Bough2007 Anticonvulsant Mechanisms of The Ketogenic Diet

Epilepsia, 48(1):4358, 2007
Blackwell Publishing, Inc.

C 2007 International League Against Epilepsy
Anticonvulsant Mechanisms of the Ketogenic Diet

Kristopher J. Bough and Jong M. Rho
Center for Drug Evaluation & Research, Food and Drug Administration, Rockville, Maryland;
Barrow Neurological Institute, Phoenix, Arizona, U.S.A.
Summary: The ketogenic diet (KD) is a broadly effective treatment for medically refractory epilepsy. Despite nearly a century
of use, the mechanisms underlying its clinical efficacy remain
unknown. In this review, we present one intersecting view of how
the KD may exert its anticonvulsant activity against the backdrop
of several seemingly disparate mechanistic theories. We summarize key insights gleaned from experimental and clinical studies
of the KD, and focus particular attention on the role that ketone
bodies, fatty acids, and limited glucose may play in seizure control. Chronic ketosis is anticipated to modify the tricarboxcylic
acid cycle to increase GABA synthesis in brain, limit reactive
oxygen species (ROS) generation, and boost energy production
in brain tissue. Among several direct neuro-inhibitory actions,
polyunsaturated fatty acids increased after KD induce the expression of neuronal uncoupling proteins (UCPs), a collective
up-regulation of numerous energy metabolism genes, and mitochondrial biogenesis. These effects further limit ROS generation
and increase energy production. As a result of limited glucose
and enhanced oxidative phosphorylation, reduced glycolytic flux
is hypothesized to activate metabolic KATP channels and hyperpolarize neurons and/or glia. Although it is unlikely that a single
mechanism, however well substantiated, will explain all of the
diets clinical benefits, these diverse, coordinated changes seem
poised to stabilize synaptic function and increase the resistance
to seizures throughout the brain. Key Words: Ketogenic diet
EpilepsyMetabolismPolyunsaturated fatty acids.
The ketogenic diet (KD) is a high-fat, low-protein, lowcarbohydrate diet that has been employed as a treatment
for medically refractory epilepsy for 86 years. The classic KD is based upon consumption of long-chain saturated triglycerides (LCTs) in a 3:14:1 ketogenic diet ratio
(KD ratio) of fats to carbohydrates + protein (by weight).
The vast majority of calories (>90%) are derived from fat.
While clinical implementation of the KD has varied from
center to center (Kossoff and McGrogan, 2005), diet treatment generally begins with a period of fasting followed by
gradual increase in calories to a target KD ratio of 3:14:1.
This is conducted in the inpatient setting over the course
of several days, where blood glucose, urine ketones, and
several other metabolic variables are closely monitored.
The hallmark feature of KD treatment is the production of
ketone bodies by the liver. Ketone bodies provide an alternative substrate to glucose for energy utilization, and in
developing brain, also constitute essential building blocks
for biosynthesis of cell membranes and lipids.
While the clinical effectiveness of the KD is widely
accepted, surprisingly little is understood about its under-
lying mechanisms of action. Although some studies suggest that dietary constituents or metabolites have direct
anticonvulsant effects, emerging evidence indicates that
adaptations to chronic administration of the KD result in
improved seizure control. These data suggest that the KD
activates several endogenous metabolic and genetic programs to stabilize and/or enhance cellular metabolism,
and that these fundamental changes help counter neuronal
dysfunction associated with seizure activity.
MECHANISTIC INSIGHTS FROM STUDIES
OF KD EFFICACY
The anticonvulsant efficacy of the KD has been examined in various acute and chronic animal models of
epilepsy over the years (Stafstrom, 1999). Clinical and
experimental studies have provided key insights into important treatment-related variables and, when considered
together, these studies have helped direct mechanistic research. Commonalities between clinical and experimental studies of efficacy are summarized in Table 1 (adapted
from Stafstrom, 2004).
Based on vast clinical experience, almost any diet that
produces ketonemia and/or diminished blood glucose levels can induce an anticonvulsant effect. Ketogenic diets
comprised of either LCTs (Freeman et al., 1998; Vining et al., 1998) or medium-chain triglycerides (MCTs;
Accepted October 24, 2006.

Address correspondence and reprint requests to Kristopher J. Bough
at FDA Center for Drug Evaluation & Research, MPN 1 Room
1345, 7520 Standish Place, Rockville, MD 20855, U.S.A. E-mail:
Kristopher.Bough@fda.hhs.gov
doi:10.1111/j.1528-1167.2007.00915.x
43
44
K. J. BOUGH AND J. M. RHO

TABLE 1. Translational correlations of ketogenic diet (KD) efficacy
Variable
Experimental findings
Clinical findings
References
Age
Young rats and mice <P40 at diet onset

Adult rats and mice >P40 at diet
onset
Infants, children and adolescents <19

years of age
Some evidence of efficacy in adults
Seizure type
Effective in a wide variety of acute and

chronic seizure models
However, protection however can be
transient; maximal seizure activity
can be prolonged
Equally efficacious in a variety of

seizure types
Dietary composition
Qualitative differences do not affect

outcome
Classic LCT = MCT = PUFA fat diet
Qualitative differences do not affect

outcome
Classic LCT, MCT, Atkins can all
diminish seizure frequency
KD ratio
Increase KD ratio = greater KD effect
Increase KD ratio = greater KD effect
Calorie restriction
CR is anticonvulsant
CR enhances KD anticonvulsant effect
NAL < NCR < KAL < KCR
CR is an integral part of KD regimen

(CR associated with seizure control)
Growth rate
Initial transient decline in body weight

Followed by, resumption of normal or
near-normal growth rate
Latency to KD effect
12 weeks
Reversal of KD effect
Seizure are behaviorally more severe

within hours, but threshold returns to
baseline over the course of 12
weeks
Males = females
Decline in weight/height over first few

months
Followed by resumption of normal or
near-normal height/weight gain
Very young children do grow poorly
Seizures can be reduced within 12
days
Fine-tuning is key to success of the
diet; at least 12 weeks are required
to see if changes are maximally
effective
Breakthrough seizure activity can
occur immediately with ingestion of
carbohydrates
Millichap (1964), Uhlemann and

Neims (1972), Appleton and DeVivo
(1974), Nakazawa et al. (1983),
Kinsman et al. (1992), Hori et al.
(1997), Bough et al. (1999b),
Muller-Schwarze et al. (1999),
Sirven et al. (1999), Nordli et al.
(2001), Coppola et al. (2002),
Kossoff et al. (2002), Mady et al.
(2003)
Livingston (1972), Appleton and
DeVivo (1974), Mahoney et al.
(1983), Otani et al. (1984), Schwartz
et al. (1989a), Hori et al. (1997),
Freeman et al. (1998), Vining et al.
(1998), Rho et al. (1999), Su et al.
(2000), Thavendiranathan et al.
(2000), Greene et al. (2001), Bough
et al. (2002)
Huttenlocher et al. (1971), Sills et al.
(1986), Schwartz et al. (1989b),
Freeman et al. (1998), Vining et al.
(1998), Mak et al. (1999), Likhodii
et al. (2000), Dell et al. (2001),
Kossoff et al. (2003), Henderson
et al. (2006), Kossoff et al. (2006)
Dekaban (1966), Livingston (1972),
Bough et al. (1999a), Bough et al.
(2000b), Freeman et al. (2000)
Bough et al.(2000a), Bough et al.
(2000b), Freeman et al. (2000),
Greene et al. (2001), Greene et al.
(2003)
Rho et al. (1999), Freeman et al.
(2000), Su et al. (2000), Vining et al.
(2002), Peterson et al. (2005), Thio
et al. (2006)
Gender
Males = females
Livingston (1972), Appleton and

DeVivo (1974), Freeman and Vining
(1999), Rho et al. (1999), Freeman
et al. (2000), Bough et al. (2006)
Appleton and DeVivo (1974),
Huttenlocher (1976), Freeman et al.
(2000), Mady et al. (2003), Bough
et al. (2006)
Millichap (1964), Nakazawa et al.
(1983), Schwartz et al. (1989b),
Freeman et al. (1998), Bough et al.
(2002), Mady et al. (2003)
Abbreviations: P40 age, postnatal day 40; LCT, long-chain triglycerides; MCT, medium-chain triglycerides; PUFA, polyunsaturated fatty acid; Atkins
diet, diet high in fats + protein, low in carbohydrates; CR, calorie-restriction; NAL, normal, ad libitum rodent diet; NCR, normal, calorie-restricted
(by 15%) rodent diet; KAL, ketogenic, ad libitum diet; KCR, ketogenic, calorie-restricted diet; KD ratio, ratio of [fats / (carbohydrates + proteins)];
KAP, ketogenic/antiketogenic potential. Table adapted from Stafstrom (2004).
Huttenlocher et al., 1971; Sills et al., 1986; Schwartz

et al., 1989b; Mak et al., 1999) can control seizures with
similar efficacy. Even the high-fat, high-protein, and lowcarbohydrate Atkins diet that produced a ketotic state, reduced seizures in epileptic patients (Kossoff et al., 2003,
2006). Similar effects have been observed experimentally.
Ketogenic diets containing myriad types of fats (i.e., with
low carbohydrate content) all produced similar levels of
Epilepsia, Vol. 48, No. 1, 2007
seizure control (Likhodii et al., 2000; Dell et al., 2001).

Thus, available evidence indicates that dietary composition per se does not appear to affect the anticonvulsant
efficacy of the diet, as long as there is a degree of sustained ketosis.
By comparison, KD ratios and calorie-restriction (CR)
appear to be important variables in enabling seizure protection. Seizure control is reportedly optimized when KDs
ANTICONVULSANT ACTIONS OF KETOGENIC DIET

are administered in ratios of 3:1 (Freeman et al., 2000),
and higher KD ratios increased both clinical (Dekaban,
1966; Livingston, 1972) and experimental anticonvulsant
efficacy (Bough et al., 2000b). Similar to the KD ratio,
increasing the extent of CR resulted in improved seizure
control in epileptic mice (Greene et al., 2001), irrespective of the type of diet that was restricted (Eagles et al.,
2003). In general, extra calories in the form of carbohydrates or proteins translate to additional metabolic substrates for gluconeogenesis and diminished KD efficacy.
Breakthrough seizures are believed to result from overestimation and administration of excess calories (Freeman
et al., 2000). As such, CR may share common anticonvulsant mechanisms and adjunctively optimize KD efficacy.
In rodents, maximal seizure control develops 12 weeks
after initiation of a KD (Appleton and DeVivo, 1974; Rho
et al., 1999; Bough et al., 2006). Similarly in humans,
clinical efficacy does not reach its zenith in many patients
until after 2 weeks (Dekaban, 1966; Freeman et al., 2000).
One notable feature of the KD is the rapid occurrence of
breakthrough seizures and loss of ketosis when carbohydrates are introduced (e.g., after a child sneaks a cookie;
Huttenlocher, 1976). As a result, the KD must be strictly
enforced in order for efficacy to be maintained. However, a
breakthrough seizure may not necessarily translate to a total loss of seizure control. Studies have shown that, despite
an abrupt discontinuation of the KD, the increased resistance to seizures waned gradually when switched back to
control (Bough et al., 2006) or even high-carbohydrate,
antiketogenic chow (Appleton and DeVivo, 1974). This
decline in seizure threshold generally occurred over 12
weeks, mirroring the onset of seizure protection (Appleton and DeVivo, 1974; Bough et al., 2006). This indicates
that a critical, minimal level of sustained ketosis is necessary but not sufficient to maintain seizure control. Thus,
it would seem that metabolic adaptations to KDs underlie
their key anticonvulsant actions.
Many studies and anecdotal observations have suggested that the KD is most effective in immature animals
or infants and children (Livingston, 1972; Uhlemann and
Neims, 1972; Otani et al., 1984; Bough et al., 1999b; Rho
et al., 1999). This is perhaps due to enhanced metabolic capacity, more efficient extraction of ketone bodies from the
blood (Morris, 2005), and/or greater compliance of KDs
in the pediatric population. However, a lack of efficacy
in older children or adults may simply reflect noncompliance or dietary intolerance rather than an inadequate response physiological (Livingston, 1972). The KD has been
demonstrated to be similarly effective in infants (Nordli
et al., 2001; Kossoff et al., 2002), adolescents (Kinsman
et al., 1992; Mady et al., 2003), and adults (Sirven et al.,
1999; Coppola et al., 2002). Furthermore, experimental
KDs are effective in both young (i.e., <P40 days; Uhlemann and Neims, 1972; Otani et al., 1984; Bough et al.,
1999b) and adult rodents (Appleton and DeVivo, 1974;
45
Muller-Schwarze et al., 1999). CR, too, is equally anticonvulsant in both juvenile and adult mice (Greene et al.,
2001). Thus, increasing evidence suggests the anticonvulsant effects of KDs do not appear to be age-dependent.
Clinical reports indicate that outcome is unrelated to
seizure type or frequency (Freeman et al., 1998; Schwartz
et al., 1989a; Vining et al., 1998). At least 50% of patients treated with a classical KD will exhibit at least a
50% reduction in seizures (Livingston, 1972; Freeman
et al., 1998). Comparatively, anticonvulsant effects of KD
in animals are more modest. Rats and mice only demonstrate a 1520% increase in seizure threshold (Appleton
and DeVivo, 1974; Bough and Eagles, 1999; Rho et al.,
1999; Bough et al., 2000b). Despite efficacy across a variety of acute and chronic seizure models (Hori et al., 1997;
Muller-Schwarze et al., 1999; Su et al., 2000), KD-induced
anticonvulsant effects have been incomplete (Bough et al.,
2002) or of limited duration (Hori et al., 1997). Further,
there is little evidence to indicate that KDs diminish severity once a seizure begins. Indeed, many studiesincluding
our ownhave shown that CR and/or KDs can even exacerbate (maximal) seizures (Mahoney et al., 1983; Otani
et al., 1984; Bough et al., 2000a; Thavendiranathan et al.,
2000; Bough et al., 2003). If one considers that seizure
activity requires large amounts of energy, seizure exacerbation may be a reflection of enhanced energy reserves
after KD treatment, a situation that would allow for prolonged ictal activity once it begins (discussed below).
In summary, the following generalizations can be made
about the KD: (1) its anticonvulsant effects appear independent of dietary formulation, but appear to be strongly
linked to the total quantity of calories consumed; (2) the
KD must be strictly adhered to, if the anticonvulsant effect
is to be maintained; (3) CR may work synergistically with
KD to limit seizures and optimize treatment; (4) maximum
efficacy is not achieved for several days or weeks after initiation, suggesting that adaptive metabolic and/or genetic
programs underlie KD-induced seizure protection; (5)
these adaptations are likely generalized throughout the
(epileptic) brain, irrespective of underlying pathology or
genetic predisposition to seizures since the KD is an effective treatment for diverse epileptic conditions; and (6)
efficacy is independent of gender and age, suggesting that
KD treatment produces seizure control via a common set
of pathways in all clinical responders.
ANTICONVULSANT MECHANISMS
OF THE KETOGENIC DIET
Since the KD was originated over 85 years ago, several
major hypotheses have been advanced, but none have been
widely accepted. Several key aspects of the KD might ultimately result in seizure protection. Ketone bodies, free
fatty acids (in particular, polyunsaturated fatty acids), or
46
glucose restriction might each lead directly or indirectly

to seizure control. While it is possible that any one of
these KD-induced changes is responsible for the anticonvulsant action of the KD, available evidence suggests that
improved seizure control, at a minimum, likely requires
all three.
Role of ketone bodies
Beta-hydroxybutyrate (BHB) is the predominant ketone
body measured in the blood, and as such, has been used as
a clinical measure of KD implementation (Fig. 1). Accordingly, nearly all KD studies have attempted to establish a
causative link between ketonemia and anticonvulsant efficacy. Although robust elevations in plasma BHB levels
have been observed during KD treatment (Bough et al.,
1999b; Thavendiranathan et al., 2000), there is no significant correlation between plasma BHB levels and seizure
protection. Optimal seizure protection generally lags days
to weeks behind the development of ketonemia, which occurs within hours of KD onset.
Nevertheless, there is some evidence that ketones other
than BHB may possess anticonvulsant properties. When
injected into animals, acetone and its parent acetoacetate
(ACA), prevent acutely provoked seizures. Seminal work
in the 1930s revealed that acute intraperitoneal administration of acetone or ethyl-acetoacetate protected rabbits from thujone-induced seizures (Helmholz and Keith,
1930; Keith, 1933). Thujone is the active constituent of
wormword oil, and is an antagonist of GABAA receptors
(Hold et al., 2000). More recent experimental studies have
shown similar results in rodents. Acetone (Likhodii et al.,
2003) and ACA (Rho et al., 2002)but not BHBwere
anticonvulsant in a variety of acute and chronic models of
epilepsy, consistent with earlier observations (Helmholz
and Keith, 1930; Yamashita et al., 1976; Vodickova et al.,

1995). Clinically, acetone levels of up to 1 millimolar
(mM) were detected in the brains of five of seven wellcontrolled epileptic patients following KD using magnetic resonance spectroscopic techniques (Seymour et al.,
1999). Although acetone could not be detected in two other
seizure-free patients, the authors concluded that acetone
contributes to the anticonvulsant effect of the KD. Interestingly, the concept that a lipophilic solvent may potently
block seizure activity is not new. The classic example of
this is valproic acid, which was initially used as a solvent
to dissolve investigational anticonvulsant compounds, but
was serendipitously discovered to possess intrinsic anticonvulsant properties.
Whereas in vivo pharmacodynamic studies have
suggested that both ACA and acetone may act as
anticonvulsant agents, there is no evidence that ketone
bodies can directly modulate synaptic transmission and/or
neuronal excitability. In vitro cellular electrophysiological experiments have failed to demonstrate an effect on the
principal ion channels that mediate neuronal excitability
and inhibition. Specifically, neither L-BHB nor ACA were
found to modulate GABAA receptors, AMPA receptors,
or NMDA receptors in both hippocampus and neocortex
of rats (Thio et al., 2000; Donevan et al., 2003). Despite
these negative observations, it remained possible that ketone bodies might affect network activity or synchrony.
However, in field potential recordings conducted in vitro,
Thio et al. (2000) demonstrated clearly that neither ACA
nor BHB modified evoked excitatory postsynaptic potentials (EPSPs) or population spikes in the CA1 subfield of
the hippocampal tissue. In summary, there is no evidence
for direct anticonvulsant effects for either ACA or BHB,
and acetone has yet to be studied in neuronal (CNS) tissue.
FIG. 1. Metabolic pathways highlighting the production of ketone bodies fatty acids during fasting or treatment with the ketogenic diet (KD).
Estimated fasting- or KD-induced concentrations of beta-hydroxybutyrate, acetoacetate, and acetone in blood are listed (large boxes).
Measures of beta-hydroxybutyrate levels in blood are most commonly used as the clinical indicator of successful KD treatment. From
Likhodii and Burnham (2004).

This may, in large part, reflect the technical difficulties in
investigating a compound that is highly volatile and can
react with perfusion systems ordinarily used in pharmacological in vitro experiments.
Recently, it has been suggested that ACA and/or its
metabolic byproduct, acetone, may activate a novel class
of potassium leak channels known as the two-pore domain or K2P channels (Vamecq et al., 2005). K2P channels
represent a diverse superfamily of channels that generally
hyperpolarize cell membranes, and regulate membrane excitability both pre- and postsynaptically (Lesage, 2003).
These channels can be modulated by changes in pH, osmolality, temperature, mechanical pressure, and certain
fatty acids (Franks and Honore, 2004). Links between KDinduced elevations in ketone bodies (and/or fatty acids, as
discussed below) and K2P channels, however, have yet to
be explored.
In conclusion, although ketone bodies have been shown
to possess anticonvulsant properties in vivo, there is no evidence to date that they mediate directly these effects. It
is clear that some degree of sustained ketosis is required
for clinical efficacy and that efficacy is maximized over a
period of weeks versus days, despite a rapid onset of ketosis within hours. Whereas it is plausible that some dietary,
pharmacokinetic factor(s) results in some level of seizure
protection, the approximate 2-week time course for optimal seizure protection suggests a pharmacodynamic effect
of the KD (e.g., parallel time course for changes in gene
expression, mitochondrial proliferation, up-regulation of
UCPs/transporters, etc) likely underlies the anticonvulsant
nature of the diet. Thus, available data suggest that adaptations to, rather than a direct effect of, ketosis underlie
the anticonvulsant nature of the KD.
Role of glucose restriction

Whereas most studies have suggested that persistent
ketosis is essential to KD-induced seizure protection, others have posited that glucose restriction is the key feature (Greene et al., 2003). In addition to ketosis, it is
clear that as ketonemia develops, another immediate consequence of CR and/or KDs is a moderate reduction
in blood glucose. Does caloric restriction simply act to
limit gluconeogenic substrates that would otherwise reduce KD ratio and counter efficacy? Or, might glucose restriction result in another metabolic adaptation that helps
quell aberrant hyperexcitability? Calorie restriction alone
was sufficient to retard seizure susceptibility in juvenile
and adult epileptic EL mice; and, blood glucose levels were inversely correlated with a decreased risk of
seizures (Greene et al., 2001). Greene et al. (2003) hypothesized that CR reduces energy production through glycolysis, which limits a neurons ability to reach (and maintain) high levels of synaptic activity necessary for seizure
genesis.
47
Others have hypothesized that glucose restriction during KD treatment activates ATP-sensitive potassium
(KATP ) channels (Schwartzkroin, 1999; Vamecq et al.,
2005). Interestingly, KATP channels are ligand-gated receptors broadly expressed throughout the central nervous
system, in both neurons and glia (Thomzig et al., 2005).
These channels act as metabolic sensors, linking cellular
membrane excitability to fluctuating levels of ADP and
ATP. Activation of this receptor by reduced ATP/ADP ratios opens the channel and leads to membrane hyperpolarization. When glucose is limited (e.g., during administration of a classic KD, which is typically CR by 25%),
KATP channels might open to hyperpolarize the cell as the
intracellular ATP concentrations fall. Conversely, when
glucose is present and ATP concentrations rise, KATP channels close. As such, KATP channels may provide a measure of protection against a variety of metabolic stressors
such as hypoxia, ischemia, and hypoglycemia, and are
believed to regulate seizure threshold (Seino and Miki,
2003).
KATP channels are particularly abundant in the substantia nigra (Hicks et al., 1994), a region of the brain thought
to act centrally in the propagation of seizure activity
(Iadarola and Gale, 1982). KATP channels would therefore
be ideally positioned to metabolically regulate the onset
of several different seizure types, as does the KD. There is
growing evidence that KATP channels may critically regulate seizure activity. Genetically engineered mice that
overexpress the sulfonylurea (SUR) subunit of KATP channels were significantly more resistant to seizures induced
by kainate, and showed no marked cell loss in hippocampus (Hernandez-Sanchez et al., 2001). Studies of KATP
channel (Kir6.2/ ) knockout mice suggested that these
channels help determine seizure threshold (Yamada et al.,
2001). Following hypoxic challenge (5% O2 ), knockout mice exhibited myoclonictonic seizure activity, and,
ultimately, death compared to controls who all recovered
without sequelae.
Despite these observations, there is one important
caveat in implicating KATP channels as mediators of a KDinduced anticonvulsant effect. Other studies have demonstrated an increase in energy reserves (specifically, ATP)
after KD treatment (DeVivo et al., 1978; Pan et al., 1999).
These data predict that KATP channels would remain
closed, not open, during diet treatment, and would thus
contribute to neuronal/glial cell membrane depolarization.
Nevertheless, several findings are consistent with the notion that KATP channels are selectively activated during
administration of a low-glucose, high-fat KD. First, KATP
channels are regulated preferentially via glycolytic energy
sources (Dubinsky et al., 1998). It has recently been shown
that the glycolytic enzyme glyceraldehyde 3-phosphate
dehydrogenase (GAPDH) serves as an accessory protein to KATP channels and regulates directly their activity
(Dhar-Chowdhury et al., 2005; Jovanovic et al., 2005).
48
The observed reduction in glycolytic processes after KD treatment (specifically, the concentration of
fructose-1,6-bisphosphate, the key regulatory enzyme
of glycolysis) is consistent with this notion (DeVivo
et al., 1978; Puchowicz et al., 2005; Melo et al., 2006).
Glycolytic flux may be further limited as a consequence
of elevated ATP (DeVivo et al., 1978; Otani et al., 1984;
Pan et al., 1999; Bough et al., 2006) and citrate (Yudkoff
et al., 2001) levels on KD treatment; both ATP and citrate
are feedback inhibitors of glycolysis.
Second, it is hypothesized that the accumulation of free
fatty acids over the course of KD administration (Dekaban,
1966) may boost KATP channel activation (Vamecq et al.,
2005). Whereas PUFAs freely cross the BBB, saturated
free fatty acids are transported across the BBB via carriermediated processes (Avellini et al., 1994). Fatty acids
that intercalate within neuronal cell membranes have been
shown to interact potently with KATP channels, specifically
reducing their affinity for (and inhibition by) ATP (Shyng
and Nichols, 1998). Overall, these findings suggest that
the unique nature of low-glucose, high-fat KDs promotes
KATP channel activation, despite observed enhancements
in oxidative energy production.
Recent experiments involving 2-deoxyglucose (2-DG)
provide further support for a glucose-restriction hypothesis of KD action. Two-deoxyglucose is a glucose analogue,
which inhibits phosphoglucose isomerase and, hence, glycolysis. Stafstrom et al. (2005) reported that the addition of 1 mM 2-DG decreased epileptiform burst frequency to 2580% of baseline in rat hippocampal slices
exposed to elevated extracellular potassium. More significantly, the same group also showed that 2-DG (250 mg/kg,
i.p) elevated the after-discharge threshold in olfactory
bulb of perforant-path kindled rats, markedly reduced the
progression of kindling, and limited the expression of
BDNF and its cognate receptor, trkB (Garriga-Canut et al.,
2006).
Interestingly, there are a number of anticonvulsant parallels between 2-DG (Stafstrom et al., 2005; GarrigaCanut et al., 2006) and KD treatment (Bough et al., 2003).
First, both 2-DG and KD elevated electrographic seizure
threshold in vivo; second, both 2-DG and KD potently retarded the progression of epileptogenesis in kindling models of epilepsy in vivo; and, third, both 2-DG (in vitro)
and KD (in vivo) diminished measures of hippocampal
hyperexcitability. These results collectively suggest that
the anticonvulsant actions of KD may work, in large part,
via an inhibition of glycolysis. Importantly, because 2-DG
is fairly well tolerated when administered orally (Pelicano
et al., 2006), this compound may represent a novel treatment strategy for epilepsy.
Role of fatty acids
Polyunsaturated fatty acids (PUFAs) such as docosahexanoic acid (DHA, C22:63), arachidonic acid (AA,
C20:46), or eicosapentanoic acid (EPA, C20:53) are

believed to affect profoundly cardiovascular function and
health (Leaf and Kang, 1996; Nordoy, 1999; Leaf et al.,
2003). In cardiac myocytes, PUFAs inhibited fast, voltagegated sodium channels (Xiao et al., 1998) and L-type calcium channels (Xiao et al., 1997). Similar findings have
been observed in neuronal tissue. For example, DHA and
EPA diminished neuronal excitability and bursting in hippocampus (Xiao and Li, 1999).
It is not surprising then that PUFAs are becoming an increasingly popular focus of KD research. After KD treatment, specific PUFAs (i.e., AA and DHA) were found to
be elevated in both serum (Cunnane et al., 2002; Fraser
et al., 2003) and brain (Taha et al., 2005) of patients and
animals. Importantly, one report documented that the rise
(or drop) in total fatty acids during KD treatment closely
paralleled clinical improvement (or loss) of seizure control
(Dekaban, 1966). An additional study found that dietary
supplementation with 5 g of (65%) n-3 PUFAs once per
day produced a marked reduction in seizure frequency
and intensity in a few epileptic patients (Schlanger et al.,
2002). These findings suggest that KD-induced elevations
in PUFAs such as DHA and/or AA might act directly to
limit neuronal excitability and dampen seizure activity.
PUFAs could ultimately block seizure activity in a number of ways (Fig. 2). First, PUFAs may inhibit directly ion
channel activity. Omega-3 (-3) PUFAs have been shown
to: (1) inhibit both voltage-gated Na+ and Ca2+ channels,
(2) increase the resistance to bursting induced by bicuculline, zero Mg2+ , pentylenetetrazole or glutamate, and
(3) prolong the recovery time from inactivation in hippocampal neurons (Vreugdenhil et al., 1996; Xiao and Li,
1999; Young et al., 2000). Second, in conjunction with ketone bodies, PUFAs may activate a lipid-sensitive class of
K2P potassium channels (Vamecq et al., 2005). And, third,
PUFAs may enhance the activity of the Na+ /K+ -ATPase
(sodium pump). Elevated -3 and diminished -6 PUFAs levels in plasma membranes significantly increased
sodium pump function (Wu et al., 2004). These findings
indicate that elevations in brain levels of PUFAs after KD
treatment (Taha et al., 2005) could help reduce neuronal
hyperexcitability via a variety of direct mechanisms.
Uncoupling proteins
In addition to their direct actions on neuronal excitability, PUFAs may also act indirectly to limit excitotoxicity and neurodegeneration. PUFAs regulate the
expression of numerous genes in brain via transcription
factors such as PPAR (peroxisome proliferator-activated
receptor-; Sampath and Ntambi, 2004). Through induction of PPAR and its coactivator PGC-1, PUFAs
induce the expression of mitochondrial uncoupling proteins (UCPs) and activate these proteins directly as well
(Jaburek et al., 1999; Diano et al., 2003). Recent evidence
49
FIG. 2. Potential pathways through which polyunsaturated fatty acids (PUFAs) may limit hyperexcitability in the brain. Acting directly,
PUFAs such as arachidonic acid (AA), docosahexanoic acid (DHA), and/or eicosapentanoic acid (EPA) might inhibit both voltage-gated
Na+ and Ca2+ channels, activate a lipid-sensitive class of K2P potassium channels, and enhance the activity of the Na+ /K+ -ATPase to
limit neuronal excitability and dampen seizure activity. Acting indirectly, PUFAs might induce the expression and activity of uncoupling
proteins (UCPs) to diminish reactive oxygen species (ROS), reduce neuronal dysfunction and induce a neuroprotective effect. Finally,
PUFAs are expected to activate PPAR and induce a coordinate up-regulation of energy transcripts leading to enhanced energy reserves,
stabilized synaptic function and limited hyperexcitability.
suggests that PUFAs are required for mitochondrial UCP

activity (Garlid et al., 2001).
Uncoupling proteins are homodimers that span the inner mitochondrial membrane and allow a proton leak from
the intermembrane space to the mitochondrial matrix.
There are three major isoforms that have been identified
in the brain, UCP2, UCP4 and UCP5 (a.k.a., BMCP-1
or brain mitochondrial carrier protein-1). UCP proteins
are increasingly implicated in the regulation of neuronal
excitability and survival (Andrews et al., 2005). The uncoupling effect, albeit of small magnitude, reduces the
proton-motive force, disassociates or uncouples electron transport from ATP production, and indirectly decreases the production of reactive oxygen species (ROS).
Although it would seem that increased levels of UCP proteins would diminish cellular energy production, Diano
et al. (2003) showed that chronic overexpression of UCP2
in neuronal tissue increased cellular ATP and ADP levels
by triggering mitochondrial biogenesis. KD appears to do
the same; that is, studies show that the KD induces UCP
expression, stimulates mitochondrial biogenesis, and enhances energy production (see also below). Seizures, by
comparison, increase ROS generation and/or mitochondrial dysfunction, which can lead to neuronal dysfunction
and excitotoxicity (Layton and Pazdernik, 1999; Kovacs
et al., 2001; Kovacs et al., 2002; Sullivan et al., 2003).
Interestingly, UCP2 is up-regulated after seizures (Diano
et al., 2003). The protective role of UCPs was recently

highlighted by Sullivan et al. (2003) who demonstrated
that dietary enhancement of UCP expression and function
in immature rats protected against kainate-induced excitotoxicity, most likely by decreasing ROS generation (Andrews et al., 2005). Further work demonstrated that mice
maintained on a high-fat KD demonstrated an increase in
the hippocampal expression and activity of all three mitochondrial UCPs and exhibited a significant reduction in
ROS generation in mitochondria isolated from the same
brain region (Sullivan et al., 2004). In conjunction with
reports that ketone bodies potently decrease ROS generation (Veech et al., 2001; Veech, 2004), these reports
suggest that the KD compensates for seizure-induced elevations in ROS generation and neuronal dysfunction to
provide a neuroprotective effect.
Energy production
Polyunsaturated fatty acids additionally regulate the
transcription of numerous genes linked to energy
metabolism (Sampath and Ntambi, 2005) through activation of PPAR, a scenario in which the KD is thought to reprogram cellular metabolism (Cullingford, 2004). Indeed,
numerous studies have described changes consistent with
an enhancement in energy production following KD treatment. First, microarray expression studies demonstrated
that KD induces a coordinated up-regulation of several
50
dozen metabolic genes associated with oxidative phosphorylation after KD (Noh et al., 2004; Bough et al., 2006).
Second, KD treatment stimulated mitochondrial biogenesis, resulting in a striking 46% increase in the number of
mitochondria in the hilar/dentate gyrus region of rat hippocampus (Bough et al., 2006). And, third, levels of energy metabolites were increased after KD. Brain glycogen
and ATP concentrations were boosted throughout rodent
brain (DeVivo et al., 1978; Otani et al., 1984) and there was
an elevation in the phosphocreatine-to-creatine (PCr:Cr)
energy-reserve ratio in both animals (Bough et al., 2006)
and humans (Pan et al., 1999). These findings are consistent with results that show ketones (4 mM BHB + 1 mM
ACA) increased hydraulic work by 14% and improved
energy status in perfused heart tissue (Sato et al., 1995).
Further, there is an overall increased metabolic efficiency
(DeVivo et al., 1978; Bough et al., 2006), decreased respiratory quotient (Bough et al., 2000b), and maximal mitochondrial respiratory rate in rodents following the KD
(Sullivan et al., 2004). Collectively, these data provide
compelling evidence that the KD enhances oxidative energy production by activating a variety of transcriptional,
translational, and biochemical mechanisms in a concerted
fashion.
Metabolic dysfunction has been identified in regions of
hyperexcitability within the brain and is associated with
several epileptic conditions. Impairment of mitochondrial
function has been observed in the seizure foci of both human and experimental epilepsies (Kunz et al., 2000). Severe metabolic dysfunction occurred in both human and
rat hippocampal tissue during periods of heightened neuronal activity (Kann et al., 2005). Kudin et al. (2002)
demonstrated that seizure activity down-regulated mitochondrial enzymes involved in oxidative phosphorylation.
In an earlier study, the same group demonstrated a specific
deficiency in complex I activity and mitochondrial ultrastructural abnormalities within the hippocampal CA3 region of epileptic tissue resected from 57 human patients
(Kunz et al., 2000). In view of previous studies demonstrating impaired oxidative phosphorylation capacity in
pilocarpine-treated rats (Kudin et al., 2002) and in patients
with epilepsy (Antozzi et al., 1995; Kunz et al., 2000), a
KD-induced augmentation in oxidative phosphorylation
and energy reserves seems likely to counter energetic deficiencies in epileptic tissue, making neuronal tissue more
resilient to aberrant neuronal activity and, in this way, contributing to the diets anticonvulsant actions.
Stabilized synaptic function
Intriguing as this argument may be, how exactly would
enhanced energy reserves lead to stabilized synaptic function and diminished seizures? One possibility is via the
sodium pump. ATP is primarily used to maintain ionic gradients, especially through actions of the transmembrane
sodium pump (Hulbert and Else, 2000). Schwartzkroin
originally hypothesized that KD-induced elevations in

ATP concentrations might enhance and/or prolong the activation of the Na+ /K+ -ATPase , perhaps via an increase in

the delta-G of ATP hydrolysis (Veech et al., 2001; Veech,
2004). In neurons, increased sodium pump activity might
hyperpolarize the cell and/or reduce the resting membrane potential to diminish firing probability. Enhanced
Na+ /K+ -ATPase function in neurons might also preserve
normal neuronal functioning and/or delay a pathological
buildup of high external K+ (Xiong and Stringer, 2000).
In glia, increased activation of the Na+ /K+ -ATPase might
slow glial depolarization and allow for prolonged uptake
of extracellular K+ during periods of intense neuronal activity (e.g., high-frequency bursting). Increases in neuronal and/or glial action of the sodium pump would be
expected to limit hyperexcitability and increase the resistance to seizures, as is noted after treatment with KD.
Although no studies have tested this sodium-pump
hypothesis directly, a recent report suggests that KD
tissue is more resistant to metabolic stress. When challenged with mild hypoglycemia, synaptic transmission
within the dentate gyrus was maintained for approximately 60% longer in tissue from KD-fed animals compared to controls (Bough et al., 2006). These data suggest
that the KD stabilizes synaptic transmission (both excitatory and inhibitory) for prolonged periods of time during
metabolic stress such as during seizure activity. Hence,
it seems likely that the KD induces seizure protection
in part by preventing neuronal dysfunction (diminution
of ROS/enhancement of energy reserves) and stabilizing
synaptic transmission (enhancement in energy reserves).
A role for neurotransmitter systems
The noradrenergic hypothesis
One of the more intriguing observations regarding KD
action involves norepinephrine, its receptors and signaling
cascades. In general, increases in noradrenergic tone result in an anticonvulsant effect. Several lines of evidence
support this view. Norepinephrine (NE) re-uptake inhibitors can prevent seizure activity in genetically epilepsy
prone rats (GEPRs; Yan et al., 1993) and pharmacological NE agonists are generally, though not always, anticonvulsant (Weinshenker and Szot, 2002); damage to
the locus coeruleusthe principal region of the brain
from which ascending and descending noradrenergic innervation originatesconverts occasional seizures into
self-sustaining status epilepticus (SSSE) in rats (Giorgi
et al., 2004); animals are more prone to seizures when NE
is chemically depleted with reserpine (Weinshenker and
Szot, 2002); and, interestingly, there are several reports of
diminished brain levels of NE in several animal models
of epilepsy, including GEPRs, kindled animals, EL mice,
seizure-sensitive Mongolian gerbils, and tottering mice
(Weinshenker and Szot, 2002).

Of significant interest is the observation that mice lacking the ability to produce NE (Dbh/ knockout mice)
do not exhibit an increased resistance to flurothyl seizures
when treated with a KD (Szot et al., 2001). These data
indicate that NE is required for the anticonvulsant effect
of KD, at least in the flurothyl seizure threshold model.
Weinshenker and Szot (2002) additionally reported an approximate twofold increase in NE levels in hippocampus
following a KD, suggesting that KD increases basal release of NE. These studies indicate the anticonvulsant action of KD may result in part from an enhancement in
noradrenergic signaling in the brain.
If the KD enhances NE release as described above, it
may also promote the corelease of anticonvulsant orexigenic peptides such as neuropeptide-Y (NPY) and galanin.
NPY has been shown to inhibit glutamatergic synaptic
transmission and epileptogenesis in vitro (Rhim et al.,
1997; Richichi et al., 2004; Vezzani and Sperk, 2004);
galanin has been shown to limit SSSE (Saar et al., 2002)
and diminish both excitatory synaptic transmission and
ictal activity in vitro (Schlifke et al., 2006). Both neuropeptides are elevated after calorie restriction. However,
there was no evidence for enhanced transcription of either
of these peptides in the brain after KD treatment, suggesting that neither NPY nor galanin contribute significantly
to the anticonvulsant actions of KD (Tabb et al., 2004).
The GABAergic hypothesis
One of the more popular hypotheses for KD action involves -aminobutyric acid (GABA), the major inhibitory
neurotransmitter in the mammalian brain. In general, the
KD is most effective against seizures evoked by GABAergic antagonists. The KD potently inhibits seizures in-
51
duced by pentylenetetrazole, bicuculline, picrotoxin, and

-butyrolactone. In contrast, the diet demonstrates little if any efficacy in acute seizure models involving activation of ionotropic glutamate receptors (e.g., kainic
acid), voltage-dependent sodium channels (e.g., maximal
electroshock [MES]), or glycine receptor inhibition (e.g.,
strychnine; Bough et al., 2002).
Yudkoff et al. (2005) have proposed that ketosis induces major shifts in brain amino acid handling favoring
the production of GABA. This results from a reduction
of aspartate relative to glutamate, the precursor to GABA
synthesis, and a shift in the equilibrium of the aspartate
aminotransferase reaction in the ketotic state. As a result, there is an increase in glutamic acid decarboxylase
(GAD) activity and GABA production (Fig. 3). Elevated
GABA levels would, in turn, be expected to dampen hyperexcitability throughout the brain. Several studies support
this possibility. First, KD and CR diet treatments both
increased GAD transcript and protein levels in inferior
and superior colliculi, cerebellar and temporal cortex, and
striatum (the latter, KD only; Cheng et al., 2004). Second, both BHB and ACA increased the rate and extent
of GABA formation in synaptosomes (Erecinska et al.,
1996; Yudkoff et al., 1997). And, finally, KD treatment
in vivo modified amino acid levels in a manner consistent
with enhanced GABA production (Yudkoff et al., 2001;
Melo et al., 2006). Although brain levels of glutamate
and GABA have not been consistently elevated in rodents
(DeVivo et al., 1978; Al-Mudallal et al., 1996; Yudkoff
et al., 2001; Bough et al., 2006), two recent clinical studies report significant increases in GABA levels following
KD treatment (Wang et al., 2003; Dahlin et al., 2005),
further substantiating this view.
FIG. 3. Metabolic modifications of glutamate and GABA synthesis as a consequence of diminished glucose and ketosis. In ketosis, betahydroxybutyrate and acetoacetate contribute heavily to brain energy needs. A variable fraction of pyruvate (1) is ordinarily converted to
acetyl-CoA via pyruvate dehydrogenase. In contrast, all ketone bodies generate acetyl-CoA, which enters the tricarboxcylic acid (TCA)
cycle via the citrate synthetase pathway (2). This involves the consumption of oxaloacetate, which is necessary for the transamination of
glutamate to aspartate. Oxaloacetate is then less available as a reactant of the aspartate aminotransferase pathway, which couples the
glutamate-aspartate interchange via transamination to the metabolism of glucose through the TCA cycle. Less glutamate is converted
to aspartate and thus, more glutamate is available for synthesis of GABA (3) through glutamic acid decarboxylase (GAD). Adapted from
Yudkoff et al. (2004).
52
In addition to biochemical measures of KD-enhanced

GABAergic inhibition, there is functional evidence as
well. Electrophysiological recordings conducted in vivo
demonstrated that network excitability was diminished in
both KD- and calorie-restricted rats (Bough et al., 2003);
greater stimulus intensities were required to evoke population spikes in both CR- and KD-fed animals compared to
ad libitum controls. Paired-pulse inhibition was increased.
Both CR and KD dietary treatments resulted in greater
paired-pulse inhibition compared to controls at the 30ms interpulse interval (Bough et al., 2003), a result consistent with an enhancement in fast, GABAA -mediated
inhibition. Additionally, KD-fed animals exhibited an elevated electrographic seizure threshold and an increased
resistance to a modified, 1-day kindling protocol (maximal dentate activation). These data suggested that both
KD and calorie-restricted diets limited network excitability and elevated seizure threshold via an enhancement of
GABAergic inhibition.
GABAergic interneurons, which at baseline have more
depolarized resting membrane potentials, endure nonaccommodating bursts of neuronal firing and must
metabolically persist (Attwell and Laughlin, 2001), lest
network inhibition becomes compromised. Previous studies have shown that a KD increases total brain [ATP]
(DeVivo et al., 1978) and PCr/Cr or PCr/ATP energy reserve (Pan et al., 1999; Bough et al., 2006). Accordingly,
KD-induced elevations in PCr are likely to play a pivotal
role in maintaining the activity of the Na+ /K+ -ATPase
during periods of intense seizure activity, in both glutamatergic and GABAergic neurons. In a recent study of
human temporal lobe epilepsy (Williamson et al., 2005),
the PCr/ATP ratio correlated with the recovery of the
membrane potential following a stimulus train, which
was inversely correlated with granule cell bursting. Because creatine kinase is predominantly localized within
GABAergic interneurons (Boero et al., 2003), Boero et al.
concluded that PCr and energy levels are especially critical to the maintenance of GABAergic inhibitory output.
In this manner, a KD-induced increase in energy reserves
might enhance GABAergic function in particular and improve seizure control.
HOW CAN THE KETOGENIC
DIET BE OPTIMIZED?
Historically, few guidelines have emerged regarding the
clinical implementation of the KD and its variants including the medium-chain triglyceride (MCT) formulation (Huttenlocher et al., 1971) and more recent options
such as the Atkins diet (Kossoff et al., 2003; Kossoff
et al., 2006). This is largely a result of the fact that, until recently, few KD centers existed throughout the world. Even
with a resurgence of interest in dietary approaches toward
epilepsy treatment in the past decade, there remains a noEpilepsia, Vol. 48, No. 1, 2007
table absence of Class I and II clinical studies. Today, few

question the clinical efficacy of the KD in both young and
older patients (Vining, 1999; Coppola et al., 2002; Mady
et al., 2003), and many successful international centers
have evolved (Kossoff and McGrogan, 2005; Freeman
et al., 2006). However, since we do not fundamentally
know how the KD prevents seizures, there exists as yet no
rational basis for optimizing the efficacy of the diet, other
than through trial and error.
When examining the accumulated clinical data, it appears seizure control can be achieved in the majority
of epileptic patients as long as there is a shift from
glycolytic flux to intermediary metabolism (resulting in
measurable ketosis), irrespective of the precise dietary
formulation (Henderson et al., 2006). On the other hand,
the experimental literature suggests that different treatment protocols may result in differential efficacy or even
lack of efficacy, despite significant ketosis (Bough et al.,
2000a; Thavendiranathan et al., 2000; Bough et al., 2002;
Thavendiranathan et al., 2003). Most of the published
studies have been based on acute seizure models, and not
on developmental epilepsy models. Hence, of course, one
must bear in mind that, despite dozens of animal models
of the KD, none recapitulate all of the essential features
in the human epileptic condition (Stafstrom, 1999).
So how can we reach the goal of developing a safer and
more effective KD? The reductionist approach posits that
were we to identify the critical mediator of the diets anticonvulsant effect, administration of this substrate alone
would likely yield a similar clinical effect as the traditional KD, and importantly, spare the patient significant
side-effects that may preclude its useeven in the face of
clear clinical efficacy. The closest we have come to this
situation is the recent use of BHB as an oral neuroprotectant. Promising results have already been demonstrated
in Phase I clinical trials (Smith et al., 2005). Nevertheless, despite increasing experimental evidence that BHB
and ACA both possess neuroprotective properties (Kashiwaya et al., 2000; Noh et al., 2006), a direct anticonvulsant
effect of ketone bodies has not yet been demonstrated in
epileptic brain, either animal or human. Intriguing, however, are animal studies indicating that ACA and acetone
are anticonvulsant in acute seizure models. Yet, there remains a perplexing lack of an acute anticonvulsant effect
of the principal ketone body, BHB.
Conversely, if we believe that certain PUFAs, in lieu of
ketone bodies, are direct mediators of an anticonvulsant effect (Cunnane et al., 2002; Cunnane, 2004), as suggested
by clinical studies (Schlanger et al., 2002; Fraser et al.,
2003; Fuehrlein et al., 2004; Yuen et al., 2005), we may be
closer to distilling the essence of the KD. However, there
is likely no single fatty acid that is necessary and sufficient
for an anticonvulsant effect. And experimentally, while it
has been straightforward to demonstrate the inhibitory effects of PUFAs on specific voltage-gated ion channels and
53
FIG. 4. Hypothetical pathways leading to the anticonvulsant effects of the ketogenic diet (KD). Elevated free fatty acids (FFA) lead to
chronic ketosis and increased concentrations of polyunsaturated fatty acids (PUFAs) in the brain. Chronic ketosis is predicted to lead
to increased levels of acetone; this might activate K2P channels to hyperpolarize neurons and limit neuronal excitability. Chronic ketosis
is also anticipated to modify the tricarboxcylic acid (TCA) cycle. This would increase glutamate and, subsequently, GABA synthesis in
brain. Among several direct inhibitory actions (see also Fig. 2), PUFAs boost the activity of brain-specific uncoupling proteins (UCPs).
This is expected to limit ROS generation, neuronal dysfunction, and resultant neurodegeneration. Acting via the nuclear transcription
factor peroxisome proliferator-activated receptor- (PPAR), PUFAs would induce the expression of UCPs and coordinately up-regulate
several dozen genes related to oxidative energy metabolism. PPAR expression is inversely correlated with IL-1 cytokine expression;
given the role of IL-1 in hyperexcitability and seizure generation (Vezzani et al., 2000), diminished expression of IL- cytokines during KD
treatment could lead to improved seizure control. Ultimately, PUFAs would stimulate mitochondrial biogenesis. Mitochondrial biogenesis is
predicted to increase ATP production capacity and enhance energy reserves, leading to stabilized synaptic function and improved seizure
control. In particular, an elevated phosphocreatine:creatine (PCr:Cr) energy-reserve ratio is predicted to enhance GABAergic output,
perhaps in conjunction with the ketosis-induced elevated GABA production, leading to diminished hyperexcitability. Reduced glucose
coupled with elevated free fatty acids are proposed to reduce glycolytic flux during KD, which would further be feedback inhibited by high
concentrations of citrate and ATP produced during KD treatment. This would activate metabolic KATP channels. Opening of KATP channels
would hyperpolarize neurons and diminish neuronal excitability to contribute to the anticonvulsant (and perhaps neuroprotective) action
of the KD. Reduced glucose is also expected to downregulate brain-derived neurotrophic factor (BDNF) and trkB signaling in brain. As
activation of TrkB pathways by BDNF have been shown to promote hyperexcitability and kindling, these potential KD-induced effects would
be expected to limit the symptom (seizures) as well as the progression of epilepsy. Boxed variables depict findings taken from KD studies;
up () or down () arrows indicate the direction of the relationship between variables as a result of KD treatment.
the resultant diminution of cellular excitability in vitro,

it is not an easy task to demonstrate that ingestion of a
specific fatty acid or fatty acid cocktail, acts directly on
relevant brain receptor targets without first undergoing
beta-oxidation. The collective data, from both animals and

humans, indicate that the critical condition necessary for
achieving seizure control is a metabolic shift toward fatty
acid oxidation from glycolysis, reflected in the variable
54
rise in blood/brain ketone levels and a concomitant (moderate) reduction in blood/brain glucose. Fatty acid composition may not ultimately matter, as long as this important
metabolic shift occurs. And, interestingly, calorie restriction (Greene et al., 2001; Bough et al., 2003; Eagles et al.,
2003; Greene et al., 2003) or intake of 2-DG (Stafstrom
et al., 2005), both of which result in mild hypoglycemia,
may be the only requirement for seizure protection, regardless of whether fats are consumed or not.
As we continue to explore putative anticonvulsant
mechanisms of KD action, we are left with many outstanding clinical questions regarding dietary treatments
for epilepsy. Well-designed, multicenter prospective- and
controlled clinical trials are essential toward developing
the optimum KD. If woven together with pharmacokinetic
and pharmacogenetic investigations, these clinical studies will not only provide further insights into mechanistic
underpinnings, but will also help differentiate responders
from non-responders and identify patients in whom the
diet is definitively contraindicated. Clinicians would be
given the tools to make evidence-based decisions rather
than rely upon a few casecontrolled studies, anecdotal reports of efficacy, or clinical folklore as has been the practice in the past. Toward this end, information regarding the
impact of pharmacogenetics on epilepsy treatment is now
beginning to emerge (Depondt and Shorvon, 2006; Spurr,
2006), although much less is known regarding the genetically determined variables influencing dietary impact on
brain function, particularly as it relates to the epileptic
brain.
CONCLUSIONS
After nearly a century of clinical use, we still do not
know how the KD works. However, much progress in KD
research has been made in the past decade. Among other
factors, current evidence indicates KD optimizes cellular
metabolism. Endogenous biochemical and genetic programs are switched on in the brain in response to ketosis, glucose restriction, and elevated free fatty acids. This
unique metabolic state, if maintained, induces a shift away
from glycolytic energy production (glucose restriction)
toward the production of energy via oxidative phosphorylation (beta-oxidation of fatty acids and production of
ketone bodies). The reduction in glycolytic energy supply may activate selectively KATP channels to increase the
resistance to onset of ictal activity. An increase in oxidative phosphorylation coupled with an induction of UCPs
and mitochondrial biogenesis can diminish ROS generation and increase energy reserves, both of which would
be expected to prevent neuronal dysfunction, seizures and
even neurodegeneration.
It is improbable that one mechanistic target or mediator will produce entirely the seizure protection associated with the KD. Rather, several factors likely contribute
mechanistically to this broadly efficacious treatment for

epilepsy. The challenge of finding key variables is made
ever more difficult by the intrinsic complexity of metabolic
effects and their resultant actions on neurons, glia and on
the epileptic condition itself. We have reviewed here a
number of seemingly disparate variables that must be sustained for a meaningful anticonvulsant effect to be rendered. These interrelationships are summarized in Fig. 4.
The fact that a fundamental modification in diet can have
such profound, therapeutic effects on neurological disease
underscores the importance of elucidating mechanisms of
KD action. Future studies will no doubt provide unique
insights into how diet can affect the brain, both in health
and disease, and likely provide the scientific basis for the
development of potent new treatment strategies for the
epilepsies.
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Epilepsia, 48(1):4358, 2007

Blackwell Publishing, Inc.

Anticonvulsant Mechanisms of the Ketogenic Diet

Accepted October 24, 2006.

K. J. BOUGH AND J. M. RHO

Young rats and mice <P40 at diet onset

Infants, children and adolescents <19

Effective in a wide variety of acute and

Equally efficacious in a variety of

Qualitative differences do not affect

Qualitative differences do not affect

Increase KD ratio = greater KD effect

Increase KD ratio = greater KD effect

CR is an integral part of KD regimen

Initial transient decline in body weight

Seizure are behaviorally more severe

Decline in weight/height over first few

Millichap (1964), Uhlemann and

Livingston (1972), Appleton and

Huttenlocher et al., 1971; Sills et al., 1986; Schwartz

seizure control (Likhodii et al., 2000; Dell et al., 2001).

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

K. J. BOUGH AND J. M. RHO

glucose restriction might each lead directly or indirectly

and Keith, 1930; Yamashita et al., 1976; Vodickova et al.,

Epilepsia, Vol. 48, No. 1, 2007

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

Role of glucose restriction

K. J. BOUGH AND J. M. RHO

C20:46), or eicosapentanoic acid (EPA, C20:53) are

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

suggests that PUFAs are required for mitochondrial UCP

et al., 2003). The protective role of UCPs was recently

K. J. BOUGH AND J. M. RHO

originally hypothesized that KD-induced elevations in

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

duced by pentylenetetrazole, bicuculline, picrotoxin, and

K. J. BOUGH AND J. M. RHO

In addition to biochemical measures of KD-enhanced

table absence of Class I and II clinical studies. Today, few

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

the resultant diminution of cellular excitability in vitro,

beta-oxidation. The collective data, from both animals and

Epilepsia, Vol. 48, No. 1, 2007

K. J. BOUGH AND J. M. RHO

mechanistically to this broadly efficacious treatment for

ANTICONVULSANT ACTIONS OF KETOGENIC DIET

Epilepsia, Vol. 48, No. 1, 2007

K. J. BOUGH AND J. M. RHO

Kashiwaya Y, Takeshima T, Mori N, Nakashima K, Clarke K, Veech

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