Many biological processes are not only determined by DNA code itself, but also regulated by activity-related chromosomal remodeling (Clapier and Cairns, 2009). Since acetylation of histone modifies the basicity of histone protein which affects histone-DNA association, histone acetylation is essential event for the epigenetic control of gene expression (Allfrey et al., 1964; Hebbes et al., 1988; Riccio, 2010). The amount of histone acetylation is controlled by histone acetyltranferases (HATs) (Brownell et al., 1996) and histone deacetylases (HDACs) (Ruiz-Carrillo et al., 1975; Annunziato and Seale, 1983). During the development, changes in the histone acetylation greatly affect the differentiation of stem cells (Lee et al., 2004), neuronal maturation (Hsieh et al., 2004), and programmed cell death (Pelzel et al., 2010) via modulation of specific gene expressions which are responsible for these biological processes. It is believed that gene transcription machineries recruit histone modification enzymes into their transcriptome complex, and in this way specific gene transcription-mediated biological processes would be promoted. For instance, histone acetylation level declines during the differentiation of embryonic stem cells, which affects the gene transcription of stemness genes, such as Oct4 and Brachyury (Lee et al., 2004). Therefore, treatments of HDAC inhibitors, such as trichostatin A (TSA) and valproic acid (VPA), or deficiency of HDAC greatly affect many aspects of brain function, including differentiation of neurons and oligodendrocytes (Marin-Husstege et al., 2002; Hsieh et al., 2004), neuronal apoptosis (Pelzel et al., 2010), motor innervation (Mejat et al., 2005), and memory consolidation (Korzus et al., 2004).

While there have been extensive investigations of the histone acetylation patterns of specific gene promoter regions, little attempt was made to explore global changes in the histone acetylation during the normal embryonic development. In this study, we found that histone H3 and H4 acetylation levels were stronger in the early differentiating neurons in the rat cortex and chick spinal cord, comparing to the proliferating neuroepithelial cells in the ventricular zone. These results suggest that transient enhancement of histone acetylation may be an important event for neuronal differentiation.

Increased acetylation of histones may promote the overall gene transcription efficiency. In this study, we found that developing neurons in the mouse cerebral cortex and chick spinal cord exhibit considerably high level of histone H3 and H4 acetylation, compared to the proliferating stem/progenitor populations. The level of histone acetylation was reported to increase during the postnatal development of rat brain (Serra et al., 1986), consistent with our current observation. Although there was little attempt to compare histone acetylation or RNA synthesis rates between stem cells and progenitor neurons, neurons appear to have higher histone acetylation level than glial cells (Hsieh et al., 2004; Shen et al., 2005; Humphrey et al., 2008). Considering that the size of neurons greatly expands during the differentiation, it appears that neurons have high demands for synthesis of many different RNA species during neuronal differentiation. However, it is noted that histone acetylation does not directly indicate the rate of RNA synthesis., Histone acetylation do not control the RNA polymerase activity, but it allows RNA polymerases to bind to the DNA. In this respect, increased histone acetylation may be linked to other biological processes.

In contrast to our in vivo data, it has been reported that cultured neural stem cells in vitro exhibit high level of global histone acetylation compared to their progenitor neurons or glial cells (Hsieh et al., 2004; Shen et al., 2005; Humphrey et al., 2008). Although it is unclear why these in vitro results are opposite to our current in vivo observations, it is important to note that epigenetic modification mainly depends on the extracellular signals, rather than cell autonomous events (Turner, 2009; Riccio, 2010). In this respect, it is no wonder that in vitro situation and in vivo environment resulted in different histone acetylation profiles. Considering that histone acetylation allows the "active" status of stem cells, and thus many stemness gene promoter regions are less acetylated, it does not mean that the global histone acetylation level should be high in less differentiated stem/progenitor cells.

Another important aspects about global histone acetylation levels identified in this study, is the heterogeneity among the same cell population. For instance, we found that only a subset of LMC chick motoneurons exhibited very strong level of histone acetylation. We failed to correlate the levels with known MN subtypes (data not shown), suggesting that these are differences among the same type of cells. This heterogeneity may be caused by highly dynamic nature of histone acetylation. In fact, histone acetylation is mainly regulated by two enzyme systems, histone acetyltransferases (HAT) and histone deacetylases (HDAC) (Riccio, 2010). Since these enzyme activities are dynamically regulated by multiple mechanisms, it is believed that the level of histone acetylation in cells are flexible and modulated by multiple factors, which may produce the diversity of histone acetylation even in the same population of cells.

In contrast to the developmental nervous system, we failed to detect any significant difference in the global histone acetylation levels among neural stem cells, neuroblasts, and mature neurons in the adult dentate gyrus. Compared to the embryonic development, neural stem cells and their progenitor neurons are similar in size and space, thus they may experience less dramatic changes in cell-autonomous gene transcription and extracellular environment. Additional studies would clarify how these differences occur, and the mechanism/significance underlying it.