In vitro preparations (defined here as cultured cells, brain slices, and isolated whole brains) offer a variety of approaches to modeling various aspects of seizures and epilepsy. Such models are particularly amenable to the application of anti-seizure compounds, and consequently are a valuable tool to screen the mechanisms of epileptiform activity, mode of action of known anti-seizure medications (ASMs), and the potential efficacy of putative new anti-seizure compounds. Despite these applications, all disease models are a simplification of reality and are therefore subject to limitations. In this review, we summarize the main types of in vitro models that can be used in epilepsy research, describing key methodologies as well as notable advantages and disadvantages of each. We argue that a well-designed battery of in vitro models can form an effective and potentially high-throughput screening platform to predict the clinical usefulness of ASMs, and that in vitro models are particularly useful for interrogating mechanisms of ASMs. To conclude, we offer several key recommendations that maximize the potential value of in vitro models in ASM screening. This includes the use of multiple in vitro tests that can complement each other, carefully combined with in vivo studies, the use of tissues from chronically epileptic (rather than naïve wild-type) animals, and the integration of human cell/tissue-derived preparations.

Department of Neuroscience Physiology and Pharmacology University College London London UK

Department of Neuroscience Tufts University School of Medicine Boston Massachusetts USA

Department of Pharmacology University of Oxford Oxford UK

Department of Physiology 2nd Medical School Motol Charles University Prague Czech Republic

Department of Physiology McGill University Montréal Quebec Canada

Discipline of Physiology School of Medicine Trinity College Dublin Dublin 2 Ireland

Division of Neuroscience Faculty of Biology Medicine and Health School of Biological Sciences Manchester Academic Health Science Centre University of Manchester Manchester UK

Epilepsy Unit Fondazione IRCCS Istituto Neurologico Carlo Besta Milan Italy

Inserm INS Institut de Neurosciences des Systèmes Aix Marseille Univ Marseille France

Laboratory of Animal and Human Physiology Department of Biological Applications and Technology Faculty of Health Sciences University of Ioannina Ioannina Greece

Montreal Neurological Institute Hospital and Departments of Neurology and Neurosurgery McGill University Montréal Quebec Canada

Neurology Department Massachusetts General Hospital Harvard Medical School Boston Massachusetts USA

Zobrazit více v PubMed

WHO. Epilepsy factsheet. 2019.

Ngugi AK, Bottomley C, Kleinschmidt I, Sander JW, Newton CR. Estimation of the burden of active and life-time epilepsy: a meta-analytic approach. Epilepsia. 2010;51(5):883-890.

Mello LEAM, Cavalheiro EA, Tan AM, Kupfer WR, Pretorius JK, Babb TL, et al. Circuit mechanisms of seizures in the pilocarpine model of chronic epilepsy: cell loss and mossy fiber sprouting. Epilepsia. 1993;34(6):985-995.

Vezzani A, French J, Bartfai T, Baram TZ. The role of inflammation in epilepsy. Nat Rev Neurol. 2011;7(1):31-40.

Devinsky O, Vezzani A, Najjar S, de Lanerolle NC, Rogawski MA. Glia and epilepsy: excitability and inflammation. Trends Neurosci [Internet]. 2013;36(3):174-184. https://doi.org/10.1016/j.tins.2012.11.008

Sutula T, Xiao-Xian H, Cavazos J, Scott G. Synaptic reorganization in the hippocampus induced by abnormal functional activity. Science. 1988;239(4844):1147-1150.

Symonds JD, Elliott KS, Shetty J, Armstrong M, Brunklaus A, Cutcutache I, et al. Early childhood epilepsies: epidemiology, classification, aetiology, and socio-economic determinants. Brain. 2021;144(9):2879-2891.

Brennan GP, Henshall DC. MicroRNAs as regulators of brain function and targets for treatment of epilepsy. Nat Rev Neurol [Internet]. 2020;16(9):506-519.

Venø MT, Reschke CR, Morris G, Connolly NMC, Su J, Yan Y, et al. A systems approach delivers a functional microRNA catalog and expanded targets for seizure suppression in temporal lobe epilepsy. Proc Natl Acad Sci USA. 2020;117(27):15977-15988.

Mula M, Kanner AM, Jetté N, Sander JW. Psychiatric comorbidities in people with epilepsy. Neurol Clin Pract [Internet]. 2021;11(2):e112-e120.

Löscher W. Single-target versus multi-target drugs versus combinations of drugs with multiple targets: preclinical and clinical evidence for the treatment or prevention of epilepsy. Front Pharmacol. 2021;12:1-22.

Chen Z, Brodie MJ, Liew D, Kwan P. Treatment outcomes in patients with newly diagnosed epilepsy treated with established and new antiepileptic drugs a 30-year longitudinal cohort study. JAMA Neurol. 2018;75(3):279-286.

Janmohamed M, Brodie MJ, Kwan P. Pharmacoresistance - epidemiology, mechanisms, and impact on epilepsy treatment. Neuropharmacology. 2020;168:107790.

Archie SR, Al SA, Cucullo L. Blood-brain barrier dysfunction in CNS disorders and putative therapeutic targets: an overview. Pharmaceutics. 2021;13(11): 1779.

Oby E, Janigro D. The blood-brain barrier and epilepsy. Epilepsia. 2006;47(11):1761-1774.

Lidster K, Jefferys JG, Blümcke I, Crunelli V, Flecknell P, Frenguelli BG, et al. Opportunities for improving animal welfare in rodent models of epilepsy and seizures. J Neurosci Methods [Internet]. 2016;260:2-25.

Percie du Sert N, Hurst V, Ahluwalia A, Alam S, Avey MT, Baker M, et al. The ARRIVE guidelines 2.0: updated guidelines for reporting animal research. PLoS Biol [Internet]. 2020;18(7):e3000410. https://doi.org/10.1371/journal.pbio.3000410

Raimondo JV, Heinemann U, de Curtis M, Goodkin HP, Dulla CG, Janigro D, et al. Methodological standards for in vitro models of epilepsy and epileptic seizures. A TASK1-WG4 report of the AES/ILAE Translational TASK Force of the ILAE. Epilepsia. 2017;58:40-52.

Dulla CG, Janigro D, Jiruska P, Raimondo JV, Ikeda A, Lin C-CK, et al. How do we use in vitro models to understand epileptiform and ictal activity? A report of the TASK1-WG4 group of the ILAE/AES Joint Translational TASK Force. Epilepsia Open. 2018;3(4):460-473.

Bernard C. Hippocampal slices: designing and interpreting studies in epilepsy research. Models of seizures and epilepsy. London: Elsevier Inc.; 2006. p. 59-72.

Colin-Le Brun I, Ferrand N, Caillard O, Tosetti P, Ben-Ari Y, Gaïarsa J-L. Spontaneous synaptic activity is required for the formation of functional GABAergic synapses in the developing rat hippocampus. J Physiol. 2004;559(Pt 1):129-139.

Khalilov I, Holmes GL, Ben-Ari Y. In vitro formation of a secondary epileptogenic mirror focus by interhippocampal propagation of seizures. Nat Neurosci. 2003;6(10):1079-1085.

Empson RM, Heinemann U. The perforant path projection to hippocampal area CA1 in the rat hippocampal-entorhinal cortex combined slice. J Physiol. 1995;484(3):707-720.

von Bohlen und Halbach O, Albrecht D. Reciprocal connections of the hippocampal area CA1, the lateral nucleus of the amygdala and cortical areas in a combined horizontal slice preparation. Neurosci Res. 2002;44(1):91-100.

Barbarosie M, Avoli M. CA3-driven hippocampal-entorhinal loop controls rather than sustains in vitro limbic seizures. J Neurosci. 1997;17(23):9308-9314.

D'Antuono M, Benini R, Biagini G, D'Arcangelo G, Barbarosie M, Tancredi V, et al. Limbic network interactions leading to hyperexcitability in a model of temporal lobe epilepsy. J Neurophysiol. 2002;87(1):634-639.

Borck C, Jefferys JG. Seizure-like events in disinhibited ventral slices of adult rat hippocampus. J Neurophysiol. 1999;82(5):2130-2142.

Traynelis SF, Dingledine R. Potassium-induced spontaneous electrographic seizures in the rat hippocampal slice. J Neurophysiol [Internet]. 1988;59:259-276.

Morris G, Jiruska P, Jefferys JGR, Powell AD. A new approach of modified submerged patch clamp recording reveals interneuronal dynamics during epileptiform oscillations. Front Neurosci. 2016;10:519.

Mody I, Lambert JDC, Heinemann U. Low extracellular magnesium induces epileptiform activity and spreading depression in rat hippocampal slices. J Neurophysiol. 1987;57(3):869-888.

Jefferys JGR, Haas HL. Synchronized bursting of CA1 hippocampal pyramidal cells in the absence of synaptic transmission. Nature. 1982;300(5891):448-450.

Fueta Y, Avoli M. Effects of antiepileptic drugs on 4-aminopyridine-induced epileptiform activity in young and adult rat hippocampus. Epilepsy Res. 1992;12(3):207-215.

Straub H, Köhling R, Speckmann EJ. Picrotoxin-induced epileptic activity in hippocampal and neocortical slices (Guinea pig): suppression by organic calcium channel blockers. Brain Res. 1994;658(1-2):119-126.

Khalilov I, Khazipov R, Esclapez M, Ben-Ari Y. Bicuculline induces ictal seizures in the intact hippocampus recorded in vitro. Eur J Pharmacol. 1997;319(2-3):5-6.

Bingmann D, Speckmann E-J. Actions of pentylenetetrazol (PTZ) on CA3 neurons in hippocampal slices of Guinea pigs. Exp Brain Res. 1986;64(1):94-104.

Fujiwara-Tsukamoto Y, Isomura Y, Nambu A, Takada M. Excitatory gaba input directly drives seizure-like rhythmic synchronization in mature hippocampal CA1 pyramidal cells. Neuroscience. 2003;119(1):265-275.

Ridler T, Matthews P, Phillips KG, Randall AD, Brown JT. Initiation and slow propagation of epileptiform activity from ventral to dorsal medial entorhinal cortex is constrained by an inhibitory gradient. J Physiol. 2018;596(11):2251-2266.

Luhmann HJ, Dzhala VI, Ben-Ari Y. Generation and propagation of 4-AP-induced epileptiform activity in neonatal intact limbic structures in vitro. Eur J Neurosci. 2000;12(8):2757-2768.

Mikroulis A, Psarropoulou C. Endogenous ACh effects on NMDA-induced interictal-like discharges along the septotemporal hippocampal axis of adult rats and their modulation by an early life generalized seizure. Epilepsia. 2012;53(5):879-887.

Chan F, Lax NZ, Voss CM, Aldana BI, Whyte S, Jenkins A, et al. The role of astrocytes in seizure generation: insights from a novel in vitro seizure model based on mitochondrial dysfunction. Brain. 2019;142(2):391-411.

Nikkanen J, Forsström S, Euro L, Paetau I, Kohnz RA, Wang L, et al. Mitochondrial DNA replication defects disturb cellular dNTP pools and remodel one-carbon metabolism. Cell Metab. 2016;23(4):635-648.

Jirsa VK, Stacey WC, Quilichini PP, Ivanov AI, Bernard C. On the nature of seizure dynamics. Brain [Internet]. 2014;137(8):2210-2230.

Graham RT, Parrish RR, Alberio L, Johnson EL, Owens L, Trevelyan AJ. Optogenetic stimulation reveals a latent tipping point in cortical networks during ictogenesis. Brain. 2022;146(7):2814-2827.

Le Duigou C, Bouilleret V, Miles R. Epileptiform activities in slices of hippocampus from mice after intra-hippocampal injection of kainic acid. J Physiol. 2008;586(20):4891-4904.

West PJ, Saunders GW, Billingsley P, Smith MD, White HS, Metcalf CS, et al. Recurrent epileptiform discharges in the medial entorhinal cortex of kainate-treated rats are differentially sensitive to antiseizure drugs. Epilepsia. 2018;59(11):2035-2048.

Jefferys JG. Chronic epileptic foci in vitro in hippocampal slices from rats with the tetanus toxin epileptic syndrome. J Neurophysiol. 1989;62(2):458-468.

Stein RE, Kaplan JS, Li J, Catterall WA. Hippocampal deletion of NaV1.1 channels in mice causes thermal seizures and cognitive deficit characteristic of Dravet syndrome. Proc Natl Acad Sci USA. 2019;116(33):16571-16576.

Jones RSG, da Silva AB, Whittaker RG, Woodhall GL, Cunningham MO. Human brain slices for epilepsy research: pitfalls, solutions and future challenges. J Neurosci Methods [Internet]. 2016;260:221-232. https://doi.org/10.1016/j.jneumeth.2015.09.021

Gabriel S, Njunting M, Pomper JK, Merschhemke M, Sanabria ERG, Eilers A, et al. Stimulus and potassium-induced epileptiform activity in the human dentate gyrus from patients with and without hippocampal sclerosis. J Neurosci. 2004;24(46):10416-10430.

Huberfeld G, Blauwblomme T, Miles R. Hippocampus and epilepsy: findings from human tissues. Rev Neurol (Paris). 2015;171(3):236-251.

Toyoda I, Bower MR, Leyva F, Buckmaster PS. Early activation of ventral hippocampus and subiculum during spontaneous seizures in a rat model of temporal lobe epilepsy. J Neurosci. 2013;33(27):11100-11115.

Sheybani L, van Mierlo P, Birot G, Michel CM, Quairiaux C. Large-scale 3-5 Hz oscillation constrains the expression of neocortical fast ripples in a mouse model of mesial temporal lobe epilepsy. eNeuro. 2019;6(1):ENEURO.0494-18.2019.

Khalilov I, Esclapez M, Medina I, Aggoun D, Lamsa K, Leinekugel X, et al. A novel in vitro preparation: the intact hippocampal formation. Neuron. 1997;19:743-749.

Quilichini PP, Diabira D, Chiron C, Ben-Ari Y, Gozlan H. Persistent epileptiform activity induced by low Mg2+ in intact immature brain structures. Eur J Neurosci. 2002;16(5):850-860.

Quilichini PP, Diabira D, Chiron C, Milh M, Ben-Ari Y, Gozlan H. Effects of antiepileptic drugs on refractory seizures in the intact immature corticohippocampal formation in vitro. Epilepsia. 2003;44(11):1365-1374.

Mühlethaler M, de Curtis M, Walton K, Llinás R. The isolated and perfused brain of the guinea-pig in vitro. Eur J Neurosci. 1993;5(7):915-926.

Hájos N, Ellender TJ, Zemankovics R, Mann EO, Exley R, Cragg SJ, et al. Maintaining network activity in submerged hippocampal slices: importance of oxygen supply. Eur J Neurosci. 2009;29(2):319-327.

de Curtis M, Takashima I, Iijima T. Optical recording of cortical activity after in vitro perfusion of cerebral arteries with a voltage-sensitive dye. Brain Res. 1999;837(1-2):314-319.

Biella G, Spaiardi P, Toselli M, de Curtis M, Gnatkovsky V. Functional interactions within the parahippocampal region revealed by voltage-sensitive dye imaging in the isolated guinea pig brain. J Neurophysiol. 2010;103(2):725-732.

Federico P, Borg SG, Salkauskus AG, MacVicar BA. Mapping patterns of neuronal activity and seizure propagation by imaging intrinsic optical signals in the isolated whole brain of the guinea-pig. Neuroscience. 1994;58(3):461-480.

Gómez-Gonzalo M, Losi G, Chiavegato A, Zonta M, Cammarota M, Brondi M, et al. An excitatory loop with astrocytes contributes to drive neurons to seizure threshold. PLoS Biol. 2010;8(4):e1000352.

de Curtis M, Librizzi L, Uva L, Gnatkovsky V. Neuronal networks in the in vitro isolated guinea pig brain. Totowa, NJ: Humana Press; 2012. p. 357-383.

Mazzetti S, Librizzi L, Frigerio S, de Curtis M, Vitellaro-Zuccarello L. Molecular anatomy of the cerebral microvessels in the isolated Guinea-pig brain. Brain Res. 2004;999(1):81-90.

Librizzi L, Janigro D, de Biasi S, de Curtis M. Blood-brain barrier preservation in the in vitro isolated Guinea pig brain preparation. J Neurosci Res. 2001;66(2):289-297.

van Vliet EA, Marchi N. Neurovascular unit dysfunction as a mechanism of seizures and epilepsy during aging. Epilepsia. 2022;63(6):1297-1313.

Uva L, Librizzi L, Marchi N, Noe F, Bongiovanni R, Vezzani A, et al. Acute induction of epileptiform discharges by pilocarpine in the in vitro isolated guinea-pig brain requires enhancement of blood-brain barrier permeability. Neuroscience. 2008;151(1):303-312.

Librizzi L, Vila Verde D, Colciaghi F, Deleo F, Regondi MC, Costanza M, et al. Peripheral blood mononuclear cell activation sustains seizure activity. Epilepsia. 2021;62(7):1715-1728.

Librizzi L, Noè F, Vezzani A, de Curtis M, Ravizza T. Seizure-induced brain-borne inflammation sustains seizure recurrence and blood-brain barrier damage. Ann Neurol. 2012;72(1):82-90.

Librizzi L, de Cutis M, Janigro D, Runtz L, de Bock F, Barbier EL, et al. Cerebrovascular heterogeneity and neuronal excitability. Neurosci Lett. 2018;667:75-83.

Librizzi L, Pastori C, de Grazia U, Croci D, de Curtis M. Rapid in vitro elimination of anesthetic doses of thiopental in the isolated Guinea pig brain. Neurosci Lett. 2005;380(1-2):66-69.

Gähwiler BH, Capogna M, Debanne D, McKinney RA, Thompson SM. Organotypic slice cultures: a technique has come of age. Trends Neurosci. 1997;20(10):471-477.

Stoppini L, Buchs PA, Muller D. A simple method for organotypic cultures of nervous tissue. J Neurosci Methods. 1991;37(2):173-182.

Humpel C. Neuroscience forefront review organotypic brain slice cultures: a review. Neuroscience. 2015;305:86-98.

Morris G, O'Brien D, Henshall DC. Opportunities and challenges for microRNA-targeting therapeutics for epilepsy. Trends Pharmacol Sci [Internet]. 2021;42(7):605-616. https://doi.org/10.1016/j.tips.2021.04.007

Snowball A, Chabrol E, Wykes RC, Shekh-Ahmad T, Cornford JH, Lieb A, et al. Epilepsy gene therapy using an engineered potassium channel. J Neurosci. 2019;39(16):3159-3169.

Kullmann DM, Schorge S, Walker MC, Wykes RC. Gene therapy in epilepsy-is it time for clinical trials? Nat Rev Neurol [Internet]. 2014;10(5):300-304.

McBain CJ, Boden P, Hill RG. Rat hippocampal slices “in vitro” display spontaneous epileptiform activity following long-term organotypic culture. J Neurosci Methods. 1989;27(1):35-49.

Lau LA, Staley KJ, Lillis KP. In vitro ictogenesis is stochastic at the single neuron level. Brain. 2022;145(2):531-541.

Berdichevsky Y, Saponjian Y, Park K, Roach B, Pouliot W, Lu K, et al. Staged anticonvulsant screening for chronic epilepsy. Ann Clin Transl Neurol. 2016;3(12):908-923.

Obergrussberger A, Rinke-Weiß I, Goetze TA, Rapedius M, Brinkwirth N, Becker N, et al. The suitability of high throughput automated patch clamp for physiological applications. J Physiol. 2022;600(2):277-297.

Vanoye CG, Desai RR, Ji Z, Adus S, Jairam N, Joshi N, et al. High-throughput evaluation of epilepsy-associated KCNQ2 variants reveals functional and pharmacological heterogeneity. JCI Insight. 2021;7:e156314.

Barilli A, Aldegheri L, Bianchi F, Brault L, Brodbeck D, Castelletti L, et al. From high-throughput screening to target validation: benzo[ d]isothiazoles as potent and selective agonists of human transient receptor potential cation channel subfamily M member 5 possessing in vivo gastrointestinal prokinetic activity in rodents. J Med Chem. 2021;64(9):5931-5955.

Bar-Yehuda D, Korngreen A. Space-clamp problems when voltage clamping neurons expressing voltage-gated conductances. J Neurophysiol. 2008;99(3):1127-1136.

Neher E, Sakmann B. Single-channel currents recorded from membrane. Nature [Internet]. 1976;260:799-802.

Marguet SL, Le-Schulte VTQ, Merseburg A, Neu A, Eichler R, Jakovcevski I, et al. Treatment during a vulnerable developmental period rescues a genetic epilepsy. Nat Med. 2015;21(12):1436-1444.

Marder E, Taylor AL. Multiple models to capture the variability in biological neurons and networks. Nat Neurosci. 2011;14(2):133-138.

Saneto RP. Preparation of highly purified populations of neurons, astrocytes, and oligodendrocytes. Methods Neurosci. 1990;2:119-133.

Furshpan EJ, Potter DD. Seizure-like activity and cellular damage in rat hippocampal neurons in cell culture. Neuron. 1989;3(2):199-207.

Segal MM. Epileptiform activity in microcultures containing one excitatory hippocampal neuron. J Neurophysiol. 1991;65(4):761-770.

Wei Y, Ullah G, Schiff SJ. Unification of neuronal spikes, seizures, and spreading depression. J Neurosci. 2014;34(35):11733-11743.

Gonzalez-Sulser A, Wang J, Motamedi GK, Avoli M, Vicini S, Dzakpasu R. The 4-aminopyridine in vitro epilepsy model analyzed with a perforated multi-electrode array. Neuropharmacology. 2011;60(7-8):1142-1153.

Gonzalez-Sulser A, Wang J, Queenan BN, Avoli M, Vicini S, Dzakpasu R. Hippocampal neuron firing and local field potentials in the in vitro 4-aminopyridine epilepsy model. J Neurophysiol. 2012;108(9):2568-2580.

Morris G, Leite M, Kullmann D, Pavlov I, Schorge S, Lignani G, et al. Activity clamp provides insights into paradoxical effects of the anti-seizure drug carbamazepine. J Neurosci [Internet]. 2017;37(22):5484-5495.

Kovac S, Domijan A-M, Walker MC, Abramov AY. Seizure activity results in calcium- and mitochondria-independent ROS production via NADPH and xanthine oxidase activation. Cell Death Dis. 2014;5(10):e1442.

Kovac S, Domijan A-M, Walker MC, Abramov AY. Prolonged seizure activity impairs mitochondrial bioenergetics and induces cell death. J Cell Sci. 2012;125:1796-1806.

Malik N, Rao MS. A review of the methods for human iPSC derivation. Methods Mol Biol. 2013;997(5):23-33.

Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126(4):663-676.

Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell. 2007;131(5):861-872.

Shi Y, Inoue H, Wu JC, Yamanaka S. Induced pluripotent stem cell technology: a decade of progress. Nat Rev Drug Discov. 2017;16(2):115-130.

Lancaster MA, Corsini NS, Wolfinger S, Gustafson EH, Phillips AW, Burkard TR, et al. Guided self-organization and cortical plate formation in human brain organoids. Nat Biotechnol. 2017;35(7):659-666.

Fair SR, Julian D, Hartlaub AM, Pusuluri ST, Malik G, Summerfied TL, et al. Electrophysiological maturation of cerebral organoids correlates with dynamic morphological and cellular development. Stem Cell Reports. 2020;15(4):855-868.

Samarasinghe RA, Miranda OA, Buth JE, Mitchell S, Ferando I, Watanabe M, et al. Identification of neural oscillations and epileptiform changes in human brain organoids. Nat Neurosci. 2021;24(10):1488-1500.

Steinberg DJ, Repudi S, Saleem A, Kustanovich I, Viukov S, Abudiab B, et al. Modeling genetic epileptic encephalopathies using brain organoids. EMBO Mol Med. 2021;13(8):e13610.

Qiu Y, O'Neill N, Maffei B, Zourray C, Almacellas-Barbanoj A, Carpenter JC, et al. On-demand cell-autonomous gene therapy for brain circuit disorders. Science. 2022;378(6619):523-532.

Morris G, Rowell R, Cunningham MO. Limitations of animal epilepsy research models: can epileptic human tissue provide translational benefit? ALTEX [Internet]. 2021;38:451-462.

Jobst BC, Cascino GD. Resective epilepsy surgery for drug-resistant focal epilepsy: a review. JAMA. 2015;313(3):285-293.

Wickham J, Brödjegård NG, Vighagen R, Pinborg LH, Bengzon J, Woldbye DPD, et al. Prolonged life of human acute hippocampal slices from temporal lobe epilepsy surgery. Sci Rep. 2018;8(1):1-13.

Morris G, Langa E, Fearon C, Conboy K, Lau E-How K, Sanz-Rodriguez A, et al. MicroRNA inhibition using antimiRs in acute human brain tissue sections. Epilepsia. 2022;63:e92-e99.

Schwarz N, Uysal B, Welzer M, Bahr JC, Layer N, Löffler H, et al. Long-term adult human brain slice cultures as a model system to study human CNS circuitry and disease. Elife. 2019;8:1-26.

Schwarz N, Hedrich UBS, Schwarz H, Harshad PA, Dammeier N, Auffenberg E, et al. Human cerebrospinal fluid promotes long-term neuronal viability and network function in human neocortical organotypic brain slice cultures. Sci Rep. 2017;7(1):1-12.

McLeod F, Dimtsi A, Marshall AC, Lewis-Smith D, Thomas R, Clowry GJ, et al. Altered synaptic connectivity in an in vitro human model of STXBP1 encephalopathy. Brain. 2022;146:850-857.

Kovács R, Raue C, Gabriel S, Heinemann U. Functional test of multidrug transporter activity in hippocampal-neocortical brain slices from epileptic patients. J Neurosci Methods. 2011;200(2):164-172.

Roopun AK, Simonotto JD, Pierce ML, Jenkins A, Nicholson C, Schofield IS, et al. A nonsynaptic mechanism underlying interictal discharges in human epileptic neocortex. Proc Natl Acad Sci USA. 2010;107(1):338-343.

Pallud J, le Van Quyen M, Bielle F, Pellegrino C, Varlet P, Labussiere M, et al. Cortical GABAergic excitation contributes to epileptic activities around human glioma. Sci Transl Med. 2014;6(244):244ra89.

Huberfeld G, Menendez de la Prida L, Pallud J, Cohen I, le Van Quyen M, Adam C, et al. Glutamatergic pre-ictal discharges emerge at the transition to seizure in human epilepsy. Nat Neurosci. 2011;14(5):627-634.

Schwartzkroin PA, Haglund MM. Spontaneous rhythmic synchronous activity in epileptic human and Normal monkey temporal lobe. Epilepsia. 1986;27(5):523-533.

Köhling R, Lücke A, Straub H, Speckmann EJ, Tuxhorn I, Wolf P, et al. Spontaneous sharp waves in human neocortical slices excised from epileptic patients. Brain. 1998;121(Pt 6):1073-1087.

Taing KD, O'Brien TJ, Williams DA, French CR. Anti-epileptic drug combination efficacy in an in vitro seizure model - phenytoin and valproate, lamotrigine and valproate. PLoS One. 2017;12(1):e0169974.

Whittington MA, Traub RD, Jefferys JG. Erosion of inhibition contributes to the progression of low magnesium bursts in rat hippocampal slices. J Physiol. 1995;486(3):723-734.

Dreier JP, Zhang C-L, Heinemann U. Phenytoin, phenobarbital, and midazolam fail to stop status epilepticus-like activity induced by low magnesium in rat entorhinal slices, but can prevent its development. Acta Neurol Scand. 1998;98(3):154-160.

Burman RJ, Selfe JS, Lee JH, van den Berg M, Calin A, Codadu NK, et al. Excitatory GABAergic signalling is associated with benzodiazepine resistance in status epilepticus. Brain. 2019;142(11):3482-3501.

D'Antuono M, Köhling R, Ricalzone S, Gotman J, Biagini G, Avoli M. Antiepileptic drugs abolish ictal but not interictal epileptiform discharges in vitro. Epilepsia. 2010;51(3):423-431.

Avoli M, de Curtis M. GABAergic synchronization in the limbic system and its role in the generation of epileptiform activity. Prog Neurobiol. 2011;95(2):104-132.

Heuzeroth H, Wawra M, Fidzinski P, Dag R, Holtkamp M. The 4-aminopyridine model of acute seizures in vitro elucidates efficacy of new antiepileptic drugs. Front Neurosci. 2019;13:677.

Morris G, Heiland M, Lamottke K, Guan H, Hill TDM, Zhou Y, et al. BICS01 mediates reversible anti-seizure effects in brain slice models of epilepsy. Front Neurol. 2022;12:791608.

Hall BJ, Satterfield-Doerr M, Parikh AR, Brodbelt JS. Determination of cannabinoids in water and human saliva by solid-phase microextraction and quadrupole ion trap gas chromatography/mass spectrometry. Anal Chem. 1998;70(9):1788-1796.

Mantegazza M, Curia G, Biagini G, Ragsdale DS, Avoli M. Voltage-gated sodium channels as therapeutic targets in epilepsy and other neurological disorders. Lancet Neurol. 2010;9:413-424.

Ogiwara I, Miyamoto H, Morita N, Atapour N, Mazaki E, Inoue I, et al. Nav1.1 localizes to axons of parvalbumin-positive inhibitory interneurons: a circuit basis for epileptic seizures in mice carrying an Scn1a gene mutation. J Neurosci. 2007;27(22):5903-5914.

Gidon A, Zolnik TA, Fidzinski P, Bolduan F, Papoutsi A, Poirazi P, et al. Dendritic action potentials and computation in human layer 2/3 cortical neurons. Science. 2020;367(6473):83-87.

Boldog E, Bakken TE, Hodge RD, Novotny M, Aevermann BD, Baka J, et al. Transcriptomic and morphophysiological evidence for a specialized human cortical GABAergic cell type. Nat Neurosci. 2018;21(9):1185-1195.

Wickham J, Corna A, Schwarz N, Uysal B, Layer N, Honegger JB, et al. Human cerebrospinal fluid induces neuronal excitability changes in resected human neocortical and hippocampal brain slices. Front Neurosci. 2020;14:1-14.

Deisseroth K, Feng G, Majewska AK, Miesenböck G, Ting A, Schnitzer MJ. Next-generation optical technologies for illuminating genetically targeted brain circuits. J Neurosci. 2006;26(41):10380-10386.

Panzera LC, Hoppa MB. Genetically encoded voltage indicators are illuminating subcellular physiology of the axon. Front Cell Neurosci. 2019;13:1-9.

Emiliani V, Cohen AE, Deisseroth K, Häusser M. All-optical interrogation of neural circuits. J Neurosci. 2015;35(41):13917-13926.

Nakai J, Ohkura M, Imoto K. A high signal-to-noise Ca(2+) probe composed of a single green fluorescent protein. Nat Biotechnol. 2001;19(2):137-141.

Marvin JS, Borghuis BG, Tian L, Cichon J, Harnett MT, Akerboom J, et al. An optimized fluorescent probe for visualizing glutamate neurotransmission. Nat Methods. 2013;10(2):162-170.

Marvin JS, Shimoda Y, Magloire V, Leite M, Kawashima T, Jensen TP, et al. A genetically encoded fluorescent sensor for in vivo imaging of GABA. Nat Methods. 2019;16(8):763-770.

Zaaimi B, Turnbull M, Hazra A, Wang Y, Gandara C, McLeod F, et al. Closed-loop optogenetic control of the dynamics of neural activity in non-human primates. Nat Biomed Eng. 2022;7(4):559-575.

Sohal VS, Zhang F, Yizhar O, Deisseroth K. Parvalbumin neurons and gamma rhythms enhance cortical circuit performance. Nature. 2009;459(7247):698-702.

Morris G, Schorge S. Gene therapy for neurological disease: state of the art and opportunities for next-generation approaches. Neuroscience. 2022;490:309-314.