Respiratory plasticity after perinatal hyperoxia is not prevented by antioxidant supplementation

Abstract

Perinatal hyperoxia attenuates the hypoxic ventilatory response in rats by altering development of the carotid body and its chemoafferent neurons. In this study, we tested the hypothesis that hyperoxia elicits this plasticity through the increased production of reactive oxygen species (ROS). Rats were born and raised in 60% O(2) for the first two postnatal weeks while treated with one of two antioxidants: vitamin E (via milk from mothers whose diet was enriched with 1000 IU vitamin E kg(-1)) or a superoxide dismutase mimetic, manganese(III) tetrakis (1-methyl-4-pyridyl) porphyrin pentachloride (MnTMPyP; via daily intraperitoneal injection of 5-10 mg kg(-1)); rats were subsequently raised in room air until studied as adults. Peripheral chemoreflexes, assessed by carotid sinus nerve responses to cyanide, asphyxia, anoxia and isocapnic hypoxia (vitamin E experiments) or by hypoxic ventilatory responses (MnTMPyP experiments), were reduced after perinatal hyperoxia compared to those of normoxia-reared controls (all P<0.01); antioxidant treatment had no effect on these responses. Similarly, the carotid bodies of hyperoxia-reared rats were only one-third the volume of carotid bodies from normoxia-reared controls (P <0.001), regardless of antioxidant treatment. Protein carbonyl concentrations in the blood plasma, measured as an indicator of oxidative stress, were not increased in neonatal rats (2 and 8 days of age) exposed to 60% O(2) from birth. Collectively, these data do not support the hypothesis that perinatal hyperoxia impairs peripheral chemoreceptor development through ROS-mediated oxygen toxicity.

Fig. 1

Carotid sinus nerve (CSN) responses to NaCN (80, 40, and 20 μg kg⁻¹) for adult, normoxia-reared (Normoxia) and hyperoxia-reared (Hyperoxia) rats. Rats were raised with mothers fed a standard laboratory diet (Control diet) or a diet enriched with vitamin E (+ Vit. E diet) through postnatal week 2. Changes in CSN activity are reported as means ± S.E.M.; sample sizes (n) are given beneath each bar. Statistical analyses were run separately at each concentration of NaCN. * P≤0.05 for Normoxia versus Hyperoxia groups (i.e., main effect for developmental F_IO₂). No differences were detected between Control and Vitamin E diets (two-way ANOVA; diet and diet × developmental F_IO₂, all P>0.05).

Fig. 2

Carotid sinus nerve (CSN) responses to (A) asphyxia, (B) 100% N₂, and (C) isocapnic hypoxia for adult, normoxia-reared (Normoxia) and hyperoxia-reared (Hyperoxia) rats. Rats were raised with mothers fed a standard laboratory diet (Control diet) or a diet enriched with vitamin E (+ Vit. E diet) through postnatal week 2. Changes in CSN activity are reported as means ± S.E.M.; sample sizes (n) are given beneath each bar. For asphyxia, * P≤0.05 for Normoxia versus Hyperoxia groups (i.e., main effect for developmental F_IO₂); no differences were detected between Control and Vitamin E diets (two-way ANOVA; diet and diet × developmental F_IO₂, both P>0.05). For 100% N₂ and isocapnic hypoxia, CSN responses were not compared by two-way ANOVA due to insufficient sample sizes; see text for details.

Fig. 3

Ventilatory response to hypoxia (12.5% O₂) for adult, normoxia-reared (Normoxia) and hyperoxia-reared (Hyperoxia) rats treated with saline or MnTMPyP through postnatal week 2. Panels A–D present respiratory frequency (f_R), tidal volume (V_T), minute ventilation (V̇_E) and ventilation-to-metabolism ratio (V̇_E/V̇_O2) for rats breathing hypoxic gas mixtures. In panels E and F, V̇_E and V̇_E/V̇_O2 responses are presented as the percentage increase from baseline. Values are means ± S.E.M.; n=16 per group. * P≤0.05 for Normoxia versus Hyperoxia groups (i.e., main effect for developmental F_IO₂). No differences were detected between saline-treated and MnTMPyP-treated groups (two-way ANOVA; drug and treatment x drug, all P>0.05).

Fig. 4

Carotid body volumes for adult, normoxia-reared (Normoxia) and hyperoxia-reared (Hyperoxia) rats. In panel A, rats were raised with mothers fed a standard laboratory diet (Control diet) or a diet enriched with vitamin E (+ Vit. E diet) through postnatal week 2. In panel B, rats were treated with saline or MnTMPyP through postnatal week 2. Values are means ± S.E.M.; n=3 per group in panel A and n=7 per group in panel B. * P≤0.05 for Normoxia versus Hyperoxia groups (i.e., main effect for developmental F_IO₂). No effects of antioxidant treatments were detected (two-way ANOVA; main effects and interactions, all P>0.05).

Fig. 5

Protein carbonyl concentrations in blood plasma of two-day-old (P2) and eight-day-old (P8) rats raised from birth in 21% O₂ (Normoxia) or 60% O₂ (Hyperoxia). Values are means ± S.E.M.; n=7 per group, except n=6 for Normoxia at P2. No effect of hyperoxic exposure was detected at either age (both P>0.05).