Sleep loss drives brain region- and cell type-specific alterations in ribosome-associated transcripts involved in synaptic plasticity and cellular timekeeping

Abstract

Sleep and sleep loss are thought to impact synaptic plasticity, and recent studies have shown that sleep and sleep deprivation (SD) differentially affect gene transcription and protein translation in the mammalian forebrain. However, much less is known regarding how sleep and SD affect these processes in different microcircuit elements within the hippocampus and neocortex - for example, in inhibitory vs. excitatory neurons. Here we use translating ribosome affinity purification (TRAP) and in situ hybridization to characterize the effects of sleep vs. SD on abundance of ribosome-associated transcripts in Camk2a-expressing (Camk2a+) pyramidal neurons and parvalbumin-expressing (PV+) interneurons in mouse hippocampus and neocortex. We find that while both Camk2a+ neurons and PV+ interneurons in neocortex show concurrent SD-driven increases in ribosome-associated transcripts for activity-regulated effectors of plasticity and transcriptional regulation, these transcripts are minimally affected by SD in hippocampus. Similarly we find that while SD alters several ribosome-associated transcripts involved in cellular timekeeping in neocortical Camk2a+ and PV+ neurons, effects on circadian clock transcripts in hippocampus are minimal, and restricted to Camk2a+ neurons. Taken together, our results indicate that SD effects on transcripts destined for translation are both cell type- and brain region-specific, and that these effects are substantially more pronounced in the neocortex than the hippocampus. We conclude that SD-driven alterations in the strength of synapses, excitatory-inhibitory balance, and cellular timekeeping are likely more heterogeneous than previously appreciated.Significance StatementSleep loss-driven changes in transcript and protein abundance have been used as a means to better understand the function of sleep for the brain. Here we use translating ribosome affinity purification (TRAP) to characterize changes in abundance of ribosome-associated transcripts in excitatory and inhibitory neurons in mouse hippocampus and neocortex after a brief period of sleep or sleep loss. We show that these changes are not uniform, but are generally more pronounced in excitatory neurons than inhibitory neurons, and more pronounced in neocortex than in hippocampus.

Hippocampal neurons’ cytosolic and membrane-bound ribosomal transcript profiles are differentially regulated by learning and subsequent sleep

“…We first validated cell type-specific transcript enrichment from Camk2a+ and pS6+ cell populations, using qPCR. Consistent with recent data (36), relative to Input mRNA, Camk2a+ TRAP produced similar mRNA levels of Arc, Cfos, Homer1a, Glua1, and Vglut1 transcripts and reduced expression of interneuron and glial cell markers. In light of hemizygosity for HA-Rpl22, this is consistent with roughly twofold enrichment of excitatory markers relative to Input.…”

“…2B and 4). Pathways affected by SD-across neuronal populations and across both subcellular compartments-were those linked to regulation of transcription, consistent with prior findings (4)(5)(6)36) and the AMPK, IL-3, IGF1, and PDGF signaling pathways. Critically, AMPK (70) and IGF1 signaling (71) have been implicated in homeostatic sleep responses-that is, changes in sleep architecture of brain oscillations-following SD.…”

“…3 B, Top). Previous transcriptome analysis has shown that mRNAs encoding transcription regulators are more abundant following brief SD in the hippocampus (Fos, Elk1, Nr4a1, Creb, and Crem1) (4-6, 36) and neocortex (Per2, Egr1, and Nr4a1) (2,36). Consistent with those findings, SD increased the abundance of multiple mRNAs encoding transcription factors and upstream regulators in all cell populations, including E2f6, Elk1, Erf, Fosl1, Fosl2, Fos, Fosb, Lmo4, Taf12, Xbp1, Atf7, Artnl2, Atoh8, Bhlhe40 (Dec1), Crebl2, Crem, Egr2, Nfil3, and Ubp1.…”

“…In other words, CFC had an apparent effect on ΔFosb abundance, even after the effects of SD alone on ΔFosb expression were accounted for. However, on ribosomes taken from all Camk2a+ neurons, both Fosb and ΔFosb transcripts increased in CFC + Sleep mice, but this increase was occluded by increases in the transcripts caused by SD alone (36) in CFC + SD mice (SI Appendix, Fig. S8 A, Bottom).…”

Sleep loss drives acetylcholine- and somatostatin interneuron–mediated gating of hippocampal activity to inhibit memory consolidation

“…This finding is consistent with our recent work, showing that expression of the immediate early gene Arc is increased after SD in the somatosensory cortex while simultaneously being suppressed in hippocampal DG (19). More recently, we have found that while SD drives higher expression of a number of activity-regulated genes in both pyramidal cells and PV+ interneurons in the neocortex, expression of the same genes in the hippocampus is refractory to SD (63). Together, these findings might suggest that SD-driven activation of Sst+ interneurons, leading to general network inhibition, could be unique to the hippocampal network.…”