Regulation of TASK potassium channels by G-protein coupled receptors
TASK potassium channels control the membrane potential in many cell types and thus affect a plethora of cellular functions such as excitability of neurons and cardiac muscle, and secretion of aldosterone in the adrenal gland. Although commonly termed ‘leak channels’, TASK channels are highly regu...
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|TASK potassium channels control the membrane potential in many cell types and thus affect
a plethora of cellular functions such as excitability of neurons and cardiac muscle, and secretion
of aldosterone in the adrenal gland. Although commonly termed ‘leak channels’, TASK channels
are highly regulated. Most importantly, they are strongly inhibited by a variety of hormones and
neurotransmitters activating Gq-protein coupled receptors (GqPCRs). Despite extensive studies
of TASK inhibition by the GqPCR-induced signaling cascade, the underlying mechanism of
channel regulation has not been elucidated. Thus I aimed to unravel the second messenger responsible
for GqPCR-mediated TASK channel inhibition and validate my findings from the heterologous
expression system in cerebellar granule neurons.
The signaling cascade induced by GqPCRs is initiated by activation of Gαq, which in turn
stimulates phospholipase Cβ to hydrolyze the membrane phospholipid phosphatidylinositol(
4,5)bisphosphate producing the second messengers 1,2-diacylglycerol (DAG) and inostol(
1,4,5)trisphosphate. Using different approaches, I first established that phospholipase C is
critical for GqPCR-mediated TASK channel inhibition. Next, I found that direct application of a
DAG analog was sufficient to inhibit TASK channels. Accordingly, experimental attenuation of
the DAG transients evoked by GqPCR stimulation diminished TASK channel inhibition, indicating
that DAG is responsible for the current reduction following receptor activation. Because
it had been previously established that a six amino acid motif within the proximal C-terminus is
important for TASK channel regulation by GqPCRs, I compared the effects of GqPCR stimulation
and DAG application on TASK channel proteins either truncated or mutated within this motif. A
correlation of the sensitivities towards DAG and GqPCR activation further supported the hypothesis
of DAG production as the underlying mechanism for the GqPCR-mediated effect. Lastly,
to test whether native TASK-mediated currents were also inhibited by DAG, I probed application
of this lipid on dissociated cerebellar granule neurons that express the TASK-mediated
standing outward potassium current (IKSO). IKSO was inhibited by muscarinic receptor agonist as
well as by direct application of DAG, producing a significant membrane depolarization.
In conclusion, my findings demonstrate that DAG mediates the GqPCR-induced inhibition of
TASK channels in an expression system as well as native, TASK-mediated currents. Thus, my
data expand the view on the signaling effects of the small membrane lipid DAG and establish a
link between DAG and cell excitability. Additionally, they may pave the way towards understanding
the mechanism of DAG action on ion channels as atypical DAG effector proteins.