Several aspects of hippocampal cell function are influenced by adrenal-secreted glucocorticoids in a delayed, genomic fashion. Previously, we used Serial Analysis of Gene Expression to identify glucocorticoid receptor (GR)-induced transcriptional changes in the hippocampus at a fixed time point. However, because changes in mRNA levels are transient and most likely precede the effects on hippocampal cell function, the aim of the current study was to assess the transcriptional changes in a broader time window by generating a time curve of GR-mediated gene expression changes. Therefore, we used rat hippocampal slices obtained from adrenalectomised rats, substituted in vivo with low corticosterone pellets, predominantly occupying the hippocampal mineralocorticoid receptors. To activate GR, slices were treated in vitro with a high (100 nM) dose of corticosterone and gene expression was profiled 1, 3 and 5 h after GR-activation. Using Affymetrix GeneChips, a striking pattern with different waves of gene expression was observed, shifting from exclusively down-regulated genes 1 h after GR-activation to both up and down regulated genes 3 h after GR-activation. After 5 h, the response was almost back to baseline. Additionally, real-time quantitative polymerase chain reaction was used for validation of a selection of responsive genes including genes involved in neurotransmission and synaptic plasticity such as the corticotropin releasing hormone receptor 1, monoamine oxidase A, LIMK1 and calmodulin 2. This permitted confirmation of GR-responsiveness of 15 out of 18 selected genes. In conclusion, direct activation of GR in hippocampal slices results in transient changes in gene expression. The pattern in which gene expression was modulated suggests that the fast genomic effects of glucocorticoids may be realised via transrepression, preceding a later wave of transactivation. Furthermore, we identified a number of interesting candidate genes which may underlie the glucocorticoid-mediated effects on hippocampal cell function.

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Expression profiling in the hippocampus is hampered by its cellular heterogeneity. The aim of this study was to evaluate the feasibility of using laser-microdissected hippocampal subregions for expression profiling to improve detection of transcripts with a subregion-specific expression. Cornu ammonis (CA)3 and dentate gyrus (DG) subregions were isolated from rat brain slices using laser microdissection, subjected to two rounds of linear amplification and hybridized to rat GeneChips containing approximately 8000 transcripts (RG_U34A; Affymetrix). Analysis of the data using significance analysis of microarrays revealed 724 genes with a significant difference in expression between CA3 and DG with a false discovery rate of 2.1%, of which 264 had higher expression in DG and 460 in CA3. Several transcripts with known differential expression between the subregions were included in the dataset, as well as numerous novel mRNAs and expressed sequence tags. Sorting of the differentially expressed genes according to gene ontology classification revealed that genes involved in glycolysis and general metabolism, neurogenesis and cell adhesion were consistently expressed at higher levels in CA3. Conversely, a large cluster of genes involved in protein biosynthesis were expressed at higher levels in DG. In situ hybridization was used to validate differential expression of a selection of genes. The results of this study demonstrate that the combination of laser microdissection and GeneChip technology is both technically feasible and very promising. Besides providing an extensive inventory of genes showing differential expression between CA3 and DG, this study supports the idea that profiling in hippocampal subregions should improve detection of genes with a subregion-specific expression or regulation.

Recent studies in rodents have shown that there are significant differences in gene expression profiles between the hippocampal subregions CA1, CA3, and DG. These differences in molecular make-up within the hippocampus most likely underlie the differences in morphology, physiology, and vulnerability to insults that exist between the subregions of the hippocampus and are as such part of the basic molecular architecture of the hippocampus. The aim of this study was to investigate at large scale whether these subregional differences in gene expression are conserved in the hippocampus of a nonhuman primate, the common marmoset. This study is very timely, given the recent development of the first marmoset-specific DNA microarray, exclusively containing sequences targeting transcripts derived from the marmoset hippocampus. Hippocampal subregions CA1, CA3, and DG were isolated by laser microdissection and RNA was isolated, amplified, and hybridized to the marmoset-specific microarray (EUMAMA) containing more than 1,500 transcripts expressed in the adult marmoset hippocampus. Large differences in expression were observed in particular between the DG region and both pyramidal subregions. Moreover, the subregion-specific patterns of gene expression showed a remarkable conservation with the rodent brain both in terms of individual genes and degree of differential expression. To our knowledge, this is the first study investigating large scale hippocampal gene expression in a nonhuman primate. The obtained expression profiles not only provide novel data on the expression of more than 1,500 transcripts per hippocampal subregion but also are of potential interest to neuroscientists interested in the role of the different subregions in learning and memory in the nonhuman primate brain.

Acute stress and corticosterone enhance 5-HT1A receptor-mediated responses in rat hippocampal CA1 cells within 1-2 h, through a process involving transcriptional regulation of unknown genes. Earlier studies showed that regulation of the 5-HT1A receptor gene cannot explain the functional effects. We here tested the hypothesis that corticosterone targets genes encoding RGS4 or SGK1, which can both affect the 5-HT1A receptor associated Kir channel, thus affecting 5-HT1A receptor function. To this end, the effect of a single corticosterone injection on hippocampal expression of RGS4 and SGK1 mRNAs, measured by in situ hybridization, was studied. Expression of RGS4 or SGK1 mRNA was not affected by the corticosterone injection, neither in the CA1 area nor in other hippocampal subregions. Strikingly, SGK1 mRNA expression was strongly up-regulated in the corpus callosum. We reject, however, the hypothesis that the effect of corticosterone on 5-HT1A responsiveness is mediated via altered RGS4 or SGK1 mRNA expression.