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Title: AMPK and the hypoxic ventilatory response : signal integration at an oxygen-sensing nucleus within the brainstem respiratory network?
Author: MacMillan, Sandy
ISNI:       0000 0004 7969 2619
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2019
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The capacity for mammals to regulate breathing is critical to match ventilation to changes in oxygen availability and demand, for example during sleep or ascent to altitude. Acute falls in arterial PO2 activate specialised oxygen-sensing chemoreceptors, which relay this information to the cardiorespiratory centre of the brainstem, which then acts to restore blood gas homeostasis. While the role of the carotid bodies as peripheral chemoreceptors is undisputed, it has been proposed that a central oxygen sensor may exist. However, this view remains controversial. The work in this thesis built on recent findings which showed that the AMP-activated protein kinase (AMPK), a ubiquitously expressed cellular energy sensor, within catecholaminergic (tyrosine hydroxylase (TH)-expressing) cells is critical to the proper regulation of breathing during acute hypoxia. It was found that targeted deletion of AMPK within this subpopulation of respiratory neurons of the mouse brainstem led to decreases in ventilation and more frequent and prolonged apnoeas. The fact that this ventilatory deficit was hypoxia-specific and rescued by addition of hypercapnia showed that the capacity for neuronal activation was not affected per se, because central and peripheral chemosensors contributing to the hypercapnic response also express TH. Importantly, the activity of the carotid bodies in response to hypoxia remained fully intact, indicating that the respiratory deficits originated centrally. In light of these findings, the experiments carried out in this thesis further investigated how the deletion of both AMPK catalytic subunits in catecholaminergic cells (TH AMPK-α1/α2 dKO) impacts on hypoxic ventilation and where within the brainstem neuronal activation may be affected. By using whole-body plethysmography, I show that AMPK within the brainstem respiratory network is not only critical for appropriate ventilatory adjustments during acute hypoxia, but also during prolonged periods of low oxygen availability. Following the severe hypoventilation observed at the onset of hypoxia, ventilation remained attenuated relative to controls, even during periods lasting up to an hour. Moreover, some TH AMPK-α1/α2 dKO mice lost all respiratory rhythmogenesis after 20-30min of hypoxia. In those mice where respiratory rhythmogenesis was retained, it was found to be interrupted at regular intervals by high frequency periods of spontaneous apnoeas. Immunohistochemical analyses of the brainstems of TH AMPK-α1/α2 dKO mice revealed a comparable number of catecholaminergic cells relative to controls, supporting the conclusion of a functional deficit within these neurons rather than hypo- or hyperplasia. This was further corroborated by the identification of a specific area within the brainstem that showed significant reductions in immediate early gene expression (cFos) and thus neuronal activation during hypoxia. Importantly, cardiovascular responses to hypoxia were unaltered in TH AMPK-α1/α2 dKO mice relative to controls, indicating that this identified brainstem nucleus may separately control the respiratory and the cardiovascular responses to hypoxia through AMPK-dependent pathways. Finally, directing AMPK deletion to the adrenergic subset of catecholaminergic neurons revealed that the ventilatory deficit originated within the noradrenergic cells of the caudal brainstem, consistent with the location of the identified area of cFos deficiency. In summary, AMPK-dependent signalling is required within a specific group of noradrenergic neurons within the caudal brainstem to ensure appropriate signal integration and transduction during hypoxia in order to appropriately regulate ventilatory adaptations and protect from hypoventilation and respiratory instability. Therefore, AMPK may be a potential new therapeutic target to protect from sleep-disordered breathing associated with metabolic syndrome-related disorders and ascent to altitude.
Supervisor: Evans, Anthony ; Livingstone, Dawn ; Larkman, Philip Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: carotid bodies ; hypoxia ; AMP-activated protein kinase ; AMPK ; AMPK deficiency ; ventilatory adjustments