NT-4 5 AND LIF, BUT NOT NT-3 AND BDNF, PROMOTE NPY MESSENGER-RNA EXPRESSION IN CORTICAL-NEURONS IN THE ABSENCE OF SPONTANEOUS BIOELECTRICALACTIVITY/

Epigenetic factors are known to influence the differentiation of neoco rtical neurons. The present study analyses the role of spontaneous bio electrical activity (SBA) and neurotrophic factors on the expression o f neuropeptide Y (NPY) in rat visual cortical neurons using organotypi c monocultures prepared from newborn animals and in situ hybridization to detect the NPY messenger ribonucleic acid (mRNA). Spontaneously ac tive cortex cultures display NPY mRNA expression in about 7% of all co rtical neurons from 10 days in vitro (DIV) on. Blocking the SBA by chr onic application of 10 mM Mg2+ for 3-30 DIV reduces the percentage of NPY neurons to about 2%. Allowing an initial phase of SBA (1-20 DIV) f ollowed by an SBA blockade (for 21-50 DIV) results in 2% labelled neur ons, indicating a dramatic reduction of NPY mRNA expression in the abs ence of SBA. Surprisingly, the reverse experiment (a period of SBA blo ckade for 1-20 DIV followed by a period of SBA recovery for 21-40 DIV) does not cause an upregulation of NPY mRNA expression. However, allow ing cultures to differentiate as spontaneously active cultures, then a pplying a transient period of SBA blockade which is followed by a seco nd period of SBA, does rescue the NPY mRNA expression in 7% of the cor tical neurons, We conclude that SBA is a main trigger for NPY mRNA exp ression and it is particularly important during an early postnatal per iod of differentiation. We then analysed whether neurotrophic factors known to modulate cortical neuropeptide expression are able to do so i n the absence of SBA. Supplementing chronically blocked cultures with the neurotrophins, brain-derived neurotrophic factor (BDNF), neurotrop hin-3 (NT-3), neurotrophin-4/5 (NT-4/5) and the cytokine, leukaemia in hibitory factor (LIF), reveals that BDNF and NT-3 are unable to increa se the percentage of NPY neurons, In contrast, LIF and NT-4/5 increase the percentage of NPY neurons to 4 and 6-7%, respectively. Moreover, neurons treated with NT-4/15 display a very high level of NPY mRNA exp ression in somata and in the dendritic trees. The data suggest a compl ex interplay and a hierarchy of epigenetic factors in regulating the n eurochemical architecture of the developing neocortex.