An Excitatory Circuit in the Perioculomotor Midbrain for Non-REM Sleep Control

Abstract

Graphical Abstract Highlights d Activity-dependent genetic labeling and gene profiling identify sleep-active neurons d CALCA-expressing glutamatergic neurons in pIII are NREM active and NREM promoting d pIII CCK neurons are NREM promoting and are reciprocally connected to CALCA neurons d Both cell types send long-range excitatory projections to other sleep-promoting areas In BriefIdentification of sleep-active excitatory midbrain neurons that are interconnected and promote sleep via long range projections to other sleep centers of the mouse brain. SUMMARYThe perioculomotor (pIII) region of the midbrain was postulated as a sleep-regulating center in the 1890s but largely neglected in subsequent studies. Using activity-dependent labeling and gene expression profiling, we identified pIII neurons that promote non-rapid eye movement (NREM) sleep. Optrode recording showed that pIII glutamatergic neurons expressing calcitonin gene-related peptide alpha (CALCA) are NREM-sleep active; optogenetic and chemogenetic activation/inactivation showed that they strongly promote NREM sleep. Within the pIII region, CALCA neurons form reciprocal connections with another population of glutamatergic neurons that express the peptide cholecystokinin (CCK). Activation of CCK neurons also promoted NREM sleep. Both CALCA and CCK neurons project rostrally to the preoptic hypothalamus, whereas CALCA neurons also project caudally to the posterior ventromedial medulla. Activation of each projection increased NREM sleep. Together, these findings point to the pIII region as an excitatory sleep center where different subsets of glutamatergic neurons promote NREM sleep through both local reciprocal connections and long-range projections.

Involvement of Centrally Projecting Edinger–Westphal Nucleus Neuropeptides in Actions of Addictive Drugs

“…On the other hand, one report indicates that under certain pathological conditions Ucn1 and TH can be co-expressed in the human embryonic EWcp [13]. More recently, fluorescent in-situ hybridization (FISH) demonstrated that almost all CCK cells also express the gene Slc17a6 [14], encoding the vesicular glutamate transporter 2 (Vglut2), strongly suggesting that they are also glutamatergic [15,16]. Although almost all CCK-positive cells in the EWcp appear to express Slc17a6, Zhang and colleagues found that there were a number of Slc17a6-positive cells that did not express CCK [14].…”

“…More recently, fluorescent in-situ hybridization (FISH) demonstrated that almost all CCK cells also express the gene Slc17a6 [14], encoding the vesicular glutamate transporter 2 (Vglut2), strongly suggesting that they are also glutamatergic [15,16]. Although almost all CCK-positive cells in the EWcp appear to express Slc17a6, Zhang and colleagues found that there were a number of Slc17a6-positive cells that did not express CCK [14]. To date it is not known what other neuropeptides these glutamatergic neurons may express, although preliminary data from the Ryabinin lab indicates that adult Vglut2 neurons in the EWcp do not express Ucn1.…”

“…Substance P and CCK afferents originating from the EWcp have also been detected in the paraventricular thalamic nucleus, where they have been hypothesized to regulate stress responses [81]. Most recently, EWcp CCK-positive cells were shown to project to the pre-optic area (POA) [14]. Using fluorescent in-situ hybridization, this study found that nearly all of the CCK-positive neurons they examined also expressed Slc17a6, indicating that they are also glutamatergic, and thus may also release glutamate in the POA.…”

A wake-active locomotion circuit depolarizes a sleep-active neuron to switch on sleep

“…This sleep switch is under the control of arousal that favors wake and inhibits sleep through the suppression of sleep-active neurons by inhibitory wake-active neurons [6,9]. It has been proposed that sleep induction is favored by disinhibition of inhibitory sleep-active neurons [10][11][12]; also, excitatory sleep-active neurons exist that might perhaps present activators of inhibitory sleep-active neurons [13]. However, the forces and mechanisms that flip the sleep switch from wake to sleep when an organism gets sleepy cannot be satisfactorily explained by the present circuit models as it is unclear how sleep-active neurons are turned on when the system is set to sleep.…”

“…These neurons reside in the anterior hypothalamus, brain stem, and cortex [6,12]. Excitatory sleep-active neurons were found in the periocular midbrain that project to inhibitory sleep-active neurons in the anterior hypothalamus, the role of which could be to activate inhibitory sleep-active neurons [13]. Studying sleep in less complex brains facilitates sleep circuit analysis.…”