Scalable assays for morphological analysis of mammalian neurons
Medium and high-throughput assays (i.e., screens) have generally not been applied tomammalian neurons because of the difficulties in culturing them in large numbers and because of the lowefficiency with which the genetic makeup of neurons can be altered. Furthermore, because many aspects ofneuronal function can only be assayed with electrophysiological assays, follow-up analysis and validation ofscreening hits is difficult. Dr. Sabatini proposes to use automated imaging approaches to analyze synapsenumber and neuronal structure in vitro in a scalable format. He has implemented tissue culture andimmunostaining approaches to monitor the number and types of synapses formed onto neurons in multi-wellplates. He will couple this analysis with lentivirus mediated introduction of short-hairpin RNAs to induce RNAinterference against genes expressed in neurons. This will be performed in concert with transcriptionalanalysis of neurons to determine the key changes in gene expression that correlate with structural andsynaptic changes. This proposal represents a significant collaboration between several groups with expertisein functional analysis of neurons, automated analysis of images, viral mediated manipulation of geneexpression, and whole-genome transcriptional analysis. The work may lead, for the first time, to a turn-keyand robust method of analysis of neuron and synapse structure suitable for scalable, whole-genomeanalysis.