RESUMO
Dravet syndrome is a catastrophic pediatric epilepsy with severe intellectual disability, impaired social development and persistent drug-resistant seizures. One of its primary monogenic causes are mutations in Nav1.1 (SCN1A), a voltage-gated sodium channel. Here we characterize zebrafish Nav1.1 (scn1Lab) mutants originally identified in a chemical mutagenesis screen. Mutants exhibit spontaneous abnormal electrographic activity, hyperactivity and convulsive behaviours. Although scn1Lab expression is reduced, microarray analysis is remarkable for the small fraction of differentially expressed genes (~3%) and lack of compensatory expression changes in other scn subunits. Ketogenic diet, diazepam, valproate, potassium bromide and stiripentol attenuate mutant seizure activity; seven other antiepileptic drugs have no effect. A phenotype-based screen of 320 compounds identifies a US Food and Drug Administration-approved compound (clemizole) that inhibits convulsive behaviours and electrographic seizures. This approach represents a new direction in modelling pediatric epilepsy and could be used to identify novel therapeutics for any monogenic epilepsy disorder.
Assuntos
Anticonvulsivantes/uso terapêutico , Benzimidazóis/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Canal de Sódio Disparado por Voltagem NAV1.1/metabolismo , Animais , Anticonvulsivantes/farmacologia , Benzimidazóis/farmacologia , Brometos/farmacologia , Diazepam/farmacologia , Dioxolanos/farmacologia , Epilepsias Mioclônicas/tratamento farmacológico , Perfilação da Expressão Gênica , Mutação , Canal de Sódio Disparado por Voltagem NAV1.1/genética , Compostos de Potássio/farmacologia , Convulsões/tratamento farmacológico , Convulsões/genética , Ácido Valproico/farmacologia , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/genética , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Febrile seizures are the most common seizure type in children under the age of five, but mechanisms underlying seizure generation in vivo remain unclear. Animal models to address this issue primarily focus on immature rodents heated indirectly using a controlled water bath or air blower. Here we describe an in vivo model of hyperthermia-induced seizures in larval zebrafish aged 3 to 7 days post-fertilization (dpf). Bath controlled changes in temperature are rapid and reversible in this model. Acute electrographic seizures following transient hyperthermia showed age-dependence, strain independence, and absence of mortality. Electrographic seizures recorded in the larval zebrafish forebrain were blocked by adding antagonists to the transient receptor potential vanilloid (TRPV4) channel or N-methyl-d-aspartate (NMDA) glutamate receptor to the bathing medium. Application of GABA, GABA re-uptake inhibitors, or TRPV1 antagonist had no effect on hyperthermic seizures. Expression of vanilloid channel and glutamate receptor mRNA was confirmed by quantitative PCR analysis at each developmental stage in larval zebrafish. Taken together, our findings suggest a role of heat-activation of TRPV4 channels and enhanced NMDA receptor-mediated glutamatergic transmission in hyperthermia-induced seizures.
Assuntos
Hipertermia Induzida , Receptores de Glutamato/fisiologia , Convulsões/etiologia , Convulsões/metabolismo , Canais de Cátion TRPV/fisiologia , Proteínas de Peixe-Zebra/fisiologia , Potenciais de Ação/genética , Envelhecimento/genética , Animais , Modelos Animais de Doenças , Embrião não Mamífero/fisiologia , Hipertermia Induzida/métodos , RNA Mensageiro/biossíntese , Receptores de Glutamato/biossíntese , Receptores de N-Metil-D-Aspartato/biossíntese , Receptores de N-Metil-D-Aspartato/fisiologia , Convulsões/fisiopatologia , Canais de Cátion TRPV/biossíntese , Canais de Cátion TRPV/genética , Peixe-Zebra , Proteínas de Peixe-Zebra/biossíntese , Proteínas de Peixe-Zebra/genéticaRESUMO
PURPOSE OF REVIEW: Animal models provide a means to investigate fundamental mechanisms of abnormal electrical discharge (i.e., seizures). Understanding the pathogenesis of epilepsy and therapy development have greatly benefited from these models. Here we review recent mouse mutants featuring spontaneous seizures and simpler organisms. RECENT FINDINGS: New genetically engineered mice provide additional insights to cellular mechanisms underlying seizure generation (BK calcium-activated potassium channels and interneuron-expressed sodium channels), genetic interactions that exacerbate seizure phenotype (Scn2a, Kcnq2 and background) and neurodevelopmental influences (Dlx transcription factors). Mutants for neuronal nicotinic acetylcholine receptors, Glut-1 deficiency and aquaporin channels highlight additional seizure phenotypes in mice. Additional models in Caenorhabditis elegans (Lis-1) and Danio rerio (pentylenetetrazole) highlight a reductionist approach. Taking further advantage of 'simple' organisms, antiepileptic drugs and genetic modifiers of seizure activity are being uncovered in Drosophila. SUMMARY: Studies of epilepsy in mutant mice provide a framework for understanding critical features of the brain that regulate excitability. These, and as yet undiscovered, mouse mutants will continue to serve as the foundation for basic epilepsy research. Interestingly, an even greater potential for analyzing epileptic phenotypes may lie in the more widespread use of genetically tractable organisms such as worms, flies and zebrafish.
Assuntos
Modelos Animais de Doenças , Epilepsia/genética , Animais , Dípteros , Avaliação Pré-Clínica de Medicamentos , Epilepsia/fisiopatologia , Peixes , Humanos , Camundongos , NematoidesRESUMO
Neuropeptides are commonly colocalized with classical neurotransmitters, yet there is little evidence for peptidergic neurotransmission in the mammalian CNS. We performed whole-cell patch-clamp recording from rodent thalamic brain slices and repetitively stimulated corticothalamic fibers to strongly activate NPY-containing GABAergic reticular thalamic (RT) neurons. This resulted in long-lasting (approximately 10 sec) feedforward slow IPSPs (sIPSPs) in RT cells, which were mimicked and blocked by NPY1 (Y1) receptor agonists and antagonists, respectively, and were present in wild-type mice but absent in NPY-/- mice. NPYergic sIPSPs were mediated via G-proteins and G-protein-activated, inwardly rectifying potassium channels, as evidenced by sensitivity to GDP-beta-S and 0.1 mm Ba2+. In rat RT neurons, NPYergic sIPSPs were also present but were surprisingly absent in the major synaptic targets of RT, thalamic relay neurons, where instead robust GABA(B) IPSPs occurred. In vitro oscillatory network responses in rat thalamus were suppressed and augmented by Y1 agonists and antagonists, respectively. These findings provide evidence for segregation of postsynaptic actions between two targets of RT cells and support a role for endogenously released NPY within RT in the regulation of oscillatory thalamic responses relevant to sleep and epilepsy.