CRYOPRESERVED GABAERGIC NEURONS IN CULTURES OF RAT CEREBRAL-CORTEX AND MESENCEPHALON - A COMPARATIVE MORPHOMETRIC STUDY WITH ANTI-GABA ANTIBODIES

Blocks of embryonic rat cerebral cortex and mesencephalon were cryopre served and stored for up to 1 year in liquid nitrogen at -196 degrees C with 7.5% dimethylsulfoxide (DMSO) as cryoprotectant. After thawing, these tissues were only mechanically dissociated and the cells were c ultured for 2-7 weeks before immunocytochemical staining with anti-GAB A (gamma-aminobutyric acid) antibodies. The freeze-stored GABA-immunor eactive (LR) mesencephalic neurons were compared, with computerized mo rphometry, to fresh mesencephalic cells and to their fresh and frozen cerebral cortical counterparts. A part of the cortical cells was treat ed with thienyl-phencyclidine (TCP) in order to assess the potential m orphological effects of this neuroprotective agent upon these cortical neurons. Two types of GABA-IR neurons (small and large neuritic field cells) could be evidenced in both structures without any difference b etween fresh and frozen materials, but with significant quantitative m orphological differences linked to their anatomical source. GABAergic phenotype is expressed similarly in fresh and frozen cultured neurons with intrinsically programmed morphological features and only minor in fluences of epigenetic factors. Small and large neuritic field GABA-TR neurons represent, respectively, local and long-range circuits of inh ibition, strongly reminiscent of those described in vivo and which rem ain unchanged in culture even after freeze-storage.