Free Floating Microcultures of Human Fetal Ventral Mesencephalon

327 In the reported clinical trials of human fetal ventral mesencephalic transplants to the brain of Parkinson patients the tissue was transplanted either as small solid pieces or as cell suspensions. In all cases the tissues were implanted immediately after explantation. Delayed surgery and the in vitro maintenance of the tissue prior to transplantation would present several advantages. For example, this would allow one to estimate the approximate number of neurons transplanted and to measure dopamine content and tyrosine-hydroxylase (TH) activity in vitro expressing the activity of the cultures. Furthermore tissues in vitro could be treated with various growth factors. Here we present a protocol to maintain human fetal ventral mesencephalic tissue in free floating microcultures. Ventral mesencephalon of human fetuses was obtained from material of induced suction abortions. Out of 38 abortions the mesencephalon could be identified in 16 cases (50% in fetuses age 8-10 weeks post mens. and 31% in fetuses age 11-13 weeks post mens.) based on macroscopic criteria. The landmarks which allowed identification were first the isthmus rhombencephali and second the IV ventricle (caudal to the mesencephalon) and the recessus ventriculi III (rostral to the mesencephalon) as well as the mesencephalic flexure. However the tissue was most often fractured at the isthmus. The ventral mesencephalon was dissected out of the identified brain tissue and divided into four longitudinal pieces. These pieces were