Poster presentation at the SIBPA 2021 Congress
In occasion of the SIBPA 2021 Congress, organized by the Italian Society of Pure and Applied Biophysics-SIBPA, IBF-CNR and ETT presented the poster entitled “Pharmacological treatment option for a genetic form of migraine assayed in mouse cortical
neuronal cultures”. The presenting Author, Dr Raffaella Barbieri – IBF CNR, was awarded the 3Brain prize dedicated to “Neuro-biophysics and biophysics of excitable systems”.
In vitro cultured neuronal networks are a valuable experimental model to unravel many aspects of brain physiology. However, it is unclear to which extent cultured neurons can reliably model genetic diseases. Indeed the establishment of an in vitro model based on cultured neurons presents some limitations needed to be taken into consideration. On the one hand, 2D-cultures cannot represent the full complexity of the brain. Another concern regards the experimental variability among different preparations of neuronal cultures. Taking these limitations into consideration, in this work, we tested the usefulness of cortical neuronal cultures obtained from genetically modified knockin-mice (KI) mimicking a human monogenetic migraine model. The KI mice recapitulate familial hemiplegic migraine of type 3 (FHM3), a genetic form of migraine with aura caused by gain-of-function mutations in the SCNA1 gene encoding the Nav1.1 channel . In particular, we employed a KI mouse model of FHM3 bearing a mutation in the Scn1a gene encoding the Nav1.1 channel; for this mouse model, neurophysiological phenomena have been recently investigated , disclosing key differences between the heterozygous and the wild-type phenotype. In the present work, brain cortices were dissected from WT and FHM3-KI embryos at E17 and dissociated neurons were cultured in defined medium for up to 25 days. Cultures were then studied by whole-cell and outside-out patch clamp recording and population excitability recording using microelectrode arrays (MEAs). By patch-clamp recording, kinetic parameters of Na current activation and inactivation were determined for single neurons, while using MEA devices electrical activity of the whole cortical network was investigated. In particular, the effect of the Na current blocker GS967 on the spontaneous electrical activity of the network was investigated for the different genotypes (WT, heterozygous KI, homozygous KI). In addition, the expression level of brain Nav channel genes, namely Scn1a, 2a, 3a, and 8a, was estimated by RT-qPCR to detect possible dysregulation caused by FHM3 disease. Overall, the results suggest that the in vitro model can recapitulate several features observed in the FHM3 mouse model and it is worth emphasizing that, despite its limitation, the use of dissociated culture may be a valid alternative method to reduce the number of animals used in experimental studies.
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