Microarray technology has been instrumental in advancing studies of altered gene expression in human disease, particularly in tissues that are modestly complex in structure and cell type. The approach is just now being applied to complex brain diseases. Recently, we used high-density cDNA microarrays to analyze gene expression changes in the prefrontal cortex in schizophrenic and control human subjects. Our hierarchy analysis of more than 250 gene groups represented on the chips revealed that genes encoding proteins involved in presynaptic function were among the most significantly changed. We have found that no commercially available microarrays contain a sufficient number of putative gene targets related to various neurotransmitter systems and synaptic machinery genes. Furthermore, current microarray sensitivity makes it difficult to routinely analyze low abundance transcripts, among which are neurotransmitter receptors. This technology-focused application proposes to enhance microarray technology that will be most suitable for analysis of complex tissues such as brain, with starting material that is more ideal for assessment of discrete regions and cell populations.
Aim 1 will focus on designing and building moderate density, custom cDNA gene chips that will contain multiple representations of a relatively complete complement of genes encoding proteins involved in synaptic function and neurotransmission. These arrays will be applied to a more detailed analysis of changes in gene expression in prefrontal cortex in schizophrenia. These arrays will be shared with the scientific community and serve as an important analytical strategy for assessing changes in gene expression in a wide variety of neuropsychiatric disorders.
Aim 2 is designed to address technical issues of improving array sensitivity. We will produce cDNA polymers similar to those used in oligonucleotide arrays to serve as targets on cDNA arrays. By creating more binding sites per target, and thus more binding per unit area on each array spot, we expect to greatly improve array sensitivity and decrease the current required amount of starting material. We expect that our proposed studies will facilitate use of the technology through the neuroscience community, and in particular improve experimental designs to allow for sensitive and reproducible assessment of changes in gene expression in neuropsychiatric diseases.
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