Histotypic mouse parietal cortex cultures: Excitation/inhibition ratio and ultrastructural analysis

Abstract

Primary cultures of mouse parietal cortex, prepared between postnatal day 3 (P3) and P9, were studied using transmission electron microscopy and HPLC of excitatory (aspartate and glutamate) and inhibitory neurotransmitters (glycine, GABA and taurine) to determine their morphological and functional development. Relations between excitation and inhibition ( E/ I) were contrasted with ultrastructural features over the time course of in vitro development. After 6 days in vitro, cultured parietal cortex neurons prepared from mice at P3 had immature morphological characteristics, whereas P5 cultures showed a more developed histological structure but still with scarce synapses. The acquirement of histotypic characteristics was seen in P7 cultures, which contained numerous symmetric and asymmetric synaptic contacts. On P9, the cultures showed signs of tissue damage. In terms of neurotransmitter levels and E/ I ratios, P7 cultures had relatively low E/ I ratio as compared with the rest of the cultures prepared before or after P7. These results demonstrated that the development of inhibitory synaptic transmission, as indicated in the fall of E/ I ratio, marked the maturation of cerebral cortical tissue and that the critical period to obtain histotypic cultures of mouse parietal cerebral cortex coincides between P5 and P7. This work provides useful information regarding the balance between excitation and inhibition as an indicative parameter for in vitro nerve cell survival, differentiation and maturation and reinforces the great value of histotypic cultures in the study of central nervous system development.