Reflexes from the carotid body have been implicated in cardiorespiratory disorders associated with chronic intermittent hypoxia (CIH). To investigate whether CIH causes functional and/or structural plasticity in the carotid body, rats were subjected to 10 days of recurrent hypoxia or normoxia. Acute exposures to 10 episodes of hypoxia evoked long-term facilitation (LTF) of carotid body sensory activity in CIH-conditioned but not in control animals. The magnitude of sensory LTF depended on the length of CIH conditioning and was completely reversible and unique to CIH, because conditioning with a comparable duration of sustained hypoxia was ineffective. Histological analysis revealed no differences in carotid body morphology between control and CIH animals. Previous treatment with superoxide anion ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} \begin{equation*}{\mathrm{O}}_{2}^{{\cdot}-}\end{equation*}\end{document} ) scavenger prevented sensory LTF. In the CIH-conditioned animals, carotid body aconitase enzyme activity decreased compared with controls. These observations suggest that increased generation of reactive oxygen species contribute to sensory LTF. In CIH animals, carotid body complex I activity of the mitochondrial electron transport is inhibited, suggesting mitochondria as one source of \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} \begin{equation*}{\mathrm{O}}_{2}^{{\cdot}-}\end{equation*}\end{document} generation. These observations demonstrate that CIH induces a previously uncharacterized form of reactive oxygen species-dependent, reversible, functional plasticity in carotid body sensory activity. The sensory LTF may contribute to persistent reflex activation of sympathetic nerve activity and blood pressure in recurrent apnea patients experiencing CIH.

Gaseous messengers, nitric oxide and carbon monoxide, have been implicated in O ₂ sensing by the carotid body, a sensory organ that monitors arterial blood O ₂ levels and stimulates breathing in response to hypoxia. We now show that hydrogen sulfide (H ₂ S) is a physiologic gasotransmitter of the carotid body, enhancing its sensory response to hypoxia. Glomus cells, the site of O ₂ sensing in the carotid body, express cystathionine γ-lyase (CSE), an H ₂ S-generating enzyme, with hypoxia increasing H ₂ S generation in a stimulus-dependent manner. Mice with genetic deletion of CSE display severely impaired carotid body response and ventilatory stimulation to hypoxia, as well as a loss of hypoxia-evoked H ₂ S generation. Pharmacologic inhibition of CSE elicits a similar phenotype in mice and rats. Hypoxia-evoked H ₂ S generation in the carotid body seems to require interaction of CSE with hemeoxygenase-2, which generates carbon monoxide. CSE is also expressed in neonatal adrenal medullary chromaffin cells of rats and mice whose hypoxia-evoked catecholamine secretion is greatly attenuated by CSE inhibitors and in CSE knockout mice.

Chronic intermittent hypoxia (CIH) occurs in patients with sleep apnoea and has adverse effects on multiple physiological functions. Previous studies have shown that reflexes arising from carotid bodies mediate CIH‐evoked cardio‐respiratory responses, and reactive oxygen species (ROS) play important roles in eliciting systemic responses to CIH. Very little is known about the molecular mechanisms underlying CIH. The transcriptional activator hypoxia‐inducible factor‐1 (HIF‐1) mediates a broad range of cellular and systemic responses to hypoxia, and HIF‐1 is activated in cell cultures exposed to IH. In the present study we examined whether CIH activates HIF‐1 and if so whether it contributes to cardio‐respiratory responses and ROS generation in mice. Experiments were performed on male littermate wild‐type (WT) and heterozygous (HET) mice partially deficient in HIF‐1α, the O₂ regulated subunit of the HIF‐1 complex. Both groups of mice were exposed to either 10 days of CIH (15 s of hypoxia followed by 5 min of normoxia, 9 episodes h⁻¹, 8 h day⁻¹) or to 10 days of 21% O₂ (controls). Carotid body response to hypoxia was augmented, and acute intermittent hypoxia (AIH) induced sensory long‐term facilitation (sLTF) of the chemoreceptor activity in CIH‐exposed WT mice. In striking contrast, hypoxic sensory response was unaffected and AIH was ineffective in eliciting sLTF in CIH‐exposed HET mice. Analysis of cardio‐respiratory responses in CIH‐exposed WT mice revealed augmented hypoxic ventilatory response, LTF of breathing, elevated blood pressures and increased plasma noradrenaline. In striking contrast these responses were either absent or attenuated in HET mice exposed to CIH. In CIH‐exposed WT mice, ROS were elevated and this response was absent in HET mice. Manganese (III) tetrakis(1‐methyl‐4‐pyridyl) porphyrin pentachloride, a potent scavenger of superoxide, not only prevented CIH‐induced increases in ROS but also CIH‐evoked HIF‐1α up‐regulation in WT mice. These results indicate that: (a) HIF‐1 activation is critical for eliciting CIH‐induced carotid body‐mediated cardio‐respiratory responses; (b) CIH increases ROS; and (c) the effects of CIH involve complex positive interactions between HIF‐1 and ROS.

To investigate whether the transcriptional activator hypoxia-inducible factor 1 (HIF-1) is required for ventilatory responses to hypoxia, we analyzed mice that were either wild type or heterozygous for a loss-of-function (knockout) allele at the Hif1a locus, which encodes the O ₂ -regulated HIF-1α subunit. Although the ventilatory response to acute hypoxia was not impaired in Hif1a ^+/− mice, the response was primarily mediated via vagal afferents, whereas in wild-type mice, carotid body chemoreceptors played a predominant role. When carotid bodies isolated from wild-type mice were exposed to either cyanide or hypoxia, a marked increase in sinus nerve activity was recorded, whereas carotid bodies from Hif1a ^+/− mice responded to cyanide but not to hypoxia. Histologic analysis revealed no abnormalities of carotid body morphology in Hif1a ^+/− mice. Wild-type mice exposed to hypoxia for 3 days manifested an augmented ventilatory response to a subsequent acute hypoxic challenge. In contrast, prior chronic hypoxia resulted in a diminished ventilatory response to acute hypoxia in Hif1a ^+/− mice. Thus partial HIF-1α deficiency has a dramatic effect on carotid body neural activity and ventilatory adaptation to chronic hypoxia.

Reflexes arising from the carotid bodies may play an important role in cardiorespiratory changes evoked by chronic intermittent hypoxia (CIH). In the present study, we examined whether CIH affects the hypoxic sensing ability of the carotid bodies and, if so, by what mechanisms. Experiments were performed on adult male rats (Sprague-Dawley, 250–300 g) exposed to two paradigms of CIH for 10 days: 1) multiple exposures to short durations of intermittent hypoxia per day (SDIH; 15sof5%O₂ + 5 min of 21% O₂, 9 episodes/h, 8 h/day) and 2) single exposure to longer durations of intermittent hypoxia per day [LDIH; 4 h of hypobaric hypoxia (0.4 atm/day) + 20 h of normoxia]. Carotid body sensory response to graded isocapnic hypoxia was examined in both groups of animals under anesthetized conditions. Hypoxic sensory response was significantly enhanced in SDIH but not in LDIH animals. Similar enhancement in hypoxic sensory response was also elicited in ex vivo carotid bodies from SDIH animals, suggesting that the effects were not secondary to cardiovascular changes. SDIH, however, had no significant effect on the hypercapnic sensory response. The effects of SDIH on the hypoxic sensory response completely reversed after SDIH animals were placed in a normoxic environment for an additional 10 days. Previous treatment with systemic administration of [ ] radical scavenger prevented SDIH-induced augmentation of the hypoxic sensory response. These results demonstrate that SDIH but not LDIH results in selective augmentation of the hypoxic response of the carotid body and [ ] radicals play an important role in SDIH-induced sensitization of the carotid body.

Intermittent hypoxia (IH) occurs in many pathological conditions including recurrent apneas. Hypoxia-inducible factors (HIFs) 1 and 2 mediate transcriptional responses to low O ₂ . A previous study showed that HIF-1 mediates some of the IH-evoked physiological responses. Because HIF-2α is an orthologue of HIF-1α, we examined the effects of IH on HIF-2α, the O ₂ -regulated subunit expression, in pheochromocytoma 12 cell cultures. In contrast to the up-regulation of HIF-1α, HIF-2α was down-regulated by IH. Similar down-regulation of HIF-2α was also seen in carotid bodies and adrenal medullae from IH-exposed rats. Inhibitors of calpain proteases (ALLM, ALLN) prevented IH-evoked degradation of HIF-2α whereas inhibitors of prolyl hydroxylases or proteosome were ineffective. IH activated calpain proteases and down-regulated the endogenous calpain inhibitor calpastatin. IH-evoked HIF-2α degradation led to inhibition of SOD2 transcription, resulting in oxidative stress. Over-expression of transcriptionally active HIF-2α prevented IH-evoked oxidative stress and restored SOD2 activity. Systemic treatment of IH-exposed rats with ALLM rescued HIF-2α degradation and restored SOD2 activity, thereby preventing oxidative stress and hypertension. These observations demonstrate that, unlike continuous hypoxia, IH leads to down-regulation of HIF-2α via a calpain-dependent signaling pathway and results in oxidative stress as well as autonomic morbidities.