gms | German Medical Science

Objective: The human brain is constantly challenged to balance the ongoing electrical activity, which is the basis of many essential processes including the integration of information. A dysregulation of this balance can shift neuronal networks into an overexcited state, ultimately leading to epileptic seizures. The cause of epileptic brain activity is very diverse and the mechanisms participating in the generation and development of an epileptic disorder are very complex. Many studies have used animal models to investigate the cellular and network changes within the CNS occurring during this critical time. However, it is only poorly investigated how far these results can be directly translated to the human brain. Toward these questions we established an organotypic slice culture system based on human access tissue from resective epilepsy surgery, amenable to immuncytochemical and electrophysiological studies.

Methods: Approval of the ethics committee of the University of Tübingen as well as written informed consent from patients was obtained, allowing spare tissue to be included in our study. In several instances removal of cortical tissue outside the epileptic focus is required in order to get access to the pathology. Even though neurons within access tissue have been part of an epileptic brain for many years, data by other groups demonstrate that intrinsic properties of neurons may remain unaffected. Therefore, cortical access tissue was carefully microdissected and resected with only minimal use of bipolar forceps to ensure tissue integrity, transferred into ice-cold aCSF equilibrated with carbogene, and immediately transported to the laboratory. After removal of the pia tissue chunks were trimmed perpendicular to the cortical surface, thick acute slices were prepared using a vibrating microtome, and tissue was cultured under subsequently optimized conditions.

Results: State-of-the-art whole cell patch clamp recordings of human cortical neurons in combination with immunocytochemistry, morphological reconstruction after biocytin filling, and extracellular field recordings were performed. Results demonstrate that optimized culture conditions preserve viability including maintenance of network activity of cortical cultures up to several weeks.

Conclusion: This model system will serve as a platform to study a) the plasticity of human neurons and b) the influence of epilepsy-causing mutations - following virus-mediated overexpression - on the excitability of human neurons in direct comparison to murine organotypic cultures.