D. Petite et Mc. Calvet, CRYOPRESERVED GABAERGIC NEURONS IN CULTURES OF RAT CEREBRAL-CORTEX AND MESENCEPHALON - A COMPARATIVE MORPHOMETRIC STUDY WITH ANTI-GABA ANTIBODIES, Brain research, 747(2), 1997, pp. 279-289
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.