Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, C2

Oral Communications

Quantification of cardiomyocyte proliferation during early development and following heart laser injury in zebrafish embryo.

G. Matrone1, K. S. Wilson1, C. S. Tucker1, J. J. Mullins1, M. A. Denvir1

1. BHF Centre for Cardiovascular Science, QMRI, University of Edinburgh, Edinburgh, United Kingdom.

Adult Zebrafish (D.rerio) can regenerate heart ventricle after partial resection (Poss et al., 2002; Raya et al., 2003). We have developed a model of cardiac injury in the zebrafish embryo using a laser microbeam, which demonstrates the ability of the embryonic ventricle to recovery functionality after damage. The aim of this study is to assess cardiomyocyte loss and proliferation in this model. Tg(cmlc2:EGFP) zebrafish embryos underwent heart laser injury at 72 hours post fertilisation (hpf), control embryos received fin injury. Heart ventricle cardiomyocyte number (CMn) was assessed by counting DAPI stained nuclei in isolated hearts, at 48, 70 (pre-laser), 74 (post-laser), 96 and 120 hpf. Ejection fraction was measured by video image capture and diastolic-systolic area subtraction method using imageJ. In normal embryos, CMn increased by approximately 30% per day between 48hpf and 96hpf (138±16 vs 234±48, p<0.001) with a further increase by 58% between 96 and 120hpf (234±48 vs 369±34, p<0.001). Laser injury at 72 hours produced a small non-significant reduction in CMn by 15% compared with controls (195±42 vs 219±45, p>0.05) but this recovered within 24 hours following laser injury (234±48 vs 252±21). Changes in CMn were mirrored by changes in ejection fraction with a significant fall after laser (20.4±0.7% vs 14.2±1.2%, p<0.001) and recovery by 24 hours (23.2±0.9% vs 23.4±1.4%, p=ns). Data are expressed as mean±SEM (n=5 per group; p=ANOVA) Cardiomyocyte proliferation occurs rapidly during development with acceleration of cell number on day 5. Loss and proliferation of cardiomyocytes contributes partailly to the fall and subsequent functional recovery in this model of laser injury.

Where applicable, experiments conform with Society ethical requirements