Stéphanie Maret

Homer1a is a core brain molecular correlate of sleep loss

Sleep is regulated by a homeostatic process that determines its need and by a circadian process that determines its timing. By using sleep deprivation and transcriptome profiling in inbred mouse strains, we show that genetic background affects susceptibility to sleep loss at the transcriptional level in a tissue-dependent manner. In the brain, Homer1a expression best reflects the response to sleep loss. Time-course gene expression analysis suggests that 2,032 brain transcripts are under circadian control. However, only 391 remain rhythmic when mice are sleep-deprived at four time points around the clock, suggesting that most diurnal changes in gene transcription are, in fact, sleep–wake-dependent. By generating a transgenic mouse line, we show that in Homer1-expressing cells specifically, apart from Homer1a, three other activity-induced genes (Ptgs2, Jph3, and Nptx2) are overexpressed after sleep loss. All four genes play a role in recovery from glutamate-induced neuronal hyperactivity. The consistent activation of Homer1a suggests a role for sleep in intracellular calcium homeostasis for protecting and recovering from the neuronal activation imposed by wakefulness.

Elevated Tribbles homolog 2–specific antibody levels in narcolepsy patients

Narcolepsy is a sleep disorder characterized by excessive daytime sleepiness and attacks of muscle atonia triggered by strong emotions (cataplexy). Narcolepsy is caused by hypocretin (orexin) deficiency, paralleled by a dramatic loss in hypothalamic hypocretin-producing neurons. It is believed that narcolepsy is an autoimmune disorder, although definitive proof of this, such as the presence of autoantibodies, is still lacking. We engineered a transgenic mouse model to identify peptides enriched within hypocretin-producing neurons that could serve as potential autoimmune targets. Initial analysis indicated that the transcript encoding Tribbles homolog 2 (Trib2), previously identified as an autoantigen in autoimmune uveitis, was enriched in hypocretin neurons in these mice. ELISA analysis showed that sera from narcolepsy patients with cataplexy had higher Trib2-specific antibody titers compared with either normal controls or patients with idiopathic hypersomnia, multiple sclerosis, or other inflammatory neurological disorders. Trib2-specific antibody titers were highest early after narcolepsy onset, sharply decreased within 2-3 years, and then stabilized at levels substantially higher than that of controls for up to 30 years. High Trib2-specific antibody titers correlated with the severity of cataplexy. Serum of a patient showed specific immunoreactivity with over 86% of hypocretin neurons in the mouse hypothalamus. Thus, we have identified reactive autoantibodies in human narcolepsy, providing evidence that narcolepsy is an autoimmune disorder.

Clinical efficacy of high-dose intravenous immunoglobulins near the onset of narcolepsy in a 10-year-old boy

Recent studies in narcoleptic children found undetectable cerebro spinal fluid (CSF) hypocretin-1 levels as early as within the first 3 weeks after the disease onset (Kubota et al., 2003). Hecht et al., (2003) also reported an 8-year-old boy with an acute onset of hypocretin-deficiency narcolepsy who did not respond to high-dose prednisone. These together with previous results suggest that clinical manifestations of narcolepsy might occur once CSF hypocretin levels are undetectable, that a putative immune process targets hypocretin neurons with irreversible damage, and that immunosuppressive treatments are ineffective even near the onset of narcolepsy. We have seen a 10-year-old boy 2 months after the onset of cataplexy. Disease onset occurred abruptly with excessive sleepiness and irresistible naps during the winter holidays in February 2003. Two months previously he was perfectly awake under the same conditions, during the Christmas holidays. No triggering factors or accompanying symptoms could be identified except for increased appetite and substantial weight gain (10 kg over the last year). The boy fell asleep regularly at school and was found napping at home. Clear-cut cataplexy with bilateral muscle weakness in lower limbs and face muscles, having duration of a few seconds, and triggered by laughter, subsequently appeared during April. These gradually increased, with daily episodes and injurious falls. He was seen first by ML on June 23. The history was considered typical for classical narcolepsy–cataplexy (in the absence of sleep paralysis or hallucinations), and neurologic and psychiatric examinations were normal. Family history was negative for sleep or neurologic disorders. A routine EEG was normal, with the absence of epileptiform activities or signs of sleepiness. The human leukocyte antigen typing was positive for DQB1*0602/ 0301. Standard brain magnetic resonance imaging did not reveal any lesion except for a non-specific, small left thalamic hyper-intense signal abnormality (gadolinium was not used). CSF hypocretin-1 level was below the detection limit (<40 pg ml). Genomic sequencing of the prepro-hypocretin and its two receptor genes did not reveal any mutation. After obtaining the parents’ consent we initiated intravenous immunoglobulin G (IVIg) perfusion at 1 g kg day for two consecutive days (July 29–30, within 3 months after cataplexy onset). Although the perfusion rate did not exceed 4.5 g h, headache, fever, and flushing were noticed that were resolved with 5 mg dexchlorpheniramine and 50 mg hydroxycortisone (intravenous). Two days after IVIg, prednisolone 1.3 mg kg day was initiated and lasted for 3 weeks. Three days after start of treatment, improvement of both sleepiness and cataplexy were noticed. In the following 3 weeks not a single cataplexy or irresistible nap was reported. The boy’s mother reported a ‘dramatic improvement’, which was confirmed by the absence of irresistible naps and falls both at school and at home. The prednisolone dosage was then reduced to half daily for 3 weeks and a second lumbar puncture was performed on October 1. CSF hypocretin-1 measurement indicated still undetectable levels. Only four partial cataplexies (only facial muscles) were reported during the 2 months following IVIg treatment, with a tendency for gradual increase in daytime sleepiness. Because of prednisolone side effects (additional 3 kg weight gain and severe acne with irritant dermatitis) the treatment was stopped and modafinil at 200 mg day was instituted. At the time of the last interview with the boy’s family, on 3 November 2003, a gradual reappearance over the last 10 days of cataplectic attacks, with few leading to falls, was reported while modafinil was considered to be effective in improving his sleepiness. This is the first report illustrating the clinical efficacy of high-dose IVIg 5 months after the onset of first narcolepsy symptoms. If confirmed in controlled trials and with objective measures of sleepiness, this finding suggests that the immunemediated part of narcolepsy symptoms is independent of hypocretin deficiency. Our finding also indicates that, in contrast to the lack of efficacy of corticosteroids (Hecht et al., 2003), also reported in such immune-mediated inflammatory neuropathies as Guillain–Barré syndrome and multifocal motor neuropathy with conduction blocks, IVIg may represent a new treatment for narcolepsy. Based on our observation we propose the IVIg treatment with an initial 400 mg kg day for 5 days repeated three times at 3–4 week intervals and followed by a similar single-day dose every month for 6 months. Correspondence: Mehdi Tafti, HUG, Belle-Idée, 2 Chemin du PetitBel-Air, 1225 Chêne-Bourg, Switzerland. Tel.: +4122 305 5302; fax: +4122 305 5309; e-mail: Mehdi.Tafti@medecine.unige.ch J. Sleep Res. (2003) 12, 347–348

Genes for normal sleep and sleep disorders

Sleep and wakefulness are complex behaviors that are influenced by many genetic and environmental factors, which are beginning to be discovered. The contribution of genetic components to sleep disorders is also increasingly recognized as important. Point mutations in the prion protein, period 2, and the prepro‐hypocretin/orexin gene have been found as the cause of a few sleep disorders but the possibility that other gene defects may contribute to the pathophysiology of major sleep disorders is worth in‐depth investigations. However, single gene disorders are rare and most common disorders are complex in terms of their genetic susceptibility, environmental effects, gene‐gene, and gene‐environment interactions. We review here the current progress in the genetics of normal and pathological sleep.

A monozygotic twin pair discordant for narcolepsy and CSF hypocretin-1

A genetic basis for narcolepsy is evidenced by a strong association with the HLA DQB1*0602 and the occasional familial occurrence.1 A major discovery was recently made through the identification of orexin/hypocretin deficiency.2,3⇓ While a single mutation in the prepro-hypocretin gene was reported in one atypical patient,3 undetectable levels of CSF hypocretin-1 were reported in most sporadic narcoleptics2 and a dramatic decrease of hypocretin neurons in few postmortem brains.3 Among the 16 monozygotic twin pairs described,1 only 6 are considered to be concordant and most of them were not completely documented. One HLA-DQB1*0602 positive twin pair concordant for narcolepsy was recently reported with normal levels of CSF hypocretin-1 and without any mutation in the hypocretin genes system.4 We studied the CSF hypocretin-1 in a HLA DQB1*0602 positive monozygotic twin pair discordant for narcolepsy. The twin sisters (brought up together until age 18 years) were the second and third born among three siblings, born in 1976. …

Monozygotic twins concordant for narcolepsy-cataplexy without any detectable abnormality in the hypocretin (orexin) pathway

Narcolepsy with cataplexy is thought to be a hypocretin ligand or hypocretin receptor deficiency syndrome caused by genetic and environmental factors. We looked for an abnormality of the hypocretin pathway in HLA-DQB1*0602-positive monozygotic twins who were concordant for narcolepsy-cataplexy. They had normal cerebrospinal fluid concentrations of hypocretin-1, and we found no mutation in the prepro-hypocretin gene or either hypocretin receptor gene. Our finding points to the existence of presumably genetic forms of narcolepsy with cataplexy without any demonstrable defect in the hypocretin pathway.

In Vivo Imaging of the Central and Peripheral Effects of Sleep Deprivation and Suprachiasmatic Nuclei Lesion on PERIOD-2 Protein in Mice.

STUDY OBJECTIVES That sleep deprivation increases the brain expression of various clock genes has been well documented. Based on these and other findings we hypothesized that clock genes not only underlie circadian rhythm generation but are also implicated in sleep homeostasis. However, long time lags have been reported between the changes in the clock gene messenger RNA levels and their encoded proteins. It is therefore crucial to establish whether also protein levels increase within the time frame known to activate a homeostatic sleep response. We report on the central and peripheral effects of sleep deprivation on PERIOD-2 (PER2) protein both in intact and suprachiasmatic nuclei-lesioned mice. DESIGN In vivo and in situ PER2 imaging during baseline, sleep deprivation, and recovery. SETTINGS Mouse sleep-recording facility. PARTICIPANTS Per2::Luciferase knock-in mice. INTERVENTIONS N/A. MEASUREMENTS AND RESULTS Six-hour sleep deprivation increased PER2 not only in the brain but also in liver and kidney. Remarkably, the effects in the liver outlasted those observed in the brain. Within the brain the increase in PER2 concerned the cerebral cortex mainly, while leaving suprachiasmatic nuclei (SCN) levels unaffected. Against expectation, sleep deprivation did not increase PER2 in the brain of arrhythmic SCN-lesioned mice because of higher PER2 levels in baseline. In contrast, liver PER2 levels did increase in these mice similar to the sham and partially lesioned controls. CONCLUSIONS Our results stress the importance of considering both sleep-wake dependent and circadian processes when quantifying clock-gene levels. Because sleep deprivation alters PERIOD-2 in the brain as well as in the periphery, it is tempting to speculate that clock genes constitute a common pathway mediating the shared and well-known adverse effects of both chronic sleep loss and disrupted circadian rhythmicity on metabolic health.