We use primary cultures of astrocytes and neurons derived from wildtype animals or animal models of MDD as far as they represent an invaluable source of information to investigate cell-autonomous and non-autonomous specific responses of these cells to pharmacological treatments.
To investigate morphologic changes occurring in astrocytes and neurons upon treatments we take advantage of time lapse imaging methods. Using GFP or RFP-tagged proteins expressed in specific cellular compartments (i.e. synapses) we can directly follow changes in distinct cellular compartments in a live imaging setting.
For the identification of the main molecular targets involved in cell-autonomous responses to pharmacologic treatments, a wide array of molecular methods are used: microarray screening, real-time PCR (qPCR), in situ hybridization, cloning for the preparation of several types of constructs (i.e. fluorescent-tagged constructs to perform live imaging or to induce target-specific knockout/knockdown via RNA interference).
Immunochemistry in either cell culture models or on brain slices is performed to either confirm findings obtained with screening approaches on protein level or to analyze differences in the distributions of candidate molecules in different cells/regions of brains from animal models used for in vivo studies.
The examination of changes occurring at the protein level is performed using Western blots, immunoprecipitation (to verify specific protein-protein interactions that may be influenced by drug treatments), ELISA (to examine proteins released by cells upon treatments).
With the help of the strong translational basics of the projects, the cooperation among groups at the Department will be used to transfer pre-clinical studies in the clinical context to verify results in a translational concept.