Recurrent apnea with intermittent hypoxia is a major clinical problem in preterm infants. Recent studies, although limited, showed that adults who were born preterm exhibit increased incidence of sleep-disordered breathing and hypertension, suggesting that apnea of prematurity predisposes to autonomic dysfunction in adulthood. Here, we demonstrate that adult rats that were exposed to intermittent hypoxia as neonates exhibit exaggerated responses to hypoxia by the carotid body and adrenal chromaffin cells, which regulate cardio-respiratory function, resulting in irregular breathing with apneas and hypertension. The enhanced hypoxic sensitivity was associated with elevated oxidative stress, decreased expression of genes encoding antioxidant enzymes, and increased expression of pro-oxidant enzymes. Decreased expression of the Sod2 gene, which encodes the antioxidant enzyme superoxide dismutase 2, was associated with DNA hypermethylation of a single CpG dinucleotide close to the transcription start site. Treating neonatal rats with decitabine, an inhibitor of DNA methylation, during intermittent hypoxia exposure prevented oxidative stress, enhanced hypoxic sensitivity, and autonomic dysfunction. These findings implicate a hitherto uncharacterized role for DNA methylation in mediating neonatal programming of hypoxic sensitivity and the ensuing autonomic dysfunction in adulthood.blood pressure | developmental programming | norepinephrine I n preterm infants, respiratory disorders with recurrent apnea and the associated intermittent hypoxemia (IH) are major clinical problems (1). Infants with recurrent apnea exhibit an enhanced hypoxic ventilatory response (2), an effect that was attributed to a heightened chemo-reflex arising from the carotid body, which is a sensory organ that detects changes in arterial blood O 2 levels (3). Carotid body sensitivity to hypoxia is reset after birth and this effect is modulated by chronic hypoxia (4-6). Neonatal rats exposed to IH showed exaggerated carotid body and ventilatory responses to hypoxia (7,8). Catecholamine secretion from the adrenal medulla is another important homeostatic mechanism that preserves cardiovascular function under hypoxia (9, 10). In neonates, hypoxia facilitates catecholamine secretion by directly affecting the excitability of adrenal chromaffin cells (11), a response that is markedly augmented in neonatal rats subjected to IH (12,13). Exaggerated hypoxic sensing of carotid body and adrenal chromaffin cells in neonates by IH is attributed to increased oxidative stress (12,14). IH also augments hypoxic responses of the carotid body and adrenal medulla in adult rats (15, 16), which is completely reversed following reoxygenation (15). In striking contrast, in neonates the augmented hypoxic sensitivity that is induced by IH persists into adulthood (8,12,14). The molecular mechanisms underlying the long-lasting effects of neonatal IH on hypoxic sensing and its physiological consequences in adult life are not known.It is being increasingly recognized that environm...

Cardiorespiratory functions in mammals are exquisitely sensitive to changes in arterial O 2 levels. Hypoxia-inducible factors (e.g., HIF-1 and HIF-2) mediate transcriptional responses to reduced oxygen availability. We demonstrate that haploinsufficiency for the O 2 -regulated HIF-2α subunit results in augmented carotid body sensitivity to hypoxia, irregular breathing, apneas, hypertension, and elevated plasma norepinephrine levels in adult Hif-2α +/− mice. These dysregulated autonomic responses were associated with increased oxidative stress and decreased mitochondrial electron transport chain complex I activity in adrenal medullae as a result of decreased expression of major cytosolic and mitochondrial antioxidant enzymes. Systemic administration of a membrane-permeable antioxidant prevented oxidative stress, normalized hypoxic sensitivity of the carotid body, and restored autonomic functions in Hif-2α +/− mice. Thus, HIF-2α-dependent redox regulation is required for maintenance of carotid body function and cardiorespiratory homeostasis.blood pressure | control of ventilation | catecholamines H ypoxia-inducible factors (HIFs) mediate transcriptional responses to reduced O 2 availability (1). HIF-1, the first identified member of the HIF family, is comprised of an O 2 -regulated α subunit and a constitutively expressed β subunit (2). Complete deficiency of HIF-1α in Hif-1α −/− mice results in embryonic lethality at midgestation with major malformations of the central nervous system, heart, and vasculature (3). Hif-1α +/− mice, which are partially deficient in HIF-1α expression, develop normally and are indistinguishable from their WT littermates in the sedentary state with respect to autonomic functions, including blood pressure (BP), ventilatory responses to hypoxia or hypercapnia, and plasma catecholamine levels (4). However, Hif-1α +/− mice exhibit impaired O 2 sensing by the carotid body (4, 5), the primary sensory organ for detecting hypoxemia (6, 7), and altered physiological adaptations to chronic hypoxia (5, 8).The HIF-2α subunit, also known as endothelial PAS domain protein-1 (EPAS-1), shares 48% amino acid sequence identity with HIF-1α (9). As in the case of HIF-1α, continuous hypoxia leads to HIF-2α accumulation and subsequent dimerization with HIF-1β. Transcriptional activation by HIF-2 regulates some target genes in common with HIF-1 as well as other genes that are uniquely regulated by HIF-2 (10-12). Homozygous deficiency of HIF-2α is often lethal with the surviving Hif-2α -/− mice exhibiting multiple organ pathology and increased oxidative stress as a result of impaired induction of genes encoding major antioxidant enzymes (AOEs) (12).Hif-2α +/− mice, like Hif-1α +/− mice, are phenotypically indistinguishable from WT littermates under sedentary conditions, but exhibit impaired responses to chronic hypoxia such as impaired pulmonary vascular remodeling, erythropoiesis, and retinal neovascularization (13-15). Although the carotid body is a prominent site of HIF-2α expression (16), the effects of Hi...

Previous studies identified NADPH oxidases (Nox) and mitochondrial electron transport chain at complex I as major cellular sources of reactive oxygen species (ROS) mediating systemic and cellular responses to intermittent hypoxia (IH). In the present study, we investigated potential interactions between Nox and the mitochondrial complex I and assessed the contribution of mitochondrial ROS in IH-evoked elevation in blood pressure. IH treatment led to stimulus-dependent activation of Nox and inhibition of complex I activity in rat pheochromocytoma (PC)12 cells. After re-oxygenation, Nox activity returned to baseline values within 3 h, whereas the complex I activity remained downregulated even after 24 h. IH-induced complex I inhibition was prevented by Nox inhibitors, Nox2 but not Nox 4 siRNA, in cell cultures and was absent in gp91(phox-/Y) (Nox2 knock-out; KO) mice. Using pharmacological inhibitors, we show that ROS generated by Nox activation mobilizes Ca(2+) flux from the cytosol to mitochondria, leading to S-glutathionylation of 75- and 50-kDa proteins of the complex I and inhibition of complex I activity, which results in elevated mitochondrial ROS. Systemic administration of mito-tempol prevented the sustained but not the acute elevations of blood pressure in IH-treated rats, suggesting that mitochondrial-derived ROS contribute to sustained elevation of blood pressure.