Evidence for a role of heme oxygenase-1 in the control of cardiac function in zebrafish (Danio rerio) larvae exposed to hypoxia

Abstract

Carbon monoxide (CO) is a gaseous neurotransmitter produced from the breakdown of heme via heme oxygenase-1 (HO-1; hypoxia inducible isoform) and 2 (HO-2; constitutively expressed isoform). In mammals, CO is involved in modulating cardiac function. The role of the HO-1/CO system in the control of heart function in fish, however, is unknown and investigating its physiological function in lower vertebrates will provide a better understanding of the evolution of this regulatory mechanism. We explored the role of the HO-1/CO system in larval zebrafish (Danio rerio) in vivo by investigating the impact of translational gene knockdown of HO-1 on cardiac function. Immunohistochemistry revealed the presence of HO-1 in the pacemaker cells of the heart at 4 days post fertilization and thus the potential for CO production at these sites. Sham zebrafish larvae (experiencing normal levels of HO-1) significantly increased heart rate (fH) when exposed to hypoxia (PwO2=30 mmHg). Zebrafish larvae lacking HO-1 expression after morpholino knockdown (morphants) exhibited significantly higher fH under normoxic (but not hypoxic) conditions when compared to shams. The increased fH in HO-1 morphants was rescued (fH was restored to control levels) after treatment of larvae with a CO releasing molecule (40 µM CORM). The HO-1 deficient larvae developed significantly larger ventricles and when exposed to hypoxia they displayed higher cardiac outputs (Q) and stroke volumes (SV). These results suggest that under hypoxic conditions, HO-1 regulates Q and SV presumably via the production of CO. Overall, this study provides a better understanding into the role of the HO-1/CO system in controlling heart function in lower vertebrates. We demonstrate for the first time the ability for CO to be produced in presumptive pacemaker cells of the heart where it plays an inhibitory role in setting the resting cardiac frequency.