Effects Of 9050 On Electroencephalogram In Mice And The Role Of Extrasynaptic GABA_A Receptor In Its Effect

Objective:Extrasynaptic GABAARs(GABAARs)is considered as a key target for sedative hypnosis.Compared with traditional synaptic GABAARs allosteric modulators,sedative hypnotics acting on extrasynaptic GABAARs have the advantages of not affecting sleep structure and light adverse reactions.In this study,a series of quinazolinones were screened,among which 9050 exhibited good and stable sedative effects.Patch clamp experiments on Xenopus oocytes showed that 9050 not only had an effect on synaptic GABAARs,but also had a strong effect on extrasynaptic GABAARs.This study was designed to evaluate the effects of compound 9050 on sleep and awakening in mice compared with zolpidem,THIP and methaqualone and its mechanism of action,providing ideas for further modification of compound 9050.Methods:1.Mice were given compound 9050(2.3,2.9,3.6,4.5,5.6,7.0 mg/kg,I.P.)and positive control drug Zolpidem(14.41,20.58,29.4,42,60 mg/kg,I.p.),methaquone(25.6,32.0,40.0,50.0,62.5 mg/kg,I.P.)and THIP(5.9,8.4,12.0,17.2,24.5,35.0,50.0 mg/kg,I.P.).The incidence,induction time and duration of righting reflex were observed and recorded.The sedative and hypnotic effect of compound 9050 was evaluated by using the highest dose of eeg recording without causing the loss of righting reflex.After 9050(0.5,1,2 mg/kg),zolpidem(2.5,5,10 mg/kg),THIP(1.25,2.5,5 mg/kg),methaquone(6,12,24 mg/kg)were administered,eeg was monitored for 24 hours,and NREM and REM sleep latency were analyzed.Wakefulness,NREM,REM sleep phase ratio,sleep structure(frequency and average duration of each phase),degree of sleep fragmentation(micro-arousal and transient wakefulness),eeg power density spectrum,slow wave activity(SWA),to evaluate the sedative and hypnotic effects of drugs.The chronic trial investigated the tolerability of chronic administration of 9050 and whether there was withdrawal after withdrawal.Mice were given 9050(1 mg/kg),THIP(5 mg/kg)and methaquidine(24 mg/kg),respectively,and eeg records were recorded for 13 days,including one day baseline(BSL),10 days administration(T1-T10)and two days withdrawal(W1,W2).Gabrd KO mice and WT mice in the same nest were given 9050(1 mg/kg),zolpidem(10 mg/kg),THIP(5 mg/kg)and methaqualone(24 mg/kg),respectively,and eeg was recorded to preliminatively explore the mechanism of compound 9050.Results:1.The ED50 value of LORR induced by compound 9050,ZPD,THIP and MTQ were 4.29 mg/kg,19.47 mg/kg,16.64 mg/kg and 40 mg/kg,respectively.2.9050,methaqualone and Zolpidem all had significant effect on promoting sleep.THIP did not shorten the latency of falling asleep or prolong the total sleep time,but it enhanced the activity of delta waves and sleep stability.The 9050 extended NREM sleep time,did not inhibit REM sleep,and reduced the frequency of short and micro awakenings,significantly improving sleep continuity.3.Tolerance of 9050,methaqualone,and Zolpidem occurred during chronic administration,but there was a rebound phenomenon after withdrawal of methaqualone and zolpidem,while 9050 withdrawal did not cause negative sleep effects within two days.THIP produced no tolerance within ten days of administration and no negative sleep effects within two days of withdrawal.4.NREM latency of 9050,methaqualone and THIP decreased in Gabrd KO mice and WT mice,but compared with WT mice,the duration of each time phase decreased,the frequency of transient wakeup increased,and the activity of delta wave decreased in KO mice after administration.Conclusion:9050 May be an effective hypnotic compound that can promote sleep,promote and improve NREM sleep without inhibiting REM sleep,enhance delta wave activity and enhance sleep stability.The enhancement of delta wave activity and sleep stability may berelated to GABAA receptor containing δ subunit.Although 9050 has no withdrawal symptoms,it is tolerated under chronic administration conditions and there is room for further modification.