Measurement of absolute blood flow velocity in outflow tract of HH18 chicken embryo based on 4D reconstruction using spectral domain optical coherence tomography

The measurement of blood-plasma absolute velocity distributions with high spatial and temporal resolution in vivo is important for the investigation of embryonic heart at its early stage of development. We introduce a novel method to measure absolute blood flow velocity based on high speed spectral domain optical coherence tomography (OCT) and apply it to measure velocities across the heart outflow tract (OFT) of a chicken embryo (stage HH18). First, we use the OCT system to acquire 4D 
[(x,y,z) + t] images of the OFT in vivo. Second, we reconstruct the 4D microstructural images and obtain the orientation of the OFT at its maximum expansion, from which the centerline of the OFT is calculated based on the OFT boundary segmentation. Assuming flow is parallel to the vessel orientation, the obtained centerline indicates the flow direction. Finally, the absolute flow velocity is evaluated based on the direction given by the centerline and the axial velocity obtained from Doppler OCT. Using this method, we compare flow velocity profiles at various positions along the chicken embryo OFT

Imaging Approaches and the Quantitative Analysis of Heart Development.

Heart morphogenesis is a complex and dynamic process that has captivated researchers for almost a century. This process involves three main stages, during which the heart undergoes growth and folding on itself to form its common chambered shape. However, imaging heart development presents significant challenges due to the rapid and dynamic changes in heart morphology. Researchers have used different model organisms and developed various imaging techniques to obtain high-resolution images of heart development. Advanced imaging techniques have allowed the integration of multiscale live imaging approaches with genetic labeling, enabling the quantitative analysis of cardiac morphogenesis. Here, we discuss the various imaging techniques used to obtain high-resolution images of whole-heart development. We also review the mathematical approaches used to quantify cardiac morphogenesis from 3D and 3D+time images and to model its dynamics at the tissue and cellular levels.Grant support PGC2018-096486-B-I00 from the Spanish Ministerio de Ciencia e Innovación and Grant H2020-MSCA-ITN-2016-722427 from the EU Horizon 2020 program to M.T. M.S. was supported by a La Caixa Foudation PhD fellowship (LCF/BQ/DE18/11670014) and The Company of Biologists travelling fellowship (DEVTF181145). The CNIC is supported by the Spanish Ministery of Science and the ProCNIC Foundation.S

Quantifying function in the zebrafish embryonic heart: a study on the role of timed mechanical cues

2014 Summer.Congenital heart defects are a relatively common problem, yet the cause is unknown in the large majority of cases. A significant amount of past research has shown that there is a link between altered blood-induced mechanical stress and heart development. However, very little research has been done to examine the effect of altered loading timing. During embryonic development, the heart undergoes a drastic change in morphology from its original valveless tube structure to a complete multi-chambered pump with valves. Blood flow dynamics are consequently altered significantly as well. Given the changes occurring through this period, it is hypothesized that significant and persistent decreases in heart function occur when cardiac loading is altered during certain time windows of early development. The main objectives of this work were to (1) develop a methodology to quantify heart function in the embryonic zebrafish from high-speed bright field images, (2) develop a model for temporary and noninvasive alteration of cardiac loading, and (3) apply the methodology to normal and treated embryos to determine whether certain time windows of altered loading are more impactful than others. Results indicated that altered loading during the tube and early looping stages of development produce persistent changes in heart morphology along with accompanying decreases in cardiac function. Altered loading during late cardiac looping resulted in temporary changes in function which did not persist through the latest time point measured. This work has produced extensive tools for quantifying heart function from high speed images and presents a new model for altered cardiac loading in the zebrafish. Results support the hypothesis that the heart is more sensitive to altered loading during certain windows in development. This provides new insight into how congenital defects may develop and sets the stage for future experiments investigating the effects of altered loading on heart development

Novel Video Imaging to Examine Cardiac Function in Xenopus Embryos

Congenital heart defects (CHDs) occur in approximately 1% of live births and the etiology has been associated with disturbances in cardiogenesis. However, the majority of research examining CHDs relies on static morphological data, which does not elucidate how defects alter cardiac function. I used Xenopus laevis embryos to examine the association between CHDs and functional alterations using a novel imaging system that can obtain high- resolution images through a non-invasive procedure. A high-speed video camera and software were used to assess cardiac function, permitting functional characterization of late Xenopus cardiogenesis. Verification of the imaging system’s ability to detect changes in function was confirmed by exposing embryos to well-established agents that alter heart rate. Additionally, significant functional changes were detected following exposure of embryos to small molecules known to disrupt morphogenesis. The imaging system will be a useful alternative to current imaging modalities for elucidating mechanisms underlying CHDs in optically transparent embryos

Quantitative analysis of the mechanical environment in the embryonic heart with respect to its relationship in cardiac development

Includes bibliographical references.2015 Fall.In order to understand the causes of congenital heart defects, which afflict at least 4 infants per 1,000 live births, research has implemented the use of animal models to study embryonic heart development. Zebrafish (Danio rerio) have become one of the more prominent of these animal models due to the fact that their heart morphology at the earliest stages of development is remarkably similar to humans, and because embryos lack pigmentation, rendering them transparent. This transparency allows for high-speed images of blood flow to be acquired in the developing heart so that the mechanotransductive relationship between the intracardiac flow environment and myocardial progenitor cell differentiation can be understood. One particular aspect of the flow environment, a cyclic retrograde flow at the junction of the forming atrium and ventricle, has been shown to be necessary for valve formation, though the mechanisms causing it to occur had previously been unknown. By comparing the results of two-dimensional spatiotemporal analysis applied to embryos both with normal retrograde flow and inhibited retrograde flow, this study shows that a particular range of pressures associated with the pumping mechanics of the heart as well as resistance due to systolic contractile closure must exist in order to maintain adequate retrograde flow to induce valve formation. The use of two-dimensional spatiotemporal analysis was sufficient to acquire these results, however when applied to analysis of other aspects of the intracardiac flow environment, this computational method is subject to critical limitations. Therefore, this study includes the development of methodology to integrate the results of spatiotemporal analysis on multiple focal planes bisecting the heart into a more accurate, three-dimensional result. The results of this study not only increase our understanding of the mechanics behind an important factor in embryonic development, but also enable future experiments pertaining to the measurement of embryonic intracardiac blood flow to be performed with increased certainty

Development of High-speed Optical Coherence Tomography for Time-lapse Non-destructive Characterization of Samples

Optical coherence tomography (OCT) is an established optical imaging modality which can obtain label-free, non-destructive 3D images of samples with micron-scale resolution and millimeter penetration. OCT has been widely adopted for biomedical researches

Arrhythmia Caused by a Drosophila Tropomyosin Mutation Is Revealed Using a Novel Optical Coherence Tomography Instrument

Background: Dilated cardiomyopathy (DCM) is a severe cardiac condition that causes high mortality. Many genes have been confirmed to be involved in this disease. An ideal system with which to uncover disease mechanisms would be one that can measure the changes in a wide range of cardiac activities associated with mutations in specific, diversely functional cardiac genes. Such a system needs a genetically manipulable model organism that allows in vivo measurement of cardiac phenotypes and a detecting instrument capable of recording multiple phenotype parameters. Methodology and Principal Findings: With a simple heart, a transparent body surface at larval stages and available genetic tools we chose Drosophila melanogaster as our model organism and developed for it a dual en-face/Doppler optical coherence tomography (OCT) instrument capable of recording multiple aspects of heart activity, including heart contraction cycle dynamics, ostia dynamics, heartbeat rate and rhythm, speed of heart wall movement and light reflectivity of cardiomyocytes in situ. We applied this OCT instrument to a model of Tropomyosin-associated DCM established in adult Drosophila. We show that DCM pre-exists in the larval stage and is accompanied by an arrhythmia previously unidentified in this model. We also detect reduced mobility and light reflectivity of cardiomyocytes in mutants. Conclusion: These results demonstrate the capability of our OCT instrument to characterize in detail cardiac activity i