Analysis of sleep disorders under pain using an optogenetic tool: possible involvement of the activation of dorsal raphe nucleus-serotonergic neurons

Abstract

BackgroundSeveral etiological reports have shown that chronic pain significantly interferes with sleep. Inadequate sleep due to chronic pain may contribute to the stressful negative consequences of living with pain. However, the neurophysiological mechanism by which chronic pain affects sleep-arousal patterns is as yet unknown. Although serotonin (5-HT) was proposed to be responsible for sleep regulation, whether the activity of 5-HTergic neurons in the dorsal raphe nucleus (DRN) is affected by chronic pain has been studied only infrequently. On the other hand, the recent development of optogenetic tools has provided a valuable opportunity to regulate the activity in genetically targeted neural populations with high spatial and temporal precision. In the present study, we investigated whether chronic pain could induce sleep dysregulation while changing the activity of DRN-5-HTergic neurons. Furthermore, we sought to physiologically activate the DRN with channelrhodopsin-2 (ChR2) to identify a causal role for the DRN-5-HT system in promoting and maintaining wakefulness using optogenetics.ResultsWe produced a sciatic nerve ligation model by tying a tight ligature around approximately one-third to one-half the diameter of the sciatic nerve. In mice with nerve ligation, we confirmed an increase in wakefulness and a decrease in non-rapid eye movement (NREM) sleep as monitored by electroencephalogram (EEG). Microinjection of the retrograde tracer fluoro-gold (FG) into the prefrontal cortex (PFC) revealed several retrogradely labeled-cells in the DRN. The key finding of the present study was that the levels of 5-HT released in the PFC by the electrical stimulation of DRN neurons were significantly increased in mice with sciatic nerve ligation. Using optogenetic tools in mice, we found a causal relationship among DRN neuron firing, cortical activity and sleep-to-wake transitions. In particular, the activation of DRN-5-HTergic neurons produced a significant increase in wakefulness and a significant decrease in NREM sleep. The duration of NREM sleep episodes was significantly decreased during photostimulation in these mice.ConclusionsThese results suggest that neuropathic pain accelerates the activity of DRN-5-HTergic neurons. Although further loss-of-function experiments are required, we hypothesize that this activation in DRN neurons may, at least in part, correlate with sleep dysregulation under a neuropathic pain-like state.

Possible involvement of activated locus coeruleus–noradrenergic neurons in pain-related sleep disorders

“…It has been reported that both ␣ and ␤ receptors are distributed in the PFC [15,16]. Although further studies are needed using an optogenetic tool with TH-cre transgenic mice in which AAV-HAL (a light-driven chloride pump) is injected into the LC, we propose that the acceleration of LC-PFC noradrenergic neurons, as well as DRN-5-HTergic neurons [5] may, at least in part, explain the sleep disturbance under a state of neuropathic pain.…”

“…In a previous study, we found that sciatic nerve-ligated mice exhibited sleep disturbance along with thermal hyperalgesia [5]. Although it has been considered that several areas in the brainstem and forebrain are important for modulation and expression of the sleep/wake cycle [8], and several neurotransmitters and neuropeptides are involved in modulation of this cycle [9][10][11], the neurophysiological and molecular mechanisms by which neuropathic pain affects sleep/arousal patterns are not fully understood.…”

“…Clinical studies have shown that most patients experience problems with sleep after they develop chronic pain [4]. In our previous study, we demonstrated that neuropathic pain accelerated the activity of dorsal raphe nucleus (DRN)-serotonergic neurons, implying that the activation of DRN neurons may play a role in sleep dysregulation under a neuropathic pain-like state [5]. However, the neurophysiological mechanism by which neuropathic pain affects sleep-arousal patterns remains unclear.…”

5-HT2A receptor activation is necessary for CO2-induced arousal

“…Serotonin (5-HT) neurons located in the rostral brain stem comprise a component of the ascending arousal system that is responsible for regulating sleep-wake transitions (Brown et al 2012;Ito et al 2013). 5-HT neurons, including those in this region, are chemosensitive (Richerson 2004;Severson et al 2003).…”

Selective Manipulation of Neural Circuits

“…Manipulation of the targeted neurons in the spinal cord and hindbrain showed that glutamatergic neurons in the spinal cord are critical for generating rhythm in the central pattern generators and the activation of glutamatergic reticulospinal neurons in hindbrain area is sufficient to activate directly the spinal locomotor network [18]. Likewise, the promoter regions of vesicular GABA transporter, choline acetyltransferase or tryptophan hydroxylase 2 have been used to target GABAergic, cholinergic, or serotonergic neurons, respectively [19,[24][25][26][27].…”