Understanding Ketogenic Diet with Rodent Epilepsy Models Do-Young Kim Barrow Neurological Institute Phoenix, Arizona, USA
Causes of Epilepsy The most common causes of epilepsy include: Brain infection, Brain tumor, Head trauma, Stroke, Drug use, Neurodevelopmental disorders, High fever, Genetic factors In more than half of people with epilepsy, the cause is unknown.
Golyala et al., 2017 Seizure Http://www.cureepilepsy.org
Alternative Therapies Surgery Vagal Nerve Stimulation Ketogenic diet o o In the early 1920 s, investigators found that a high-fat diet could control seizures. The KD experienced tremendous attention in the 1990 s, following the successful treatment of Charlie who was with refractory epilepsy and a long track record of success, particularly at Johns Hopkins.
Freeman et al., Epilepsy Research (2006) Li et al., Iran J. Pediatr (2013)
Ketogenic diet Nutrient Daily intake (g) Grams per meal (3 meals/d) Fat 130 43.3 Protein 22 7.3 CHO 10.5 3.5 % of calories 4:1 KD TD.96355 6:1 KD TD.7797 Normal Chow TD.6319 Fat 90.5 93.2 17.3 Protein 9.1 6.4 18.9 CHO 0.4 0.5 63.9 Kcal/g 6.66 6.69 3.77 For example, the TD 96355 formula contains (in g/100g) 67.43 fat, 15.08 protein, and 0.54 carbohydrate, for a [fat]:[protein+carbohydrate] Ratio of 4.3:1, approximating the classic 4:1 ketogenic diet formation. Stafstrom & Rho, Epilepsy and the Ketogenic diet Humana Press 2004
Maximal Electroshock (MES) Corneal or auricular stimulation with a high-frequency (60 Hz), short-duration (0.2 s) stimulus. Generalized tonic-clonic seizure. Model Mouse KD Efficacy of the Reference KD MES CD1 2.5:1 y Uhlemann and Neims, 1972 DDY 2.8:1 y Nakazawa et al., 1983 C57BL/6 6.3:1 (8.6:1) y Martillotti et al., 2006 NIH Swiss 6.3:1 (8.6:1) n Zarnowska et al., 2017 Model Rat KD Efficacy of the KD Reference MES Male Wistar MCT diet n Thavendiranathan et al, 2000 Sprague Dawley 6:1 n Bough et al., 2000 Long-Evans/Wister 3.5:1 n Likhodii et al., 2000
6Hz Model Corneal stimulation. Complex partial seizure. Model Mouse KD Efficacy of the KD Reference 6Hz NIH Swiss 6.3:1 y Hartman et al., 2008 CD1 4:1 y/n Samala et al., 2008 CD1 6:1 y Samala et al., 2008 CD1 4.3:1 y/n Lucchi et al., 2017 NIH Swiss 6.3:1 y Dolce et al., 2018 Swiss Webster 6.3:1 y Olson et al., 2018 The KD significantly elevates the seizure threshold in the 6-Hz test depending on treatment period and KD composition, potentially through inhibition of AMPA receptor
Pentylenetetrazole (PTZ) Model Mouse KD Efficacy of Reference composition the KD PTZ (85 mg/kg) CD1 2.5:1 n Uhlemann and Neims, 1972 PTZ (Tail Vein Injection) GABA A antagonist. Generalized seizure. CD1 4:1/6:1 n Samala et al., 2008 PTZ Kindling CD1 6:1 y Hansen et al., 2009 PTZ Kindling CD1 6:1 y Lusardi et al., 2016 PTZ (85 mg/kg) C57Bl/6 6:1 y Knowles et al., 2018 The KD suppresses kindling-epileptogenesis, potentially through a mechanism involving regulation adenosine and DNA methylation.
Model Rat KD composition Efficacy of the KD Reference PTZ (10 mg Tail vein) SD 6:1 y Bough et al, 1999 PTZ (10 mg Tail vein) SD 6:1 y Bough et al, 2000 PTZ Wistar MCT diet n Thavendiranathan et al, 2000 PTZ (50mg SC) Wistar 3.5:1 y Likhodii et al., 2000 PTZ (Tail vein) SD 6:1 y Eagles et al., 2003 PTZ Wister 6:1 y Ziegler et al., 2004 PTZ (70mg/kg IP) SD 6:1 y Rhodes et al., 2005 PTZ (10 mg Tail vein) Wistar 4:1/6:1 n/y Nylen et al, 2005 PTZ (20 mg Tail vein) Wistar 4:1 y Raffo et al., 2008 PTZ (Tail vein) Wistar 4:1 y Porta et al, 2009 PTZ Kindling Wistar 6:1 y Lusardi et al., 2016 PTZ (Tail vein) SD 4:1 y Wang et al., 2016 PTZ (35 mg/kg IP) SD 4:1 y Jiang et al., 2016 PTZ Wister? y Sanya et al., 2017 PTZ Kindling SD 4:1 y Wang et al., 2018
Flurothyl GABA antagonist. Multiple acute seizures. Model Mouse KD Efficacy of the KD Reference Flurothyl C3Heb/Fej 4:1 y Rho et al., 1999 a mixed 129/SvEv and C57BL/6J 4:1 y Szot et al., 2001 CD1 4:1/6:1 n Samala et al., 2008 a mixed 129svC57Bl/6 6:1 y Simeone et al., 2017 C57Bl/6 1:1, 3:1, 6:1 y Simeone et al., 2017
Kainate L-glutamate analog. Injected rodents show recurrent seizure. Model Rat KD Efficacy of the KD Reference KA SD 5:1 y Muller-Schwarze et al., 1999 KA SD 4:1 n Ko et al., 1999 KA SD 5:1 y Su et al.,2002 KA SD 4:1 n Bough et al., 2002 KA SD 4.1 y Xu et al., 2008 KA SD 7.6:1 y Jeong et al.,2010 KA SD 6:1 y McDaniel et al., 2011 KA SD 4.1 y Dustin et al., 2016
Model Mouse KD Efficacy of the KD Reference KA ICR 4:1 y Noh et al.,2003 KA ICR 4:1 y Noh et al.,2005 KA ICR 9.5: 1 y Kwon et al.,2007 KA C3H 4.1/6.1 n Samala et al., 2008 KA ICR 9.5:1 y Jeon et al.,2009 KA NIH Swiss 6.5:1 n Hartman et al., 2010 KA ICR 4:1 n Jeong et al., 2011 KA C57BL/6 11.5:1 y Luan et al.,2012 KA NIH Swiss 6.5: 1 n Dolce et al,. 2018
Pilocarpine Acetylcholine receptor agonist. A Iimbic SE. Model Rat KD Efficacy Reference of thae KD Pilocarpine SD 4:1 y Zhoa., 2004 Wister 4:1 n Porta et al., 2009 Wister 6.4:1 y Lusardi et al., 2015 Wister 9:1 y Chwiej et al., 2017
Genetic Models KD Efficacy Reference EL mice Idiopathic Model 4.75:1 y Todorova et al, 2000 Aldh5a1 absence seizures 4:1 y Nylen et al., 2009 SCN1A Kcna1 absence seizures 4:1 y Nylen et al., 2010 Dravet syndrome GEFS+ limbic system seizures 6:1 y Dutton et al, 2011 6:1 y Fenoglio-Simeone et al., 2009 6:1 y Simeone et al., 2014 6:1 y Kim et al, 2015 6:1 y Simeone et al., 2016 6:1 y Simeone et al., 2017 6:1 y Chun et al., 2018 6:1 y Iyer et al., 2018
The KD appears to render anticonvulsant effects in all models of epilepsy, and its action is more favorable with higher fat composition. However, the KD is not anticonvulsant in MES, particularly in rats. Further research will be needed to evaluate the effect of the KD against kainate, particularly in mice. The KD increased seizure thresholds in the 6Hz test (a model of drugresistant seizures), potentially through inhibition of AMPA receptor. The quest or antiseizure mechanisms by the KD can provide insights into new therapeutic interventions in patients with pharmacoresistant epilepsy, and rodent models of epilepsy will continue to be critical in expanding our understanding of how the KD can help patients with epilepsy.
Ketogenic Mechanisms Neurotransmission Inhibitory Neurotransmitters Inhibition of AMPA receptor mtor signaling Regulation of K ATP channel Adenosine Metabolism Regulation Altered gut microbiota Glucose uptake/glycolysis Ketone metabolism Metabolic sensors Neuroprotection/Inflamm ation NLRP3 inflammasome (?) Immune response (?) Anti-inflammation (?) Genomic Effect HDACs inhibition PPARgamma DNA methylation Mitochondrial regulation mpt Biogenesis
Kcna1-null mice appear relatively normal at birth at P14-16, exhibit spontaneous limbic seizures (epilepsy) Learning and memory deficits death associated with seizures Inflammation Social Persistent Conse Seizures -quences Refractory Epilepsy Increased Cognitive Mortality & SUDEP Adverse Effects of ASDs Problems
Kim et al, 2012
Given the rising concerns for a critical mechanism of neuronal insult and neuroinflammation in epileptogenesis, we hypothesize that the anti-seizure and functional protection of the KD might be in part a direct consequence of ketones, specifically via the regulation of inflammation. We ask whether chronic administration of the KD can render anticonvulsant properties and preservation of hippocampal structural integrity in epileptic Kcna1-null mice; and these actions may affect the alternation of astrocyte and microglial activation, more specific expression profiling of glial fibrillary acidic protein (GFAP), microglia marker (Iba), and pro-inflammatory mediators in spontaneous epileptic Kcna1-null mice.
Unpublished data
Unpublished data
Elucidation mechanisms of dietary effects on functional protection will enhance knowledge of, and possibly treatment for, other pathological conditions that may respond to the KD and its clinical variants. Ultimately, such information will likely provide new insights for future therapies for pharmacoresistant epilepsy and for important related comorbidities such as SUDEP and cognitive dysfunction.