Grant: Research Tools and Instruments - Category 1 (<$150,000)

Understanding the cellular basis of cognition critically requires the assessment of patterns of neural activity over widespread regions of the brain during multiple epochs of behaviour in the same animal at cellular resolution. In fact, a sensitive technique, termed cellular compartment analysis of temporal activity by fluorescence in situ hybridization (catFISH) permits such discrimination by visualizing the transcription of immediate-early genes (IEGs). Neurons engaged in information processing immediately transcribe IEGs that are visible for several minutes within the nucleus. Subsequently, processed mRNA is trans-located to the cytoplasm, peaking in concentration at ~30 min. Because mRNA can be resolved in two independent cellular compartments, it is possible to identify active neurons during two behavioural epochs in the same animal. Moreover, the proportion of cells transcribing IEGs during behaviour equals estimates generated by electrophysiological recording. Thus, like fMRI or PET, catFISH provides large-scale repeated neuronal activity measurements in the same subject, but with the cellular resolution to examine signalling and plasticity in neural networks mediating cognition. This new experimental system allows us to examine signalling and plasticity in neural networks that underlie information processing and storage.  In fact, using this method it has been possible to visualize neurons associated with spatial processing in the hippocampus, movement processing in posterior parietal cortex, and feeding behaviour in the amygdala and orbitofrontal cortex.  Further work in this area, however, critically requires an ultra-low freezer.  The practical constraints inherent in  minimizing experimentally-induced variation in hybridization yield make the storage of tissue for days or weeks unavoidable. During these periods, the availability of storage at temperatures that maintain the integrity of RNA is critical to the quality of the data that is generated.