Abstract Body (Do not enter title and authors here): Background/Introduction
Delayed bedtime following stress disorder is prevalent in waves of pandemics and modern life. Considered to be a specific and important contributor to cardiovascular health, stress-related sleep disturbance has an unmet need in steady preclinical models. We previously found exciting corticotropin-releasing hormone (CRH) neurons in the paraventricular nucleus of the hypothalamus (PVH) area of mice could induce 3-hour-long wakefulness.
Methods
The chemogenetic method of designer receptors exclusively activated by designer drugs (DREADD) system was adopted to mimic stress-related sleep disturbance. We transfected PVH CRH neurons with rAAV-hSyn-DIO-hM3Dq-mCherry and rAAV-Crh-CRE. Prolonged CRH neuron activation was induced by daily intraperitoneal injection of clozapine N-oxide (CNO, 3mg/kg) at 9 am. Bulk RNA-sequencing and bioinformatics analysis were conducted for mechanistic exploration.
Results
2-week repeated chemogenetic activation of PVH CRH neurons induced a 5-fold corticosterone release, consistent with increased daily 3-hour wakefulness and corresponding decreases in both rapid eye movement (REM) as well as non-REM sleep. Over 30% of chronic CRH activation mice displayed difficulties in maintaining balance and experienced premature mortality. Mice subjected to prolonged CRH activation showed impaired left ventricular ejection fraction (67.9% versus 48.2%, p=0.0011), and immune cell infiltration demonstrated by histological staining. Intriguingly, the number of circulating monocytes increased. Then, we performed bulk RNA-sequencing of heart and bone marrow from CRH-activated and control mice. Differential gene expression and gene set enrichment analysis (GSEA) indicated marked activation of interferon-beta-related pathways in both tissues. Cytosolic DNA-sensing pathway and related key effector genes (cGAS, Cxcl10, Ccl5) were found up-regulated in the heart, while the mitochondrial oxidative phosphorylation pathway was suppressed. We further adopted the CIBERSORT tool to estimate immune infiltration in heart tissues and characterized M1 macrophage as the main pro-inflammatory cell. In our stress-related sleep disturbance mouse model, macrophages in the heart and bone marrow shared similar properties inducing interferon-stimulated genes.
Conclusion(s)
Taken together, we report a failing heart in a mouse model of stress-related sleep disturbance. The neuro-immune axis involvement and molecular mechanisms merit in-depth explorations.