Christopher N. Wyatt, Ph.D. (Advisor); Eric S. Bennett, Ph.D. (Committee Member); Paula A. Bubulya, Ph.D. (Committee Member); Kathy Engisch, Ph.D. (Committee Member); Robert M. Lober, M.D., Ph.D. (Committee Member)
Doctor of Philosophy (PhD)
The carotid bodies (CB) are peripheral chemoreceptors that detect changes in arterial oxygenation and, via afferent inputs to the brainstem, correct the pattern of breathing to restore blood gas homeostasis. Elucidating the “signal” that couples carotid body sensory type I cell (CBSC) hypoxic mitochondrial inhibition with potassium channel closure has proven to be an arduous task; to date, a multitude of oxygen-sensing chemotransduction mechanisms have been described and altercated (Varas, Wyatt & Buckler, 2007; Gao et al, 2017; Rakoczy & Wyatt, 2018). Herein, we provide preliminary evidence supporting a novel oxygen-sensing hypothesis suggesting CBSC hypoxic chemotransductive signaling may in part be mediated by mitochondria-generated thermal transients in TASK-channel-containing microdomains. Confocal microscopy measured distances between antibody-labeled mitochondria and TASK-potassium channels in primary rat CBSCs. Sub-micron distance measurements (TASK-1: 0.33 ± 0.04µm, n = 47 vs. TASK-3: 0.32 ± 0.03µm, n = 54) provided the first direct evidence for CBSC oxygen-sensing microdomains. Using a temperature-sensitive dye (ERthermAC), hypoxic-inhibition of mitochondrial oxidative phosphorylation in CBSCs was suggested to cause a rapid and reversible inhibition of mitochondrial thermogenesis and thus temperature in these microdomains. Whole-cell perforated-patch current-clamp electrophysiological recordings demonstrated CBSC sensitivity of resting-Vm to temperature: lowering bath temperature from 37°C to 24°C induced consistent and reversible depolarizations (Vm at 37°C: -48.4 ± 4.11mV vs. Vm 24°C: -31.0 ± 5.69mV; n = 5; p<0.01) in isolated, primary rat CBSCs. We propose that hypoxic inhibition of mitochondrial thermogenesis may play a critical role in hypoxic chemotransduction in the carotid body. A reduction in temperature within cellular microdomains will inhibit plasma membrane ion channels, influence the balance of cellular phosphorylation–dephosphorylation, and extend the half-life of reactive oxygen species. Furthermore, characterizing a thermosensory mechanism, that may also be used by other oxygen-sensitive cell types, would identify therapeutic targets for alleviating a host of respiratory disorders. Thus, a consideration of sub-cellular temperature gradients is critical if we are to fully understand how CBSCs, and potentially other oxygen-sensitive cells, respond to hypoxia.
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