Noriaki Sakai

Muscleblind-like 2-Mediated Alternative Splicing in the Developing Brain and Dysregulation in Myotonic Dystrophy

The RNA-mediated disease model for myotonic dystrophy (DM) proposes that microsatellite C(C)TG expansions express toxic RNAs that disrupt splicing regulation by altering MBNL1 and CELF1 activities. While this model explains DM manifestations in muscle, less is known about the effects of C(C)UG expression on the brain. Here, we report that Mbnl2 knockout mice develop several DM-associated central nervous system (CNS) features including abnormal REM sleep propensity and deficits in spatial memory. Mbnl2 is prominently expressed in the hippocampus and Mbnl2 knockouts show a decrease in NMDA receptor (NMDAR) synaptic transmission and impaired hippocampal synaptic plasticity. While Mbnl2 loss did not significantly alter target transcript levels in the hippocampus, misregulated splicing of hundreds of exons was detected using splicing microarrays, RNA-seq, and HITS-CLIP. Importantly, the majority of the Mbnl2-regulated exons examined were similarly misregulated in DM. We propose that major pathological features of the DM brain result from disruption of the MBNL2-mediated developmental splicing program.

Basal forebrain circuit for sleep-wake control

The mammalian basal forebrain (BF) has important roles in controlling sleep and wakefulness, but the underlying neural circuit remains poorly understood. We examined the BF circuit by recording and optogenetically perturbing the activity of four genetically defined cell types across sleep-wake cycles and by comprehensively mapping their synaptic connections. Recordings from channelrhodopsin-2 (ChR2)-tagged neurons revealed that three BF cell types, cholinergic, glutamatergic and parvalbumin-positive (PV+) GABAergic neurons, were more active during wakefulness and rapid eye movement (REM) sleep (wake/REM active) than during non-REM (NREM) sleep, and activation of each cell type rapidly induced wakefulness. By contrast, activation of somatostatin-positive (SOM+) GABAergic neurons promoted NREM sleep, although only some of them were NREM active. Synaptically, the wake-promoting neurons were organized hierarchically by glutamatergic→cholinergic→PV+ neuron excitatory connections, and they all received inhibition from SOM+ neurons. Together, these findings reveal the basic organization of the BF circuit for sleep-wake control.

Greatly increased numbers of histamine cells in human narcolepsy with cataplexy.

To determine whether histamine cells are altered in human narcolepsy with cataplexy and in animal models of this disease.

A PERIOD3 variant causes a circadian phenotype and is associated with a seasonal mood trait

Significance It has long been thought that sleep and mood are intimately connected in humans, but at present there are no known molecular links. We identified rare variants in the PERIOD3 gene in persons with both altered sleep behavior and features of seasonal affective disorder. We show that these variants recapitulate circadian and mood phenotypes in mouse models. Although we were not able to test mood in fruit flies, we did uncover a sleep trait similar to that seen in humans in flies carrying the human variants. Our molecular studies reveal that the variants led to less stable PER3 protein and reduced the stabilizing effect of PER3 on PER1/PER2, providing a mechanistic explanation for the circadian trait. In humans, the connection between sleep and mood has long been recognized, although direct molecular evidence is lacking. We identified two rare variants in the circadian clock gene PERIOD3 (PER3-P415A/H417R) in humans with familial advanced sleep phase accompanied by higher Beck Depression Inventory and seasonality scores. hPER3-P415A/H417R transgenic mice showed an altered circadian period under constant light and exhibited phase shifts of the sleep-wake cycle in a short light period (photoperiod) paradigm. Molecular characterization revealed that the rare variants destabilized PER3 and failed to stabilize PERIOD1/2 proteins, which play critical roles in circadian timing. Although hPER3-P415A/H417R-Tg mice showed a mild depression-like phenotype, Per3 knockout mice demonstrated consistent depression-like behavior, particularly when studied under a short photoperiod, supporting a possible role for PER3 in mood regulation. These findings suggest that PER3 may be a nexus for sleep and mood regulation while fine-tuning these processes to adapt to seasonal changes.

The sleep-promoting and hypothermic effects of glycine are mediated by NMDA receptors in the suprachiasmatic nucleus.

The use of glycine as a therapeutic option for improving sleep quality is a novel and safe approach. However, despite clinical evidence of its efficacy, the details of its mechanism remain poorly understood. In this study, we investigated the site of action and sleep-promoting mechanisms of glycine in rats. In acute sleep disturbance, oral administration of glycine-induced non-rapid eye movement (REM) sleep and shortened NREM sleep latency with a simultaneous decrease in core temperature. Oral and intracerebroventricular injection of glycine elevated cutaneous blood flow (CBF) at the plantar surface in a dose-dependent manner, resulting in heat loss. Pretreatment with N-methyl-D-aspartate (NMDA) receptor antagonists AP5 and CGP78608 but not the glycine receptor antagonist strychnine inhibited the CBF increase caused by glycine injection into the brain. Induction of c-Fos expression was observed in the hypothalamic nuclei, including the medial preoptic area (MPO) and the suprachiasmatic nucleus (SCN) shell after glycine administration. Bilateral microinjection of glycine into the SCN elevated CBF in a dose-dependent manner, whereas no effect was observed when glycine was injected into the MPO and dorsal subparaventricular zone. In addition, microinjection of D-serine into the SCN also increased CBF, whereas these effects were blocked in the presence of L-701324. SCN ablation completely abolished the sleep-promoting and hypothermic effects of glycine. These data suggest that exogenous glycine promotes sleep via peripheral vasodilatation through the activation of NMDA receptors in the SCN shell.

Familial narcolepsy in the Lipizzaner horse: a report of three fillies born to the same sire

The occurrence of sleep disorder in three half sibling Lipizzaner is described. Sleepiness, swaying, stumbling, carpal joints buckling and falling down onto the carpal joints had been present since early foal age in all of them. Clinical signs had gradually reduced since the age of 2 years in cases 1 and 3. Sleepiness was induced by going out from the stable in adulthood. A physostigmine test was performed in all three affected horses and produced positive results in cases 1 and 3. The result of the test in case 2 was unclear due to the almost continuous sleepiness of the foal. Hypocretin-1 concentration in the cerebrospinal fluid was established using a standardised radioimmunoassay in case 1 (317.85 pg/mL), case 2 (303.43 pg/mL) and five adult control horses (275.2 ± 47.9 [SD] pg/mL) and was considered as normal in all horses. The sire of the affected horses has had 19 other registered offspring who did not show clinical signs of sleep disorder and also dams of all three cases produced healthy foals. Based on the demographic and clinical data together with the responses to the physostigmine challenges, the diagnosis of familial equine narcolepsy was made.

An overview of hypocretin based therapy in narcolepsy

ABSTRACT Introduction: Narcolepsy with cataplexy is most commonly caused by a loss of hypocretin/orexin peptide-producing neurons in the hypothalamus (i.e., Narcolepsy Type 1). Since hypocretin deficiency is assumed to be the main cause of narcoleptic symptoms, hypocretin replacement will be the most essential treatment for narcolepsy. Unfortunately, this option is still not available clinically. There are many potential approaches to replace hypocretin in the brain for narcolepsy such as intranasal administration of hypocretin peptides, developing small molecule hypocretin receptor agonists, hypocretin neuronal transplantation, transforming hypocretin stem cells into hypothalamic neurons, and hypocretin gene therapy. Together with these options, immunotherapy treatments to prevent hypocretin neuronal death should also be developed. Areas covered: In this review, we overview the pathophysiology of narcolepsy and the current and emerging treatments of narcolepsy especially focusing on hypocretin receptor based treatments. Expert opinion: Among hypocretin replacement strategies, developing non-peptide hypocretin receptor agonists is currently the most encouraging since systemic administration of a newly synthesized, selective hypocretin receptor 2 agonist (YNT-185) has been shown to ameliorate symptoms of narcolepsy in murine models. If this option is effective in humans, hypocretin cell transplants or gene therapy technology may become realistic in the future.

Noninvasive detection of sleep/wake changes and cataplexy-like behaviors in orexin/ataxin-3 transgenic narcoleptic mice across the disease onset.

Sleep and behavioral monitoring of young mice is necessary for understating the progress of symptoms in congenital and acquired diseases associated with sleep and movement disorders. In the current study, we have developed a non-invasive sleep monitoring system that identifies wake and sleep patterns of newborn mice using a simple piezoelectric transducer (PZT). Using this system, we have succeeded in detecting age-dependent occurrences and changes in sleep fragmentation of orexin/ataxin-3 narcoleptic mice (a narcoleptic mouse model with postnatal hypocretin/orexin cell death) across the disease onset. We also detected REM sleep/cataplexy patterns (i.e., immobility with clear heartbeat [IMHB] signals due to the flaccid posture) by the PZT system, and found that sudden onset of REM sleep-like episodes specifically occur in narcoleptic, but not in wild type mice, suggesting that these episodes are likely cataplexy. In contrast, gradual onset of IMHB likely reflects occurrence of REM sleep. In summary, we have shown that the PZT system is useful as a non-invasive sleep and behavior monitoring system to analyze the developmental aspects of sleep and movement disorders in mice models.

Modulations of Ventral Tegmental Area (VTA) Dopaminergic Neurons by Hypocretins/Orexins: Implications in Vigilance and Behavioral Control

In this review, our current understandings of interactions of the hypothalamic hypocretin/orexin (hcrt/ox) and midbrain VTA dopaminergic (DAergic) systems will be discussed. We will first overview the (1) neurobiology of wakefulness and (2) symptoms of narcolepsy, followed by discoveries of the hcrt/ox system and hcrt/ox deficiency in narcolepsy. We will then discuss the functional links between the VTA DAergic and hcrt/ox systems, by introducing the results of anatomical and functional (electrophysiological /pharmacological results) studies. Many neuroscientists are also interested in functional roles of the VTA DAergic and hcrt/ox systems in reward-motivational behavior, food intake, and nociception, and some of these results will also be briefly introduced.

The pathogenesis of narcolepsy, current treatments and prospective therapeutic targets

Introduction: Narcolepsy is a chronic sleep disorder characterized by excessive daytime sleepiness (EDS), cataplexy and abnormal manifestations of rapid eye movement (REM) sleep such as sleep paralysis and hypnagogic hallucinations. Recent studies have demonstrated that narcolepsy–cataplexy is caused by the postnatal loss of hypocretin neurons (Type 1 narcolepsy), possibly because of autoimmune mechanisms. Although several new therapeutic options have recently been introduced, these new findings have not yet been incorporated to the therapies. Areas covered: In this review, we first describe clinical symptoms and pathogenesis of narcolepsy. Second, we summarize the current knowledge about pharmacological treatments of narcolepsy and the modes of action of each drug. Finally, we discuss prospective therapeutic targets for narcolepsy. Expert opinion: Current treatments are mostly symptomatic, but it is necessary to initiate the appropriate pharmacological treatment for each symptom with the understanding of pharmacological mechanisms. Development of synthetic hypocretin receptor agonists is the first and most urgent step for the improved treatment of Type 1 narcolepsy. Successes in this step will likely lead to developments of cell transplantation and gene therapies.