Download A fatty acid in the MCT ketogenic diet for epilepsy treatment blocks

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A fatty acid in the MCT ketogenic diet for
epilepsy treatment blocks AMPA receptors
This scientific commentary refers to
‘Seizure control by decanoic acid
through direct AMPA receptor inhibition’, by Chang et al. (doi:10.1093/
brain/awv325).
Dietary therapies can provide control
of seizures in patients with drugrefractory epilepsy. There are several
types of dietary therapies, all of
which are high in fat, restrict carbohydrates to some extent, and are
associated with ketosis. In the classical ketogenic diet introduced into
clinical practice in the 1920s, the fat
component is provided by long-chain
triglycerides (LCTs). Recognizing that
medium chain triglycerides (MCTs)
are more ketogenic than LCTs,
Huttenlocher et al. (1971) created
the MCT oil diet, which permits
greater amounts of carbohydrate
and protein, and therefore allows a
more flexible meal plan. The efficacy
of the MCT oil ketogenic diet is
equivalent to that of the classic LCT
ketogenic diet, and the tolerability is
also comparable (Neal et al., 2009).
MCTs, which are abundant in coconut and palm kernel oil, have a glycerol backbone and three fatty acid
esters with 6 to 12 carbons arranged
in a straight chain. The major fatty
acids in MCT oil are n-octanoic
acid (C8; caprylic acid; 50–80%)
and n-decanoic acid (C10; capric
acid; 20–50%). Following ingestion,
MCTs are rapidly absorbed into the
portal circulation to the liver, in contrast to LCTs which are absorbed by
chylomicrons in the lymph. It was
previously believed that medium
chain fatty acids derived from
MCTs are immediately oxidized in
the liver to form ketone bodies, but
it is now known that appreciable
amounts appear in the circulation of
patients receiving the MCT diet (Sills
et al., 1986a). For example, children
on the MCT diet had plasma concentrations of decanoic acid in the range
of 0.1–0.2 mM. Decanoic acid readily
crosses the blood–brain barrier, probably by a combination of diffusion
and saturable carrier-mediated transport via a medium-chain fatty acid
transporter that also mediates brain
uptake of the antiseizure drug valproic acid, itself a medium chain
fatty acid (Adkison and Shen, 1996).
In this issue of Brain, Chang et al.
show for the first time that decanoic
acid is an antagonist of AMPA receptors, providing a possible basis for
the antiseizure activity of the MCT
diet (Chang et al., 2016).
A number of theories have been
advanced to explain the antiseizure
activity of high fat ketogenic diets
but none has as yet received broad
acceptance (Hartman et al., 2007).
Early on, it was proposed that
ketone bodies are responsible for seizure protection and indeed all of the
ketone bodies (acetone, b-hydroxybutyrate and acetoacetate) have been reported to have antiseizure effects in
one or another animal model.
However, as confirmed by Chang
et al. the ketones are not generally
active in in vitro seizure models.
Moreover, the activity profile of ketogenic diets in animal models does not
correspond with that of the ketones
and the degree of ketosis does not
correlate with the extent of seizure
protection. Other investigators have
concluded that the diets enhance
GABA synthesis, correcting deficits
in
inhibitory
neurotransmission.
More recent studies have implicated
increased production of brain bioenergetic substrates such as ATP, creatine and phosphocreatine and
increased mitochondrial energy metabolism, which is postulated to
Scientific Commentaries
BRAIN 2016: 139; 304–316
| 307
NH2
Aminoterminal
domain
Competitive
antagonists
D1
Ligand
binding
domain
Linker
Glutamate
D2
S2-M4
S1-M1
Extracellular side
M1
M3
Transmembrane
domain
Noncompetitive
antagonists
N
M4
N
O
N
M2
Perampanel
Cytoplasmic side
Decanoic acid
O
O–
HOOC
Figure 1 AMPA receptor protein structure illustrating putative locations of binding sites for decanoic acid and non-fatty acid
non-competitive antagonists such as perampanel. The AMPA receptor consists of four subunits, each of which is rendered in the structure
on the left in a different colour; one of the subunits is shown as a ribbon diagram. Small molecule binding sites are highlighted in the subunit
rendered in black lines; similar binding sites exist in the other identical subunits but are not highlighted. Non-competitive antagonists are believed
to bind to the S1-M1 linker (residues FLDPLAYE) and the S2-M4 linker; the S1-M1 residues are highlighted as light grey spheres. By molecular
modelling, Chang et al. determined that a likely site for binding of decanoic acid is to the M3 segment in the transmembrane domain; key residues
near the cytoplasmic side (PRSL_GR) are highlighted as dark grey spheres. Additional likely residues for binding of decanoic acid to the cytoplasmic sides of the M1 (LFLVS) and M2 (NS_WF) segments are not highlighted. Binding of decanoic acid at these sites could block the channel
pore, which is formed by the transmembrane domains of all four subunits. Evidence in favour of a pore-blocking mechanism is the demonstration
of increased block at depolarized membrane potentials, since the more positive electrostatic field at the cytoplasmic side of the membrane at
depolarized potentials would tend to more strongly attract into the pore the negatively charged fatty acid approaching the channel from the
extracellular solution. The dashed arrow indicates the axis of symmetry of the transmembrane domain and the path for ion flow. The structure is
3KG2 (homomeric rat GluA2 in closed antagonist bound state) from the RCSB Protein Data Bank as was used by Chang et al. The illustration on
the right schematically represents the various channel domains and the locations of the small molecule binding sites in a single subunit.
stabilize neuronal ion gradients,
thereby resisting depolarizing influences and consequent seizure generation. In addition, the increased ATP
has been proposed to enhance the
production of the inhibitory mediator
adenosine, which is well recognized to
have antiseizure properties. It has also
been noted that the diets enhance mobilization of polyunsaturated fatty
acids, which may protect against seizures through effects on ion channels.
In sum, many of the hypothesized
mechanisms of the dietary therapies
implicate effects on intermediary metabolism. This is in contrast to the
mechanisms of antiseizure drugs,
which—with a few exceptions—are
believed to act through direct effects
on ion channels, leading to reduced
neuronal excitation, or on components of the GABA system, leading
to
enhanced
GABA-mediated
inhibition.
Fast (millisecond timescale) synaptic
excitation in the brain is mediated by
ionotropic glutamate receptors (a
family of ligand-gated ion channels),
which are membrane-spanning protein complexes localized at synapses,
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| BRAIN 2016: 139; 304–316
usually
on
dendritic
spines.
Neurotransmitter glutamate released
from presynaptic excitatory terminals
diffuses across the synaptic cleft to activate ionotropic glutamate receptors
in the postsynaptic membrane, leading to entry of cations (Na + and in
some cases Ca2 + ), which generates
the depolarizing excitatory postsynaptic potential (EPSP). AMPA receptors,
which are assembled as heteromeric
or homomeric tetramers of the protein subunits GluA1, GluA2, GluA3
and GluA4 (each consisting of 900
amino acids), are responsible for the
bulk of the EPSP and there is a variable contribution of NMDA receptors. AMPA receptors are also key
mediators of pathological electrochemical events in focal epilepsies,
including the paroxysmal depolarization shift (the basis of the EEG spike)
and the electrographic seizure discharge. In addition to mediating synchronous discharges in epileptic foci,
AMPA receptors are critically important in the spread of seizure activity
locally and to distant brain regions.
Since the early 1980s, it has been
known that pharmacological inhibition of ionotropic glutamate receptors can protect against seizures in
experimental models. Early efforts to
develop antiseizure agents that inhibit
glutamate excitation focused on
NMDA receptor antagonists, but disappointing clinical results dampened
enthusiasm. There has recently been
a resurgence of interest in ionotropic
glutamate receptor antagonists with
the demonstration in clinical trials
that perampanel, a potent selective
orally-active non-competitive AMPA
receptor antagonist, is effective in
the treatment of focal seizures and
primary and secondarily generalized
tonic-clonic seizures.
Medium chain fatty acids, including
decanoic acid, have long been known
to have acute antiseizure actions in
animal models. Valproic acid is a
branched medium chain fatty acid
that was demonstrated to be effective
in clinical trials in the 1960s. The
mechanism of action of valproic acid
is obscure. Chang et al. now confirm
the antiseizure activity of decanoic
Scientific Commentaries
acid (1 mM) in in vitro rat hippocampal slice models, including a model
generated by exposure to the GABAA
receptor antagonist pentylenetetrazol
(PTZ) and another model in which
extracellular Mg2 + is reduced, leading
to unblock of NMDA receptors. At
concentrations within the range of
those present in the blood of patients
maintained on the MCT diet
(0.3 mM), Chang et al. further demonstrate that decanoic acid selectively
blocks excitatory synaptic currents
without affecting inhibitory synaptic
currents. These results are then followed by the key observation of their
study, which is that decanoic acid
causes a selective non-competitive inhibition of recombinant AMPA receptors expressed in Xenopus oocytes, but
with variable potency depending upon
the subunit combination. The effect of
decanoic acid was selective in that neither valproic acid nor octanoic acid
significantly affected AMPA receptor
currents at a concentration of 1 mM.
The authors argue that perampanellike effects of decanoic acid are responsible for the antiseizure actions
of the MCT diet and that it may not
be necessary to invoke any other
mechanism.
How reasonable is this proposal? If
the antiseizure activity of the MCT diet
is entirely due to block of AMPA receptors, then the diet could be replaced
by perampanel or another AMPA receptor antagonist. There are no headto-head comparisons of perampanel
with other antiseizure agents or with
the MCT diet. Nevertheless, it is
believed by many epileptologists that
ketogenic diets, including the MCT
diet, are often effective in patients
who do not respond to antiseizure
drugs (Sills et al., 1986b). These physicians would be surprised if the diet was
no more efficacious than an antiseizure
drug. The present study suggests that a
comparative trial between the MCT
diet and an AMPA receptor antagonist
would be worthwhile. If it were possible to replace the diet with an
AMPA receptor antagonist, this would
enormously simplify therapy, avoid the
poor palatability and gastrointestinal
side effects of the diet, and therefore
make treatment available to a broader
group of patients.
A randomized trial comparing the
LCT and MCT ketogenic diets in children with drug-refractory epilepsy
failed to find any difference in the
number of children who responded to
the diet with a reduction in seizures or
who achieved seizure freedom, as
occurred in a small number of subjects
(Neal et al., 2009). This observation is
difficult to reconcile with the idea that
the MCT diet has a unique mechanism
of action. It seems more plausible that
the LCT and MCT diets act through a
similar set of mechanisms, perhaps with
the addition of AMPA receptor blockade by decanoic acid in the case of the
MCT diet. There are other observations
in the literature that raise further questions. In animal models, decanoic acid
does elevate the seizure threshold in
some situations but the effect is
modest (minimal effective dose is
1723 mg/kg, orally) and there is no
effect on PTZ-induced seizures (Wlaź
et al., 2015). In contrast, perampanel
has potent, broad-spectrum antiseizure
activity in diverse models including the
PTZ model (ED50, 0.94 mg/kg, orally).
Octanoic acid, a quantitatively greater
constituent of MCT oil than decanoic
acid, did not affect epileptiform discharges in vitro at a concentration of
1 mM (Chang et al., 2013) and also
failed to affect AMPA receptor currents
in Xenopus oocytes (Chang et al.,
2016). However, octanoic acid has
long been known to have sedative
and antiseizure activities (Liu and
Pollack, 1994) and a recent study indicates that it is at least as potent as
decanoic acid (Wlaź et al., 2012).
Surprisingly, octanoic acid does raise
the PTZ threshold (Wlaź et al., 2012).
Understanding the antiseizure action of
other medium chain fatty acids including valproic acid represents a major scientific challenge. In sum, Chang et al.
for the first time invoke AMPA receptors as a potential molecular target in a
dietary therapy for epilepsy. AMPA receptors are a clinically validated antiseizure drug target, thus it is plausible that
an action on AMPA receptors could
contribute to the activity of the MCT
diet in epilepsy therapy. Whether this is
Scientific Commentaries
BRAIN 2016: 139; 304–316
| 309
Glossary
AMPA receptor: Ligand-gated non-selective cation channel that allows the flow of Na + , K + and in some cases Ca2 + in response to binding of
glutamate; ligands, such as glutamate, that gate (open) the channel are referred to as agonists.
Channel block: Functional inhibition of an ion channel by a blocking drug that binds inside the ion conduction pore preventing flow of ions
through the channel.
Heteromeric receptor/homomeric receptor: Heteromeric applies when at least one subunit is non-identical to the others; homomeric
applies when all subunits are identical.
Ketone body: Three molecules (acetone, b-hydroxybutyrate and acetoacetate) produced in the liver during the metabolism of fats that serve as
an alternative source of energy to glucose.
MCT diet: A dietary treatment for epilepsy more commonly used in Europe than in the United States that is high in fats containing predominantly medium chain triglycerides; causes ketosis, a state where there are elevated concentrations of ketone bodies in the blood.
Non-competitive antagonist: In the case of ionotropic glutamate receptors, an inhibitor whose blocking action is not reduced progressively as
the agonist (glutamate) concentration is increased as occurs for competitive antagonists. Non-competitive antagonists allosterically inhibit the
channel by binding to a site other than the ligand binding domain.
Triglyceride: The storage form of fatty acids in the body; specifically, an ester derived from glycerol and three fatty acids, such as decanoic acid
(capric acid) triglyceride, which is composed of glycerol and three decanoic acid molecules covalently linked.
the only mechanism of the MCT diet
requires further investigation.
Michael A. Rogawski
Department of Neurology,
School of Medicine, University of
California, Davis Sacramento,
California, USA
E-mail: [email protected]
doi:10.1093/brain/awv369
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When is temporal lobe epilepsy not temporal
lobe epilepsy?
This scientific commentary refers to
‘Temporal plus epilepsy is a major
determinant of temporal lobe surgery
failures’, by Barba et al. (doi:10.1093/
brain/awv372).
Stereotactic
electroencephalography
(SEEG) originated in France in the
1950s. Bancaud and Talairach, at St.
Anne’s Hospital in Paris, introduced
this approach to define the 3D extent
of dysfunctional brain tissue surrounding intraparenchymal brain tumours,
and more specifically, to define epileptogenic brain tissue in patients with
pharmacoresistant epileptic seizures
(Bancaud et al., 1975). The approach
required a hypothesis that would
enable the placement of multiple,
mostly unilateral, depth electrodes to
sample a reasonably limited number
of regions of interest. The results
then guided a tailored surgical resection. This technique was adapted by
Crandall et al. (1963) at UCLA for
patients with suspected mesial temporal
lobe
epilepsy.
Crandall,