Rhythmic forces shaping the zebrafish cardiac system.

The structural development of the heart depends heavily on mechanical forces, and rhythmic contractions generate essential physical stimuli during morphogenesis. Cardiac cells play a critical role in coordinating this process by sensing and responding to these mechanical forces. In vivo, cells experience rhythmic spatial and temporal variations in deformation-related stresses throughout heart development.

Calcium Signal Analysis in the Zebrafish Heart via Phase Matching of the Cardiac Cycle.

Calcium signalling in the endocardium is critical for heart valve development. Calcium ion pulses in the endocardium are generated in response to mechanical forces due to blood flow and can be visualised in the beating zebrafish heart using a genetically encoded calcium indicator such as GCaMP7a. Analysing these pulses is challenging because of the rapid movement of the heart during heartbeat.

Role of tissue biomechanics in the formation and function of myocardial trabeculae in zebrafish embryos.

Cardiac trabeculae are uneven ventricular muscular structures that develop during early embryonic heart development at the outer curvature of the ventricle. Their biomechanical function is not completely understood, and while their formation is known to be mechanosensitive, it is unclear whether ventricular tissue internal stresses play an important role in their formation. Here, we performed imaging and image-based cardiac biomechanics simulations on zebrafish embryonic ventricles to investigate these issues.

Highly specific and non-invasive imaging of Piezo1-dependent activity across scales using GenEPi.

Mechanosensing is a ubiquitous process to translate external mechanical stimuli into biological responses. Piezo1 ion channels are directly gated by mechanical forces and play an essential role in cellular mechanotransduction. However, readouts of Piezo1 activity are mainly examined by invasive or indirect techniques, such as electrophysiological analyses and cytosolic calcium imaging.

TissUExM protocol for ultrastructure expansion microscopy of zebrafish larvae and mouse embryos.

Expansion microscopy of millimeter-large mechanically heterogeneous tissues, such as whole vertebrate embryos, has been limited, particularly when combined with post-expansion immunofluorescence. Here, we present a protocol to perform ultrastructure expansion microscopy of whole vertebrate embryos, optimized to perform post-expansion labeling. We describe steps for embedding and denaturing zebrafish larvae or mouse embryos.

TissUExM enables quantitative ultrastructural analysis in whole vertebrate embryos by expansion microscopy.

Super-resolution microscopy reveals the molecular organization of biological structures down to the nanoscale. While it allows the study of protein complexes in single cells, small organisms, or thin tissue sections, there is currently no versatile approach for ultrastructural analysis compatible with whole vertebrate embryos. Here, we present tissue ultrastructure expansion microscopy (TissUExM), a method to expand millimeter-scale and mechanically heterogeneous whole embryonic tissues, including wing discs, whole zebrafish, and mouse embryos.

Fluid mechanics of the zebrafish embryonic heart trabeculation.

Embryonic heart development is a mechanosensitive process, where specific fluid forces are needed for the correct development, and abnormal mechanical stimuli can lead to malformations. It is thus important to understand the nature of embryonic heart fluid forces. However, the fluid dynamical behaviour close to the embryonic endocardial surface is very sensitive to the geometry and motion dynamics of fine-scale cardiac trabecular surface structures.

Mechanical control of tissue shape: Cell-extrinsic and -intrinsic mechanisms join forces to regulate morphogenesis.

During vertebrate development, cells must proliferate, move, and differentiate to form complex shapes. Elucidating the mechanisms underlying the molecular and cellular processes involved in tissue morphogenesis is essential to understanding developmental programmes. Mechanical stimuli act as a major contributor of morphogenetic processes and impact on cell behaviours to regulate tissue shape and size.

Extracellular mechanical forces drive endocardial cell volume decrease during zebrafish cardiac valve morphogenesis.

Organ morphogenesis involves dynamic changes of tissue properties while cells adapt to their mechanical environment through mechanosensitive pathways. How mechanical cues influence cell behaviors during morphogenesis remains unclear. Here, we studied the formation of the zebrafish atrioventricular canal (AVC) where cardiac valves develop.

Cardiac forces regulate zebrafish heart valve delamination by modulating Nfat signaling.

In the clinic, most cases of congenital heart valve defects are thought to arise through errors that occur after the endothelial-mesenchymal transition (EndoMT) stage of valve development. Although mechanical forces caused by heartbeat are essential modulators of cardiovascular development, their role in these later developmental events is poorly understood. To address this question, we used the zebrafish superior atrioventricular valve (AV) as a model.

Bioelectric signaling and the control of cardiac cell identity in response to mechanical forces.

Developing cardiovascular systems use mechanical forces to take shape, but how ubiquitous blood flow forces instruct local cardiac cell identity is still unclear. By manipulating mechanical forces in vivo, we show here that shear stress is necessary and sufficient to promote valvulogenesis. We found that valve formation is associated with the activation of an extracellular adenosine triphosphate (ATP)–dependent purinergic receptor pathway, specifically triggering calcium ion (Ca) pulses and nuclear factor of activated T cells 1 (Nfatc1) activation.

Notch and Bmp signaling pathways act coordinately during the formation of the proepicardium.

Background: The epicardium is the outer mesothelial layer of the heart. It encloses the myocardium and plays key roles in heart development and regeneration. It derives from the proepicardium (PE), cell clusters that appear in the dorsal pericardium (DP) close to the atrioventricular canal and the venous pole of the heart, and are released into the pericardial cavity.

Intraflagellar Transport Complex B Proteins Regulate the Hippo Effector Yap1 during Cardiogenesis.

Cilia and the intraflagellar transport (IFT) proteins involved in ciliogenesis are associated with congenital heart diseases (CHDs). However, the molecular links between cilia, IFT proteins, and cardiogenesis are yet to be established. Using a combination of biochemistry, genetics, and live-imaging methods, we show that IFT complex B proteins (Ift88, Ift54, and Ift20) modulate the Hippo pathway effector YAP1 in zebrafish and mouse.

Blood Flow Forces in Shaping the Vascular System: A Focus on Endothelial Cell Behavior.

The endothelium is the cell monolayer that lines the interior of the blood vessels separating the vessel lumen where blood circulates, from the surrounding tissues. During embryonic development, endothelial cells (ECs) must ensure that a tight barrier function is maintained whilst dynamically adapting to the growing vascular tree that is being formed and remodeled. Blood circulation generates mechanical forces, such as shear stress and circumferential stretch that are directly acting on the endothelium.

Blood Flow Limits Endothelial Cell Extrusion in the Zebrafish Dorsal Aorta.

Blood flow modulates endothelial cell (EC) response during angiogenesis. Shear stress is known to control gene expression related to the endothelial-mesenchymal transition and endothelial-hematopoietic transition. However, the impact of blood flow on the cellular processes associated with EC extrusion is less well understood.

How to define and optimize axial resolution in light-sheet microscopy: a simulation-based approach.

"How thick is your light sheet?" is a question that has been asked frequently after talks showing impressive renderings of 3D data acquired by a light-sheet microscope. This question is motivated by the fact that most of the time the thickness of the light-sheet is uniquely associated to the axial resolution of the microscope. However, the link between light-sheet thickness and axial resolution has never been systematically assessed and it is still unclear how both are connected.

Mechanotransduction in cardiovascular morphogenesis and tissue engineering.

Cardiovascular morphogenesis involves cell behavior and cell identity changes that are activated by mechanical forces associated with heart function. Recently, advances in in vivo imaging, methods to alter blood flow, and computational modelling have greatly advanced our understanding of how forces produced by heart contraction and blood flow impact different morphogenetic processes. Meanwhile, traditional genetic approaches have helped to elucidate how endothelial cells respond to forces at the cellular and molecular level.

Mechanically activated piezo channels modulate outflow tract valve development through the Yap1 and Klf2-Notch signaling axis.

Mechanical forces are well known for modulating heart valve developmental programs. Yet, it is still unclear how genetic programs and mechanosensation interact during heart valve development. Here, we assessed the mechanosensitive pathways involved during zebrafish outflow tract (OFT) valve development in vivo.

The cilium as a force sensor-myth versus reality.

Cells need to sense their mechanical environment during the growth of developing tissues and maintenance of adult tissues. The concept of force-sensing mechanisms that act through cell-cell and cell-matrix adhesions is now well established and accepted. Additionally, it is widely believed that force sensing can be mediated through cilia.

Hemodynamic-mediated endocardial signaling controls in vivo myocardial reprogramming.

Lower vertebrate and neonatal mammalian hearts exhibit the remarkable capacity to regenerate through the reprogramming of pre-existing cardiomyocytes. However, how cardiac injury initiates signaling pathways controlling this regenerative reprogramming remains to be defined. Here, we utilize in vivo biophysical and genetic fate mapping zebrafish studies to reveal that altered hemodynamic forces due to cardiac injury activate a sequential endocardial-myocardial signaling cascade to direct cardiomyocyte reprogramming and heart regeneration.

Chiral Cilia Orientation in the Left-Right Organizer.

Chirality is a property of asymmetry between an object and its mirror image. Most biomolecules and many cell types are chiral. In the left-right organizer (LRO), cilia-driven flows transfer such chirality to the body scale.

Three-dimensional microscopy and image analysis methodology for mapping and quantification of nuclear positions in tissues with approximate cylindrical geometry.

Organogenesis involves extensive and dynamic changes of tissue shape during development. It is associated with complex morphogenetic events that require enormous tissue plasticity and generate a large variety of transient three-dimensional geometries that are achieved by global tissue responses. Nevertheless, such global responses are driven by tight spatio-temporal regulation of the behaviours of individual cells composing these tissues.

Hippo signaling determines the number of venous pole cells that originate from the anterior lateral plate mesoderm in zebrafish.

The differentiation of the lateral plate mesoderm cells into heart field cells constitutes a critical step in the development of cardiac tissue and the genesis of functional cardiomyocytes. Hippo signaling controls cardiomyocyte proliferation, but the role of Hippo signaling during early cardiogenesis remains unclear. Here, we show that Hippo signaling regulates atrial cell number by specifying the developmental potential of cells within the anterior lateral plate mesoderm (ALPM), which are incorporated into the venous pole of the heart tube and ultimately into the atrium of the heart.