Paradoxical (REM) sleep genesis: The switch from an aminergic–cholinergic to a GABAergic–glutamatergic hypothesis

In the middle of the last century, Michel Jouvet discovered paradoxical sleep (PS), a sleep phase paradoxically characterized by cortical activation and rapid eye movements and a muscle atonia. Soon after, he showed that it was still present in “pontine cats” in which all structures rostral to the brainstem have been removed. Later on, it was demonstrated that the pontine peri-locus coeruleus α (peri-LCα in cats, corresponding to the sublaterodorsal nucleus, SLD, in rats) is responsible for PS onset. It was then proposed that the onset and maintenance of PS is due to a reciprocal inhibitory interaction between neurons presumably cholinergic specifically active during PS localized in this region and monoaminergic neurons. In the last decade, we have tested this hypothesis with our model of head-restrained rats and functional neuroanatomical studies. Our results confirmed that the SLD in rats contains the neurons responsible for the onset and maintenance of PS. They further indicate that (1) these neurons are non-cholinergic possibly glutamatergic neurons, (2) they directly project to the glycinergic premotoneurons localized in the medullary ventral gigantocellular reticular nucleus (GiV), (3) the main neurotransmitter responsible for their inhibition during waking (W) and slow wave sleep (SWS) is GABA rather than monoamines, (4) they are constantly and tonically excited by glutamate and (5) the GABAergic neurons responsible for their tonic inhibition during W and SWS are localized in the deep mesencephalic reticular nucleus (DPMe). We also showed that the tonic inhibition of locus coeruleus (LC) noradrenergic and dorsal raphe (DRN) serotonergic neurons during sleep is due to a tonic GABAergic inhibition by neurons localized in the dorsal paragigantocellular reticular nucleus (DPGi) and the ventrolateral periaqueductal gray (vlPAG). We propose that these GABAergic neurons also inhibit the GABAergic neurons of the DPMe at the onset and during PS and are therefore responsible for the onset and maintenance of PS.     

Localization of the Brainstem GABAergic Neurons Controlling Paradoxical (REM) Sleep

Paradoxical sleep (PS) is a state characterized by cortical activation, rapid eye movements and muscle atonia. Fifty years after its discovery, the neuronal network responsible for the genesis of PS has been only partially identified. We recently proposed that GABAergic neurons would have a pivotal role in that network. To localize these GABAergic neurons, we combined immunohistochemical detection of Fos with non-radioactive in situ hybridization of GAD67 mRNA (GABA synthesis enzyme) in control rats, rats deprived of PS for 72 h and rats allowed to recover after such deprivation. Here we show that GABAergic neurons gating PS (PS-off neurons) are principally located in the ventrolateral periaqueductal gray (vlPAG) and the dorsal part of the deep mesencephalic reticular nucleus immediately ventral to it (dDpMe). Furthermore, iontophoretic application of muscimol for 20 min in this area in head-restrained rats induced a strong and significant increase in PS quantities compared to saline. In addition, we found a large number of GABAergic PS-on neurons in the vlPAG/dDPMe region and the medullary reticular nuclei known to generate muscle atonia during PS. Finally, we showed that PS-on neurons triggering PS localized in the SLD are not GABAergic. Altogether, our results indicate that multiple populations of PS-on GABAergic neurons are distributed in the brainstem while only one population of PS-off GABAergic neurons localized in the vlPAG/dDpMe region exist. From these results, we propose a revised model for PS control in which GABAergic PS-on and PS-off neurons localized in the vlPAG/dDPMe region play leading roles.     

Control of REM sleep: an aspect of the regulation of physiological homeostasis.

Since REM sleep is characterized by a suspension of the hypothalamic integration of homeostatic regulations, it has been assumed that the duration of both REM sleep episodes and of the time interval between the end of one episode and the beginning of the following episode may be regulated according to sleep related processes and the homeostatic needs of the organism. A series of studies performed on the rat has shown that REM sleep episodes occur as two basic types: single REM sleep episodes, that are separated by intervals > 3 min and sequential episodes, that are separated by intervals < or = 3 min and appear in a cluster. Moreover, it has been observed that, in this species, a change in REM sleep occurrence is caused by a modification in the number of episodes and not in their duration. With respect to this, sleep deprivation and recovery are characterized by a decrease and an increase, respectively, in the number of sequential REM sleep episodes, but the number of single episodes tends to be kept constant. The central aspects of this kind of regulation have been examined biochemically in the preoptic-anterior hypothalamus, an area involved in the control of autonomic and sleep related processes. The results show that the accumulation of adenosine 3':5'-cyclic monophosphate (cAMP) is impaired, in this region, during sleep deprivation and appears to return to the control levels, during the recovery, with a rate inversely related to the degree of the previous deprivation. Moreover, it has been observed that the systemic administration of DL-propranolol and LiCl reduces cAMP accumulation mainly in the preoptic-anterior hypothalamus; this condition is concomitant with a reduction in REM sleep occurrence.     

Neural reapportionment: an hypothesis to account for the function of sleep

Sleep is a ubiquitous component of animal life, and prolonged sleep deprivation is fatal in both vertebrates and invertebrates. The physiologic function of sleep, however, is not known. We propose here that sleep provides a period of time necessary to reapportion resources within neurons and neural systems that become sub-optimally distributed during active waking. Three specific examples of such reapportionment during sleep are suggested: (1) the return of the neurotransmitter, glutamate, to synaptic vesicles at presynaptic sites most active during waking, (2) the intracellular movement of mitochondria from neuronal processes to the cells soma where mitochondrial replication can occur, and (3) the readjustment of the level and distribution of neurotransmitters within the brainstem modulatory systems and elsewhere that must function in an integrated fashion during waking. Experimental approaches that might be utilized to test these hypotheses are suggested.     

Phenomenology of rapid eye movement (REM) sleep behavior disorder in Parkinson’s disease: A descriptive study using the REM sleep behavior disorder severity scale (RBDSS)

Using the rapid eye movement (REM) sleep behavior disorder severity scale (RBDSS) as a reliable clinical tool we further assessed the phenomenology of REM sleep behavior disorder (RBD) in 56 Parkinson’s disease patients with RBD. The scale uses phenomenological categories based on the localization of movements in the distal or proximal extremities and/or involving the trunc and on the presence or absence of vocalizations. Interrater reliability has been published previously. In the current study we looked into the distribution of motor and vocal phenomena. We identified a small percentage of patients with only vocalizations (4/56), and 8/56 patients with and 10/56 patients without vocalizations and small nonviolent movements of the distal extremities or facial jerks. Ten patients showed more forceful movements of the proximal extremities, 11 patients with movements involving the proximal extremities had additional vocalizations, 5 patients were recorded with axial involvement, and 12 patients with axial movements and vocalizations. These findings underline the clinical variety of RBD manifestation in Parkinson’s disease patients without a certain pattern. However, both extreme variants, the mild one and the violent one, are rare. The RBDSS facilitates the risk estimate for harmful behaviors and may be recommended for comparative studies on RBD and its pharmacotherapy.

     

Paradoxical sleep deprivation: effects on brain energy metabolism.

A study was made of brain nucleotides and glycolytic intermediates in paradoxical sleep (PS)-deprived and recovery-sleeping rats. It was observed that PS deprivation of 24 h produced a fall in glucose, glucose 6-phosphate and pyruvate in cerebral frontal lobes. After three hours of recovery sleep all values returned toward their predeprivational levels. In cerebellar hemispheres ATP was increased, while glucose 6-phosphate and pyruvate were decreased. After three hours of recovery sleep, glucose 6-phosphate was increased and pyruvate decreased, indicating restoration of glycogen and creatine phosphate respectively.     

Reduction of REM sleep by a tryptophan-free amino acid diet.

The acute administration of a tryptophan-free amino acid diet to rats has previously been reported to produce a marked reduction in brain serotonin concentrations. The present study examined the effects of such a diet on electroencephalographic sleep measures. There was a decrease in REM sleep and a small increase in nonREM sleep, with no change in total sleep time. In view of these and other observations, the hypothesis that the serotonergic system plays an important role in the maintenance of nonREM sleep should be carefully reevaluated.     

Cerebral blood flow regulation in REM sleep: a model for flow-metabolism coupling.

The pattern of metabolic and circulatory changes occurring during REM sleep in the whole brain is also observed at a regional level in different instances of functional activation. This pattern is characterized by an increase in metabolic rate, blood flow, glucose and oxygen uptake, the increase in glucose uptake generally exceeding oxygen uptake. A model of interpretation is presented, based on the assumption that substantial limitation to oxygen diffusion exists in the brain. According to the model, microregions lying at mid-distance between capillaries may become hypoxic, depending on metabolic rate and blood-cell PO2 difference. At increasing metabolic rates, O2 consumption in pericapillary microregions increases and the PO2 drop becomes steeper. As a consequence, in microregions far from capillaries a decrease in O2 availability occurs, in concomitance with the increase in metabolic rate, so that non-oxidative glucose metabolism develops locally. A similar spatial PO2 pattern forms in the case of arterial hypoxia, when capillary PO2, and then blood-cell PO2 difference, is reduced. The hypoxic microregions are the source of vasodilatatory messages, the consequent vasodilatation increasing average capillary PO2 and then favoring O2 diffusion to the tissue. Oxygen thus appears to be a better candidate than glucose as a mediator of blood flow-metabolism coupling. This is supported by its higher extraction fraction and by the fact that, in physiologic conditions, arterial hypoxia (and not hypoglycemia) acts on cerebral blood flow. Moreover, the diffusion capacity of glucose in the brain is higher than that of oxygen, so that diffusion limitation is more likely to occur for oxygen. The present model allows consistent organization of the stereotyped changes in cerebral blood flow and glucose and oxygen uptake occurring both in REM sleep and in other instances of brain activation.     

Diabetic impotence: studies of nocturnal erection during REM sleep.

Nocturnal erections were studied in 30 diabetic patients who complained of impotence and in 11 healthy volunteers. The maximum increase in penile circumference was measured by a penile strain-gauge and recorded on a portable tape-recorder; an external oculogram was recorded simultaneously to identify periods of rapid-eye-movement sleep. The technique gave reproducible results, was acceptable to patients, and was suitable for use in an oridnary hospital ward. Only six diabetics showed a maximum increase in penile circumference of under 15 mm, whereas all but one of the healthy subjects showed maximum increases above this value. Of the six diabetics, five complained of total impotence and had other features of autonomic neuropathy that suggested an organic basis for their impotence. The other patient complained of partial importence, which was probably caused by psychological factors. These findings suggest that the prevalence of organic impotence among diabetics has been overestimated.     

Minireview. Catecholamines and the sleep-wake cycle. II. REM sleep

The exact role of catecholamines (CA) on REM sleep is still controversial. Lesion studies suggest that norepinephrine plays a neuromodulatory role in REM sleep. Support for this view is provided by pharmacological studies in which noradrenergic neurons are activated or inhibited. Thus, disturbances in the dynamic balance between neurochemical systems may alter the conditions under which optimal REM sleep takes place. Discrete radiofrequency lesions to the pontine giganto-cellular tegmental field (which includes the nuclei reticularis pontis oralis and caudalis and where cholinergic and cholinoceptive neurons have been described), result in the elimination of REM sleep. Circumscribed, electrolytic lesions of the locus coeruleus (IC) area, which only minimally extend beyond it, eliminate atonia and reduce PGO activity in REM sleep. Selective destruction of the LC or ascending noradrenergic axons with 6-hydroxydopamine does not result in significant changes of tonic or phasic components of desynchronized sleep. These results indicate that noradrenergic neurons are not necessary for the initiation and maintenance of REM sleep. Most probably, many of the effects attributed to noradrenergic structures are due to destruction of non-noradrenergic neurons and fibers of passage in the lesioned area.Inhibition of CA synthesis with α-methyl-p-tyrosine has resulted in conflicting effects on REM sleep, which could be related to factors other than NE depletion. Systemic administration of dopamine-β-hydroxylase inhibitors (disulfiram, diethyldithiocarbamate, FLA-63, fusaric acid) produced consistent reductions of REM sleep. However, the simultaneous increase of 5-HT and DA levels complicates the interpretation of these results. Selective pharmacological stimulation of presynaptic α-adrenergic (α2) receptors with clonidine, xylazine or α-methyl-dopa decreases REM sleep. Specific blockade of α 2-receptors with yohimbine, piperoxane or tolazoline also reduces desynchronized sleep, but increases wakefulness. In contrast, drugs with similar affinity for pre and postsynaptic (α1) adrenoceptors (phentolamine) markedly increase REM sleep. Compounds Compounds with agonistic activity at postsynaptic α-adrenergic sites (methoxamine) consistently reduce REM sleep, while derivatives with inhibitory activity restricted to these receptors (thymoxamine, prazosin) produce REM sleep increments. Results from studies where propranolol and isoproterenol were administered to laboratory animals point to an involvement of β-adrenergic mechanisms in REM sleep modulation.Although there is no direct evidence to support a dopaminergic influence upon REM sleep executive mechanisms, indirect pharmacological data suggests a neuromodulatory role for dopaminergic neurons. Direct dopaminergic agonists and antagonists show biphasic effects on REM sleep. Low dosages of apomorphine increase, while large doses decrease, REM sleep. Opposite effects are observed after the dopaminergic antagonist pimozide. These dose-dependent effects seem to be related to the activation or blockade of different receptors.     

Role of the lateral paragigantocellular nucleus in the network of paradoxical (REM) sleep: an electrophysiological and anatomical study in the rat

The lateral paragigantocellular nucleus (LPGi) is located in the ventrolateral medulla and is known as a sympathoexcitatory area involved in the control of blood pressure. In recent experiments, we showed that the LPGi contains a large number of neurons activated during PS hypersomnia following a selective deprivation. Among these neurons, more than two-thirds are GABAergic and more than one fourth send efferent fibers to the wake-active locus coeruleus nucleus. To get more insight into the role of the LPGi in PS regulation, we combined an electrophysiological and anatomical approach in the rat, using extracellular recordings in the head-restrained model and injections of tracers followed by the immunohistochemical detection of Fos in control, PS-deprived and PS-recovery animals. With the head-restrained preparation, we showed that the LPGi contains neurons specifically active during PS (PS-On neurons), neurons inactive during PS (PS-Off neurons) and neurons indifferent to the sleep-waking cycle. After injection of CTb in the facial nucleus, the neurons of which are hyperpolarized during PS, the largest population of Fos/CTb neurons visualized in the medulla in the PS-recovery condition was observed in the LPGi. After injection of CTb in the LPGi itself and PS-recovery, the nucleus containing the highest number of Fos/CTb neurons, moreover bilaterally, was the sublaterodorsal nucleus (SLD). The SLD is known as the pontine executive PS area and triggers PS through glutamatergic neurons. We propose that, during PS, the LPGi is strongly excited by the SLD and hyperpolarizes the motoneurons of the facial nucleus in addition to local and locus coeruleus PS-Off neurons, and by this means contributes to PS genesis.     

REM sleep related increase in brain temperature: a physiologic problem

The roles of metabolic heat production, arterial blood flow and temperature in the genesis of the brain temperature increase related to REM sleep occurrence in several mammalian species are discussed on the basis of available experimental evidence. The experimental data show that only changes in arterial blood flow and temperature consistently underlie the rise in brain temperature in presence (cat) or absence (rabbit) of the carotid rete. The alteration of cardiovascular regulation in REM sleep is the remote cause of such rise. The proximate causes are decrease in carotid blood supply and increase in vertebral blood supply to the brain and related depression of systemic and selective brain cooling.     

Tuberal Hypothalamic Neurons Secreting the Satiety Molecule Nesfatin-1 Are Critically Involved in Paradoxical (REM) Sleep Homeostasis

The recently discovered Nesfatin-1 plays a role in appetite regulation as a satiety factor through hypothalamic leptin-independent mechanisms. Nesfatin-1 is co-expressed with Melanin-Concentrating Hormone (MCH) in neurons from the tuberal hypothalamic area (THA) which are recruited during sleep states, especially paradoxical sleep (PS). To help decipher the contribution of this contingent of THA neurons to sleep regulatory mechanisms, we thus investigated in rats whether the co-factor Nesfatin-1 is also endowed with sleep-modulating properties. Here, we found that the disruption of the brain Nesfatin-1 signaling achieved by icv administration of Nesfatin-1 antiserum or antisense against the nucleobindin2 (NUCB2) prohormone suppressed PS with little, if any alteration of slow wave sleep (SWS). Further, the infusion of Nesfatin-1 antiserum after a selective PS deprivation, designed for elevating PS needs, severely prevented the ensuing expected PS recovery. Strengthening these pharmacological data, we finally demonstrated by using c-Fos as an index of neuronal activation that the recruitment of Nesfatin-1-immunoreactive neurons within THA is positively correlated to PS but not to SWS amounts experienced by rats prior to sacrifice. In conclusion, this work supports a functional contribution of the Nesfatin-1 signaling, operated by THA neurons, to PS regulatory mechanisms. We propose that these neurons, likely releasing MCH as a synergistic factor, constitute an appropriate lever by which the hypothalamus may integrate endogenous signals to adapt the ultradian rhythm and maintenance of PS in a manner dictated by homeostatic needs. This could be done through the inhibition of downstream targets comprised primarily of the local hypothalamic wake-active orexin- and histamine-containing neurons.