The progression of epileptogenesis in cultured juvenile rodent slices

Abstract

The investigation of the brain’s physiological activity is subject to a range of compromises; the fidelity and spatial resolution of data from cells and networks must be balanced with maintaining as natural an environment and as much connectivity as possible. This is further complicated when measuring aberrant cellular and network activity in models of neurological conditions such as epilepsy or performing longitudinal studies looking at drug treatment or the development of recurrent seizure- like events. This thesis will present the deployment and optimisation of an organotypic slice culture preparation, using both rodent tissue and slices created from tissue blocks taken from human paediatric epilepsy patients experiencing refractory seizures. The naïve rodent tissue can be maintained relatively easily in a physiological state or provoked into exhibiting spontaneous recurrent seizure-like events which are responsive to a variety of pharmacological treatments both acutely and over days/weeks of treatment. This approach in rodent slice cultures allows for experimental manipulation of brain tissue rapidly, repeatably and over a longer time-period than an acute slice preparation would allow as slices were viable for testing for up to 2 weeks post extraction. However, human tissue slices were more difficult to obtain and to maintain in culture, making them significantly more difficult to obtain results of pharmacological activity from. The process of optimisation, including the assessment of various culture mediums, different ages of animal, antibiotic testing, and experimenting with new materials for culture substrates, is discussed and the preparation tested with a range of common anti-epileptic drugs, as well as the development of epileptiform activity being followed via cellular and network-level in vitro electrophysiology. The novel antiepileptic drugs that will be tested the tricyclic antidepressant tianeptine and the anorectic dexfenfluramine, as they have both shown potential to be. These drugs were tested both acutely and at a low concentration applied chronically to the organotypic cultures. The results obtained from these studies illustrate the advantage of the culture approach with longitudinal dosing showing promising results in our hands. The finding in this thesis suggest that dexfenfluramine shows promise as a therapeutic treatment for epilepsy, results obtained when testing tianeptine were much more variable, but they do indicate that at administered chronically at low doses tianeptine suppresses seizure activity.