Proceedings of The Physiological Society

Kings College London (2005) J Physiol 565P, C170

Communications

Regulation of a TASK-like potassium channel in rat arterial chemoreceptor cells by ATP

Varas, Rodrigo ; Buckler, Keith J;

1. University Laboratory of Physiology, Oxford, United Kingdom.


Carotid body type I cells respond to hypoxia with membrane depolarisation, voltage-gated calcium entry and neurosecretion. The inhibition of TASK-like background potassium channels by hypoxia plays a key role in initiating this response (Buckler et al. 2000). The mechanism by which hypoxia modulates TASK-like channel activity is however unknown. We have previously reported that TASK-like K-channel activity is also markedly reduced by inhibitors of mitochondrial energy metabolism (Williams & Buckler, 2004) suggesting that cytosolic ATP may play a role in regulating channel activity. We have therefore investigated the effects of ATP upon background K-channel activity. Type I cells were obtained as previously described (Buckler et al. 2000). Briefly, carotid bodies were excised from 19 anaesthetised (4% halothane) 11- to 15-day-old Sprague-Dawley rat pups and disassociated by collagenase-trypsin digestion. All animals were killed by decapitation whilst anaesthetised. Isolated cells were maintained in HAMs F-12 culture media under 5 % CO2 in air at 37 °C on glass coverslips coated with poly-D-lysine and used within 2-6 h. Channel activity was recorded in excised inside out patches taken from type-1 cells at a membrane potential of -70 mV and is reported as NPo values ((N = number of channels, Po= open probability, mean ± SEM). Upon patch excision channel activity ran down abruptly. Following rundown, application of ATP (0.1-20 mM) to the cytosolic side of the membrane patch resulted in a dose dependent activation of channel activity. Responses to ATP were apparent at concentrations as low as 0.5 mM (control = 0.02 ± 0.01, 0.5 mM ATP = 0.03 ± 0.01, p <0.05, paired t test, n=8) and appeared to reach a maximum at 10 mM where an 8-fold increase in NPo was observed (0.17 ± 0.05, p <0.01, n = 10). The dose response was adequately described by a Michaelis-Menten function with an EC50 of 2.3 mM (R2= 0.99). The non-hydrolysable ATP analogue AMP-PCP (10 mM) also increased channel activity (control = 0.02 ± 0.004, AMP-PCP = 0.08 ± 0.02, p<0.01, n = 8) suggesting that ATP sensitivity may be mediated through nucleotide binding to some form of regulatory domain rather than through phosphorylation. In conclusion, the TASK-like oxygen sensitive K-channel of rat carotid body type-1 cells has the potential to respond to changes in cytosolic ATP levels within the physiological range. Thus direct ATP sensing by the channel, or associated regulatory protein, could account for the potent stimulatory effects of metabolic poisons on chemoreceptor activity. These data are also consistent with the metabolic theory of oxygen sensing wherein oxygen is indirectly sensed through changes in mitochondrial ATP synthesis.

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