RESUMO
Lightsheet microscopy is a form of fluorescence microscopy that can be used to visualize specimen with high resolution, a large depth-of-field, and minimal photodamage and photobleaching as compared to traditional confocal microscopy. As this technology becomes much more readily available, it will be useful in revealing new findings in the cardiovascular development field that may be hidden or difficult to image. In this manuscript, we describe an approach for mounting and culturing postimplantation mouse embryos to visualize blood vessel development with a lightsheet microscope.
Assuntos
Angiografia/métodos , Vasos Sanguíneos/diagnóstico por imagem , Técnicas de Cultura/métodos , Embrião de Mamíferos/diagnóstico por imagem , Desenvolvimento Embrionário , Microscopia de Fluorescência/métodos , Neovascularização Fisiológica , Animais , Vasos Sanguíneos/crescimento & desenvolvimento , Vasos Sanguíneos/metabolismo , Meios de Cultura/química , Dissecação/métodos , Embrião de Mamíferos/irrigação sanguínea , Camundongos , Camundongos Transgênicos , Microscopia ConfocalRESUMO
Development of the embryonic heart involves an intricate network of biochemical and genetic cues to ensure its proper growth and morphogenesis. However, studies from avian and teleost models reveal that biomechanical force, namely hemodynamic loading (blood pressure and shear stress), plays a significant role in regulating heart development. To study how hemodynamic loading impacts development of the mammalian embryonic heart, we utilized mouse embryo culture and manipulation techniques and performed optical projection tomography imaging followed by morphometric analysis to determine how reduced-loading affects heart volume, myocardial thickness, trabeculation and looping. Our results reveal that hemodynamic loading can regulate these features at different thresholds. Intermediate levels of hemodynamic loading are sufficient to promote proper myocardial growth and heart size, but insufficient to promote looping and trabeculation. Whereas, low levels of hemodynamic loading fails to promote proper growth of the myocardium and heart size. These results reveal that the regulation of heart development by biomechanical force is conserved across many vertebrate classes, and this study begins to elucidate how these specific forces regulate development of the mammalian heart.
