| Introduction: Sleep includes rapid eye movement sleep (REMS) and non-rapideye movement sleep (NREMS). Between the neonatal and adult periods, not only does the regulation of sleep/wake cycle change, but the percentage of each sleep stage is very different. An interesting feature of neonatal REMS is that it is at the highest percentage shortly after birth and then decreases as maturation proceeds. In the adult, cholinergic neurons in the laterodorsal and pedunculopontine tegmental (LDT/PPT) have been implicated in the generation and maintenance of REMS. But their effect on neonatal REMS remains to be explored. Brain levels of ACh and acetylcholinesterase (AChE) are low in neonates and gradually increase, displaying a development pattern that is dissimilar to that of REMS. However, the neurons containing corticotropin-releasing factor (CRF) have the highest level of activity and secretion at the beginning of life, which parallels the development of REMS. Moreover, increasing evidence suggests that CRF may drive the high percentage of neonatal REMS. Based on the facts mentioned above, we hypothesize that blockage of CRF R1 receptor will suppress REMS in developing rats.Objective: To determine the role of NBI27914, a CRF R1 receptor antagonist, in the regulation of neonatal sleep/wake cycle by using polysomnographic recording combined with fruorometric assay for brain ACh.Methods: This study consisted of three parts.In the first part, rat pups were surgically implanted with electrodes on postnatal day (PN) 13. On PN 14, 6 hours of polysomnographic recording were continuously collected before and after administration of various doses of NBI 27914 (NBI, 12.5, 25 and 50mg/kg), a CRF R1 receptor antagonist. The same amount of saline was injected as a vehicle control.In the second part, the effect of atropine (6 mg/kg) on REMS in two-week old rats (n=8) was investigated.In the third part, brain tissues from a second set of rat pups that were given vehicle or a medium dose of NBI on PN 14 were analyzed to determine ACh levels. Two hours after treatment, rats were killed by decapitation. Brain tissues were dissected for quantification of ACh using fluorometric assay.All of data were expressed as mean±SE. The changes in sleep/wake cycles were assessed by one way ANOVA analysis followed by F test or evaluated by a Student't test. While P value less than 0.05 was considered as significant.Results: Compared with the baseline, REMS was significantly reduced in all groups treated with NBI in a dose-dependent manner, but not with vehicle. The reduction of REMS was primarily compensated by NREMS at lower doses, while compensated by wakefulness at the highest dose of NBI. NBI induced only a small amount, insignificant decrease of ACh in the thalamus and mid brain (7.3 pg/mg for the NBI group vs. 8.2 pg/mg for the control group). In addition, atropine, an M1 receptor blocker, also significantly suppressed REMS. However, the reduction of REMS was only compensated by wakefulness but not NREMS.Conclutions: Our data, for the first time, revealed that blockage of CRF R1 receptor deprives neonatal REMS, suggesting an imoportant role of CRF in the onset and maintenance of REMS for neonatal rat. Atropine induced a similar effect on neonatal REMS, but with a different alteration pattern of sleep/wake cycle. It appears that the regulatory mechanisms of CRF and ACh on REMS during the neonatal period might act through distinct pathways.
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