How do general anaesthetics cause loss of consciousness?
anaesthetics have been used for over 100 years and continue to be used daily
for surgeries around the world. But how do they induce changes to
consciousness? This article explores the research that suggests the
anaesthetics chloroform and isoflurane disrupt the organisation of lipid rafts
within cellular membranes to cause downstream loss of consciousness.
The first record of inhaled general anaesthetics being used in
surgery occurred in 1846 when William Morton used diethyl ether to anaesthetise
a patient.1 Since then, the use of inhaled anaesthetics has
become widespread, with thousands of surgeries now using them every day.
However, despite speculation that they somehow impacted cellular membranes to
activate TWIK-related potassium (TREK) channels and reversibly induce loss of
consciousness, the precise mechanism through which this chemically diverse
group of hydrophobic molecules acts has been unknown.
It has been reported that
lipophilicity – the ability of a drug to distribute or dissolve into fats – is
a major indicator of potency for general anaesthetics. This property is known
as the Meyer-Overton correlation2 and has led researchers to
investigate whether general anaesthetics may perturb the organisation of lipid
rafts within cellular membranes to cause a patient to become unconscious.
Lipid rafts are a region of ordered fats within the cellular
membrane, in which certain proteins, receptors or channels may be isolated. To
establish whether general anaesthetics could disrupt these organised regions,
or their associations with other components in the cell membrane, researchers
from The Scripps Research Institute in the US studied the effect of chloroform
and isoflurane on monosialotetrahexosylganglioside1 (GM1) lipid rafts.3 GM1 is a well characterised type of lipid raft,
composed of cholesterol and saturated fats such as sphingomyelin.
Pavel et al. hypothesised that anaesthetics could indirectly
activate TREK-1 potassium channels by disrupting GM1 rafts. The disruption
would activate the phospholipase D2 (PLD2) enzyme localised to GM1 rafts and
allow it to colocalise to rafts containing phosphatidylinositol 4,5-bisphosphate
(PIP2). The resultant interaction between PLD2 and PIP2 would
facilitate the production of phosphatidic acid (PA), which could then activate
TREK-1 potassium channels, causing the channels to release potassium. The
result would be hyperpolarised neurons that have limited ability to fire and
thus a loss of consciousness.
Techniques and findings
The researchers used direct stochastical optical reconstruction
microscopy (dSTORM) and assays to establish how general anaesthetics caused
TREK-1 channel activation.
In previous work, they had demonstrated that PLD2 activates
TREK-1 by binding to a disordered loop in the channel’s C terminus. To test if
the anaesthetic sensitivity of TREK-1 is through PLD2, they blocked the
enzyme’s catalytic activity and observed the effect.
The team overexpressed a mutant TREK-1 channel (K758R PLD2
mutant [xPLD2]) which expresses a PLD2 enzyme that is catalytically inactive
and so cannot produce PA. When treated with chloroform, these xPLD2 TREK-1
channels did not open, nor was there any detectable current as a result of
administering the chloroform anaesthetic, even at high concentrations. This led
the researchers to conclude that aesthetics activate TREK-1 channels in
cultured cells almost entirely through a PLD2-dependent process.
Moreover, the researchers demonstrated that PLD2 localisation
and the production of PA could render an anaesthetic-insensitive homologue to
the TREK-1 channel (TWIK-related arachidonic acid-stimulated K+ [TRAAK]
channel) sensitive to chloroform. The authors also suggest that direct binding
of the anaesthetic to the transmembrane domain of channels alone is
insufficient to reversibly induce unconsciousness. They found that PA
production was required for this sensitivity in both modified TRAAK and
endogenous TREK-1 channels.
They further confirmed their
hypothesis that disrupting PLD2 localisation leads to TREK-1 activation using
dSTORM to monitor the translocation of fluorescently-tagged PLD2 away from GM1
lipid rafts. In N2A cells (which naturally express both PLD2 and TREK-1
channels), administration of chloroform or isoflurane caused PLD2 to
translocate from GM1 rafts. Interestingly, the team observed that the
disruption caused by inhaled anaesthetics was similar to when cholesterol had
been removed from the rafts; they concluded as a result, that anaesthetics inhibit
the ability of cholesterol to localise PLD2 to GM1 rafts.
In whole brain studies of
chloroform-treated fruit flies, the authors identified that PLD2-mediated
modulation of PA production shifted the anaesthetic threshold in animals. In
more recent studies, PA has emerged as a signalling lipid that can regulate ion
channels and thus modulate neuronal excitability and pain signalling in vivo.
From these findings, the team concluded that PLD2 signalling (and therefore
inhaled anaesthetics) contribute to the central regulation of neuronal
They also explained that
the mechanism of action they propose for inhaled anaesthetics “relies on a
non-uniform (heterogeneous) distribution of lipids in the membrane” and that
decreasing the longevity of GM1 rafts is likely to shift the equilibrium of
PLD2 distribution, such that PLD2 interacts more with PIP2.
The three major findings of the study were:
anaesthetics do not directly activate TREK-1 channels to
induce a loss of consciousness
instead, anaesthetics inhibit the ability of cholesterol
to localise PLD2 to GM1 rafts
however, PLD2 translocation alone is not sufficient to
activate TREK-1 channels; the increase in PA as a result of PLD2’s interaction
with PIP2-containing lipid rafts is required for this.
Overall, the researchers suggest that the
mechanism of action for inhaled anaesthetics chloroform and isoflurane is to
activate TREK-1 channels indirectly by disrupting PLD2 localisation to GM1
lipid rafts. The translocalisation of PLD2 to rafts containing PIP2 and
the resultant increase in production of the signalling lipid PA creates a local
concentration of PA within neurons sufficient to activate TREK-1 channels to
release potassium. As a result, neurons become hyperpolarised, are unable to
fire and this causes the temporary loss of consciousness associated with anaesthetics.