The Effect And Mechanism Of Chimeric Peptide DN-9 On Sleep-wake States

Opioids are the preferred medicine for alleviating moderate to severe pain,which can lead to dependence,addiction and cause opioid-induced side effects,i.e.digestion,respiration,cardiovascular and immune system side effects.DN-9 is a chimeric peptide,it is synthesized with neuropeptides FF and biphalin,DN-9 is formed by ?-N methylation modification,substitution or configuration of its amino acids.DN-9 has powerful analgesic effect,and low tolerance,addiction or other side effects.Opioid receptors mainly include ?,?,and ? opioid receptors,? and ? opioid receptors are distributed in amygdala,bed nucleus of stria terminalis,nucleus accumbens,locus coeruleus,parabrachial nucleus,periaqueductal gray and dorsal raphe nucleus,which involve in mood and sleep-wake states.Studies suggested that opioid and opioid receptor system play key roles in sleep-wake,yet,the effects and the mechanism remain inconclusive.It has been confirmed that DN-9 has an analgesic effect by acting on ? and ? opioid receptors,but its effect and mechanism on sleep-wake states have not been studied.In order to reveal the effects and mechanism of DN-9 on sleep-wake,and to further clarify the regulatory effect of opioids on sleep-wake states,we performed this study.Methods:(1)Electroencephalogram(EEG),electromyography(EMG)recording,analysis of sleep-wake states and EEG power combined with the i.c.v.injection of saline,0.01,0.1,1 nmol DN-9 and 2 nmol morphine as a positive control were employed to evaluate the effects of DN-9 on sleep-wake states.(2)EEG,EMG recording,sleep-wake states and EEG power spectrum analysis were employed to reveal the possible mechanism of which DN-9 regulates the sleepwake states through the opioid receptor system.Non-selective opioid receptor antagonist naloxone,selective ? opioid receptor antagonist ?-funaltrexamine(?-FNA),? opioid receptor antagonist nor-binaltorphimine(nor-BNI)and ? opioid receptor antagonist naltrindole hydrochloride(NTI)were injected with 0.1 nmol DN-9.(3)Immunohistochemistry of c-Fos were performed to remark the activated neurons induced by DN-9.Results: Intracerebroventricular injection of 0.1 and 1 nmol DN-9 significantly increased the latency of NREM and REM sleep.The increase in wakefulness time was due to the extended mean duration of wakefulness instead of episode number,The reduction in NREM and REM sleep was credited to reduce the episode number of NREM and REM sleep,which accompanied with increase of EEG beta(14.5-30 Hz)activities and reduction of decreased delta(0.5-4 Hz)and theta(4.5-8.5 Hz)activities.1 nmol DN-9 was equivalent to 2 nmol morphine.Naloxone,?-FNA and nor-BNI antagonized the alterations of sleep-wake states induced by 0.1 nmol DN-9,but NTI did not.Intraventricular injection of 0.1 nmol DN-9 caused a significant increase in the number of Fos-ir neurons in the tuberomammillary nucleus,nucleus accumbens,and lateral hypothalamus.Conclusion: Intraventricular injection of DN-9 increases wakefulness and inhibits NREM and REM sleep through acting on ? and ? opioid receptors.DN-9 remarkably increases the number of Fos immunoreactive neurons in tuberomammillary nucleus,nucleus accumbens,and lateral hypothalamus.Our studies indicate that DN-9 promotes wakefulness through ? and ? opioid receptors acting on the neurons in the tuberomammillary nucleus,nucleus accumbens,and lateral hypothalamus.