AMP-activated Protein Kinase Mediates Carotid Body Excitation by Hypoxia

Christopher N Wyatt,Kirsty J Mustard,Selina A Pearson,Mark L Dallas,Lucy Atkinson,Prem Kumar,Chris Peers,D Grahame Hardie,A Mark Evans

doi:10.1074/jbc.m608742200

Abstract

Early detection of an O2 deficit in the bloodstream is essential to initiate corrective changes in the breathing pattern of mammals. Carotid bodies serve an essential role in this respect; their type I cells depolarize when O2 levels fall, causing voltage-gated Ca2+ entry. Subsequent neurosecretion elicits increased afferent chemosensory fiber discharge to induce appropriate changes in respiratory function (1). Although depolarization of type I cells by hypoxia is known to arise from K+ channel inhibition, the identity of the signaling pathway has been contested, and the coupling mechanism is unknown (2). We tested the hypothesis that AMP-activated protein kinase (AMPK) is the effector of hypoxic chemotransduction. AMPK is co-localized at the plasma membrane of type I cells with O2-sensitive K+ channels. In isolated type I cells, activation of AMPK using 5-aminoimidazole-4-carboxamide riboside (AICAR) inhibited O2-sensitive K+ currents (carried by large conductance Ca2+-activated (BKCa) channels and TASK (tandem pore, acid-sensing potassium channel)-like channels, leading to plasma membrane depolarization, Ca2+ influx, and increased chemosensory fiber discharge. Conversely, the AMPK antagonist compound C reversed the effects of hypoxia and AICAR on type I cell and carotid body activation. These results suggest that AMPK activation is both sufficient and necessary for the effects of hypoxia. Furthermore, AMPK activation inhibited currents carried by recombinant BKCa channels, whereas purified AMPK phosphorylated thealpha subunit of the channel in immunoprecipitates, an effect that was stimulated by AMP and inhibited by compound C. Our findings demonstrate a central role for AMPK in stimulus-response coupling by hypoxia and identify for the first time a link between metabolic stress and ion channel regulation in an O2-sensing system.

Highlights

Is necessary to understand the homeostatic mechanisms that monitor O2 supply to the body and elicit corrective changes in respiratory and circulatory function to maintain O2 levels
BKCa channel currents were selectively inhibited in acutely isolated type I cells by 5-aminoimidazole-4-carboxamide riboside (AICAR), a nucleoside that is taken up by cells and metabolized to the AMP mimetic, ZMP, activating AMPK in the absence of ATP depletion [22]
AICAR is not known to activate any protein kinases other than AMPK, so the finding that compound C reverses carotid body type I cell activation induced by AICAR, but not that induced by depolarization, provides strong support for the view that its effects are mediated by inhibition of the AMPK pathway

Summary

Introduction

Is necessary to understand the homeostatic mechanisms that monitor O2 supply to the body and elicit corrective changes in respiratory and circulatory function to maintain O2 levels. Consistent with the hypothesis that AMPK serves a key role in O2 sensing by the carotid body, we found that in isolated carotid body type I cells, the AMPK␣1 catalytic subunit isoform and O2-sensitive BKCa channels co-localize at the plasma membrane (Fig. 1A, panels i–iv).

Results

Conclusion