Cytosolic and Membrane-Associated Ribosomes’ mRNA Profiles are Differentially Regulated by Learning and Sleep in Hippocampal Neurons

The hippocampus plays an essential role in consolidating transient experiences into long-lasting memories. Memory consolidation can be facilitated by post-learning sleep, although the underlying cellular mechanisms are largely unknown. We took an unbiased approach to this question using a mouse model of hippocampally-mediated, sleep-dependent memory consolidation (contextual fear memory; CFM), which is known to be disrupted by post-learning sleep loss. Translating ribosome affinity purification (TRAP) was used to quantify ribosome-associated RNAs in different subcellular compartments (cytosol and membrane) and in different hippocampal cell populations (either whole hippocampus, Camk2a+ excitatory neurons, or highly active neurons expressing phosphorylated ribosomal subunit S6 [pS6+]). Using RNA-seq, we examined how these transcript profiles change as a function of sleep vs. sleep deprivation (SD), and as a function of prior learning (contextual fear conditioning; CFC). To our surprise, we found that while many mRNAs on cytosolic ribosomes were altered by sleep loss, almost none were altered by learning. Of the few changes in cytosolic ribosomal transcript abundance following CFC, almost all were occluded by subsequent SD. This effect was particularly pronounced in pS6+ neurons with the highest level of neuronal activity following CFC, suggesting SD-induced disruption of post-learning transcript changes in putative “engram” neurons. In striking contrast, far fewer transcripts on membrane-bound (MB) ribosomes were altered by SD, and many more mRNAs (and lncRNAs) were altered MB ribosomes as a function of prior learning. For hippocampal neurons, cellular pathways most significantly affected by CFC were involved in structural remodeling. Comparisons of post-CFC MB transcript profiles between freely-sleeping and SD mice implicated changes in cellular metabolism in Camk2a+ neurons, and increased protein synthesis capacity in pS6+ neurons, as biological processes disrupted by post-learning sleep loss. Together these findings suggest that disruptive effects of sleep loss on hippocampal memory consolidation are both cell type- and subcellular compartment-specific.