Background: Transitions into conscious states are partially mediated by inactivation of sleep networks and activation of arousal networks. Pharmacologic hastening of emergence from general anesthesia has largely focused on activating subcortical monoaminergic networks, with little attention on antagonizing the γ-aminobutyric acid type A receptor (GABAAR). As the GABAAR mediates the clinical effects of many common general anesthetics, the authors hypothesized that negative GABAAR modulators would hasten emergence, possibly via cortical networks involved in sleep. Methods: The authors investigated the capacity of the benzodiazepine rescue agent, flumazenil, which had been recently shown to promote wakefulness in hypersomnia patients, to alter emergence. Using an in vivo rodent model and an in vitro GABAAR heterologous expression system, they measured flumazenil's effects on behavioral, neurophysiologic, and electrophysiologic correlates of emergence from isoflurane anesthesia. Results: Animals administered intravenous flumazenil (0.4 mg/kg, n = 8) exhibited hastened emergence compared to saline-treated animals (n = 8) at cessation of isoflurane anesthesia. Wake-like electroencephalographic patterns occurred sooner and exhibited more high-frequency electroencephalography power after flumazenil administration (median latency ± median absolute deviation: 290 ± 34 s) compared to saline administration (473 ± 186 s; P = 0.042). Moreover, in flumazenil-treated animals, there was a decreased impact on postanesthesia sleep. In vitro experiments in human embryonic kidney-293T cells demonstrated that flumazenil inhibited isoflurane-mediated GABA current enhancement (n = 34 cells, 88.7 ± 2.42% potentiation at 3 μM). Moreover, flumazenil exhibited weak agonist activity on the GABAAR (n = 10 cells, 10.3 ± 3.96% peak GABA EC20 current at 1 μM). Conclusions: Flumazenil can modulate emergence from isoflurane anesthesia. The authors highlight the complex role GABAARs play in mediating consciousness and provide mechanistic links between emergence from anesthesia and arousal.

The hypothalamic peptide hypocretin 1 (orexin A) may be assayed in cerebrospinal fluid to diagnose narcolepsy type 1. This testing is not commercially available, and factors contributing to assay variability have not previously been comprehensively explored. In the present study, cerebrospinal fluid hypocretin concentrations were determined in duplicate in 155 patient samples, across a range of sleep disorders. Intra-assay variability of these measures was analyzed. Inter-assay correlation between samples tested at Emory and at Stanford was high (r=0.79, p<0.0001). Intra-assay correlation between samples tested in duplicate in our center was also high (r=0.88, p<0.0001); intra-assay variability, expressed as the difference between values as a percentage of the higher value, was low at 9.4% (SD=7.9%). Although both time the sample spent in the freezer (r=0.16, p=0.04) and age of the kit used for assay (t=3.64, p=0.0004) were significant predictors of intra-kit variability in univariate analyses, only age of kit was significant in multivariate linear regression (F=4.93, p=0.03). Age of radioimmunoassay kit affects intra-kit variability of measured hypocretin values, such that kits closer to expiration exhibit significantly more variability.

BACKGROUND: The prevalence of optic nerve and retinal vascular changes within the obstructive sleep apnea (OSA) population are not well-known, although it has been postulated that optic nerve ischemic changes and findings related to an elevated intracranial pressure may be more common in OSA patients. We prospectively evaluated the ocular fundus in unselected patients undergoing overnight diagnostic polysomnography (PSG). METHODS: Demographic data, medical/ocular history, and nonmydriatic fundus photographs were prospectively collected in patients undergoing PSG at our institution and reviewed for the presence of optic disc edema for which our study was appropriately powered a priori. Retinal vascular changes were also evaluated. OSA was defined using the measures of both sleep-disordered breathing and hypoxia. RESULTS: Of 250 patients evaluated in the sleep center, fundus photographs were performed on 215 patients, among whom 127 patients (59%) had an apnea/hypopnea index (AHI) ≥15 events per hour, including 36 with severe OSA. Those with AHI <15 served as the comparison group. None of the patients had optic disc edema (95% confidence interval [CI]: 0%-3%). There was no difference in rates of glaucomatous appearance or pallor of the optic disc among the groups. Retinal arteriolar changes were more common in severe OSA patients (odds ratio: 1.09 per 5 unit increase in AHI; 95% CI, 1.02-1.16; P = 0.01), even after controlling for mean arterial blood pressure. CONCLUSIONS: We did not find an increased prevalence of optic disc edema or other optic neuropathies in our OSA population. However, retinal vascular changes were more common in patients with severe OSA, independent of blood pressure.

Correspondence

The data we report in this study concern the types, location, numbers, forms, and composition of microscopic huntingtin aggregates in brain tissues from humans with different grades of Huntington's disease (HD). We have developed a fusion protein antibody against the first 256 amino acids that preferentially recognizes aggregated huntingtin and labels many more aggregates in neuronal nuclei, perikarya, and processes in human brain than have been described previously. Using this antibody and human brain tissue ranging from presymptomatic to grade 4, we have compared the numbers and locations of nuclear and neuropil aggregates with the known patterns of neuronal death in HD. We show that neuropil aggregates are much more common than nuclear aggregates and can be present in large numbers before the onset of clinical symptoms. There are also many more aggregates in cortex than in striatum, where they are actually uncommon. Although the striatum is the most affected region in HD, only 1-4% of striatal neurons in all grades of HD have nuclear aggregates. Neuropil aggregates, which we have identified by electron microscopy to occur in dendrites and dendritic spines, could play a role in the known dendritic pathology that occurs in HD. Aggregates increase in size in advanced grades, suggesting that they may persist in neurons that are more likely to survive. Ubiquitination is apparent in only a subset of aggregates, suggesting that ubiquitin-mediated proteolysis of aggregates may be late or variable.

Divergent results and misinterpretation of non-significant findings remain problematic in science – especially in retrospective, hypothesis generating, translational research.1 When such divergence occurs, it is imperative that the cause of the divergence be established. In their recent paper in Annals of Neurology, Dauvilliers et al2 challenged our earlier finding that cerebrospinal fluid (CSF) from some patients with unexplained excessive daytime sleepiness enhances the activation of GABAA receptors (GABAA-R)3. They present data from 15 subjects in which they were unable to find evidence of enhanced activation of GABAA receptors. Here we: 1) establish how flaws in Dauvilliers’ experimental design account for this difference; 2) present new data demonstrating the robustness and reproducibility of our methods and 3) summarize the clinical promise of GABAA-R antagonism in treating IH and related disorders.

Symptoms of the central disorders of hypersomnolence extend beyond excessive daytime sleepiness to include non-restorative sleep, fatigue and cognitive dysfunction. They share much in common with myalgic encephalomyelitis/chronic fatigue syndrome, recently renamed systemic exertion intolerance disease, whose additional features include post-exertional malaise and orthostatic intolerance. We sought to determine the frequency and correlates of systemic exertion intolerance disease in a hypersomnolent population. One-hundred and eighty-seven hypersomnolent patients completed questionnaires regarding sleepiness and fatigue; questionnaires and clinical records were used to assess for systemic exertion intolerance disease. Sleep studies, hypocretin and cataplexy were additionally used to assign diagnoses of hypersomnolence disorders or sleep apnea. Included diagnoses were idiopathic hypersomnia (n = 63), narcolepsy type 2 (n = 25), persistent sleepiness after obstructive sleep apnea treatment (n = 25), short habitual sleep duration (n = 41), and sleepiness with normal sleep study (n = 33). Twenty-one percent met systemic exertion intolerance disease criteria, and the frequency of systemic exertion intolerance disease was not different across sleep diagnoses (p =.37). Patients with systemic exertion intolerance disease were no different from those without this diagnosis by gender, age, Epworth Sleepiness Scale, depressive symptoms, or sleep study parameters. The whole cohort reported substantial fatigue on questionnaires, but the systemic exertion intolerance disease group exhibited more profound fatigue and was less likely to respond to traditional wake-promoting agents (88.6% versus 67.7%, p =.01). Systemic exertion intolerance disease appears to be a common co-morbidity in patients with hypersomnolence, which is not specific to hypersomnolence subtype but may portend a poorer prognosis for treatment response.

Cocaine- and amphetamine-regulated transcript (CART) peptides modulate anxiety, food intake, endocrine function, and mesolimbic dopamine related reward and reinforcement. Each of these disparate behaviors takes place during the state of wakefulness. Here, we identify a potential wake promoting role of CART by characterizing its effects upon sleep/wake architecture in rats. Dose-dependent increases in wake were documented following intracerebroventricular CART 55–102 administered at the beginning of the rat’s major sleep period. Sustained wake was observed for up to 4 hours following delivery of 2.0 μg of CART peptide. The wake promoting effect was specific to active CART 55–102 because no effect on sleep/wake was observed with the inactive form of the peptide. Increased wake was followed by robust rebound in NREM and REM sleep that extended well into the subsequent lights-off, or typical wake period, of the rat. These findings point to a potential novel role for CART in regulating wakefulness.

Background: Iron is crucial for proper functioning of all organs including the brain. Deficiencies and excess of iron are common and contribute to substantial morbidity and mortality. Whereas iron's involvement in erythropoiesis drives clinical practice, the guidelines informing interventional strategies for iron repletion in neurological disorders are poorly defined. The objective of this study was to determine if peripheral iron status is communicated to the brain. Methods: We used a bi-chamber cell culture model of the blood-brain-barrier to determine transcytosis of iron delivered by transferrin as a metric of iron transport. In the apical chamber (representative of the blood) we placed transferrin complexed with iron59 and in the basal chamber (representative of the brain) we placed human cerebrospinal fluid. Cerebrospinal fluid (CSF) samples (N = 24) were collected via lumbar puncture. The integrity of the tight junctions were monitored throughout the experiments using RITC-Dextran. Results: We demonstrate that iron transport correlates positively with plasma hemoglobin concentrations but not serum ferritin levels. Conclusions: The clinical ramifications of these findings are several-fold. They suggest that erythropoietic demands for iron take precedence over brain requirements, and that the metric traditionally considered to be the most specific test reflecting total body iron stores and relied upon to inform treatment decisions-i.e., serum ferritin-may not be the preferred peripheral indicator when attempting to promote brain iron uptake. The future direction of this line of investigation is to identify the factor(s) in the CSF that influence iron transport at the level of the BBB.

Narcolepsy type 1 (NT1) is caused by a loss of hypocretin/orexin transmission. Risk factors include pandemic 2009 H1N1 influenza A infection and immunization with Pandemrix®. Here, we dissect disease mechanisms and interactions with environmental triggers in a multi-ethnic sample of 6,073 cases and 84,856 controls. We fine-mapped GWAS signals within HLA (DQ0602, DQB1*03:01 and DPB1*04:02) and discovered seven novel associations (CD207, NAB1, IKZF4-ERBB3, CTSC, DENND1B, SIRPG, PRF1). Significant signals at TRA and DQB1*06:02 loci were found in 245 vaccination-related cases, who also shared polygenic risk. T cell receptor associations in NT1 modulated TRAJ*24, TRAJ*28 and TRBV*4-2 chain-usage. Partitioned heritability and immune cell enrichment analyses found genetic signals to be driven by dendritic and helper T cells. Lastly comorbidity analysis using data from FinnGen, suggests shared effects between NT1 and other autoimmune diseases. NT1 genetic variants shape autoimmunity and response to environmental triggers, including influenza A infection and immunization with Pandemrix®.