Oxygen chemoreceptor inhibition by dopamine D₂ receptors in isolated zebrafish gills

Dopamine is an essential modulator of oxygen sensing and control of ventilation and is the most well described and abundant neurotransmitter in the mammalian carotid body. Little is known of the evolutionary significance of dopamine in oxygen sensing, or whether it plays a similar role in anamniotes. In the model vertebrate, zebrafish (Danio rerio), presynaptic dopamine D₂ receptor (D₂R) expression was demonstrated in gill neuroepithelial cells (NECs), analogues of mammalian oxygen chemoreceptors; however, a mechanism for dopamine and D₂R in the gills had not been defined. The present study tested the hypothesis that presynaptic D₂Rs provide a feedback mechanism attenuating the chemoreceptor response to hypoxia. Using an isolated gill preparation from Tg(elavl3:GCaMP6s) zebrafish, we measured hypoxia-induced changes in intracellular Ca²⁺ concentration ([Ca²⁺]_i) in NECs and postsynaptic neurons. Activation of D₂R with dopamine or specific D₂R agonist, quinpirole, decreased hypoxic responses in NECs; whereas D₂R antagonist, domperidone, had the opposite effect. Addition of SQ22536, an adenylyl cyclase (AC) inhibitor, decreased the effect of hypoxia on [Ca²⁺]_i, similar to dopamine. Activation of AC by forskolin partially recovered the suppressive effect of dopamine on the Ca²⁺ response to hypoxia. Furthermore, we demonstrate that the response to hypoxia in postsynaptic neurons was dependent upon innervation with NECs, and was subject to modulation by activation of presynaptic D₂R. Our results provide the first evidence of neurotransmission of the hypoxic signal at the NEC-nerve synapse in the gill and suggest that a presynaptic, modulatory role for dopamine in oxygen sensing arose early in vertebrate evolution. KEY POINTS: For the first time, we present an experimental model that permits imaging of intracellular Ca²⁺ in identified oxygen chemoreceptors in zebrafish using GCaMP in a whole/intact sensing organ. The hypoxic response of zebrafish chemoreceptors is attenuated by dopamine through a mechanism involving D₂ receptors and adenylyl cyclase. Zebrafish oxygen chemoreceptors send a hypoxic signal to postsynaptic (sensory) neurons. Postsynaptic neuronal responses to hypoxia are modulated by presynaptic D₂ receptors, suggesting a link between chemoreceptor inhibition by dopamine and modulation of the hypoxic ventilatory response. Our results suggests that a modulatory role for dopamine in oxygen sensing arose early in vertebrate evolution.