Selective Vulnerability of Embryonic Cell Populations to Ethanol‐Induced Apoptosis: Implications for Alcohol‐Related Birth Defects and Neurodevelopmental Disorder

Dunty, William C.; Chen, Shao Yu; Zucker, Robert M.; Dehart, Deborah B.; Sulik, Kathleen K.

doi:10.1111/j.1530-0277.2001.tb02156.x

Cited by 205 publications

(199 citation statements)

References 90 publications

(56 reference statements)

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“…We have previously demonstrated that apoptosis (cell death) can be visualized in 8-9 days old mouse embryos, rat fetal limbs, and other tissues using a fluorescent dye (LysoTracker Red, LT), which stains lysosomes and acidic regions of cells/tissues (16)(17)(18)(19)(20)(21)(22)(23). The current study extends our previous work to visualize apoptosis within follicles of a whole ovary (17).…”

supporting

confidence: 66%

“…From previous studies on embryos, the appearance of LT in tissue is strongly suggestive of apoptosis (16)(17)(18)(19)(20)(21)(22)(23). The LT has a fluorescence peak at 610 nm (Leica SP1), while the AIF has a fluorescence peak around 471 nm with UV or cyan GFP excitation.…”

Section: Discussionmentioning

confidence: 95%

“…In the case of the ovary, the LT was excited with a 543 nm laser to indicate the regions of specific apoptosis that existed in the tissue (16)(17)(18)(19)(20)(21)(22)(23) while the background fluorescence consisting primarily of aldehyde autofluorescence that was optimally excited using the 488 nm laser. The combination of two lasers achieved the specificity of the LT probe for apoptotic cells and the morphology of the tissue was achieved by the Glut probe.…”

Section: Discussionmentioning

confidence: 99%

“…This change in fixation chemicals increased the tissue fluorescence that was very beneficial for good morphological visualization. The modifications of this fixation preparation procedure on ovaries have also been applicable to the study of embryos (21).…”

mentioning

confidence: 99%

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Confocal laser scanning microscopy of whole mouse ovaries: Excellent morphology, apoptosis detection, and spectroscopy

Zucker

Jeffay

2006

Cytometry Pt A

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Background:Ovaries consist of numerous follicles, oocytes, and granulosa cells in different stages of development. Many of these follicles will undergo an apoptotic process during the lifetime of the animal. By using proper tissue preparation methods, the events within the whole ovary can be observed by using 3D confocal microscopy.Methods:Whole ovaries were stained with LysoTracker Red (LT), fixed with 4% paraformaldehyde (PF) and 1% glutaraldehyde (Glut), dehydrated with methanol (MEOH), and cleared with benzyl alcohol and benzyl benzoate (BABB). Using this tissue preparation technique, the ovary becomes relatively transparent, allowing its morphology to be observed with confocal microscopes. A spectral imaging system (PARISS) located on a conventional microscope was used to interpret the LT dye spectra and fixation products in the tissues with different excitation wavelengths.Results:Apoptosis in the follicle was detected as clusters of intensely stained granulosa cells located in close proximity to the oocytes. The fixation with Glut and PF preserved morphological details, increased tissue fluorescence, thus increased the signal to noise of the background image.Conclusions:Thick tissues can be imaged after they are properly stained, aldehyde fixed, and BABB cleared. LT intensely stained single cells or clusters of apoptotic cells in the follicles and the nucleolus. Spectral differences between LT as an indicator of apoptosis and Glut‐PF fixation was used to visualize ovarian morphology and apoptosis. The PARISS spectrophotometer revealed spectral peaks for LT at 609.6 nm and for Glut‐PF at 471.3 nm. The proper use of the spectra from these fluorescence molecules is the foundation for high quality morphological images of apoptosis. By sequentially imaging the two probes with a 488 nm laser and a 543/568 nm laser, there was a reduction in fluorescent cross talk and an increase in image quality. © 2006 International Society for Analytical Cytology

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supporting

confidence: 66%

Section: Discussionmentioning

confidence: 95%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Confocal laser scanning microscopy of whole mouse ovaries: Excellent morphology, apoptosis detection, and spectroscopy

Zucker

Jeffay

2006

Cytometry Pt A

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“…In contrast, ethanol can cause death of post-mitotic neurons (Miller, 1995;Jacobs and Miller, 2001;Mooney and Miller, 2001a;Heaton et al, 2003;Mooney et al, 2006). Embryos exposed to ethanol during gastrulation display increased expression of markers associated with cell death (Dunty et al, 2001;2002). Such labeling is most apparent in the cranial neural crest, which may underlie the craniofacial changes, and the rostral hindbrain, i.e., rhombomeres 1-3, inclusive.…”

Section: Mechanisms Of Ethanol Toxicitymentioning

confidence: 99%

Time-specific effects of ethanol exposure on cranial nerve nuclei: Gastrulation and neuronogenesis

Mooney

Miller

2007

Experimental Neurology

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During the development of the central nervous system, neurons pass through critical periods or periods of vulnerability. We explored periods of vulnerability for cranial nerve nuclei by determining the effects of acute exposure to ethanol during development on the number of neurons in mature brainstem. Long-Evans rats were injected with 2.9 g ethanol/kg body weight on one day between gestational day (G) 7 and G13, inclusive. Two hours later, animals received a second injection of 1.45 g/kg. Controls were injected with equivalent volumes of saline. Brainstems of 31-day-old offspring were cryosectioned and stained with cresyl violet. Stereological methods were used to determine the volume and numerical density of neurons in three trigeminal sensory nuclei (the principal sensory nucleus of the trigeminal nerve, and the oral and interpolar subnuclei of the spinal trigeminal nuclear complex) and three motor nuclei (the trigeminal, facial, and hypoglossal nuclei). The numbers of neurons in most nuclei were lower following early (on G7 and/or G8) or later (on G12 and/or G13) exposure. Only the trigeminal interpolar nucleus was affected by neither early nor late ethanol exposure. Thus, prenatal exposure to ethanol affects the number of neurons in brainstem nuclei in a time-dependent manner. Windows of vulnerability coincide with gastrulation (G7/G8) and the period of neuronal generation (G12/G13).

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Fetal Alcohol Syndrome and Fetal Alcohol Spectrum Disorder

Chudley

Longstaffe

2005

Management of Genetic Syndromes

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Fetal alcohol spectrum disorder is an “umbrella” term that describes the spectrum of ethanol teratogenesis in humans. At one end of the spectrum are the subset of individuals with fetal alcohol syndrome, and at the other end of the spectrum are those individuals with behavioral and cognitive deficits who exhibit minimal or no physical stigmata as a consequence of ethanol‐induced prenatal brain injury. It was not until the past 3 decades that the medical community was convinced of the devastating effects of excessive alcohol use in pregnancy on the children. Although the diagnosis of fetal alcohol syndrome and its spectrum is a medical diagnosis, its effects on society are far‐reaching. Fetal alcohol spectrum disorder is distinct from many genetic syndromes described in this book in that the disorder is potentially entirely preventable, and the diagnosis points toward two affected individuals: the drinking mother and the alcohol‐exposed child. Diagnosis, intervention, and prevention offer some of the greatest challenges to health care providers, families, and their communities. Multidisciplinary diagnostic teams are developing throughout North America and other parts of the world that provide accurate and comprehensive assessments and treatment options for affected individuals with this common disorder.

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Selective Vulnerability of Embryonic Cell Populations to Ethanol‐Induced Apoptosis: Implications for Alcohol‐Related Birth Defects and Neurodevelopmental Disorder

Abstract: The results of this study revealed developmental stage-specific cell populations of the developing brain and craniofacial region that are vulnerable to ethanol-induced apoptosis and provide new insight relative to the genesis of alcohol-related birth defects.

Cited by 205 publications

References 90 publications

Confocal laser scanning microscopy of whole mouse ovaries: Excellent morphology, apoptosis detection, and spectroscopy

Confocal laser scanning microscopy of whole mouse ovaries: Excellent morphology, apoptosis detection, and spectroscopy

Time-specific effects of ethanol exposure on cranial nerve nuclei: Gastrulation and neuronogenesis

Fetal Alcohol Syndrome and Fetal Alcohol Spectrum Disorder

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