A Larval Zebrafish Model for Assessing Hypoxic‐Induced In Vivo Cardiomyocyte Damage: Time Course for Induction and Cardiac Output Recovery

Abstract

Larval zebrafish are used to study regenerative properties of cardiac muscle and treatments that may enhance this process. Cardiac injury models in zebrafish larva currently include cryoinjury, laser ablation, pharmacological treatment and cardiac dysfunction mutations. Effective in damaging cardiomyocytes, these models lack the critical element of tissue hypoxia, which induces molecular cascades within cardiac muscle. We have developed a potentially more realistic model for studying cardiac arrest and recovery in larval zebrafish (Danio rerio) by using acute, severe hypoxic exposure to briefly induce cardiac arrest. Exposure to PO2s of 6–9 mmHg induces loss of mobility followed by cardiac arrest within 120 min in 5 days post fertilization (dpf) and within 40 min at 10 dpf. Approximately 90% of 5dpf larvae survived acute, severe hypoxic exposure, but survival fell to 30% by 10 dpf. Heart beat resumed in surviving larvae and returned to air‐saturated water at 4 min in 5 dpf and 6–8 min at 8–10 dpf. Heart rate, stroke volume and cardiac output in control 6–7 dpf larvae before hypoxic exposure were 188±5 bpm, 0.20±0.001nl and 35.5.3±2.2 nl/min (n=35), respectively. In larvae with hypoxia‐induced cardiac arrest, heart rate returned to control values by 24 h of recovery, but stroke volume and cardiac output remained ~50% depressed from control values at 24 h of recovery. Full restoration of cardiac performance was evident at 72h, indicating that regenerative properties of the zebrafish heart extend to critical physiological measures. Molecular markers for apoptosis (Caspase‐3) were elevated >5‐fold immediately after resumption of heart beat, peaking after 4 hours into the recovery period. A cell regeneration marker (PCNA) increased >4‐fold beginning after a lag of 2 hours, and was still elevated ~6 hours after resumption of heart beat, presumably contributing to recovery of heart function. The highly reproducible cardiac effects of acute hypoxia‐induced cardiac arrest in the larval zebrafish presents an alternative, realistic model for studying cardiac tissue hypoxia injury and recovery of function.Support or Funding InformationAstraZenecaTime course of changes in heart rate, stroke volume, and cardiac output in 5–7 day larval zebrafish following induction of cardiac arrest by acute hypoxic exposure (PO2 ~ 7–9 mmHg).Figure 1