(1) Instituto de Medicina Molecular, Portugal;
(2) Neuropediatrics Department, Hospital de D. Estefânia, CHLC;
(3) Neurology Department, Hospital San Joan de Déu, Barcelona, Spain;
Champalimaud Neuroscience Program Retreat, Beja, 18-22 Junho
VII PhD Students Meeting CAML Lisbon -‐December 5-‐6, 2013
Best Poster Presentation Award
Epileptic encephalopathies (EE) are characterized by refractory seizures and cognitive impairment. The coincidence of developmental delay and epileptic activity, at early developmental time windows, points to a genetic perturbation underlying the neuronal dysfunction. Mutations in genes involved in either GABAergic forebrain development or synaptic function have been identified in EE. Moreover, genes associated with Rett syndrome (RTT), have also been implicated in EE, like Cyclin dependent Kinase-Like 5, CDKL5 and forkhead box G1, FOXG1, suggesting an overlap between these disorders.
We are investigating whether other single gene mutations can be identified within EE/RTT patients, through whole exome sequencing in 50 families.
Starting from results of DNA sequencing in patients with early epileptic encephalopathies, we collect fibroblasts from these patients and obtain induced pluripotent stem cells. Human induced pluripotent stem cells are cultured in special conditions to obtain cortical neural rosettes that are further differentiated into neural progenitors. After 120 days in culture, neurons and glial cells are obtained. Through patch clamp techniques we can detect spontaneous activity and a firing pattern in these neurons.
γ-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in the adult brain. During early development, activation of the chloride- permeable, postsynaptic, GABAA receptors can induce depolarization (excitatory response) and the basal intracellular chloride concentration is determinant for the action of GABA in the developing neurons. Two major contributors to intracellular chloride concentration are NKCC1 (Na+, K+, 2Cl−cotransporter, that accumulates chloride in the cell), and KCC2 (K+, Cl−cotransporter, that extrudes chloride). We found a reduced KCC2/NKCC1 ratio in the cerebrospinal fluid of Rett Syndrome patients suggesting a disturbed process of GABAergic neuronal maturation. We are now investigating the same mechanism in the RTT animal model and in neurons derived from RTT iPS cells.
Key-words: epileptic encephalopathy, autism