REM Sleep Control Research Articles

Dopamine Control of REM Sleep and Cataplexy.

Dopamine Control of REM Sleep and Cataplexy.

Running speed and REM sleep control two distinct modes of rapid interhemispheric communication.

SUMMARYRhythmic gamma-band communication within and across cortical hemispheres is critical for optimal perception, navigation, and memory. Here, using multisite recordings in both rats and mice, we show that even faster ~140 Hz rhythms are robustly anti-phase across cortical hemispheres, visually resembling splines, the interlocking teeth on mechanical gears. Splines are strongest in superficial granular retrosplenial cortex, a region important for spatial navigation and memory. Spline-frequency interhemispheric communication becomes more coherent and more precisely anti-phase at faster running speeds. Anti-phase splines also demarcate high-activity frames during REM sleep. While splines and associated neuronal spiking are anti-phase across retrosplenial hemispheres during navigation and REM sleep, gamma-rhythmic interhemispheric communication is precisely in-phase. Gamma and splines occur at distinct points of a theta cycle and thus highlight the ability of interhemispheric cortical communication to rapidly switch between in-phase (gamma) and anti-phase (spline) modes within individual theta cycles during both navigation and REM sleep.

Dopaminergic Regulation of Serotonergic Dorsal Raphe Neurons: Role in the Control of Wakefulness and REM Sleep

Dopaminergic Regulation of Serotonergic Dorsal Raphe Neurons: Role in the Control of Wakefulness and REM Sleep

A Focal Inactivation and Computational Study of Ventrolateral Periaqueductal Gray and Deep Mesencephalic Reticular Nucleus Involvement in Sleep State Switching and Bistability.

Neurons of the ventrolateral periaqueductal gray (vlPAG) and adjacent deep mesencephalic reticular nucleus (DpMe) are implicated in the control of sleep-wake state and are hypothesized components of a flip-flop circuit that maintains sleep bistability by preventing the overexpression of non-rapid eye movement (NREM)/REM sleep intermediary states (NRt). To determine the contribution of vlPAG/DpMe neurons in maintaining sleep bistability we combined computer simulations of flip-flop circuitry with focal inactivation of vlPAG/DpMe neurons by microdialysis delivery of the GABAA receptor agonist muscimol in freely behaving male rats (n = 25) instrumented for electroencephalographic and electromyographic recording. REM sleep was enhanced by muscimol at the vlPAG/DpMe, consistent with previous studies; however, our analyses of NRt dynamics in vivo and those produced by flop-flop circuit simulations show that current thinking is too narrowly focused on the contribution of REM sleep-inactive populations toward vlPAG/DpMe involvement in REM sleep control. We found that much of the muscimol-mediated increase in REM sleep was more appropriately classified as NRt. This loss of sleep bistability was accompanied by fragmentation of REM sleep, as evidenced by an increased number of short REM sleep bouts. REM sleep fragmentation stemmed from an increased number and duration of NRt bouts originating in REM sleep. By contrast, NREM sleep bouts were not likewise fragmented by vlPAG/DpMe inactivation. In flip-flop circuit simulations, these changes could not be replicated through inhibition of the REM sleep-inactive population alone. Instead, combined suppression of REM sleep active and inactive vlPAG/DpMe subpopulations was required to replicate the changes in NRt dynamics.

Melanin-concentrating hormone (MCH) in the median raphe nucleus: Fibers, receptors and cellular effects

Serotonergic neurons of the median raphe nucleus (MnR) and hypothalamic melanin-concentrating hormone (MCH)-containing neurons, have been involved in the control of REM sleep and mood.In the present study, we examined in rats and cats the anatomical relationship between MCH-containing fibers and MnR neurons, as well as the presence of MCHergic receptors in these neurons. In addition, by means of in vivo unit recording in urethane anesthetized rats, we determined the effects of MCH in MnR neuronal firing.Our results showed that MCH-containing fibers were present in the central and paracentral regions of the MnR. MCHergic fibers were in close apposition to serotonergic and non-serotonergic neurons. By means of an indirect approach, we also analyzed the presence of MCHergic receptors within the MnR. Accordingly, we microinjected MCH conjugated with the fluorophore rhodamine (R-MCH) into the lateral ventricle. R-MCH was internalized into serotonergic and non-serotonergic MnR neurons; some of these neurons were GABAergic. Furthermore, we determined that intracerebroventricular administration of MCH induced a significant decrease in the firing rate of 53 % of MnR neurons, while the juxtacellular administration of MCH reduced the frequency of discharge in 67 % of these neurons. Finally, the juxtacellular administration of the MCH-receptor antagonist ATC-0175 produced an increase in the firing rate in 78 % of MnR neurons. Hence, MCH produces a strong regulation of MnR neuronal activity. We hypothesize that MCHergic modulation of the MnR neuronal activity may be involved in the promotion of REM sleep and in the pathophysiology of depressive disorders.

Dynamic REM Sleep Modulation by Ambient Temperature and the Critical Role of the Melanin-Concentrating Hormone System

Ambient temperature (Ta) warming toward the high end of the thermoneutral zone (TNZ) preferentially increasesrapid eye movement (REM) sleep over non-REM (NREM) sleep across species. The control and function of this temperature-induced REM sleepexpression have remained unknown. Melanin-concentrating hormone (MCH) neurons play an important role in REM sleep control. We hypothesize that the MCH system may modulate REM sleep as a function of Ta. Here, we show that wild-type (WT) mice dynamically increased REM sleep durations specifically during warm Ta pulsing within the TNZ, compared to both the TNZ cool and baseline constant Ta conditions, without significantly affecting either wake or NREM sleep durations. However, genetically engineered MCH receptor-1 knockout (MCHR1-KO) mice showed no significant changes in REM sleep as a function of Ta, even with increased sleep pressure following a 4-h sleep deprivation. Using MCH-cre mice transduced with channelrhodopsin, we then optogenetically activated MCH neurons time locked with Ta warming, showing an increase in REM sleep expression beyond what Ta warming in yellow fluorescent protein (YFP) control mice achieved. Finally, in mice transduced with archaerhodopsin-T, semi-chronic optogenetic MCH neuronal silencing during Ta warming completely blocked the increase in REM sleep seen in YFP controls. These data demonstrate a previously unknown role for the MCH system in the dynamic output expression of REM sleep during Ta manipulation. These findings are consistent with the energy allocation hypothesis of sleep function, suggesting that endotherms have evolved neural circuits to opportunistically express REM sleep when the need for thermoregulatory defense is minimized.

0064 Lateral Hypothalamic Control of REM Sleep Expression During Ambient Temperature Warming

Sleep and thermoregulation are tightly integrated. Moreover, ambient temperature (Ta) warming preferentially increases REM sleep over NREM sleep across species. Although the role of the anterior hypothalamus in thermoregulation and NREM sleep expression are well documented, the hypothalamic control of REM sleep during ambient temperature manipulation remains to be determined. We examined the role of the posterior lateral hypothalamus (LH) melanin concentrating hormone (MCH) system in the modulation of REM sleep during Ta manipulation. Genetically engineered MCH-R1-knockout (KO) mice or MCH::cre mice implanted with cortical EEG and neck EMG electrodes were used for these experiments. Mice were housed at 23.0 ± 1.0°C in their home cages in a temperature-controlled cabinet. During the light period at two hour intervals, four bouts of rapid Ta warming to a maximum of 31.7°C were achieved for 30 minutes with a thermostatically controlled combination of convection heater and indirect infrared lamp. To optogenetically stimulate MCH neurons, we stereotactically injected AAV2-ChR2-eYFP or AAV2-eYFP (controls) in the LH of MCH::Cre mice. In wild type (WT) controls, REM sleep was significantly increased in the warm (27.5–31°C) versus cool (24.0–27.5˚C) condition. However, MCH-R1-KO mice showed no significant changes in REM sleep as a function of Ta. Random semi-chronic optical activation of MCH neurons significantly increased NREM-to-REM sleep transitions, and prolonged REM sleep durations compared to YFP controls, but did not affect NREM sleep. Additional Ta warming had an additive effect on increasing REM sleep, demonstrating that optical stimulation of MCH neurons in combination with Ta warming was able to “over-drive” the expression of REM sleep beyond what optical stimulation or Ta warming alone could achieve. Indeed, optical stimulation had the same effect on increasing REM sleep expression as Ta warming in the YFP control mice. These data support a key role for the MCH system in the output expression of REM sleep during Ta manipulation, and show that concomitant MCH activation with Ta warming can over-express this effect. Ohio Sleep Medicine and Neuroscience Institute, Inselspital Bern University Hospital.

Distribution of MCH-containing fibers in the feline brainstem: Relevance for REM sleep regulation

Neurons that utilize melanin-concentrating hormone (MCH) as a neuromodulator are localized in the postero-lateral hypothalamus and incerto-hypothalamic area. These neurons project diffusely throughout the central nervous system and have been implicated in critical physiological processes, such as sleep. Unlike rodents, in the order carnivora as well as in humans, MCH exerts its biological functions through two receptors: MCHR-1 and MCHR-2. Hence, the cat is an optimal animal to model MCHergic functions in humans.In the present study, we examined the distribution of MCH-positive fibers in the brainstem of the cat. MCHergic axons with distinctive varicosities and boutons were heterogeneously distributed, exhibiting different densities in distinct regions of the brainstem. High density of MCHergic fibers was found in the dorsal raphe nucleus, the laterodorsal tegmental nucleus, the periaqueductal gray, the pendunculopontine tegmental nucleus, the locus coeruleus and the prepositus hypoglossi. Because these areas are involved in the control of REM sleep, the present anatomical data support the role of this neuropeptidergic system in the control of this behavioral state.

0135 GABAERGIC NEURONS IN THE PREOPTIC HYPOTHALAMUS PROJECT TO MIDBRAIN STRUCTURES INVOLVED IN REM SLEEP CONTROL

GABAergic, sleep-active neurons in the preoptic hypothalamus are key components of hypothalamic-brainstem circuits that regulate sleep and arousal. Many sleep-active neurons recorded in the median preoptic nucleus (MnPO) and the ventrolateral preoptic area (VLPO) exhibit elevated discharge rates during both nonREM and REM sleep compared to waking. A population of neurons in the dorsal lateral preoptic area (DLPO) exhibit REM-active discharge. Preoptic nonREM/REM-active neurons may participate in control of the nonREM-REM cycle. To evaluate this, we examined projections of GABAergic neurons in the preoptic area to midbrain nuclei implicated in REM sleep control, the ventrolateral periaqueductal gray (vlPAG) and the dorsal raphe nucleus (DRN). Adult Sprague-Dawley rats received unilateral injections of the retrograde anatomical tracer, cholera toxin subunit-b Alexa Fluor 594 conjugate (CTb), targeting the vlPAG or DRN. After a 14-day survival period to allow for retrograde transport of tracer, rats were euthanatized and tissue sections through the preoptic hypothalamus were processed for visualization of CTb and of glutamic acid decarboxylase (GAD), a marker of GABAergic neurons. CTb injections that successfully targeted the vlPAG resulted in a moderate density of CTb-labeled cell bodies in the VLPO with co-localization of GAD occurring in ~30% CTb+ neurons. The density of CTb labeling in the MnPO and DLPO was somewhat lower, with co-localization of GAD occurring in ~15–20% of CTB+ neurons. CTb injections in the DRN yielded moderate to high density of retrogradely labeled neurons in the VLPO, with co-localization of GAD in ~40–50% of CTb+ neurons. Compared to VLPO, co-localization of CTb and GAD in the MnPO and DLPO was low. This study confirmed anatomical connections between GABAergic neurons in sleep-regulatory regions of the preoptic area to REM sleep-regulatory regions in the midbrain, with the highest density of GABAergic projection neurons originating in the VLPO. These connections may functionally integrate neuronal systems that control sleep onset and nonREM sleep to brainstem REM sleep generating circuits. Supported by the Department of Veterans Affairs (Grant BX00155605) and the National Institutes of Health (Grant R01-NS092383).

Novel method for evaluation of eye movements in patients with narcolepsy

BackgroundNarcolepsy causes abnormalities in the control of wake-sleep, non-rapid-eye-movement (non-REM) sleep and REM sleep, which includes specific eye movements (EMs). In this study, we aim to evaluate EM characteristics in narcolepsy as compared to controls using an automated detector. MethodsWe developed a data-driven method to detect EMs during sleep based on two EOG signals recorded as part of a polysomnography (PSG). The method was optimized using the manually scored hypnograms from 36 control subjects. The detector was applied on a clinical sample with subjects suspected for central hypersomnias. Based on PSG, multiple sleep latency test and cerebrospinal fluid hypocretin-1 measures, they were divided into clinical controls (N = 20), narcolepsy type 2 (NT2, N = 19), and narcolepsy type 1 (NT1, N = 28). We investigated the distribution of EMs across sleep stages and cycles. ResultsNT1 patients had significantly less EMs during wake, N1, and N2 sleep and more EMs during REM sleep compared to clinical controls, and significantly less EMs during wake and N1 sleep compared to NT2 patients. Furthermore, NT1 patients showed less EMs during NREM sleep in the first sleep cycle and more EMs during NREM sleep in the second sleep cycle compared to clinical controls and NT2 patients. ConclusionsNT1 patients show an altered distribution of EMs across sleep stages and cycles compared to NT2 patients and clinical controls, suggesting that EMs are directly or indirectly controlled by the hypocretinergic system. A data-driven EM detector may contribute to the evaluation of narcolepsy and other disorders involving the control of EMs.

On the Role of the Pedunculopontine Nucleus and Mesencephalic Reticular Formation in Locomotion in Nonhuman Primates.

The mesencephalic reticular formation (MRF) is formed by the pedunculopontine and cuneiform nuclei, two neuronal structures thought to be key elements in the supraspinal control of locomotion, muscle tone, waking, and REM sleep. The role of MRF has also been advocated in modulation of state of arousal leading to transition from wakefulness to sleep and it is further considered to be a main player in the pathophysiology of gait disorders seen in Parkinson's disease. However, the existence of a mesencephalic locomotor region and of an arousal center has not yet been demonstrated in primates. Here, we provide the first extensive electrophysiological mapping of the MRF using extracellular recordings at rest and during locomotion in a nonhuman primate (NHP) (Macaca fascicularis) model of bipedal locomotion. We found different neuronal populations that discharged according to a phasic or a tonic mode in response to locomotion, supporting the existence of a locomotor neuronal circuit within these MRF in behaving primates. Altogether, these data constitute the first electrophysiological characterization of a locomotor neuronal system present within the MRF in behaving NHPs under normal conditions, in accordance with several studies done in different experimental animal models. We provide the first extensive electrophysiological mapping of the two major components of the mesencephalic reticular formation (MRF), namely the pedunculopontine and cuneiform nuclei. We exploited a nonhuman primate (NHP) model of bipedal locomotion with extracellular recordings in behaving NHPs at rest and during locomotion. Different MRF neuronal groups were found to respond to locomotion, with phasic or tonic patterns of response. These data constitute the first electrophysiological evidences of a locomotor neuronal system within the MRF in behaving NHPs.

Melanin-Concentrating Hormone (MCH): Role in REM Sleep and Depression.

The melanin-concentrating hormone (MCH) is a peptidergic neuromodulator synthesized by neurons of the lateral sector of the posterior hypothalamus and zona incerta. MCHergic neurons project throughout the central nervous system, including areas such as the dorsal (DR) and median (MR) raphe nuclei, which are involved in the control of sleep and mood. Major Depression (MD) is a prevalent psychiatric disease diagnosed on the basis of symptomatic criteria such as sadness or melancholia, guilt, irritability, and anhedonia. A short REM sleep latency (i.e., the interval between sleep onset and the first REM sleep period), as well as an increase in the duration of REM sleep and the density of rapid-eye movements during this state, are considered important biological markers of depression. The fact that the greatest firing rate of MCHergic neurons occurs during REM sleep and that optogenetic stimulation of these neurons induces sleep, tends to indicate that MCH plays a critical role in the generation and maintenance of sleep, especially REM sleep. In addition, the acute microinjection of MCH into the DR promotes REM sleep, while immunoneutralization of this peptide within the DR decreases the time spent in this state. Moreover, microinjections of MCH into either the DR or MR promote a depressive-like behavior. In the DR, this effect is prevented by the systemic administration of antidepressant drugs (either fluoxetine or nortriptyline) and blocked by the intra-DR microinjection of a specific MCH receptor antagonist. Using electrophysiological and microdialysis techniques we demonstrated also that MCH decreases the activity of serotonergic DR neurons. Therefore, there are substantive experimental data suggesting that the MCHergic system plays a role in the control of REM sleep and, in addition, in the pathophysiology of depression. Consequently, in the present report, we summarize and evaluate the current data and hypotheses related to the role of MCH in REM sleep and MD.

Hypoxia and Hypercapnia Impairs Cardiac Vagal Control: the Role of Orexin

Patients with obstructive sleep apnea (OSA) have an increased risk of cardiovascular diseases and altered plasma levels of orexin. REM sleep further increases the incidence of adverse cardiovascular events. The mechanisms underlying REM sleep‐related reduction in parasympathetic activity include activation of inhibitory input to cardiac vagal neurons (CVNs) in the brainstem originating from the lateral paragigantocellular nucleus (LPGi), a nucleus which plays a role in REM sleep control. This study tests the hypotheses that (1) chronic intermittent hypoxia and hypercapnia (CIHH) inhibits CVNs due to exaggeration of GABAergic pathway from the LPGi to CVNs; (2) orexinergic signaling in CVNs is diminished by CIHH; and (3) identified orexin neurons (orexin‐EGFP‐2A‐Cre rat) are inhibited by acute hypoxia/hypercapnia. GABAergic neurotransmission to CVNs evoked by electrical stimulation of the LPGi was examined using whole‐cell patch‐clamp recordings in rats exposed to CIHH and control rats. GABAergic synaptic events were enhanced following 4 weeks of CIHH. Orexin‐A elicited inhibition of LPGi‐evoked GABAergic current in control animals but evoked no significant changes in CIHH rats. Acute hypoxia/hypercapnia inhibited the firing activity of orexin neurons and diminished glutamatergic neurotransmission to orexin neurons. In summary, repetitive episodes of hypoxia/hypercapnia would diminish activity of orexin neurons, including the orexinergic neurotransmission to CVNs. Diminished orexinergic signaling, along with exaggerated inhibitory neurotransmission from the LPGi to CVNs would reduce cardio‐protective parasympathetic activity and increase risk of cardiovascular diseases.Support: Christian Gillin Research Grant 004GN12 to O.D. and NIH grants HL49965, HL59895 and HL72006 to D.M.

Inhibitory and excitatory amino acid neurotransmitters are utilized by the projection from the dorsal deep mesencephalic nucleus to the sublaterodorsal nucleus REM sleep induction zone

The sublaterodorsal nucleus (SLD) in the pons of the rat is a locus supporting short-latency induction of a REM sleep-like state following local application of a GABAA receptor antagonist or kainate, glutamate receptor agonist. One putatively relevant source of these neurotransmitters is from the region of the deep mesencephalic nucleus (DpMe) just ventrolateral to the periaquiductal gray, termed the dorsal DpMe (dDpMe). Here, the amino acid neurotransmitter innervation of SLD from dDpMe was studied utilizing anterograde tract-tracing with biotinylated dextranamine (BDA) and fluorescence immunohistochemistry visualized with laser scanning confocal microscopy. Both markers for inhibitory and excitatory amino acid neurotransmitters were found in varicose axon fibers in SLD originating from dDpMe. Vesicular glutamate transporter2 (VGLUT2) represented the largest number of anterogradely labeled varicosities followed by vesicular GABA transporter (VGAT). Numerous VGAT and VGLUT2 labeled varicosities were observed apposed to dDpMe-labeled axon fibers indicating both excitatory and inhibitory presynaptic, local modulation within the SLD. Some double-labeled BDA/VGAT varicosities were seen apposed to small somata labeled for glutamate consistent with being presynaptic to the phenotype of REM sleep-active SLD neurons. Results found support the current theoretical framework of the interaction of dDpMe and SLD in control of REM sleep, while also indicating operation of mechanisms with a greater level of complexity.

Cholinergic involvement in control of REM sleep paralysis

REM sleep, also known as dreaming sleep, is marked by intense cortical activation and absence of skeletal muscle tone, so-called REM sleep paralysis (atonia). It is commonly believed that REM sleep paralysis functions to prevent movement during vivid dreams. Indeed, REM sleep behaviour disorder – a neurological condition marked by violent dream enactment – results from loss of REM sleep paralysis. For the last 50 years, biologists have focused on the identification of brain mechanisms responsible for REM sleep. A majority of evidence suggests that a brainstem region known as the sublaterodorsal nucleus (SLD), also called the subcoeruleus, is important for REM sleep generation (Jouvet 1962). However, there is uncertainty concerning the chemical mechanisms by which the SLD triggers REM sleep phenomena. For example, some data suggest that cholinergic modulation of SLD cells underlies REM sleep generation, whereas, other data suggest that GABAergic disinhibition and glutamatergic excitation of SLD cells are critical for REM sleep control (Boissard et al. 2002; Lu et al. 2006). The recent study by Weng et al. (2014) provides a potentially new framework for understanding REM sleep control by showing that both cholinergic and glutamatergic processes operating within the SLD could be important for triggering REM sleep paralysis.

GABAA receptors implicated in REM sleep control express a benzodiazepine binding site

It has been reported that non-subtype-selective GABAA receptor antagonists injected into the nucleus pontis oralis (PnO) of rats induced long-lasting increases in REM sleep. Characteristics of these REM sleep increases were identical to those resulting from injection of muscarinic cholinergic agonists. Both actions were blocked by the muscarinic antagonist, atropine. Microdialysis of GABAA receptor antagonists into the PnO resulted in increased acetylcholine levels. These findings were consistent with GABAA receptor antagonists disinhibiting acetylcholine release in the PnO to result in an acetylcholine-mediated REM sleep induction. Direct evidence has been lacking for localization in the PnO of the specific GABAA receptor-subtypes mediating the REM sleep effects. Here, we demonstrated a dose-related, long-lasting increase in REM sleep following injection (60nl) in the PnO of the inverse benzodiazepine agonist, methyl-6,7-dimethoxy-4-ethyl-β-carboline (DMCM, 10−2M). REM sleep increases were greater and more consistently produced than with the non-selective antagonist gabazine, and both were blocked by atropine. Fluorescence immunohistochemistry and laser scanning confocal microscopy, colocalized in PnO vesicular acetylcholine transporter, a presynaptic marker of cholinergic boutons, with the γ2 subunit of the GABAA receptor. These data provide support for the direct action of GABA on mechanisms of acetylcholine release in the PnO. The presence of the γ2 subunit at this locus and the REM sleep induction by DMCM are consistent with binding of benzodiazepines by a GABAA receptor-subtype in control of REM sleep.

Hypocretinergic and non-hypocretinergic projections from the hypothalamus to the REM sleep executive area of the pons

Within the postero-lateral hypothalamus neurons that utilize hypocretin or melanin-concentrating hormone (MCH) as neuromodulators are co-distributed. These neurons have been involved in the control of behavioral states, and a deficit in the hypocretinergic system is the pathogenic basis of narcolepsy with cataplexy.In this report, utilizing immunohistochemistry and retrograde tracing techniques, we examined the hypocretinergic innervation of the nucleus pontis oralis (NPO), which is the executive site that is responsible for the generation of REM sleep in the cat.The retrograde tracer cholera toxin subunit b (CTb) was administered in pontine regions where carbachol microinjections induced REM sleep. Utilizing immunohistochemical techniques, we found that approximately 1% of hypocretinergic neurons in the tuberal area of the hypothalamus project to the NPO. In addition, approximately 6% of all CTb+ neurons in this region were hypocretinergic. The hypocretinergic innervation of the NPO was also compared with the innervation of the same site by MCH-containing neurons. More than three times as many MCHergic neurons were found to project to the NPO compared with hypocretinergic cells; both neuronal types exhibited bilateral projections. We also identified a group of non-hypocretinergic non-MCHergic neuronal group of neurons that were intermingled with both hypocretinergic and MCHergic neurons that also projected to this same brainstem region. These neurons were grater in number that either hypocretin or MCH-containing neurons; their soma size was also smaller and their projections were mainly ipsilateral.The present anatomical data suggest that hypocretinergic, MCHergic and an unidentified companion group of neurons of the postero-lateral hypothalamus participate in the regulation of the neuronal activity of NPO neurons, and therefore, are likely to participate in the control of wakefulness and REM sleep.

Periaqueductal gray

AD= : Alzheimer disease; DBS= : deep brain stimulation; GABA= : γ-aminobutyric acid; NO= : nitric oxide; PAG= : periaqueductal gray; PVG= : periventricular gray matter; RVM= : rostral ventromedial medulla; TRPV1= : transient receptor potential vanilloid type 1; VLM= : ventrolateral medulla The periaqueductal gray (PAG) is an anatomic and functional interface between the forebrain and the lower brainstem and has a major role in integrated behavioral responses to internal (e.g., pain) or external (e.g., threat) stressors. The PAG consists of distinct columns that receive selective inputs from the prefrontal cortex, amygdala, hypothalamus, and nociceptive pathways. Via its connections with different brainstem nuclei, the PAG coordinates specific patterns of cardiovascular, respiratory, motor, and pain modulatory responses. These responses vary according to the type of stress and the subject's perception of the threatening stimulus. The PAG is also involved in vocalization, micturition, and thermoregulation, and contributes to mechanisms of arousal and control of REM sleep. The PAG is affected in neurodegenerative disorders, such as Alzheimer disease (AD) and multiple system atrophy. Stimulation of the PAG has been utilized for management of chronic neuropathic pain and recent evidence suggests that it may also be used to relieve refractory hypertension. The organization of the PAG and its multiple functions have been the subjects of several reviews.1,–,11 ### Anatomic subdivisions and neurochemistry. The PAG is continuous with the periventricular gray matter (PVG) and surrounds the midbrain aqueduct except for ventral part, which contains oculomotor-related nuclei rostrally and the dorsal raphe nucleus caudally (figure). Based on cytoarchitecture, chemoarchitecture, and connectivity patterns, the PAG has been subdivided into 4 columns: dorsomedial, dorsolateral, lateral, and ventrolateral.2,–,5 The PAG contains different types of neurons that utilize l-glutamate, γ-aminobutyric acid (GABA), opioids (particularly enkephalin), substance P, neurotensin, and other neurotransmitters. The dorsolateral PAG also contains neurons that express NADPH-diaphorase and synthesize nitric oxide (NO)12; the ventrolateral PAG contains a group of dopaminergic neurons.13 There is also abundant expression of NMDA,14 GABAA,15 μ-opioid,16 neurokinin-1,17 and transient receptor …

Cholinergic Modulation of Narcoleptic Attacks in Double Orexin Receptor Knockout Mice

To investigate how cholinergic systems regulate aspects of the sleep disorder narcolepsy, we video-monitored mice lacking both orexin (hypocretin) receptors (double knockout; DKO mice) while pharmacologically altering cholinergic transmission. Spontaneous behavioral arrests in DKO mice were highly similar to those reported in orexin-deficient mice and were never observed in wild-type (WT) mice. A survival analysis revealed that arrest lifetimes were exponentially distributed indicating that random, Markovian processes determine arrest lifetime. Low doses (0.01, 0.03 mg/kg, IP), but not a high dose (0.08 mg/kg, IP) of the cholinesterase inhibitor physostigmine increased the number of arrests but did not alter arrest lifetimes. The muscarinic antagonist atropine (0.5 mg/kg, IP) decreased the number of arrests, also without altering arrest lifetimes. To determine if muscarinic transmission in pontine areas linked to REM sleep control also influences behavioral arrests, we microinjected neostigmine (50 nl, 62.5 µM) or neostigmine + atropine (62.5 µM and 111 µM respectively) into the nucleus pontis oralis and caudalis. Neostigmine increased the number of arrests in DKO mice without altering arrest lifetimes but did not provoke arrests in WT mice. Co-injection of atropine abolished this effect. Collectively, our findings establish that behavioral arrests in DKO mice are similar to those in orexin deficient mice and that arrests have exponentially distributed lifetimes. We also show, for the first time in a rodent narcolepsy model, that cholinergic systems can regulate arrest dynamics. Since perturbations of muscarinic transmission altered arrest frequency but not lifetime, our findings suggest cholinergic systems influence arrest initiation without influencing circuits that determine arrest duration.

Cholinergic modulation of GABAergic and glutamatergic transmission in the dorsal subcoeruleus: mechanisms for REM sleep control.

Dorsal subcoeruleus (SubCD) neurons are thought to promote PGO waves and to be modulated by cholinergic afferents during REM sleep. We examined the differential effect of the cholinergic agonist carbachol (CAR) on excitatory and inhibitory postsynaptic currents (PSCs), and investigated the effects of CAR on SubCD neurons during the developmental decrease in REM sleep. Whole-cell patch clamp recordings were conducted on brainstem slices of 7- to 20-day-old rats. CAR acted directly on 50% of SubCD neurons by inducing an inward current, via both nicotinic and muscarinic M1 receptors. CAR induced a potassium mediated outward current via activation of M2 muscarinic receptors in 43% of SubCD cells. Evoked stimulation established the presence of NMDA, AMPA, GABA, and glycinergic PSCs in the SubCD. CAR was found to decrease the amplitude of evoked EPSCs in 31 of 34 SubCD cells, but decreased the amplitude of evoked IPSCs in only 1 of 13 SubCD cells tested. Spontaneous EPSCs were decreased by CAR in 55% of cells recorded, while spontaneous IPSCs were increased in 27% of SubCD cells. These findings indicate that CAR exerts a predominantly inhibitory role on fast synaptic glutamatergic activity and a predominantly excitatory role on fast synaptic GABAergic/glycinergic activity in the SubCD. We hypothesize that during REM sleep, cholinergic "REM-on" neurons that project to the SubCD induce an excitation of inhibitory interneurons and inhibition of excitatory events leading to the production of coordinated activity in SubCD projection neurons. The coordination of these projection neurons may be essential for the production of REM sleep signs such as PGO waves.