, and the brainstem region containing both rostral and caudal raphé nuclei was removed, and serum-free cultures were prepared by standard procedures (40,41). Dissected tissue was placed in Hepes-buffered Ringer solution (130 mM NaCl, 4 mM KCl, 1 mM MgCl 2 , 1.5 mM CaCl 2 , 10 mM Hepes, 10 mM dextrose, 3 mM NaOH, pH 7.3) and then digested for 15 min with papain solution (Hepes-buffered Ringer solution with 1.5 mM CaCl 2 , 0.5 mM EDTA, 9 units/ml papain, 0.2 mg/ml cysteine).…”

Section: Methodsmentioning

confidence: 99%

Molecular Cloning and Functional Identification of Mouse Vesicular Glutamate Transporter 3 and Its Expression in Subsets of Novel Excitatory Neurons

Schäfer¹,

Varoqui²,

Defamie³

et al. 2002

Journal of Biological Chemistry

372

392

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We have cloned and functionally characterized a third isoform of a vesicular glutamate transporter (VGLUT3) expressed on synaptic vesicles that identifies a distinct glutamatergic system in the brain that is partly and selectively promiscuous with cholinergic and serotoninergic transmission. Transport activity was specific for glutamate, was H ؉ -dependent, was stimulated by Cl ؊ ion, and was inhibited by Rose Bengal and trypan blue. Northern analysis revealed higher mRNA levels in early postnatal development than in adult brain. Restricted patterns of mRNA expression were observed in presumed interneurons in cortex and hippocampus, and projection systems were observed in the lateral and ventrolateral hypothalamic nuclei, limbic system, and brainstem. Double in situ hybridization histochemistry for vesicular acetylcholine transporter identified VGLUT3 neurons in the striatum as cholinergic interneurons, whereas VGLUT3 mRNA and protein were absent from all other cholinergic cell groups. In the brainstem VGLUT3 mRNA was concentrated in mesopontine raphé nuclei. VGLUT3 immunoreactivity was present throughout the brain in a diffuse system of thick and thin beaded varicose fibers much less abundant than, and strictly separated from, VGLUT1 or VGLUT2 synapses. Co-existence of VGLUT3 in VMAT2-positive and tyrosine hydroxylase -negative varicosities only in the cerebral cortex and hippocampus and in subsets of tryptophan hydroxylase-positive cell bodies and processes in differentiating primary raphé neurons in vitro indicates selective and target-specific expression of the glutamatergic/serotoninergic synaptic phenotype.

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Section: Methodsmentioning

confidence: 99%

Molecular Cloning and Functional Identification of Mouse Vesicular Glutamate Transporter 3 and Its Expression in Subsets of Novel Excitatory Neurons

Schäfer¹,

Varoqui²,

Defamie³

et al. 2002

Journal of Biological Chemistry

372

392

View full text Add to dashboard Cite

show abstract

“…Briefly, cells were grown in minimum essential medium supplemented with Earle's salts, penicillin (100 U/mL), streptomycin (100 lg/ml/mL) and 10 % fetal calf serum (FCS; Biochrome, Berlin, Germany). Primary neuronal cultures of cerebral cortex were obtained from embryos (E17) of Wistar rats (FEM, Charité) as described in detail elsewhere (Lautenschlager et al 2000). Experiments were begun after 9 days in vitro (DIV).…”

Section: Cell Culturesmentioning

confidence: 99%

Direct inhibition of retinoic acid catabolism by fluoxetine

et al. 2015

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Recent evidence from animal and human studies suggests neuroprotective effects of the SSRI fluoxetine, e.g., in the aftermath of stroke. The underlying molecular mechanisms remain to be fully defined. Because of its effects on the cytochrome P450 system (CYP450), we hypothesized that neuroprotection by fluoxetine is related to altered metabolism of retinoic acid (RA), whose CYP450-mediated degradation in brain tissue constitutes an important step in the regulation of its site-specific auto- and paracrine actions. Using traditional pharmacological in vitro assays, the effects of fluoxetine on RA degradation were probed in crude synaptosomes from rat brain and human-derived SH-SY5Y cells, and in cultures of neuron-like SH-SY5Y cells. Furthermore, retinoid-dependent effects of fluoxetine on neuronal survival following glutamate exposure were investigated in rat primary neurons cells using specific retinoid receptor antagonists. Experiments revealed dose-dependent inhibition of synaptosomal RA degradation by fluoxetine along with dose-dependent increases in RA levels in cell cultures. Furthermore, fluoxetine's neuroprotective effects against glutamate excitotoxicity in rat primary neurons were demonstrated to partially depend on RA signaling. Taken together, these findings demonstrate for the first time that the potent, pleiotropic antidepressant fluoxetine directly interacts with RA homeostasis in brain tissue, thereby exerting its neuroprotective effects.

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“…Cultures of medullary raphe neurons were prepared from neonatal (P2) mice from four mutants and four WT animals, following a protocol for serum-free cultures (Brewer, 1995), which was modified and adapted to raphe neurons (Lautenschlager et al, 2000). The dissection included neurons from all three medullary raphe nuclei: raphe obscurus, raphe magnus, and raphe pallidus.…”

Section: Primary Serum-free Cultures Of 5-ht Raphe Neurons and Immunomentioning

confidence: 99%

Necdin Plays a Role in the Serotonergic Modulation of the Mouse Respiratory Network: Implication for Prader-Willi Syndrome

Zanella¹,

Watrin²,

Mébarek³

et al. 2008

J. Neurosci.

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Prader-Willi syndrome is a neurogenetic disease resulting from the absence of paternal expression of several imprinted genes, including NECDIN. Prader-Willi children and adults have severe breathing defects with irregular rhythm, frequent sleep apneas, and blunted respiratory regulations. For the first time, we show that Prader-Willi infants have sleep apneas already present at birth. In parallel, in wild-type and Necdin-deficient mice, we studied the respiratory system with in vivo plethysmography, in vitro electrophysiology, and pharmacology. Because serotonin is known to contribute to CNS development and to affect maturation and function of the brainstem respiratory network, we also investigated the serotonergic system with HPLC, immunohistochemistry, Rabies virus tracing approaches, and primary culture experiments. We report first that Necdin-deficiency in mice induces central respiratory deficits reminiscent of Prader-Willi syndrome (irregular rhythm, frequent apneas, and blunted respiratory regulations), second that Necdin is expressed by medullary serotonergic neurons, and third that Necdin deficiency alters the serotonergic metabolism, the morphology of serotonin vesicles in medullary serotonergic neurons but not the number of these cells. We also show that Necdin deficiency in neonatal mice alters the serotonergic modulation of the respiratory rhythm generator. Thus, we propose that the lack of Necdin expression induces perinatal serotonergic alterations that affect the maturation and function of the respiratory network, inducing breathing deficits in mice and probably in Prader-Willi patients.

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Serotonin uptake and release mechanisms in developing cultures of rat embryonic raphe neurons: age- and region-specific differences

Cited by 23 publications

References 29 publications

Molecular Cloning and Functional Identification of Mouse Vesicular Glutamate Transporter 3 and Its Expression in Subsets of Novel Excitatory Neurons

Molecular Cloning and Functional Identification of Mouse Vesicular Glutamate Transporter 3 and Its Expression in Subsets of Novel Excitatory Neurons

Direct inhibition of retinoic acid catabolism by fluoxetine

Necdin Plays a Role in the Serotonergic Modulation of the Mouse Respiratory Network: Implication for Prader-Willi Syndrome

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