A cDNA microarray analysis of gene expression profiles in rat hippocampus following a ketogenic diet.

The ketogenic diet (KD) is an effective therapy for medically intractable epilepsy, but its anticonvulsant mechanisms are unknown. Few studies to date have addressed the molecular changes following treatment with a KD. In the present study, we fed juvenile rats either a standard diet or a KD for 1 month, and then determined changes in hippocampal gene expression using cDNA microarray analysis (Clontech). To validate the microarray expression results, we also performed Northern blot and RT-PCR analysis on a small subset of affected genes. Among a total of 1176 cDNAs, 42 genes were strongly up- or down-regulated (>2-fold change over controls) by a KD. We found that the expression of mitochondrial ATP synthase beta subunit, mitochondrial ATP synthase D subunit (ATP5H) and mitochondrial ATP synthase beta subunit precursor (ATP5F) were especially increased in KD-treated group, whereas the KD down-regulated protein kinase C (PKC) beta and epsilon isoforms. Thus, the most prominent changes were seen in genes encoding proteins involved in mitochondrial metabolic and intracellular signal transduction pathways. Our data provide some insights into the complex cascade of cellular changes in the hippocampus induced by a KD, some of which may contribute to its anticonvulsant effects.

[1]  T. Seyfried,et al.  Environmental Risk Factors for Multifactorial Epilepsy in EL Mice , 1999, Epilepsia.

[2]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.

[3]  Hitoshi Sato,et al.  The ketogenic diet upregulates expression of the gene encoding the key ketogenic enzyme mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase in rat brain , 2002, Epilepsy Research.

[4]  C. Gonçalves,et al.  A ketogenic diet increases protein phosphorylation in brain slices of rats. , 2002, The Journal of nutrition.

[5]  J. LaManna,et al.  Diet‐Induced Ketosis Does Not Cause Cerebral Acidosis , 1996, Epilepsia.

[6]  I. Nissim,et al.  Brain amino acid metabolism and ketosis , 2001, Journal of neuroscience research.

[7]  D. Terrian,et al.  Identification and localization of an actin-binding motif that is unique to the epsilon isoform of protein kinase C and participates in the regulation of synaptic function , 1996, The Journal of cell biology.

[8]  J M Freeman,et al.  The efficacy of the ketogenic diet-1998: a prospective evaluation of intervention in 150 children. , 1998, Pediatrics.

[9]  Richard Weindruch,et al.  Gene-expression profile of the ageing brain in mice , 2000, Nature Genetics.

[10]  A. Neims,et al.  Anticonvulsant properties of the ketogenic diet in mice. , 1972, The Journal of pharmacology and experimental therapeutics.

[11]  T. Speed,et al.  Design issues for cDNA microarray experiments , 2002, Nature Reviews Genetics.

[12]  Joseph c. Madara,et al.  Effects of acute inhibition of fatty acid oxidation on latency to seizure and concentrations of β hydroxybutyrate in plasma of rats maintained on calorie restriction and/or the ketogenic diet , 2002, Epilepsy Research.

[13]  Rm. Wilder,et al.  The effects of ketonemia on the course of epilepsy , 1921 .

[14]  C. Stafstrom Animal models of the ketogenic diet: what have we learned, what can we learn? , 1999, Epilepsy Research.

[15]  Jullie W Pan,et al.  Ketosis and Epilepsy: 31P Spectroscopic Imaging at 4.1 T , 1999, Epilepsia.

[16]  D. D. Fraser,et al.  Potential role of polyunsaturates in seizure protection achieved with the ketogenic diet. , 2002, Prostaglandins, leukotrienes, and essential fatty acids.

[17]  Bradley E. Enerson,et al.  Diet-induced ketosis increases monocarboxylate transporter (MCT1) levels in rat brain , 2001, Neurochemistry International.

[18]  H. Prydz,et al.  Binding of Factor VIIa to Tissue Factor on Keratinocytes Induces Gene Expression* , 2000, The Journal of Biological Chemistry.

[19]  G. Holmes,et al.  Ketogenic Diet: Effects on Expression of Kindled Seizures and Behavior in Adult Rats , 1997, Epilepsia.

[20]  A. Mahoney,et al.  Fasting and ketogenic diet effects on audiogenic seizure susceptibility of magnesium deficient rats , 1983, Pharmacology Biochemistry and Behavior.

[21]  S. Kang,et al.  The protective effect of a ketogenic diet on kainic acid-induced hippocampal cell death in the male ICR mice , 2003, Epilepsy Research.

[22]  M. Rattray,et al.  Selective up-regulation of protein kinase C epsilon in granule cells after kainic acid-induced seizures in rat. , 1997, Brain research. Molecular brain research.

[23]  C. Bondy,et al.  A ketogenic diet increases brain insulin-like growth factor receptor and glucose transporter gene expression. , 2003, Endocrinology.

[24]  D. Devivo,et al.  An Animal Model for the Ketogenic Diet , 1974, Epilepsia.

[25]  B. Meldrum,et al.  The protein kinase C activators, phorbol 12-myristate,13-acetate and phorbol 12,13-dibutyrate, are convulsant in the pico-nanomolar range in mice. , 1992, European journal of pharmacology.

[26]  P. Schwartzkroin Mechanisms underlying the anti-epileptic efficacy of the ketogenic diet , 1999, Epilepsy Research.

[27]  A. Yamatodani,et al.  [Effect of ketogenic diet on the convulsive threshold and brain amino acid and monoamine levels in young mice]. , 1984, No to hattatsu = Brain and development.

[28]  R. Huganir,et al.  Regulation of GABAA receptor function by protein kinase C phosphorylation , 1994, Neuron.

[29]  K. Bough,et al.  Higher ketogenic diet ratios confer protection from seizures without neurotoxicity , 1999, Epilepsy Research.

[30]  A. Feinberg,et al.  A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. , 1983, Analytical biochemistry.

[31]  D. Nicholls,et al.  Protein kinase C and the regulation of glutamate exocytosis from cerebrocortical synaptosomes. , 1993, The Journal of biological chemistry.

[32]  E. Vining,et al.  Efficacy of the Ketogenic Diet for Intractable Seizure Disorders: Review of 58 Cases , 1992, Epilepsia.

[33]  P. Schwartzkroin,et al.  Age-dependent differences in flurothyl seizure sensitivity in mice treated with a ketogenic diet , 1999, Epilepsy Research.

[34]  I. Nissim,et al.  Ketogenic diet, amino acid metabolism, and seizure control , 2001, Journal of neuroscience research.

[35]  S. Kodama,et al.  Effects of ketogenic diet on electroconvulsive threshold and brain contents of adenosine nucleotides , 1983, Brain and Development.

[36]  D. Devivo,et al.  Chronic ketosis and cerebral metabolism , 1978 .

[37]  Y. Ben-Ari,et al.  Limbic seizure and brain damage produced by kainic acid: Mechanisms and relevance to human temporal lobe epilepsy , 1985, Neuroscience.

[38]  G. Holmes,et al.  Ketogenic diet reduces spontaneous seizures and mossy fiber sprouting in the kainic acid model. , 1999, Neuroreport.

[39]  P. Schwartzkroin,et al.  Norepinephrine is required for the anticonvulsant effect of the ketogenic diet. , 2001, Brain research. Developmental brain research.

[40]  T. Seyfried,et al.  Perspectives on the metabolic management of epilepsy through dietary reduction of glucose and elevation of ketone bodies , 2003, Journal of neurochemistry.

[41]  G. Shulman,et al.  Liver fatty acid‐binding protein is required for high rates of hepatic fatty acid oxidation but not for the action of PPAR‐α in fasting mice , 2004, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[42]  A. Nehlig,et al.  Glucose and ketone body utilization by the brain of neonatal rats , 1993, Progress in Neurobiology.

[43]  M. Erecińska,et al.  Regulation of GABA Level in Rat Brain Synaptosomes: Fluxes Through Enzymes of the GABA Shunt and Effects of Glutamate, Calcium, and Ketone Bodies , 1996, Journal of neurochemistry.

[44]  C. Stafstrom,et al.  The Ketogenic Diet Inhibits Epileptogenesis in EL Mice: A Genetic Model for Idiopathic Epilepsy , 2000, Epilepsia.

[45]  E. Vining Clinical efficacy of the ketogenic diet , 1999, Epilepsy Research.

[46]  K. Bough,et al.  Seizure resistance is dependent upon age and calorie restriction in rats fed a ketogenic diet , 1999, Epilepsy Research.

[47]  J M Freeman,et al.  A multicenter study of the efficacy of the ketogenic diet. , 1998, Archives of neurology.