Abstract
BACKGROUND: Rapid eye movement sleep (REMS) loss affects almost all physiological processes, while it itself is affected in disorders. REMS maintains optimum level of noradrenaline (NA) in a healthy individual, while increased NA during disturbed REMS is associated with diseases. The synthesis, release, and degradation of neurotransmitter are modulated by biomolecules, which are genetically encoded. The aim of this study is to understand the transcriptional and translational changes of those biomolecules in locus coeruleus (LC) and pedunculo-pontine tegmentum (PPT) in association with REMS and its loss, which is expected to help us explain associated acute and chronic pathophysiological changes. METHODS: In this study, male inbred Wistar rats were deprived of REMS for 96 h using classical flowerpot method; free-moving-, large platform- and recovery-control sets were also conducted (n = 5 per group). Brain areas related to REMS regulation, namely the LC and PPT, as well as an area unrelated to REMS regulation, namely the hippocampus, were dissected out for evaluation. Animals were grouped based on similar traits (age, weight, etc.) and then randomly by random table assigned within those matched sets. Dopamine β-hydroxylase (DBH), tyrosine hydroxylase (TH), and monoamine oxidase-A (MAO-A) protein, their gene expressions and associated histone modifications were evaluated using western blot analysis, quantitative polymerase chain reaction (qPCR) and chromatin immunoprecipitation (ChIP) assays, respectively. One-way analysis of variance (ANOVA) followed by Holm Sidak multiple comparison test was applied to evaluate the significance level between the experimental and control groups using GraphPad Prism (version 9.0.0; GraphPad Software, San Diego, California, USA, www.graphpad.com) and Sigma Stat Statistical Software (version 12; Jandel Scientific Software, CA, USA). RESULTS: Upon rapid eye movement sleep deprivation (REMSD), although TH and DBH protein expressions altered significantly in all the brain areas, the latter was highest in LC (F((5,30)) = 11.320, p < 0.001); MAO-A decreased in LC (F((5,30)) = 9.286, p < 0.001). In LC, DBH (F((8,44)) = 7.138, p < 0.001) and TH (F((8,44)) = 5.813, p < 0.001) gene expressions and histone H3 at lysine 14 (H3K14)-acetylation of TH (F((11,59)) = 25.290, p < 0.001) and DBH (F((11,59)) = 11.610, p < 0.001) increased, while lysine K9 in histone H3 (H3K9)-dimethylation tended to decrease, whereas opposite modifications were seen in MAO-A gene expression (F((11,59)) = 16.970, p < 0.001). The altered gene- and protein-expressions returned or tended to return to normal levels after recovery, as in post-REMSD prazosin treated rat brains. CONCLUSION: The differential expressions of the genes and corresponding proteins (enzymes) responsible for synthesis and degradation of NA support sustained increase in NA upon REMSD that explains underlying causes of REMSD associated chronic effects, which may be exploited for amelioration of REMSD-associated disorders.