Treatment of restless legs syndrome and periodic limb movement disorder: an American Academy of Sleep Medicine systematic review, meta-analysis, and GRADE assessment.

INTRODUCTION: This systematic review provides supporting evidence for the accompanying clinical practice guideline on the treatment of restless legs syndrome (RLS) and periodic limb movement disorder (PLMD). METHODS: The American Academy of Sleep Medicine commissioned a task force of experts in sleep medicine. A systematic review was conducted to identify studies that compared the use of pharmacological or nonpharmacological treatment to no treatment to improve patient-important outcomes. Statistical analyses were performed to determine the clinical significance of using various interventions to treat RLS and PLMD in adults and children. The Grading of Recommendations Assessment, Development, and Evaluation (GRADE) process was used to assess the evidence for making recommendations. RESULTS: The literature search resulted in 3631 studies out of which 148 studies provided data suitable for statistical analyses. The task force provided a detailed summary of the evidence along with the certainty of evidence, the balance of benefits and harms, patient values and preferences, and resource use considerations.

Treatment of restless legs syndrome and periodic limb movement disorder: an American Academy of Sleep Medicine clinical practice guideline.

INTRODUCTION: This guideline establishes clinical practice recommendations for Treatment of Restless Legs Syndrome (RLS) and Periodic Limb Movement Disorder (PLMD) in adults and pediatric patients. METHODS: The American Academy of Sleep Medicine (AASM) commissioned a task force of experts in sleep medicine to develop recommendations and assign strengths based on a systematic review of the literature and an assessment of the evidence using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology. The task force provided a summary of the relevant literature and the certainty of evidence, the balance of benefits and harms, patient values and preferences, and resource use considerations that support the recommendations. The AASM Board of Directors approved the final recommendations. GOOD PRACTICE STATEMENT: The following good practice statement is based on expert consensus, and its implementation is necessary for the appropriate and effective management of patients with RLS.1. In all patients with clinically significant RLS, clinicians should regularly test serum iron studies including ferritin and transferrin saturation (calculated from iron and total iron binding capacity, TIBC). The test should ideally be administered in the morning avoiding all iron-containing supplements and foods at least 24 hours prior to blood draw. Analysis of iron studies greatly influences the decision to use oral or intravenous (IV) iron treatment. Consensus guidelines, which have not been empirically tested, suggest that supplementation of iron in adults with RLS should be instituted with oral or IV iron if serum ferritin ≤75 ng/mL or transferrin saturation < 20%, and only with IV iron if serum ferritin is between 75 ng/mL and 100 ng/mL. In children, supplementation of iron should be instituted for serum ferritin < 50 ng/mL with oral or IV formulations. These iron supplementation guidelines are different than for the general population.2. The first step in the management of RLS should be addressing exacerbating factors, such as alcohol, caffeine, antihistaminergic, serotonergic, anti-dopaminergic medications, and untreated obstructive sleep apnea (OSA).3. RLS is common in pregnancy; prescribers should consider the pregnancy-specific safety profile of each treatment being considered. RECOMMENDATIONS: The following recommendations are intended as a guide for clinicians in choosing a specific treatment for RLS and PLMD in adults and children. Each recommendation statement is assigned a strength ("Strong" or "Conditional"). A "Strong" recommendation (i.e., "We recommend…") is one that clinicians should follow under most circumstances. The recommendations listed below are ranked in the order of strength of recommendations and grouped by class of treatments within each PICO question. Some recommendations include remarks that provide additional context to guide clinicians with implementation of this recommendation.Adults with RLS.1. In adults with RLS, the AASM recommends the use of gabapentin enacarbil over no gabapentin enacarbil (Strong recommendation, moderate certainty of evidence).2. In adults with RLS, the AASM recommends the use of gabapentin over no gabapentin (Strong recommendation, moderate certainty of evidence).3. In adults with RLS, the AASM recommends the use of pregabalin over no pregabalin (Strong recommendation, moderate certainty of evidence).4. In adults with RLS, the AASM recommends the use of IV ferric carboxymaltose over no IV ferric carboxymaltose in patients with appropriate iron status (see good practice statement for iron parameters) (Strong recommendation, moderate certainty of evidence).5. In adults with RLS, the AASM suggests the use of IV low molecular weight (LMW) iron dextran over no IV LMW iron dextran in patients with appropriate iron status (see good practice statement for iron parameters) (Conditional recommendation, very low certainty of evidence).6. Recommendation 6: In adults with RLS, the AASM suggests the use of IV ferumoxytol over no IV ferumoxytol in patients with appropriate iron status (see good practice statement for iron parameters) (Conditional recommendation, very low certainty of evidence).7. In adults with RLS, the AASM suggests the use of ferrous sulfate over no ferrous sulfate in patients with appropriate iron status (see good practice statement for iron parameters) (Conditional recommendation, moderate certainty of evidence).8. In adults with RLS, the AASM suggests the use of dipyridamole over no dipyridamole (Conditional recommendation, low certainty of evidence).9. In adults with RLS, the AASM suggests the use of extended-release oxycodone and other opioids over no opioids (Conditional recommendation, moderate certainty of evidence).10. In adults with RLS, the AASM suggests the use of bilateral high-frequency peroneal nerve stimulation over no peroneal nerve stimulation (Conditional recommendation, low certainty of evidence).11. In adults with RLS, the AASM suggests against the standard use of levodopa (Conditional recommendation, very low certainty of evidence). Remarks: Levodopa may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). 12. In adults with RLS, the AASM suggests against the standard use of pramipexole (Conditional recommendation, moderate certainty of evidence). Remarks: Pramipexole may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). 13. In adults with RLS, the AASM suggests against the standard use of transdermal rotigotine (Conditional recommendation, low certainty of evidence). Remarks: Transdermal Rotigotine may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). 14. In adults with RLS, the AASM suggests against the standard use of ropinirole (Conditional recommendation, moderate certainty of evidence). Remarks: Ropinirole may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). 15. In adults with RLS, the AASM suggests against the use of bupropion for the treatment of RLS (Conditional recommendation, moderate certainty of evidence).16. In adults with RLS, the AASM suggests against the use of carbamazepine (Conditional recommendation, low certainty of evidence).17. In adults with RLS, the AASM suggests against the use of clonazepam (Conditional recommendation, very low certainty of evidence).18. In adults with RLS, the AASM suggests against the use of valerian (Conditional recommendation, low certainty of evidence).19. In adults with RLS, the AASM suggests against the use of valproic acid (Conditional recommendation, low certainty of evidence).20. In adults with RLS, the AASM recommends against the use of cabergoline (Strong recommendation, moderate certainty of evidence).Special adult populations with RLS.21. In adults with RLS and end-stage renal disease (ESRD), the AASM suggests the use of gabapentin over no gabapentin (conditional recommendation, very low certainty of evidence).22. In adults with RLS and ESRD, the AASM suggests the use of IV iron sucrose over no IV iron sucrose in patients with ferritin < 200 ng/mL and transferrin saturation < 20% (Conditional recommendation, moderate certainty of evidence).23. In adults with RLS and ESRD, the AASM suggests the use of vitamin C over no vitamin C (conditional recommendation, low certainty of evidence).24. In adults with RLS and ESRD, the AASM suggests against the standard use of levodopa (Conditional recommendation, low certainty of evidence). Remarks: Levodopa may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). 25. In adults with RLS and ESRD, the AASM suggests against the standard use of rotigotine (Conditional recommendation, very low certainty of evidence). Remarks: Rotigotine may be used to treat RLS in patients who place a higher value on the reduction of restless legs symptoms with short-term use and a lower value on adverse effects with long-term use (particularly augmentation). Adults with PLMD.26. In adults with PLMD, the AASM suggests against the use of triazolam (Conditional recommendation, very low certainty of evidence).27. In adults with PLMD, the AASM suggests against the use of valproic acid (Conditional recommendation, very low certainty of evidence).Children with RLS.28. In children with RLS, the AASM suggests the use of ferrous sulfate over no ferrous sulfate in patients with appropriate iron status (see good practice statement for iron parameters) (Conditional recommendation, very low certainty of evidence).

Overnutrition in persons with cystic fibrosis on modulator therapy and the relationship to obstructive sleep apnea.

Cystic fibrosis (CF) care is evolving with the ubiquitous use of modulator therapy and resultant increase in lifespan. It is important for CF clinicians to monitor the pathologic weight gain that is concomitantly being seen as obesity is a known risk factor for multiple other diseases. In this review we focus on obesity in CF, discuss screening and lifestyle considerations, outline CF-specific concerns with weight loss medications, and describe the vicious cycle of obesity and obstructive sleep apnea (OSA). We discuss screening and treatment for OSA, as it directly correlates with weight fluctuation. We offer interim recommendations for CF teams as they continue to care for this population.

Sex differences in variables associated with excessive daytime sleepiness in patients with epilepsy.

OBJECTIVE: Excessive daytime sleepiness (EDS) is common in patients with epilepsy (PWE). The Epworth Sleepiness Scale (ESS) is a self-reported measure of sleepiness in widespread use. The purpose of this study was to identify contributors to the ESS score in PWE and to identify variables associated with a high score indicative of EDS. METHODS: A cross-sectional study was performed on 115 PWE presenting to the epilepsy clinic. Self-reported questionnaires were administered and demographic and clinical information was gathered from the electronic medical record. Regression analyses were performed. RESULTS: A high ESS score was found in nearly 20% of the cohort. Obstructive sleep apnea (OSA) risk, standardized anti-seizure drug (ASD) dose, and female sex were associated with an increased likelihood of a high ESS score. Assessment of the ESS without the use of a cutpoint showed that standardized ASD dose and OSA risk were associated with the ESS in men, but standardized ASD dose was not associated with the ESS in women. Higher use of valproic acid and oxcarbazepine in men and higher use of lamotrigine in women may be contributing factors. SIGNIFICANCE: Sex is likely to be a key factor in determining contributors to EDS in PWE.

Reliability and Efficacy of the Epworth Sleepiness Scale: Is There Still a Place for It?

The Epworth sleepiness scale (ESS) is a commonly used questionnaire to evaluate patients for excessive daytime sleepiness (EDS). The ESS has been validated as a measure of EDS, but a number of studies have shown more test-retest variability in clinical settings compared to the original validation study. This observation of higher-than-expected test-retest variability has called into question the utility of the ESS as a clinical tool to assess EDS. The purpose of this review article is to summarize how studies of test-retest variability in clinical populations compare to the original validation study of Johns and to highlight where they differ. Furthermore, use of the ESS as a continuous variable (with no specified cutoff value) versus a categorical variable (normal versus high) is described. These observations are put into a clinical context by comparing the test-retest variability observed on the ESS with that of the multiple sleep latency test (MSLT). Finally, how contributors to ESS scores differ within certain subpopulations is described. The ESS remains an important tool to measure EDS in patient populations, but an awareness of its limitations needs to be considered.

Sex differences in Epworth Sleepiness Scale normalization with continuous positive airway pressure.

STUDY OBJECTIVES: Continuous positive airway pressure (CPAP) improves sleepiness in patients with obstructive sleep apnea, but some patients remain sleepy. The objective of this study was to identify determinants that are associated with improvements in self-reported sleepiness in patients with obstructive sleep apnea on CPAP therapy. METHODS: A retrospective cohort study was performed in a clinic-based population to determine which variables contributed to the improvement in the Epworth Sleepiness Scale (ESS) in patients on CPAP therapy for OSA, stratified by baseline ESS score (< 11 or ≥ 11). Variables associated with ESS scores normalizing with CPAP were also assessed. RESULTS: Patients with a baseline high ESS score showed greater improvements in the ESS with CPAP. When looking at interactions between baseline ESS classification and changes in ESS, we found that a higher apnea-hypopnea index was only associated with improvement in the ESS among patients with a high baseline ESS. Other assessed factors or covariates were not significantly different. When looking at ESS normalization, we found that female sex and lower body mass index were associated with a lower likelihood of ESS normalization. The difference in the rate of ESS normalization between females and males was higher with more days on CPAP. CONCLUSIONS: Of all the assessed factors and covariates, only the apnea-hypopnea index was associated with the change in the ESS differently in patients with a high or normal baseline ESS score. ESS normalization rates were lower in females than in males, and this disparity was amplified by more days on CPAP. CITATION: Scharf MT, Zhang P, Walker NA, et al. Sex differences in Epworth Sleepiness Scale normalization with continuous positive airway pressure. J Clin Sleep Med. 2022;18(9):2273-2279.

Circadian Disruption in Critical Illness.

Circadian rhythms play a vital role in metabolic, hormonal, and immunologic function and are often disrupted in patients in the ICU. Circadian rhythms modulate the molecular machinery that responds to injury and illness which can impact recovery. Potential factors contributing to the alteration in circadian rhythmicity in intensive care unit (ICU) patients include abnormal lighting, noise, altered feeding schedules, extensive patient care interactions and medications. These alterations in circadian rhythms in ICU patients may affect outcomes and therefore, normalization of circadian rhythmicity in critically ill patients may be an important part of ICU care.

Obstructive sleep apnea risk in patients with focal versus generalized epilepsy.

OBJECTIVE: Obstructive sleep apnea (OSA) is common in patients with epilepsy (PWE), and treatment may improve seizure control. However, OSA is often undiagnosed in PWE, and understanding of the risk profile for OSA is important. In this study, we sought to determine if OSA risk is similar in patients with generalized versus focal epilepsy. METHODS: We recruited 115 patients presenting to the Rutgers-Robert Wood Johnson Epilepsy Clinic with focal or generalized epilepsy. Obstructive sleep apnea risk was assessed using the Sleep Apnea Scale of the Sleep Disorders Questionnaire (SA-SDQ). Sleepiness was assessed using the Epworth Sleepiness Scale (ESS). Demographic and clinical information was gathered from the electronic medical record. Unadjusted and adjusted analyses were carried out to assess differences in the SA-SDQ between patients with generalized versus focal epilepsy. Further analyses were done to assess the relationship between seizure frequency, epilepsy type, and the SA-SDQ. RESULTS: Unadjusted mean SA-SDQ scores, as well as scores high enough to represent likely OSA, were similar in patients with generalized versus focal epilepsy. However, in adjusted analyses, patients with generalized epilepsy had a significantly higher mean SA-SDQ score. Older age, higher body mass index (BMI), and a history of hypertension (HTN) were also associated with higher SA-SDQ scores. Sleep Apnea Scale of the Sleep Disorders Questionnaire scores were not significantly affected by the presence of a seizure within the prior one month or six months. Average ESS scores and the percentage of scores consistent with an abnormal degree of sleepiness were statistically similar in patients with generalized versus focal epilepsy. SIGNIFICANCE: Our study suggests that patients with generalized epilepsy have a higher risk of OSA. Further studies measuring OSA directly as well as assessing potential benefits of treatment are needed.

Clinical utility of the Epworth sleepiness scale.

PURPOSE: The Epworth sleepiness scale (ESS) is a widely used tool which has been validated as a measure of sleepiness. However, the scores within individual patients referred for clinical sleep services vary considerably which may limit the clinical use of the ESS. We sought to determine the test-retest reliability of the ESS if scores were classified as either normal or sleepy. METHODS: We measured the ESS in patients presenting to our sleep center at a clinical visit and again when a sleep study was done. Demographic and clinical information was extracted from the electronic medical record. RESULTS: Average ESS scores were similar on 2 administrations, mean (SD) of 9.8 (5.4) and 10.2 (6.2). Bland-Altman analysis showed upper and lower limits of agreement of 7.5 and - 6.7, respectively. No demographic or clinical variables were identified which contributed to the intra-individual variability. Of the patients who presented with an initial ESS < 11, 80% had a second ESS < 11. Of the patients who presented with an initial ESS ≥ 11, 89% had a second ESS ≥ 11. Cohen's kappa for the two administrations of the ESS was 0.67 (95% CI of 0.51-0.83). Using previously published reports, we calculated Cohen's kappa for polysomnographic determination of the apnea-hypopnea index (AHI) with values ranging from 0.26 to 0.69. CONCLUSIONS: Individual ESS scores varied considerably within individual patients, but with classification into either normal or sleepy, the test-retest reliability was substantial and in line with other clinical measures including polysomnographic determination of the AHI.

Mask Refills as a Measure of PAP Adherence.

STUDY OBJECTIVES: Positive airway pressure (PAP) adherence data are a routine aspect of clinical care for obstructive sleep apnea (OSA), but not uniformly available. We hypothesized that mask refills are a measure of PAP adherence. METHODS: We measured PAP use over the first 90 days of treatment in 123 patients with OSA. The number and timing of mask refills was assessed over 18 months. Demographic and medical information was obtained from the electronic medical record. RESULTS: Average PAP use in the first week of more than 4 h/d was a predictor of adherence over the first 90 days (P < .0001). PAP use over 90 days was greater in datacard-derived data dependent on patients presenting a datacard to the clinic compared to modem-derived data (P = .0006). A mask refill within the first 30 days of treatment was associated with a 1.3 h/d lower PAP usage in the first 90 days (P = .0044). Conversely, the number of mask refills between 30 days and 18 months was associated with higher PAP adherence during the first 90 days, with an additional 0.61 h/d of use for each additional refill (P = .0015). CONCLUSIONS: In a retrospective cohort of veterans with OSA, first week PAP use was a strong predictor of 90-day use. Use of autonomously transmitted modem data avoided potential selection bias in adherence estimates. Mask refills in the first month were associated with less 90-day PAP use, whereas more mask refills after 30 days were associated with greater PAP use.

Sleep and Anesthesia Interactions: A Pharmacological Appraisal.

Anesthetics have been used in clinical practice for over a hundred years, yet their mechanisms of action remain poorly understood. One tempting hypothesis to explain their hypnotic properties posits that anesthetics exert a component of their effects by "hijacking" the endogenous arousal circuitry of the brain. Modulation of activity within sleep- and wake-related neuroanatomic systems could thus explain some of the varied effects produced by anesthetics. There has been a recent explosion of research into the neuroanatomic substrates affected by various anesthetics. In this review, we will highlight the relevant sleep architecture and systems and focus on studies over the past few years that implicate these sleep-related structures as targets of anesthetics. These studies highlight a promising area of investigation regarding the mechanisms of action of anesthetics and provide an important model for future study.

Changes in components of energy regulation in mouse cortex with increases in wakefulness.

STUDY OBJECTIVES: Increases in ATP production machinery have been described in brain after 3 h of sleep deprivation. Whether this is sustained with longer durations of extended wakefulness is unknown. We hypothesized that energy depletion could be a mechanism leading to difficulty maintaining wakefulness and assessed changes in components of the electron transport chain. DESIGN: Protein levels of key subunits of complexes IV and V of the electron transport chain (COXI, COXIV, ATP5B) and uncoupling protein 2 (UCP2) in isolated mitochondria by Westerns in mouse cerebral cortex after 3 and 12 h of sleep deprivation were compared to that in control mice. Activity of complex IV enzyme and relevant transcription factors-Nrf1, Nrf2 (Gabp), and phosphorylation of AMP-dependent kinase (AMPK)-were also assessed. PARTICIPANTS: 8-10 week old C57BL/6J male mice (n = 91). INTERVENTIONS: 3, 6, and 12 h of sleep deprivation. MEASUREMENTS AND RESULTS: After both 3 and 12 h of sleep deprivation, complex IV proteins and enzyme activity were significantly increased. The complex V catalytic subunit was significantly increased after 12 h of sleep deprivation only. Increased levels of UCP2 protein after 12 h of sleep deprivation suggests that there might be alterations in the ATP/AMP ratio as wakefulness is extended. That phosphorylation of AMPK is increased after 6 h of sleep deprivation supports this assertion. The increase in Nrf1 and Nrf2 (Gabp) mRNA after 6 h of sleep deprivation provides a mechanism by which there is up-regulation of key proteins. CONCLUSIONS: There are complex dynamic changes in brain energy regulation with extended wakefulness.

The energy hypothesis of sleep revisited.

One of the proposed functions of sleep is to replenish energy stores in the brain that have been depleted during wakefulness. Benington and Heller formulated a version of the energy hypothesis of sleep in terms of the metabolites adenosine and glycogen. They postulated that during wakefulness, adenosine increases and astrocytic glycogen decreases reflecting the increased energetic demand of wakefulness. We review recent studies on adenosine and glycogen stimulated by this hypothesis. We also discuss other evidence that wakefulness is an energetic challenge to the brain including the unfolded protein response, the electron transport chain, NPAS2, AMP-activated protein kinase, the astrocyte-neuron lactate shuttle, production of reactive oxygen species and uncoupling proteins. We believe the available evidence supports the notion that wakefulness is an energetic challenge to the brain, and that sleep restores energy balance in the brain, although the mechanisms by which this is accomplished are considerably more complex than envisaged by Benington and Heller.

AMP-activated protein kinase phosphorylation in brain is dependent on method of killing and tissue preparation.

AMP-activated protein kinase (AMPK) is activated when the catalytic alpha subunit is phosphorylated on Thr172 and therefore, phosphorylation of the alpha subunit is used as a measure of activation. However, measurement of alpha subunit of AMPK (alpha-AMPK) phosphorylation in vivo can be technically challenging. To determine the most accurate method for measuring alpha-AMPK phosphorylation in the mouse brain, we compared different methods of killing and tissue preparation. We found that freeze/thawing samples after homogenization on ice dramatically increased alpha-AMPK phosphorylation in mice killed by cervical dislocation. Killing of mice by focused microwave irradiation, which rapidly heats the brain and causes enzymatic inactivation, prevented the freeze/thaw-induced increase in alpha-AMPK phosphorylation and similar levels of phosphorylation were observed compared with mice killed with cervical dislocation without freeze/thawing of samples. Sonication of samples in hot 1% sodium dodecyl sulfate blocked the freeze/thaw-induced increase in alpha-AMPK phosphorylation, but phosphorylation was higher in mice killed by cervical dislocation compared with mice killed by focused microwave irradiation. These results demonstrate that alpha-AMPK phosphorylation is dependent on method of killing and tissue preparation and that alpha-AMPK phosphorylation can increase in a manner that does not reflect biological alterations.

Gut peptides and other regulators in obesity.

Obesity has reached epidemic levels in industrialized countries and is increasing worldwide. This trend has serious public health consequences, since obesity increases the risk of diabetes, hypertension, heart disease, sleep apnea, cancer, arthritis, cholelithiasis, fatty liver disease, and other complications. Obesity is the result of an imbalance between energy intake and expenditure; hence, an understanding of how gastrointestinal function is integrated with the hormonal regulation of energy balance is pertinent to the pathophysiology of obesity. Nutrients, peptides, and neural afferents from the gut influence the size and frequency of meals and satiety. The long-term regulation of energy stores is mediated primarily through the actions of adiposity hormones, such as leptin and insulin, in the hypothalamus and other neuronal circuits in the brain. Efforts are underway to determine how these peripheral and central pathways may be targeted for treatment of obesity and related diseases.

Impact of interrupted leptin pathways on ventilatory control.

Leptin deficiency in ob/ob mice produces marked depression of the hypercapnic ventilatory response, particularly during sleep. We now extend our previous findings to determine whether 1) leptin deficiency affects the hypoxic ventilatory response and 2) blockade of the downstream excitatory actions of leptin on melanocortin 4 receptors or inhibitory actions on neuropeptide Y (NPY) pathways has an impact on hypercapnic and hypoxic sensitivity. We have found that leptin-deficient ob/ob mice have the same hypoxic ventilatory response as weight-matched wild-type obese mice. There were no differences in the hypoxic sensitivity between agouti yellow mice and weight-matched controls, or NPY-deficient mice and wild-type littermates. Agouti yellow mice, with blocked melanocortin pathways, exhibited a significant depression of the hypercapnic sensitivity compared with weight-matched wild-type controls during non-rapid eye movement sleep (5.8 +/- 0.7 vs. 8.9 +/- 0.7 ml x min(-1) x %CO(2)(-1), P < 0.01), but not during wakefulness. NPY-deficient transgenic mice exhibited a small increase in the hypercapnic ventilatory response compared with wild-type littermates, but this was only present during wakefulness. We conclude that interruption of leptin pathways does not affect hypoxic sensitivity during sleep and wakefulness but that melanocortin 4 blockade is associated with depressed hypercapnic sensitivity in non-rapid eye movement sleep.

Protein synthesis is required for the enhancement of long-term potentiation and long-term memory by spaced training.

Spaced training is generally more effective than massed training for learning and memory, but the molecular mechanisms underlying this trial spacing effect remain poorly characterized. One potential molecular basis for the trial spacing effect is the differential modulation, by distinct temporal patterns of neuronal activity, of protein synthesis-dependent processes that contribute to the expression of specific forms of synaptic plasticity in the mammalian brain. Long-term potentiation (LTP) is a type of synaptic modification that may be important for certain forms of memory storage in the mammalian brain. To explore the role of protein synthesis in the trial spacing effect, we assessed the protein synthesis dependence of hippocampal LTP induced by 100-Hz tetraburst stimulation delivered to mouse hippocampal slices in either a temporally massed (20-s interburst interval) or spaced (5-min interburst interval) fashion. To extend our studies to the behavioral level, we trained mice in fear conditioning using either a massed or spaced training protocol and examined the sensitivity of long-term memory to protein synthesis inhibition. Larger LTP was induced by spaced stimulation in hippocampal slices. This improvement of synaptic potentiation following temporally spaced synaptic stimulation in slices was attenuated by bath application of an inhibitor of protein synthesis. Further, the maintenance of LTP induced by spaced synaptic stimulation was more sensitive to disruption by anisomycin than the maintenance of LTP elicited following massed stimulation. Temporally spaced behavioral training improved long-term memory for contextual but not for cued fear conditioning, and this enhancement of memory for contextual fear was also protein synthesis dependent. Our data reveal that altering the temporal spacing of synaptic stimulation and behavioral training improved hippocampal LTP and enhanced contextual long-term memory. From a broad perspective, these results suggest that the recruitment of protein synthesis-dependent processes important for long-term memory and for long-lasting forms of LTP can be modulated by the temporal profiles of behavioral training and synaptic stimulation.