Effects Of Nicotinic Acetylcholine Receptor α4 Mutations On Sleep In Aging Flies

With the increasing trend of population aging in China,sleep problems become especially prominent among the elderly populations.It is noteworthy that sleep deprivation may lead to organismal aging,while the related mechanism remains unclear.In this study,the nicotinic acetylcholine receptorα4（Dα4）mutant sleep model of Drosophila melanogaster was used to explore the effect and mechanism of sleep deprivation on aging.Firstly,the effect of Dα4 mutation on sleep duration in flies was investigated.Three strains of Drosophila with different Dα4 loci（V12441,V11392,12414R3）mutations were selected,and the strain with the most reduced sleep duration was screened out.Finally,effects of extremely-low frequency electromagnetic field（ELF-EMF）on locomotion and sleep in Dα4 mutant flies were elucidated,with a view to discovering physical techniques to improve sleep in aging flies.The results obtained in this study are as follows:（1）Compared with W₁₁₁₈wild-type flies,three mutant strains showed significantly increased locomotion,remarkably decreased sleep time,fragmented sleep,decreased sleep quality,and reduced sleep time,both during the day and at night.The V12441,V11392,and 12414R3 in descending order results,indicated that the 12414R3 mutation has the greatest effect on sleep duration..Mechanism analysis suggested that the Dα4 mutation reduced sleep quality by triggering abnormal expression of biological clock genes and decreased acetylcholine metabolism.（2）As revealed by comparative analysis of the lifespan of W₁₁₁₈wild-type and12414R3 mutant,the Dα4 mutation induced a decrease in the average lifespan from approximately 68 days to 47 days,a decrease of approximately 30%.Based on the the behavioral monitoring results of neonatal and 30-day-old flies,no significant change of 30-day-old W₁₁₁₈was found in locomotion and sleep,while 30-day-old 12414R3mutants presented a significant increase in locomotion,a remarkable decrease in sleep duration and showed poor sleep quality,which indicated that the Dα4 mutation caused a significant acceleration of aging in Drosophila melanogaster.According to the analysis of the expression levels of genes related to the acetylcholine metabolic pathway,for neonatal Drosophila melanogaster,the Dα4 mutation diminished the newborn flies’metabolism of acetylcholine,while enhanced the metabolism of acetylcholine of 30-day-old.It indicated that the Dα4 mutation may exert different effects on the ability of acetylcholine metabolism in Drosophila at different ages.（3）By comparing the gut flora assays of W₁₁₁₈wild type and 12414R3 mutant,this study found that the species abundance of gut flora increased in newborn mutant flies,but decreased in 30-day-old 12414R3 mutant.Based on the comparison of the gut flora assays of newborn and 30-day-old Drosophila melanogaster,species abundance of gut flora decreased in both W₁₁₁₈wild type and 12414R3 mutant,as well as declined at 30-day-old flies.The abundance of Enterobacteriaceae was significantly increased and the abundance of Commensalibacter.was remarkably decreased in 12414R3 mutant,both in newborn and 30-day-old flies.The abundance of Lactobacillus was significantly increased in 30-day-old flies in comparison to neonates,both in W₁₁₁₈wild-type and 12414R3 mutant.Especially,mutant flies’gut species abundance increase twice as large as the wild-type Drosophila.That is to say,the Dα4 mutation increases the species abundance of the newborns’gut flora,but decreases significantly with aging The Dα4 mutation may reduce sleep duration and accelerate aging by altering the gut species abundance.（4）According to the analysis of the effects of ELF-EMF on sleep in Drosophila,short-term ELF-EMF exposure triggered a further reduction in sleep duration,decreased sleep quality,and weakened acetylcholine metabolism in newborn 12414R3mutant flies.In 30-day-old flies,it was found that ELF-EMF exposure was a facilitator,which not only enhanced the metabolism of acetylcholine but also slowed the decrease of sleep duration and sleep quality.Given to negative effects of ELF-EMF on the sleep of newborn Drosophila,the effects of ELF-EMF exposure on development of Drosophila larvae were performed and results suggested that ELF-EMF exposure promoted the development of Drosophila melanogaster larvae,but shortened sleep duration,deteriorated sleep quality of adult flies furtherly,and weakened acetylcholine metabolism in their bodies.This study speculated that effects of ELF-EMF exposure has deep relations to the age of flies,and can improve sleep to a certain extent for aging adults,while its effect on the newborns’sleep is poorly negative.In summary,the Dα4 mutation accelerates aging and shortens the lifespan of Drosophila melanogaster by affecting acetylcholine metabolism,triggering reduced sleep duration,sleep fragmentation,decreased sleep quality,as well as decreased abundance and structural changes in the intestinal flora.In contrast,for aging flies with chronic sleep deprivation,despite a delaying effect of ELF-EMF exposure on their sleep duration and sleep quality decline,it negatively affects the sleep of newborn and developing flies.Therefore,this study suggests that sleep deprivation accelerates aging and ELF-EMF exposure can alleviate the sleep problems of aging individuals to some extent.It is expected that this study can not only provide a basis for the aging problem caused by chronic sleep deprivation,but also offer a reference for the evaluation and scientific utilization of the health effects of ELF-EMF exposure.