| Pheromones are chemical substances that communicate information between biological individuals.Alarm pheromones(APs)are mainly produced by threatened or injured animals to show dangerous situation.APs will be received by other individuals of the same species,and then cause special physiological behaviors,such as prolonged freezing time or shortened moving distance.It is important for the survival and germline maintenance of animals in the downstream food chain,such as rodents.Our previous results have demonstrated that APs and its major components 2-sec-buty-4,5-dihydrothiazole(SBT)had arousal effect on mice during non-rapid eye movement(NREM)sleep and rapid eye movement(REM)sleep through vomeronasal organ(VNO)and Grüeneberg ganglion(GG),and also caused mice exhibiting anxiety-like behaviors.However,the central mechanism of this process is still unknown.Objective: To reveal the central mechanism of APs’ arousal effect.Methods: The olfactory bulb was firstly injected with the trans-synaptic herpes simplex virus(HSV)combined with the immunofluorescence double labeling to find the possible neural circuit of APs’ arousal effect.Then c-fos protein labeling was used to determine whether neurons in this circuit were activated after SBT stimulation.The Noradrenaline(NA)neurons were specific destroyed by injecting 6-hydroxydopamine(6-OHDA)into locus coeruleus(LC),and with accurate nasal odor delivery and sleep recording system,the effect of locus coeruleus noradrenalinergic neurons lesion on arousal effect of SBT during NREM sleep were determined.Results: After the right olfactory bulb injected with HSV-GFP,HSV-GFP labeled neurons can be seen in the arousal associated nucleus: the basal forebrain(BF)and the locus coeruleus(LC).There were not any HSV-GFP and choline acetyltransferase(ChAT)co-labeled neurons in BF region.In the LC region,HSV-GFP and Tyrosine hydroxylase(TH)co-labeled neurons accounted for 75.70 ± 3.20% on the right side and about 72.04 ± 2.82% on the left.Compared with the Air group,the expression of c-Fos protein in LC was significantly increased with the nasal delivery of SBT,the expression of c-Fos protein was significantly increased in TH(+)neurons,and the activation rate of TH(+)neurons was also improved.The effect of LC damage was evaluated by TH labeling.After 6-OHDA was injected,the number of TH(+)neurons was significantly reduced.After LC damaged,the wake-up effect of SBT in NREM was weakened,which showed that the latency of arousal prolonged,the amount of W time decreased,the amount of REM and NREM time increased during the gas supply period.The W time decreased,and the REM time increased after the gas supply period;and the LC damaged mice were given with SBT and AIR respectively,and there was no significant difference in the total amount of the phases of the mice during the gas supply period and after the gas supply period.Conclusion: LC receives information from olfactory system.SBT stimulation via olfactory pathway can activate LC-NA neurons and increase the expression of c-fos protein in LC.6-OHDA specific damage to LC-NA neurons significantly weakens the arousal effect of SBT on mice in NREM sleep,suggesting that APs’ arousal effect is determined by activation of LC-NA neurons via olfactory pathway. |
