Ethanol Induces Morphological and Dynamic Changes on In Vivo and In Vitro Neural Crest Cells

Rovasio, Roberto A.; Battiato, Natalia Laura

doi:10.1097/00000374-200208000-00023

Cited by 17 publications

(34 citation statements)

References 41 publications

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“…However, our results are not consistent with this view because at the critical period of ethanol teratogenicity, namely prim-6 and prim-16 , the neural crest cells of the zebrafish have already completed migration into the pharyngeal arches [58]–[66]. Thus it is possible that at least some of the hypoplasia of the pharyngeal arches seen in our study could be due to effects on postmigratory neural crest cells, in contrast to studies in chick and mouse embryos that suggest ethanol to have a major effect on migratory crest cells [9], [67]–[69]. Whether these differences are due to fundamental differences in the responses of these model species remains to be determined.…”

Section: Discussioncontrasting

confidence: 54%

Large-Scale Analysis of Acute Ethanol Exposure in Zebrafish Development: A Critical Time Window and Resilience

et al. 2011

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BackgroundIn humans, ethanol exposure during pregnancy causes a spectrum of developmental defects (fetal alcohol syndrome or FAS). Individuals vary in phenotypic expression. Zebrafish embryos develop FAS-like features after ethanol exposure. In this study, we ask whether stage-specific effects of ethanol can be identified in the zebrafish, and if so, whether they allow the pinpointing of sensitive developmental mechanisms. We have therefore conducted the first large-scale (>1500 embryos) analysis of acute, stage-specific drug effects on zebrafish development, with a large panel of readouts.Methodology/Principal FindingsZebrafish embryos were raised in 96-well plates. Range-finding indicated that 10% ethanol for 1 h was suitable for an acute exposure regime. High-resolution magic-angle spinning proton magnetic resonance spectroscopy showed that this produced a transient pulse of 0.86% concentration of ethanol in the embryo within the chorion. Survivors at 5 days postfertilisation were analysed. Phenotypes ranged from normal (resilient) to severely malformed. Ethanol exposure at early stages caused high mortality (≥88%). At later stages of exposure, mortality declined and malformations developed. Pharyngeal arch hypoplasia and behavioral impairment were most common after prim-6 and prim-16 exposure. By contrast, microphthalmia and growth retardation were stage-independent.ConclusionsOur findings show that some ethanol effects are strongly stage-dependent. The phenotypes mimic key aspects of FAS including craniofacial abnormality, microphthalmia, growth retardation and behavioral impairment. We also identify a critical time window (prim-6 and prim-16) for ethanol sensitivity. Finally, our identification of a wide phenotypic spectrum is reminiscent of human FAS, and may provide a useful model for studying disease resilience.

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Section: Discussioncontrasting

confidence: 54%

Large-Scale Analysis of Acute Ethanol Exposure in Zebrafish Development: A Critical Time Window and Resilience

et al. 2011

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“…Cell death in the frontonasal process, which is mainly derived from CNCC, and the forebrain is one of the most accepted mechanism underlying FAS craniofacial and nervous system phenotypes [43]. Primary cultures of CNCC from mouse [13] and chick [12] show increased cell death in EtOH treated cultures. Additionally, experiments performed in ovo , where EtOH was injected directly into the chick egg, also reported increased cell death of CNCC [9], [11].…”

Section: Discussionmentioning

confidence: 99%

“…Additional studies in chick have reported cell death in neural crest cells [11]. In vitro analysis of CNCC has shown that EtOH exposure produces permanent changes in cell shape, surface morphology, migration, and cell death [12], [13]. Interestingly no analysis of migration defects have been reported in vivo , although histological studies have suggested the occurrence of EtOH-induced migration defects, based on the pattern of immunostained neural crest in treated chick embryos [14].…”

Section: Introductionmentioning

confidence: 99%

Quantitative Analysis of Cell Migration Using Optical Flow

et al. 2013

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Neural crest cells exhibit dramatic migration behaviors as they populate their distant targets. Using a line of zebrafish expressing green fluorescent protein (sox10:EGFP) in neural crest cells we developed an assay to analyze and quantify cell migration as a population, and use it here to characterize in detail the subtle defects in cell migration caused by ethanol exposure during early development. The challenge was to quantify changes in the in vivo migration of all Sox10:EGFP expressing cells in the visual field of time-lapse movies. To perform this analysis we used an Optical Flow algorithm for motion detection and combined the analysis with a fit to an affine transformation. Through this analysis we detected and quantified significant differences in the cell migrations of Sox10:EGFP positive cranial neural crest populations in ethanol treated versus untreated embryos. Specifically, treatment affected migration by increasing the left-right asymmetry of the migrating cells and by altering the direction of cell movements. Thus, by applying this novel computational analysis, we were able to quantify the movements of populations of cells, allowing us to detect subtle changes in cell behaviors. Because cranial neural crest cells contribute to the formation of the frontal mass these subtle differences may underlie commonly observed facial asymmetries in normal human populations.

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“…β-Catenin also has important roles in later neural crest development, through its interactions with cadherin to promote neural crest migration (de Melker et al 2004; De Calisto et al 2005) and though its transcriptional stimulation of melanocyte and sensory neuron specification (Dorsky et al 1998; Lee et al 2004). Ethanol’s dysregulation of β-catenin may not be limited to the premigratory neural crest period and could potentially explain the impaired neural crest migration (Rovasio and Battiato, 1995, 2002) and sensory deficits associated with prenatal alcohol exposure. It will be important to determine if the effects upon β-catenin reported here are stage and cell type dependent or instead reflect a broader impact of ethanol upon β-catenin activity.…”

Section: Discussionmentioning

confidence: 99%

Calcium-mediated repression of β-catenin and its transcriptional signaling mediates neural crest cell death in an avian model of fetal alcohol syndrome

Flentke

Garic

Amberger

et al. 2011

Birth Defects Research Part A: Clinical and Molecular Teratolog

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Fetal Alcohol Syndrome (FAS) is a common birth defect in many societies. Affected individuals have neurodevelopmental disabilities and a distinctive craniofacial dysmorphology. These latter deficits originate during early development from the ethanol-mediated apoptotic depletion of cranial facial progenitors, a population known as the neural crest. We showed previously that this apoptosis is caused because acute ethanol exposure activates a G protein-dependent intracellular calcium within cranial neural crest progenitors, and this calcium transient initiates the cell death. The dysregulated signals that reside downstream of ethanol’s calcium transient and effect neural crest death are unknown. Here we show that ethanol’s repression of the transcriptional effector β-catenin causes the neural crest losses. Clinically-relevant ethanol concentrations (22–78 mM) rapidly deplete nuclear β-catenin from neural crest progenitors, with accompanying losses of β-catenin transcriptional activity and downstream genes that govern neural crest induction, expansion and survival. Using forced expression studies we show that β-catenin loss of function (via dominant-negative TCF) recapitulates ethanol’s effects on neural crest apoptosis, whereas β-catenin gain-of-function in ethanol’s presence preserves neural crest survival. Blockade of ethanol’s calcium transient using Bapta-AM normalizes β-catenin activity and prevents the neural crest losses, whereas ionomycin treatment is sufficient to destabilize β-catenin. We propose that ethanol’s repression of β-catenin causes the neural crest losses in this model of FAS. β-Catenin is a novel target for ethanol’s teratogenicity. β-Catenin/Wnt signals participate in many developmental events and its rapid and persistent dysregulation by ethanol may explain why the latter is such a potent teratogen.

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Ethanol Induces Morphological and Dynamic Changes on In Vivo and In Vitro Neural Crest Cells

Cited by 17 publications

References 41 publications

Large-Scale Analysis of Acute Ethanol Exposure in Zebrafish Development: A Critical Time Window and Resilience

Large-Scale Analysis of Acute Ethanol Exposure in Zebrafish Development: A Critical Time Window and Resilience

Quantitative Analysis of Cell Migration Using Optical Flow

Calcium-mediated repression of β-catenin and its transcriptional signaling mediates neural crest cell death in an avian model of fetal alcohol syndrome

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