Late onset of NMDA receptor-mediated ventilatory control during early development in zebrafish (Danio rerio)

Increased ventilation frequency (fV) in response to hypoxia in adult fish depends on ionotropic N-methyl-D-aspartate (NMDA) receptors. Nonetheless, the ontogeny of central control mechanisms mediating hypoxic ventilatory chemoreflexes in lower vertebrates has not been studied. Therefore, the aim of this study was to determine when the hypoxic ventilatory response during zebrafish (Danio rerio) development is mediated via NMDA receptors, by performing physiological experiments and western blot analysis of NMDA receptor subunits. Zebrafish larvae at stages 4-16 days post-fertilisation (dpf) were exposed to an hypoxic pulse in control groups and in groups treated with MK801 (NMDA receptor antagonist). The hypoxic increase in fV was present at all larval stages, and it matured during development. The reflex became MK801 sensitive at 8 dpf, but did not completely rely on a glutamatergic transmission until 13 dpf. This, together with changing subunit composition during the different stages (increasing amounts of NMDAR1 subunits and appearance of NMDAR2A subunits in adults), suggests that the amount of functional NMDA receptors needed to achieve a fully developed reflex is not attained until later stages. Furthermore, our results suggest that other non-NMDA receptor mechanisms are responsible for the hypoxia-induced increase in fV during the earlier developmental stages.