Cardiac responses to hypercapnia in larval zebrafish (Danio rerio): The links between CO2 chemoreception, catecholamines and carbonic anhydrase

Abstract

The ontogeny of carbon dioxide (CO 2 ) sensing in zebrafish (Danio rerio) has not been examined. In this study, CO 2 -mediated increases in heart rate were used to gauge the capacity of zebrafish larvae to sense CO 2 . CO 2 is thought to be detected via neuroepithelial cells (NECs), which are homologous to mammalian carotid body glomus cells. Larvae at 5 days post-fertilization (d.p.f.) exhibited tachycardia when exposed for 30 min to 0.75% CO 2 (~5.63 mmHg); at 7 d.p.f., tachycardia was elicited by 0.5% CO 2 (~3.75 mmHg). Based on pharmacological evidence using β-adrenergic receptor (β-AR) antagonists, and confirmed by β 1 -AR translational gene knockdown using morpholinos, the reflex tachycardia accompanying hypercapnia was probably mediated by the interaction of catecholamines with cardiac β 1 receptors. Because the cardiac response to hypercapnia was abolished by the ganglionic blocker hexamethonium, it is probable that the reflex cardio-acceleration was mediated by catecholamines derived from sympathetic adrenergic neurons. Owing to its likely role in facilitating intracellular acidification during exposure to hypercapnia, it was hypothesized that carbonic anhydrase (CA) is involved in CO 2 sensing, and that inhibition of CA activity would blunt the downstream responses. Indeed, the cardiac response to hypercapnia (0.75% CO 2 ) was reduced in fish at 5 d.p.f. exposed to acetazolamide, a CA inhibitor, and in fish experiencing zCAc (CA2-like a) knockdown. Successful knockdown of zCAc was confirmed by CA activity measurements, western blotting and immunocytochemistry. Co-injection of embryos with zCAc morpholino and mRNA modified at the morpholino binding site restored normal levels of CA activity and protein levels, and restored (rescued) the usual cardiac responses to hypercapnia. These data, combined with the finding that zCAc is expressed in NECs located on the skin, suggest that the afferent limb of the CO 2 -induced cardiac reflex in zebrafish larvae is initiated by coetaneous CO 2 -sensing neuroepithelial cells.

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

“…Under hypoxic conditions, however, the HO/CO system has been shown to regulate cardiovascular function by modulating sympathetic tone in mammals (Hirakawa and Hayashida, 2006;Ding et al, 2008). In addition, it has been reported that the sympathetic nervous system is responsible for evoking the tachycardic response observed in larval zebrafish exposed to high environmental CO 2 (Miller et al, 2014) and therefore this may also be the mechanism responsible for the hypoxic tachycardia reported in this study. This notion was supported by the further increase of f H observed in hypoxic shamtreated larvae in the presence of adrenaline (from 155.0±5.7 to 187.0±5.7 beats min −1 ; data not shown).…”

Introducing a novel mechanism to control heart rate in the ancestral pacific hagfish

“…Therefore, as this is the first time that sAC has been implicated in the control of heart rate, new roles for sAC in vertebrate hearts become a possibility. For example, hypercapnia produced tachycardia in embryonic zebrafish, a response blocked by atenolol, a β-adrenergic antagonist, and by a carbonic anhydrase inhibitor (Miller et al, 2014). Thus, it would be interesting to investigate whether KH7 might block this tachycardia, which would implicate a sAC-mediated control of heart rate in this case.…”

Acid–base physiology, neurobiology and behaviour in relation to CO2-induced ocean acidification

“…Interestingly, the ventilatory response was blunted in zebrafish chronically exposed to hypercapnia (Vulesevic et al, 2006), suggesting potential acclimation. And in zebrafish larvae, the tachycardia response to hypercapnia was absent in 7-day-old larval zebrafish (Miller et al, 2014), showing life-stage-specific differences. Thus, as reported in freshwater zebrafish, OA-relevant CO 2 disturbances could potentially affect CO 2 -sensing neurons in marine fish and trigger physiological responses, which could be compensatory or maladaptive.…”

“…Zebrafish neuroepithelial cells have been proposed to regulate important cardiorespiratory processes such as increasing ventilatory amplitude in adult zebrafish (Vulesevic et al, 2006) and tachycardia in 7-day-old larval zebrafish (Miller et al, 2014). Interestingly, the ventilatory response was blunted in zebrafish chronically exposed to hypercapnia (Vulesevic et al, 2006), suggesting potential acclimation.…”