Transgenic zebrafish illuminate the dynamics of thyroid morphogenesis and its relationship to cardiovascular development

Abstract

Among the various organs derived from foregut endoderm, the thyroid gland is unique in that major morphogenic events such as budding from foregut endoderm, descent into subpharyngeal mesenchyme and growth expansion occur in close proximity to cardiovascular tissues. To date, research on thyroid organogenesis was missing one vital tool-a transgenic model that allows to track the dynamic changes in thyroid size, shape and location relative to adjacent cardiovascular tissues in live embryos. In this study, we generated a novel transgenic zebrafish line, tg(tg:mCherry), in which robust and thyroid-specific expression of a membrane version of mCherry enables live imaging of thyroid development in embryos from budding stage throughout formation of functional thyroid follicles. By using various double transgenic models in which EGFP expression additionally labels cardiovascular structures, a high coordination was revealed between thyroid organogenesis and cardiovascular development. Early thyroid development was found to proceed in intimate contact with the distal ventricular myocardium and live imaging confirmed that thyroid budding from the pharyngeal floor is tightly coordinated with the descent of the heart. Four-dimensional imaging of live embryos by selective plane illumination microscopy and 3D-reconstruction of confocal images of stained embryos yielded novel insights into the role of specific pharyngeal vessels, such as the hypobranchial artery (HA), in guiding late thyroid expansion along the pharyngeal midline. An important role of the HA was corroborated by the detailed examination of thyroid development in various zebrafish models showing defective cardiovascular development. In combination, our results from live imaging as well es from 3D-reconstruction of thyroid development in tg(tg:mCherry) embryos provided a first dynamic view of late thyroid organogenesis in zebrafish-a critical resource for the design of future studies addressing the molecular mechanisms of these thyroid-vasculature interactions.

Benzotriazole ultraviolet stabilizers alter the expression of the thyroid hormone pathway in zebrafish (Danio rerio) embryos

“…A member of this family, nkx2.5, is essential for myogenic and morphogenetic differentiation (Chen and Fishman, 1996). Although there is no evidence that nkx 2.1 directly regulates cardiac function, previous studies indicate that it may be involved in cardiovascular formation (Porazzi et al, 2009;Opitz et al, 2012). Early thyroid development has been found to proceed in intimate contact with cardiovascular development; thyroid budding from the pharyngeal floor is tightly coordinated with the descent of the heart (Opitz et al, 2012).…”

“…Although there is no evidence that nkx 2.1 directly regulates cardiac function, previous studies indicate that it may be involved in cardiovascular formation (Porazzi et al, 2009;Opitz et al, 2012). Early thyroid development has been found to proceed in intimate contact with cardiovascular development; thyroid budding from the pharyngeal floor is tightly coordinated with the descent of the heart (Opitz et al, 2012). In the zebrafish, nkx2.1 is expressed close to the heart primordium, and cells expressing nkx2.1 comprise the region corresponding to the thyroid bud (Porazzi et al, 2009).…”

Light-sheet microscopy: a tutorial

“…These combined effects offer a technique that allows the user to minimize both photobleaching and phototoxicity [10,[12][13][14], while allowing for long-term imaging studies (up to several days). This is an essential characteristic required for all sorts of developmental biology studies [15][16][17][18], such as organogenesis [19], cell migration [20,21], cardiac development [22][23][24][25], vascular development [26,27], neuro-development [28,29], and generally any kind of in vivo studies. Drosophila melanogaster [2,[30][31][32] and zebrafish [19,26,30,[33][34][35][36] have traditionally been the most widely used samples in this field, but it fits well many others, such as worm embryos [28,37] and other small organisms or plants [38][39][40].…”

“…Therefore, despite the fact that 2D cells placed in petri dishes have been widely used to study biological process, LSFM allows the use of agarose or fluorinated ethylene propylene (FEP) tubes [17,42,43] for mounting 3D living samples. Thus, 2D models have been constantly replaced by 3D cell cultures [44][45][46][47], spheroids [48][49][50][51][52], organs and organoids [26,[53][54][55], and model organisms such as Drosophila melanogaster, zebrafish and C. elegans [2,30,33,56].…”

Imaging and 3D Reconstruction of Cerebrovascular Structures in Embryonic Zebrafish

“…Modifications to this system can incorporate transgenic lines of zebrafish that express EGFP or other fluorescent proteins in a variety of tissue, organ or region specific patterns 10 . Further, analysis of neovascular changes in the zebrafish retina has recently been published 5 .…”