DEVELOPMENT AND ACTIVITY-DEPENDENT EXPRESSION OF NEURONAL MARKER PROTEINS IN ORGANOTYPIC CULTURES OF RAT VISUAL-CORTEX

We are interested in activity-dependent mechanisms which govern the st ructural and functional maturation of neurons in the visual cortex. We have asked whether the expression of neuronal markers microtubule-ass ociated proteins tau, MAP-2, synaptophysin (p38), and the growth-assoc iated protein GAP-43 are dependent on cortical afferents or spontaneou s activity. As a model system we have employed organotypic monoculture s of rat visual cortex (OTCs, isolated from subcortical structures) in comparison with visual cortex in vivo (innervated by thalamic and oth er afferents) at different postnatal ages. We know from previous work that the OTCs, like the cortex in vivo, display a high rate of spontan eously generated action potentials. Therefore, as a third objective, w e have analysed OTCs grown as monocultures under chronic blockade of s pontaneous action potentials. Protein expression was detected by prote in blots and/or immunohistochemistry. The proteins examined in this st udy are expressed in OTCs, even when grown under activity blockade. Ho wever, the pattern of expression differs from the cortex in vivo. Tau is expressed much weaker in OTCs than in cortex in vivo. The expressio n of the major band of about 50 kDa increases over time in vivo and in OTCs. Smaller isoforms of tau are dramatically downregulated, and lar ger (adult) isoforms do not appear within 35 days in vitro (DIV). Unde r activity blockade the expression of tau reaches a maximum by 21 DIV and decreases dramatically, so that the protein is hardly detectable b y 47 DIV. MAP-2-immunoreactive proteins are localized in somata and de ndrites, but also persist in axons. The expression in OTCs of p38 and GAP-43 correlates well with the expression observed in vivo. Synaptoph ysin (p38) occurs with a similar time course and amount in OTCs as in cortex in vivo. Synaptic boutons appear in all layers, and specialized terminal elements have been observed. Activity blockade slightly affe cts the p38 expression, although the late postnatal decline in p38 imm unoreattivity observed on protein blots from cortex in vivo and in nor mal OTCs appears more accentuated in activity-blocked OTCs. The GAP-43 expression is prominent from birth onwards in vivo and in OTCs. Howev er, in normal OTCs GAP-43 is not declining as it is in vivo, although it is downregulated in activity-blocked OTCs. As a major finding we re port that neuronal markers which are normally expressed in immature ne urons and axons during the period of differentiation and structural pl asticity are continuously expressed in OTCs, suggesting that a monocul tured cortex retains the ability for growth and structural changes lon ger than the cortex in vivo.