Synergistic and independent roles for Nodal and FGF in zebrafish CPC migration and asymmetric heart morphogenesis

‘These signals, they’re made for jogging’: Nodal and FGF signals regulate early zebrafish cardiac morphogenesis, lead to asymmetry in migration patterns, contribute to cardiac jogging and cardiac looping.

Figure 1 ‘Synergistic and independent roles for Nodal and FGF in zebrafish CPC migration and asymmetric heart morphogenesis’ Study Summary

Background: The zebrafish (Danio rerio), member of the teleost class, e.g., fish with a bony skeleton (as opposed to chondrichthyes, e.g., sharks and rays which possess a cartilaginous skeleton) [1] [2] is an important model organism for studying developmental mechanisms, including development of the heart. As opposed to other mammals, zebrafish development is rapid and external with many of the embryonic and extraembryonic membranes exhibiting transparent morphology, facilitating study and observation. The adult zebrafish heart consists of the sinus venosus (SV), the posterior-inferior placed atrium (A), the anteriorly placed ventricle (V) and the superiorly placed bulbus arteriosus (BA) [3]. Evolutionary conservation in developmental mechanisms between zebrafish and higher mammals, including humans, allows researchers to use zebrafish for the study of developmental processes that can be observed in humans as well.

Zebrafish development is usually annotated as hours post-fertilization (hpf) or days post-fertilization (dpf). It comprises several embryonic stages occurring after external egg fertilisation; completion of embryonic development and subsequent hatching from the egg enclosure occurs around 72 hpf. However, this timeline may change dependent on the thickness of the surrounding membranes. At this stage, the embryo now becomes a larva, and growth continues with the help of yolk-derived nutrients, for about 3 days. At this point, the animal can be considered a juvenile, a stage that lasts for up to the 90 dpf timepoint and at which point, animals are considered adults [4]. Heart development commences around the period of gastrulation, a period characterised by emergence of the embryonic germ layers, ectoderm derived from the epiblast and mesoderm along with endoderm, derived from mesendoderm, all layers originating from the initial blastoderm [5].

In zebrafish, the heart undergoes several distinct morphological stages; in addition, it exhibits left-right asymmetry, driven by molecular networks which are evolutionarily conserved between different organisms [6].

Table 1 Characteristic morphogenetic stages in zebrafish heart development. CPC, Cardiac progenitor cell; LPM, Lateral plate mesoderm; SHF, Second heart field; hpf, hours post fertilization; FGF, Fibroblast growth factor.

Epithelial Sheets – When: Gastrulation	CPC present bilaterally in the LPM of the developing zebrafish embryo
Cardiac Cone/Heart tube – When: Segmentation (21 to 25-Somite Stage)	CPC sheets converge medially, towards the embryonic midline and form a disk (19.5 hpf) which then forms a cardiac cone (21.5 hpf) that eventually extends into a tube (28 hpf) [7] Cardiac jogging: CPC migration follows a leftward and anterior motion, with progenitors on the left migrating faster than progenitors on the right – FGF, Nodal signals cooperate to regulate heart tube length after jogging (24 hpf)
Heart tube elongation – When: Pharyngula (24-30 hpf)	SHF populations are added to either pole of the heart tube (24-48 hpf), contributing to elongation and outflow tract formation
Cardiac looping – When: Pharyngula (30-48 hpf)	Heart tube loops towards the right, chamber formation and alignment start to occur; at the end of cardiac looping atrium and ventricle are positioned next to each other

The authors of the study use zebrafish embryos to characterise the cooperation between Nodal and FGF signalling during early heart development.

Key results of the study:

1. Nodal and FGF signals act synergistically during heart development; Nodal upregulates FGF signaling components, although FGF does not directly participate in the establishment of left–right (L-R) heart asymmetry (any FGF signaling effect must occur downstream of L-R axis development) or laterality during cardiac jogging. However, both Nodal and FGF signaling are implicated in CPC migration during this process.
2. Nodal and FGF signals regulate the direction and velocity of migrating CPC during cardiac jogging; effects from these signals are additive. Regarding trajectory direction, both Nodal and FGF contribute to anterior bound migration. However, only asymmetric Nodal signals contribute to lateral migration and in particular, left bound migration. On the other hand, FGF acts as a permissive factor in cell migration and further cooperates with Nodal signaling to regulate overall CPC migration.
3. While loss of either Nodal, FGF reduces velocity of migration during cardiac jogging, loss of both severely disrupts CPC migration and even formation of the heart tube itself.
4. Nodal signals affect the dynamic polymerization and depolymerization events that underpin CPC migration; the asymmetry in Nodal signaling results in asymmetry in F-actin formation and as a result, asymmetry during atrial CPC migration. In particular, Nodal signaling induces an increase in F-actin in migrating left atrial CPC only; as a result, these cells migrate faster than their right-sided counterparts.
5. Nodal, FGF signals both promote cardiac looping; FGF also promotes addition of SHF populations to the arterial pole of the zebrafish heart tube with absence of FGF leading to shorter heart tubes [8].

Why this work is interesting:

This study highlights the differential effects on cellular migration and velocities stemming from differential Nodal activity, which in turn cooperates with FGF regulates cardiac jogging. Cardiac jogging is the first instance of L-R asymmetry during heart development. Later down the line, Nodal and FGF also cooperate to regulate another event of L-R asymmetry, heart looping. It is collective cell migrations such as these, which differ across the L-R axes, that contribute to the complex heart forms of vertebrate organisms.

References:

[1] Witten PE, Harris MP, Huysseune A, Winkler C. Chapter 13 – Small teleost fish provide new insights into human skeletal diseases. In: Detrich HW, Westerfield M, Zon LI, editors. Methods in Cell Biology, vol. 138, Academic Press; 2017, p. 321–46. https://doi.org/10.1016/bs.mcb.2016.09.001.

[2] Stedman NL, Garner MM. Chapter 40 – Chondrichthyes. In: Terio KA, McAloose D, Leger JSt, editors. Pathology of Wildlife and Zoo Animals, Academic Press; 2018, p. 1003–18. https://doi.org/10.1016/B978-0-12-805306-5.00040-7.

[3] Sedmera D, Reckova M, deAlmeida A, Sedmerova M, Biermann M, Volejnik J, et al. Functional and morphological evidence for a ventricular conduction system in zebrafish and Xenopus hearts. American Journal of Physiology-Heart and Circulatory Physiology 2003;284:H1152–60. https://doi.org/10.1152/ajpheart.00870.2002.

[4] Singleman C, Holtzman NG. Growth and Maturation in the Zebrafish, Danio Rerio: A Staging Tool for Teaching and Research. Zebrafish 2014;11:396–406. https://doi.org/10.1089/zeb.2014.0976.

[5] Shah G, Thierbach K, Schmid B, Waschke J, Reade A, Hlawitschka M, et al. Multi-scale imaging and analysis identify pan-embryo cell dynamics of germlayer formation in zebrafish. Nat Commun 2019;10:5753. https://doi.org/10.1038/s41467-019-13625-0.

[6] Desgrange A, Le Garrec J-F, Meilhac SM. Left-right asymmetry in heart development and disease: forming the right loop. Development 2018;145:dev162776. https://doi.org/10.1242/dev.162776.

[7] Kidokoro H, Saijoh Y, Schoenwolf GC. Nodal signaling regulates asymmetric cellular behaviors, driving clockwise rotation of the heart tube in zebrafish. Commun Biol 2022;5:996. https://doi.org/10.1038/s42003-022-03826-7.

[8] Gonzalez V, Grant MG, Suzuki M, Christophers B, Williams JR, Burdine RD. Synergistic and independent roles for Nodal and FGF in zebrafish CPC migration and asymmetric heart morphogenesis 2025:2024.01.05.574380. https://doi.org/10.1101/2024.01.05.574380.

[9] Wolton M, Davey MG, Dietrich S. At early stages of heart development, the first and second heart fields are a continuum of lateral head mesoderm-derived, cardiogenic cells. Developmental Biology 2025;520:200–23. https://doi.org/10.1016/j.ydbio.2025.01.009.

 

Tags: biology, cardiovascular, development, heart

doi: https://doi.org/10.1242/prelights.41108

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