Chronic hypoxic incubation blunts a cardiovascular reflex loop in embryonic American alligator (Alligator mississippiensis)

Hypoxia is a naturally occurring environmental challenge for embryonic non-avian reptiles, and this study is the first to investigate the impact of chronic hypoxia on a possible chemoreflex loop in a developing non-avian reptile. We measured heart rate and blood pressure in normoxic and hypoxic-incubated (10% O₂) American alligator embryos (Alligator mississippiensis) at 70 and 90/95% of development. We hypothesized that hypoxic incubation would blunt embryonic alligators’ response to a reflex loop stimulated by phenylbiguanide (PBG), a 5-HT₃ receptor agonist that stimulates vagal pulmonary C-fiber afferents. PBG injection caused a hypotensive bradycardia in 70 and 95% of development embryos (paired t tests, P < 0.05), a response similar to mammals breathing inspired air (all injections made through occlusive catheter in tertiary chorioallantoic membrane artery). Hypoxic incubation blunted the bradycardic response to PBG in embryos at 95% of development (two-way ANOVA, P < 0.01). We also demonstrated that the vagally mediated afferent limb of this reflex can be partially or completely blocked in ovo with a 5-HT₃ receptor blockade using ondansetron hydrochloride dihydrate (OHD), with a ganglionic blockade using hexamethonium, or with a cholinergic blockade using atropine. Atropine eliminated the hypotensive and bradycardic responses to PBG, and OHD and hexamethonium significantly blunted these responses. This cardiovascular reflex mediated by the vagus was affected by hypoxic incubation, suggesting that reptilian sympathetic and parasympathetic reflex loops have the potential for developmental plasticity in response to hypoxia. We suggest that the American alligator, with an extended length of time between each developmental stage relative to avian species, may provide an excellent model to test the cardiorespiratory effects of prolonged exposure to changes in atmospheric gases. This extended period allows for lengthy studies at each stage without the transition to a new stage, and the natural occurrence of hypoxia and hypercapnia in crocodilian nests makes this stress ecologically and evolutionarily relevant.