Isolation and Characterizations of Neurons and Astrocytes From Rat Brains

Objective. We devised a method to isolate and culture neurons and astrocytes from the same newborn rat pups. Background. The ability to isolate different cells from the brain offers advantages for studying the complexity of the brain and the effects of angiotensin peptides on this system. Culturing of astrocytes and neurons from the same animals will allow us to study the interplay between these cells, in the control of central effects of the renin angiotensin peptides. Methods. Astrocytes and neurons were cultured from 2-3 days old rats by physical dissociation. The cell mixture destined for astrocyte cultures were plated in T-75 flasks while the cell mixtures destined for neurons were cultured in plates/coverslips previously coated with poly-L-lysine. Immunostaining techniques were used to determine the type of cells that were cultured. Results. We successfully cultured astrocytes using this method. Astrocyte cultures stained positive with glial fibrillary acidic protein, a selective marker for astrocytes but, stained negative with an antibody against neurofilament protein, a neuronal marker. These findings suggest that our current cell isolation methods for astrocytes yield relatively pure astrocyte cultures. However, the technique that was used to isolate pure neurons was unsuccessful. Only a few neurons survived the isolation method and were discernible under the microscope in culture. While we have successfully shown that growing of relatively pure astrocytes cultures could be illustrated with immunostaining techniques, the growing of neuronal cultures was unsuccessful, requiring a different protocol. Grants. This study was supported by a HPD Grant.