In-vitro models for studying neurodegenerative diseases based on SH-SY5Y cell line

Abstract

The main objective of this study was to use a variety of biophysical assays such as electrophysiological recordings and flourescent microscopy to evaluate an invitro model based on differentiated SH-SY5Y neuroblastoma cell lines. To validate the modelademonstrationstudywasconductedtoinvestigateeffectsofBrainDerivedNeurotropic Factor (BDNF) on state of transmembrane currents during neurodegeneration caused by Amyloid beta peptides. Two protocols were developed to use BDNF as an adjacent drug to a control group differentiated by Retinoic Acid (RA). Morphological and fluorescent assays, including F-actin staining, were used to verify and characterize cell differentiation and uptake of amyloid beta peptides by the cells. Upon uptake of amyloid beta, whole cell patch clamp technique was used to observe changes in transmembrane current in control and BDNF groups. Furthermore a Hodgkin Huxley based stochastic search engine was employed to extract ion channel composition data from whole cell recording experiments. The results demonstrated that however BDNF significantly increase cell survivability against amyloid beta, the transmembrane currents would be highly altered, indicating possible functional alterations of survived cells in Alzheimers disease. Lipid coated recording which has been presented in this thesis uses abilayerofsynthesizedlipidssuchas1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) to obtain gigaseals before touching the cells with glass micro pipettes. results presented in this thesis represent an initial optimization phase and proof of concept for further development of the technology and increase in its applications.|Keywords : Neurodegeneration, Electrophysiology, Neuroblastoma, Brain Derived Neurotrophic Factor.