Setting Up of Primary Cultures from Fetal Dog Brain: An Alternative Method in Line with the 3Rs Principles

In the last years, alternatives to animal testing were applied, aimed at reduce the laboratory use of animals, in agreement with the 3Rs principles (Reduction, Refinement and Replacement). In the approach to the study of neurological disorders, several in vitro models and alternative animal species like mammals with a more complex nervous system have been proposed as translational model for human. Lately, canines have been considered an excellent model for study on neurodegenerative disorders since they may develop central nervous system disease close to those observed in humans. In the present report, we describes a method to obtaining primary cultures from dog fetuses brain collected accidentally and cryopreserved until the use. The brain tissues were dissociated through a mild enzymatic papain system and cultured, and the development, morphology and the amount of neuronal cells were assessed. We appreciated the capability of cryopreserved brain tissues to provide a vital and proliferating neuronal primary culture. Moreover, we noticed which explants from early gestation days fetuses (25 days) yielded the higher number of neurons in comparison with those obtained from older fetuses (45days). Our data may suggest a good alternative experimental approach to obtain a primary neuronal culture, without necessarily having to use experimental animals.