Evaluating Hr38 as a Potential Engram Marker in Drosophila

Long-term memory formation requires transcriptional changes within engram cells, yet labelling and manipulating engrams remains a challenge in several model species, including Drosophila melanogaster. In flies, olfactory memories are encoded in the Kenyon cells, which respond sparsely to odour cues. In this thesis, I explore whether expression of the activity-regulated gene Hr38, the fly orthologue of the mammalian NR4A family, reflects odour-evoked Kenyon cell activation. I first describe an automated, fluorescence based pipeline to quantify expression of fluorescence-tagged Hr38 across 3D image stacks. Using this method, I measured a peak of Hr38 expression ~6h after odour exposure. However, this increase was not reproducible after controlling for the time of day when samples were collected. Further experiments found no increase in Hr38 expression after paired learning, unpaired learning, sugar consumption, and various odour combinations. These experiments, alongside odour-free controls, confirmed that endogenous Hr38 expression in Kenyon cells could be driven by circadian rhythms, rather than odour exposure or learning. Therefore, genetic strategies relying on Hr38 expression are unlikely to provide a reliable way to identify odour-responsive or engram Kenyon cells.