Cadherin2 (N-cadherin) plays an essential role in zebrafish cardiovascular development
© Bagatto et al; licensee BioMed Central Ltd. 2006
Received: 21 March 2006
Accepted: 23 May 2006
Published: 23 May 2006
Cadherins are cell surface adhesion molecules that play important roles in development of vertebrate tissues and organs. We studied cadherin2 expression in developing zebrafish heart using in situ hybridization and immunocytochemical methods, and we found that cadherin2 was strongly expressed by the myocardium of the embryonic zebrafish. To gain insight into cadherin2 role in the formation and function of the heart, we analyzed cardiac differentiation and performance in a cadherin2 mutant, glass onion (glo).
We found that the cadherin2 mutant had enlarged pericardial cavity, disorganized atrium and ventricle, and reduced expression of a ventricular specific marker vmhc. Individual myocardiocytes in the glo mutant embryos became round shaped and loosely aggregated. In vivo measurements of cardiac performance revealed that the mutant heart had significantly reduced heart rate, stroke volume and cardiac output compared to control embryos. Formation of the embryonic vascular system in the glo mutants was also affected.
Our results suggest that cadherin2 plays an essential role in zebrafish cardiovascular development. Although the exact mechanisms remain unknown as to the formation of the enlarged pericardium and reduced peripheral blood flow, it is clear that myocardiocyte differentiation and physiological cardiovascular performance is impaired when cadherin2 function is disrupted.
Zebrafish (Danio rerio) has emerged as an important model system in the study of vertebrate development due to its external development, transparency of embryos, and its demonstrated utility as a genetic model. The process of heart formation occurs in a similar way in all vertebrates . Heart development in the zebrafish is also rapid, which allows for numerous short-term studies. The primitive heart tube is formed by 22 hours post-fertilization (hpf). The heart is beating and the circulation becomes evident at 24 hpf. By 30 hpf, the heart tube starts to loop to the right side of the embryo. By 36 hpf, chamber boundaries are evident, although molecular markers can distinguish them in the primitive heart tube [2, 3]. Looping places both the atrium and the ventricle toward the left of the embryo, however the atrium is further to the left than the ventricle. By 60 hpf, the valves are present and by the fifth day, the heart has assumed its adult configuration, with the atrium sitting dorsally with respect to the ventricle [1, 4].
The fundamental plan of the vascular system as it develops in the zebrafish is similar to that of other vertebrates. The overall form of the zebrafish vasculature is established early, before the initiation of circulation, and the pattern of major vessel tracts is reproducible from embryo to embryo . Circulation begins at approximately 24–26 hpf and initially flows through a simple single circulatory loop. Cells from the dorsal margin of the lateral plate migrate medially to form the intermediate cell mass . This mass gives rise to both the endothelia and the major trunk vessels and the first cohort of blood cells . However, many early vasculogenic vessels first appear as a network or plexus of smaller vessels, with little apparent pattern or differentiated identity [8, 9]. By 2.5–3 days postfertilization (dpf), the trunk and tail intersegmental vessels are fully formed and by 6 dpf, the overall basic pattern of the vaculature is in place .
Although vertebrate cardiovascular development has been well described morphologically, the molecular and physiological mechanisms underlying these events are only beginning to be understood. Results from gene expression pattern and/or functional studies suggest that a variety of molecules, including transcription factors (e.g. Gata4, Nkx2.5), morphogenetic regulators (e.g. Hand2, Pitx2, Xin), endothelial growth factors (e.g. VEGF-A), cardiac specific proteins (e.g. cmlc1, cmlc2, and vmhc), cell adhesion molecule (e.g. cadherin2 and cadherin5, see below), are involved in the cardiac patterning and morphogenesis of the vertebrate heart [3, 11–20].
The cadherins are a family of Ca++-dependent transmembrane molecules that mediate cell adhesion mainly through homophilic interactions [21–23]. Cadherin2, the first cadherin discovered in the vertebrate nervous system , has been shown to be of critical importance in the early differentiation of the vertebrate central and peripheral nervous structures [18, 25–30]. Unlike the wide expression of cadherin2, expression of cadherin5 (also called VE-cadherin), is confined to the endothelial cells of both developing and adult vasculature [20, 31, 32]. Despite the importance of cardiovascular tissue itself and cadherin molecules in animal development, there are only a few published reports on cadherins function in vertebrate cardiovascular development. Zebrafish with mutations in cadherin2 (parachute, or pac mutant, ; glass onion, or glo mutant, ) have recently been identified, and their phenotypes studied. However, most of the analysis was concentrated on the central nervous system. So far, detailed information on the developmental profile of cadherin2 expression in the vertebrate cardiac tissue has been performed on only the chick and mice. Moreover, functional analysis was limited almost exclusively to descriptions of anatomical defects in the heart of embryos whose cadherin2 function was blocked [18, 33]. We propose to study cardiac differentiation, cardiac performance and formation of the vascular system in the glo mutant in order to elucidate the role(s) of cadherin2 plays in the formation and function of the cardiovascular system.
Cadherin2 expression in developing zebrafish heart
Analysis of cardiac performance in glomutant embryos
Mean stroke volume in control and glo mutant hearts was similar at 30 hpf, and it became larger in both groups as development proceeded (Fig. 2B). In control embryos, stroke volume increased more rapidly during development with the difference showing statistical significance at 72 hpf. This late significance was due, at least partially, to large variations of the stroke volume in both groups. Multiplying stroke volume by heart rate produces cardiac output (Fig. 2C). Mean cardiac output increased in both groups during the 42-hour measurement period with the slope of the increase significantly higher in the control group. As with stroke volume, cardiac output was similar between the two groups at 30 hpf, and the control embryos had higher cardiac output than the glo mutant embryos at the remaining recording time points, although their values were not significantly different until 72 hpf. Again, cardiac output variability was high in both groups.
For the majority of the developmental window investigated, the contraction time (time from diastole to systole) for glo mutant hearts was slower than controls (Fig. 2D). This difference was statistically significant during the first two measurement times (30 and 36 hpf), but by 72 hpf, the contraction time became similar between the glo and control hearts. Moreover, the variation in contraction times was also significantly higher in the glo mutant hearts, reflecting the irregular nature of cardiac contractions in this group.
Gross morphological defects in the heart of the glo mutant and cadherin2morphants
Differentiation of the glomutant heart is affected
Cardiac differentiation of the glo mutants and cadherin2 morphants was analyzed using cardiac specific markers nkx2.5, cmlc2, vmhc , cdh5  or Zn-5 immunostaining. Nkx2.5 is a transcription factor crucial for vertebrate cardiac development , and it is expressed mainly by the ventricle at 48–52 hpf (Fig. 5A). Its mRNA expression in the glo mutant heart appeared to be similar to the control heart (Fig. 5A and 5B). cmlc2 labels the cardiac myosin light chain 2 present in both the embryonic zebrafish atrium and ventricle (; Fig. 5C). Similar to cmlc2 expression in the control heart, cmlc2 was expressed by both the atrium and ventricle of the glo mutant heart (Fig. 5C and 5D). Zebrafish vmhc stands for the ventricular myosin heavy chain gene, and it labels the ventricular myocardium and skeletal muscles of the body (; Fig. 5E). vmhc expression was moderately or greatly reduced in the glo mutant heart, while its expression in their trunk muscle appeared to be less affected (Fig. 5F). cdh5 is expressed by zebrafish endothelial layer of the heart throughout embryonic development, with stronger expression in the ventricle than the atrium in two-day old embryos (; Fig. 5G). Expression of cdh5 in the glo mutant heart was similar to the control (Fig. 5H).
Formation of the vascular system in glomutants is affected
In this study, we show that zebrafish cadherin2 is expressed by the myocardium of both the atrium and ventricle during critical periods of zebrafish cardiac development, and that loss of cadherin2 function disrupts differentiation of myocardiocytes, normal functioning of the zebrafish heart and formation of the intersegmental vasculature. There are only a few systems (e.g. visual and cardiac systems) and organisms in which one can readily perform in vivo studies of gene function in cell and tissue differentiation together with measurement of organ physiological performance affected by the loss of the gene function. This study further demonstrates the usefulness of zebrafish as a model organism to study gene function in vertebrate cardiac differentiation and function. The reduced cardiac performance in the glo mutant embryos likely results from altered atrial and ventricular morphology, which in turn is likely caused by the changes in the myocardiocyte differentiation.
Pericardial and cardiac gross morphology is greatly altered in cadherin2 morphants and glomutant embryos
Our finding that the pericardial cavity was greatly enlarged in the vast majority of the cadherin2 morphants and in all glo mutants suggests that cadherin2 function could be required for developing a normal pericardial cavity. However, it cannot be ruled out that altered kidney function could be causing pericardial enlargement via fluid accumulation. Thus, the actual cause(s) for the formation of such an enlarged pericardial cavity, likely due to an accumulation of fluid in the cavity, in these embryos is unclear. It may have more to do with the much-weakened cardiac function in the mutant embryos than the development of the vascular system. This idea is supported by our finding that most of the major blood vessels were present in the glo mutant embryos, but there was little labeling of the blood vessels using the FITC-Dextran injection (Fig. 8).
Enlarged pericardial cavity was reported in cadherin2 mutant mice , but not seen in the pac mutants . This difference likely results from different degrees of knockdown/knockout of cadherin2 function in these mutants. It is possible that a small amount of functional cadherin2 protein is produced in the pac mutants due to alternative splicing , therefore defects in the pac mutants are not as severe as in the glo mutants.
The gross morphological defects observed in the glo mutant hearts are likely caused by changes in the morphology of individual myocardiocytes. Although defects in the myocardiocytes were observed in both glo and pac mutant embryos, the gross cardiac morphological defects were obvious only in the former. This again, is likely due to differences in the degree of cadherin2 function disruption, with a complete loss of cadherin2 function in the glo mutant embryos, while perhaps some cadherin2 function remaining in the pac mutant embryos (see above). The enlarged pericardial cavity is unlikely one of the major causes of the atrial and ventricular disorganization, because the gross morphological defects can be detected at 36 hpf, when there is no obvious change in the size of the pericardial cavity.
Cadherin2 plays an important role in myocardiocyte differentiation, cardiac morphogenesis and performance
During cardiac development, myocardiocytes express cardiac specific markers such as nkx2.5, cmlc2, vmhc and cdh5[3, 20]. Despite greatly altered gross cardiac morphology and myocardiocytes morphology in glo mutant embryos, expression of nkx2.5, cmlc2 and cdh5 is largely unchanged, suggesting that cadherin2 is not required for normal expression of these genes. However, expression of vmhc, a ventricle specific gene, in the glo mutant heart is reduced compared to control embryos. Although vmhc role in cardiac formation and function has not being studied in vertebrates, loss of function in an atrium specific myosin heavy chain has been linked to disruption in atrial function and altered ventricular morphogenesis in zebrafish , suggesting that vmhc may play a similar role in ventricular myocardiocyte differentiation and function. Therefore, it is reasonable to speculate that cadherin2 function in vertebrate cardiac development and the function may be mediated, at least partially, by its effect on vmhc expression.
Myocardiocytes undergo extensive morphological changes during cardiac morphogenesis . For example, cuboidal shaped chicken myocardiocytes become flattened and tightly associated when the heart begins to contract, and the myocardiocytes become fusiform shaped and arranged in the circumferential direction during looping , similar to the zebrafish myocardiocytes (Fig. 6A and 6E). Many myocardiocytes in the glo mutant become round shaped and lack tight association, which may have contributed to poor electrical conduction between cells, thus reducing the contraction rate shown in glo mutant embryos. This also may have contributed to the gross cardiac morphological defects including the disrupted heart looping observed in both the cadherin2 mutant mice  and glo mutant embryos.
The loss of cadherin2 function does not affect all aspects of myocardiocyte differentiation because myocardiocytes in the cadherin2 mutant mice and glo mutant embryos still contract and/or express cardiac specific genes (see above). Other cell adhesion molecules such as N-CAM and other cadherin molecules may still function in these animals. Although most of the blood vessels were present as indicated by cdh5 labeling, it is unclear whether or not these cdh5-positive vessels were normal. As was shown in the heart, cdh5 staining in the glo mutants was similar to controls. It is not surprising that the formation of the major trunk vessels is not greatly affected in the glo mutant embryos, since the zebrafish vasculature system express cdh5, instead of cadherin2. It is possible that there are defects on those blood vessels, but we have no other markers (e.g. Zn-5, vmhc) to assess their integrity. Additionally, the intersegmental vessels and all other vessels formed via angiogenic remodeling are poorly formed or not present in glo mutant embryos. This is likely a proximal effect of the lack of pressure and flow generated by the glo mutant embryo heart, and/or due to cadherin2 function on the trunk and tail muscle development .
Our results suggest that cadherin2 plays an essential role in zebrafish cardiovascular development. Although the exact mechanisms remain unknown as to the formation of the enlarged pericardium and reduced peripheral blood flow, it is clear that myocardiocyte differentiation and physiological performance is impaired.
Zebrafish (Danio rerio) were maintained at 28.5°C as described in the Zebrafish Book . The glo heterozygous mutant carriers and their wildtype siblings from a single breeding, obtained from the Zebrafish International Resource Center at the University of Oregon (Eugene, OR) as embryos, were raised to reproductive maturity in the animal care facility at the University of Akron. Pair-wise breeding was performed to identify glo heterozygous mutant carriers, and the glo mutant embryos were identified by gross morphological phenotype. Their wildtype and heterozygous siblings were used as controls. Embryos for cadherin2 morpholino oligonucleotides (MO) experiments were obtained from breeding of wildtype adult zebrafish. Zebrafish embryos homozygous for the pac mutation (pac tm101B ) were obtained from Max-Planck Institute for Developmental Biology (Tübingen, Germany). Embryos for whole mount in situ hybridization were raised in PTU (1-phenyl-2-thiourea, 0.003%) in order to reduce optical interference of pigments. All animal-related procedures were approved by the Care and Use of Animals in Research Committee at the University of Akron.
Measurement of cardiac performance in control and glomutant zebrafish embryos
glo mutant (N = 10) and control zebrafish embryos (heterozygous and wildtype siblings of the glo embryos, N = 8) were used for measuring cardiac function. Mutant embryos were identified and separated from wild type embryos using a Leica dissecting microscope, based on morphological differences such as curved spines and tail blisters . The embryos were dechorionated at approximately 24 hpf and were kept in a small plastic compartmentalized container (2.5 ml for each compartment) with tank water at 28.5°C for the duration of the experiment. Immediately before each measurement, the embryos were immobilized using MS-222 (0.02%). At this early stage of development and at this low concentration of MS-222, there are no measurable effects of this anesthesia on the cardiovascular system (unpublished data). The embryos were digitally recorded at 30 hpf, 36 hpf, 48 hpf, 58 hpf, and 72 hpf using a temperature controlled inverted microscope (Leica, DMIRB) equipped with a digital video camera (Redlake MASD, San Diego, CA). For each embryo at each selected period of development, a 10 sec digital video was captured at 0.008 second intervals (125 frames per second). At the completion of each video, the individual fish were returned to their respective compartments and were tracked throughout the experiment.
Videos were analyzed for heart rate, end diastolic volume, end systolic volume, stroke volume, and cardiac output using ImagePro Plus® imaging software (Media Cybernetics, Silver Spring, MD). Heart rate was calculated by counting the number of sequential contractions, beginning and ending at end diastole, occurring in the video file and dividing by the exact time interval. End diastolic volume was determined by measuring the perimeter of the ventricle at diastole (obtained by tracing the ventricle in a single frame of the cardiac cycle video stopped where the ventricle was at its largest point) along with the length and width of the ventricle at diastole. End systolic volume was determined by measuring the perimeter of the ventricle at systole (where the ventricle was at its narrowest width following diastole) in addition to the length and width of the ventricle at systole. The resulting ventricular volumes were calculated using the formula (8/3π) a/L, where a is the area of the traced ventricle and L is the length of the ventricle at either diastole or systole . The stroke volume was calculated by subtracting the end systolic volume from the end diastolic volume and the cardiac output was calculated by multiplying the stroke volume by the heart rate. Data were analyzed for differences between the glo and control embryos and over development time by using a two-way repeated measure ANOVA. Post hoc comparisons were performed using Tukey's multiple comparisons procedure. All physiological data presented are means ± S.E.M.
MO and FITC-dextran Injections
A cadherin2 translation blocking morpholino oligonucleotide (5'-TCTGTATAAAGAAACCGATAGAGTT-3', , or a standard control (5'-CCTCTTACCTCAGTTACAATTTATA-3'), gifts from Dr. James Marrs (Indiana University) who purchased it from Gene Tools, Corvallis OR, was microinjected into either blastomeres and/or the yolk immediately below the blastomeres of 1–4 cell stage wild type embryos. Injected embryos were allowed to develop at 28.5° C until desired stages. Fluorescein isothiocyanate (FITC)-dextran (Sigma) was injected into the common cardinal vein of anesthetized embryos (56 hpf) according to .
Zebrafish embryos were euthanized in 0.02% methane tricaine sulfonate (MS-222, Sigma, St Louis, MO) and fixed in 4% paraformaldehyde in 0.1 M phosphate buffered saline (PBS) overnight at 4°C. To prepare tissue for whole mount in situ hybridization or immunohistochemistry, the tissue was rinsed in PBS, followed by 70% methanol and 100% methanol. The tissue was stored in 100% methanol at -20°C. Preparation of tissues for immunohistochemical staining on sections was described previously . Briefly, the fixed tissue was processed through a graded series of increasing sucrose concentrations, placed in 20% sucrose in PBS overnight, then embedded and frozen in a mixture of OCT embedding compound and 20% sucrose (1:1, v/v). A cryostat was used to obtain 12–14 μm sections. Some glo and control embryos processed for whole mount in situ hybridization or immunostaining were embedded and sectioned as described above. Tissue sections were collected on pretreated glass slides (Fisher Scientific, Pittsburgh, PA), dried at room temperature and stored at -80°C.
A cDNA containing the presequence, the extracellular and transmembrane domains of zebrafish cadherin2, obtained by RT-PCR from 24 hpf embryonic zebrafish total RNA, was used as a template to generate anti-sense and sense cadherin2 cRNA probes . cDNAs used to generate zebrafish nkx2.5, cardiac myosin light chain 2 (cmlc2), and ventricular myosin heavy chain (vmhc) cRNA probes were kindly provided by Deborah Yelon at the New York University . Zebrafish cdh5 cDNA used to generate cdh5 cRNA probes was kindly provided by Jon Larson at the Discovery Genomics . Synthesis of digoxigenin-labeled cRNA probes, procedures for whole mount in situ hybridization were described previously . Anti-digoxigenin Fab fragment antibodies conjugated to alkaline phosphatase were used for immunocytochemical detection of the cRNA probes, and this was followed by an NBT/BCIP color reaction step (Roche Molecular Biochemicals, Indianapolis, IN).
Procedures for whole mount immunohistochemistry and immunostaining on tissue sections were described in detail previously [43, 44]. Primary antibodies used were affinity purified zebrafish cadherin2 antibody (1:80, ) and Zn-5 (1:1000, Zebrafish International Resource Center, University of Oregon, Eugene, OR). Biotinylated secondary antibodies (Vector Laboratories, Burlingame, CA) were used at 1:200. Visualization of the reaction was achieved by using a DAB kit (Vector Laboratories). For immunofluorescent double labeling experiments, the secondary antibodies were Cy3-conjugated anti-rabbit IgG and FITC-conjugated anti-mouse IgG (Jackson ImmunoResearch Laboratories, West Grove, PA) for the detection of cadherin-2 and Zn-5, respectively.
first branchial arch
second branchial arch
third branchial arch
posterior cerebral vein
ventral aorta. Other abbreviations are the same as in figure
The authors thank Dr. Deborah Yelon for providing nkx2.5, cmlc2 and vmhc cDNAs, Dr. Jon Larson for providing cdh5 cDNA, and Dr. James Marrs for providing us with the cdh2 MO. We thank the Zebrafish International Resource Center at the University of Oregon for providing the glo mutant embryos, and the Max-Planck Institute for Developmental Biology (Tübingen, Germany) for providing the pac mutant embryos. This work was supported by grants from NIH HL78169 (B. Bagatto) and NIH EY13879 (Q. Liu).
- Fishman MC, Chien KR: Fashioning the vertebrate heart: earliest embryonic decisions. Development. 1997, 124 (11): 2099-2117.PubMedGoogle Scholar
- Stainier DY, Fishman MC: Patterning the zebrafish heart tube: acquisition of anteroposterior polarity. Dev Biol. 1992, 153 (1): 91-101. 10.1016/0012-1606(92)90094-W.View ArticlePubMedGoogle Scholar
- Yelon D: Cardiac patterning and morphogenesis in zebrafish. Developmental Dynamics. 2001, 222 (4): 552-563. 10.1002/dvdy.1243.View ArticlePubMedGoogle Scholar
- Hu N, Sedmera D, Yost HJ, Clark EB: Structure and function of the developing zebrafish heart. Anatomical Record. 2000, 260 (2): 148-157. 10.1002/1097-0185(20001001)260:2<148::AID-AR50>3.0.CO;2-X.View ArticlePubMedGoogle Scholar
- Isogai S, Al. E: Angiogenic network formation in the developing vertebrate trunk. Development and disease. 2003, 130: 5281-5290.Google Scholar
- Weinstein BM, Fishman MC: Cardiovascular morphogenesis in zebrafish. Cardiovasc Res. 1996, 31 Spec No: E17-24. 10.1016/0008-6363(95)00139-5.View ArticlePubMedGoogle Scholar
- Childs S, Al. E: Paterning of angiogenesis in the zebrafish embryo. Development. 2002, 129: 973-982.PubMedGoogle Scholar
- Weinstein BM: What guides early embryonic blood vessel formation?. Dev Dyn. 1999, 215 (1): 2-11. 10.1002/(SICI)1097-0177(199905)215:1<2::AID-DVDY2>3.0.CO;2-U.View ArticlePubMedGoogle Scholar
- Weinstein BM: Plumbing the mysteries of vascular development using the zebrafish. Cell & Developmental Biology. 2002, 13: 515-522. 10.1016/S1084952102001052.View ArticleGoogle Scholar
- Isogai S, Horiguchi M, Weinstein BM: The vascular anatomy of the developing zebrafish: and atlas of embryonic and early larval development. Developmental Biology. 2001, 230: 278-301. 10.1006/dbio.2000.9995. (For complete vascular anatomy, see http://mgchd1.nichd.nih.gov:8000/zfatlas/Intro%20Page/intro1.html)View ArticlePubMedGoogle Scholar
- Wang DZ, Reiter RS, Lin JLC, Wang Q, Williams HS, Krob SL, Schultheiss TM, Evans S, Lin JJC: Requirement of a novel gene, Xin, in cardiac morphogenesis. Development. 1999, 126 (6): 1281-1294.PubMedGoogle Scholar
- Wilting J, Christ B, Bokeloh M, Weich HA: In-Vivo Effects of Vascular Endothelial Growth-Factor on the Chicken Chorioallantoic Membrane. Cell and Tissue Research. 1993, 274 (1): 163-172. 10.1007/BF00327997.View ArticlePubMedGoogle Scholar
- Biben C, Harvey RP: Homeodomain factor Nkx2-5 controls left-right asymmetric expression of bHLH gene eHand during murine heart development. Genes and Development. 1997, 11: 1357-1367.View ArticlePubMedGoogle Scholar
- Ferrara N, CarverMoore K, Chen H, Dowd M, Lu L, Oshea KS, PowellBraxton L, Hillan KJ, Moore MW: Heterozygous embryonic lethality induced by targeted inactivation of the VEGF gene. Nature. 1996, 380 (6573): 439-442. 10.1038/380439a0.View ArticlePubMedGoogle Scholar
- Ferrara N: Molecular and biological properties of vascular endothelial growth factor. Journal of Molecular Medicine-Jmm. 1999, 77 (7): 527-543. 10.1007/s001099900019.View ArticleGoogle Scholar
- Linask KK, Yu XY, Chen YP, Han MD: Directionality of heart looping: Effects of Pitx2c misexpression on flectin asymmetry and midline structures. Developmental Biology. 2002, 246 (2): 407-417. 10.1006/dbio.2002.0661.View ArticlePubMedGoogle Scholar
- Lyons I, Parsons LM, Hartley L, Li RL, Andrews JE, Robb L, Harvey RP: Myogenic and Morphogenetic Defects in the Heart Tubes of Murine Embryos Lacking the Homeo Box Gene Nkx2-5. Genes & Development. 1995, 9 (13): 1654-1666.View ArticleGoogle Scholar
- Radice GL, Rayburn H, Matsunami H, Knudsen KA, Takeichi M, Hynes RO: Developmental defects in mouse embryos lacking N-cadherin. Developmental Biology. 1997, 181 (1): 64-78. 10.1006/dbio.1996.8443.View ArticlePubMedGoogle Scholar
- Srivastava D, Olson EN: A genetic blueprint for cardiac development. Nature. 2000, 407 (6801): 221-226. 10.1038/35025190.View ArticlePubMedGoogle Scholar
- Larson JD, Wadman SA, Chen E, Kerley L, Clark KJ, Eide M, Lippert S, Nasevicius A, Ekker SC, Hackett PB, Essner JJ: Expression of VE-cadherin in zebrafish embryos: A new tool to evaluate vascular development. Developmental Dynamics. 2004, 231 (1): 204-213. 10.1002/dvdy.20102.View ArticlePubMedGoogle Scholar
- Gumbiner BM: Cell adhesion: The molecular basis of tissue architecture and morphogenesis. Cell. 1996, 84 (3): 345-357. 10.1016/S0092-8674(00)81279-9.View ArticlePubMedGoogle Scholar
- Nollet F, Kools P, van Roy F: Phylogenetic analysis of the cadherin superfamily allows identification of six major subfamilies besides several solitary members. Journal of Molecular Biology. 2000, 299 (3): 551-572. 10.1006/jmbi.2000.3777.View ArticlePubMedGoogle Scholar
- Takeichi M: Cadherin Cell-Adhesion Receptors as a Morphogenetic Regulator. Science. 1991, 251 (5000): 1451-1455.View ArticlePubMedGoogle Scholar
- Hatta K, Takeichi M: Expression of N-Cadherin Adhesion Molecules Associated with Early Morphogenetic Events in Chick Development. Nature. 1986, 320 (6061): 447-449. 10.1038/320447a0.View ArticlePubMedGoogle Scholar
- Detrick RJ, Dickey D, Kintner CR: The Effects of N-Cadherin Misexpression on Morphogenesis in Xenopus Embryos. Neuron. 1990, 4 (4): 493-506. 10.1016/0896-6273(90)90108-R.View ArticlePubMedGoogle Scholar
- Fujimori T, Miyatani S, Takeichi M: Ectopic Expression of N-Cadherin Perturbs Histogenesis in Xenopus Embryos. Development. 1990, 110 (1): 97-&.PubMedGoogle Scholar
- Kerstetter AE, Azodi E, Marrs JA, Liu Q: Cadherin-2 function in the cranial ganglia and lateral line system of developing zebrafish. Developmental Dynamics. 2004, 230 (1): 137-143. 10.1002/dvdy.20021.View ArticlePubMedGoogle Scholar
- Lele Z, Folchert A, Concha M, Rauch GJ, Geisler R, Rosa F, Wilson SW, Hammerschmidt M, Bally-Cuif L: parachute/n-cadherin is required for morphogenesis and maintained integrity of the zebrafish neural tube. Development. 2002, 129 (14): 3281-3294.PubMedGoogle Scholar
- Malicki J, Jo H, Pujic Z: Zebrafish N-cadherin, encoded by the glass onion locus, plays an essential role in retinal patterning. Developmental Biology. 2003, 259 (1): 95-108. 10.1016/S0012-1606(03)00181-7.View ArticlePubMedGoogle Scholar
- Matsunaga M, Hatta K, Takeichi M: Role of N-Cadherin Cell-Adhesion Molecules in the Histogenesis of Neural Retina. Neuron. 1988, 1 (4): 289-295. 10.1016/0896-6273(88)90077-3.View ArticlePubMedGoogle Scholar
- Lampugnani MG, Resnati M, Raiteri M, Pigott R, Pisacane A, Houen G, Ruco LP, Dejana E: A Novel Endothelial-Specific Membrane-Protein Is a Marker of Cell Cell Contacts. Journal of Cell Biology. 1992, 118 (6): 1511-1522. 10.1083/jcb.118.6.1511.View ArticlePubMedGoogle Scholar
- Breier G, Breviario F, Caveda L, Berthier R, Shnurch H, Gotsch U, Vestweber D, Risau W, Dejana E: Molecular cloning and expression of murine vascular endothelial-cadherin in early stage development of cardiovascular system. Blood. 1996, 87: 630-641.PubMedGoogle Scholar
- Nakagawa S, Takeichi M: N-cadherin is crucial for heart formation in the chick embryo. Development Growth & Differentiation. 1997, 39 (4): 451-455. 10.1046/j.1440-169X.1997.t01-3-00006.x.View ArticleGoogle Scholar
- Yelon D, Horne SA, Stainier DYR: Restricted expression of cardiac myosin genes reveals regulated aspects of heart tube assembly in zebrafish. Developmental Biology. 1999, 214 (1): 23-37. 10.1006/dbio.1999.9406.View ArticlePubMedGoogle Scholar
- Berdougo E, Coleman H, Lee DH, Stainier DY, Yelon D: Mutation of weak atrium/atrial myosin heavy chain disrupts atrial function and influences ventricular morphogenesis in zebrafish. Development. 2003, 130: 6121-6129. 10.1242/dev.00838.View ArticlePubMedGoogle Scholar
- Manasek FJ: Embryonic development of the heart. I. A light and electron miscroscopic study of myocardial development in the early chick embryo. Journal of Morphology. 1968, 125: 329-366. 10.1002/jmor.1051250306.View ArticlePubMedGoogle Scholar
- Cortes F, Daggett D, Bryson-Richardson RJ, Neyt C, Maule J, Gautier P, Hollway GE, Keenan D, Currie PD: Cadherin-mediated differential cell adhesion controls slow muscle cell migration in the developing zebrafish myotome. Developmental Cell. 2003, 5 (6): 865-876. 10.1016/S1534-5807(03)00362-9.View ArticlePubMedGoogle Scholar
- Westerfield M: The Zebrafish Book: A Guide for the Laboratory Use of Zebrafish (Danio rerio). 1995, Eugene , University of Oregon PressGoogle Scholar
- Bagatto B, Burggren WW: A three-dimensional functional assessment of heart and vessel development in the larva of the zebrafish (Danio rerio). Physiological and Biochemical Zoology. 2006, 79 (1): 194-201. 10.1086/498185.View ArticlePubMedGoogle Scholar
- Nasevicius A, Larson J, Ekker SC: Distinct requirements for zebrafish angiogenesis revealed by a VEGF-A morphant. Yeast. 2000, 17 (4): 294-301. 10.1002/1097-0061(200012)17:4<294::AID-YEA54>3.0.CO;2-5.PubMed CentralView ArticlePubMedGoogle Scholar
- Liu Q, Sanborn KL, Cobb N, Raymond PA, Marrs JA: R-cadherin expression in the developing and adult zebrafish visual system. Journal of Comparative Neurology. 1999, 410 (2): 303-319. 10.1002/(SICI)1096-9861(19990726)410:2<303::AID-CNE11>3.0.CO;2-B.View ArticlePubMedGoogle Scholar
- Liu Q, Kerstetter AE, Azodi E, Marrs JA: Cadberin-1,-2, and-11 expression and cadherin-2 function in the pectoral limb bud and fin of the developing zebrafish. Developmental Dynamics. 2003, 228 (4): 734-739. 10.1002/dvdy.10401.View ArticlePubMedGoogle Scholar
- Liu Q, Babb SG, Novince ZM, Doedens AL, Marrs J, Raymond PA: Differential expression of cadherin-2 and cadherin-4 in the developing and adult zebrafish visual system. Visual Neuroscience. 2001, 18 (6): 923-933.PubMedGoogle Scholar
- Babb SG, Kotradi SM, Shah B, Chiappini-Williamson C, Bell LN, Schmeiser G, Chen E, Liu Q, Marrs JA: Zebrafish R-cadherin (Cdh4) controls visual system development and differentiation. Developmental Dynamics. 2005, 233 (3): 930-945. 10.1002/dvdy.20431.View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.