Acute sleep deprivation induces synaptic remodeling at the soleus muscle neuromuscular junction in rats.

Sleep is important for cognitive and physical performance. Sleep deprivation not only affects neural functions but also results in muscular fatigue. A good night's sleep reverses these functional derangements caused by sleep deprivation. The role of sleep in brain function has been extensively studied. However, its role in neuromuscular junction (NMJ) or skeletal muscle morphology is sparsely addressed although skeletal muscle atonia and suspended thermoregulation during rapid eye movement sleep possibly provide a conducive environment for the muscle to rest and repair; somewhat similar to slow-wave sleep for synaptic downscaling. In the present study, we have investigated the effect of 24 h sleep deprivation on the NMJ morphology and neurochemistry using electron microscopy and immunohistochemistry in the rat soleus muscle. Acute sleep deprivation altered synaptic ultra-structure viz. mitochondria, synaptic vesicle, synaptic proteins, basal lamina, and junctional folds needed for neuromuscular transmission. Further acute sleep deprivation showed the depletion of the neurotransmitter acetylcholine and the overactivity of its degrading enzyme acetylcholine esterase at the NMJ. The impact of sleep deprivation on synaptic homeostasis in the brain has been extensively reported recently. The present evidence from our studies shows new information on the role of sleep on the NMJ homeostasis and its functioning.

Electrophysiological Evidence of Local Sleep During Yoga Nidra Practice.

Background and Objectives: Yoga nidra is a technique sages use to self-induce sleep. Classically, sleep is characterized by three cardinal electrophysiological features, namely, electroencephalography (EEG), electromyography (EMG), and electrooculography (EOG). As the literature on electrophysiological characterization of Yoga nidra is lacking, it is not known whether it is a sleep or awake state. The objective of the study was to electrophysiologically characterize yoga nidra practice. Materials and Methods: Thirty subjects underwent five initial supervised yoga nidra sessions and then continued practice on their own. The subjects completed their sleep diaries for 2 weeks before and during the intervention. The electrophysiological characterization was done after 2 weeks of yoga nidra practice using 19 EEG channels polysomnography for pre-yoga nidra, yoga nidra practice and post-yoga nidra. Polysomnographic data were scored for sleep-wake stages as per standard criteria. Power spectral density (PSD) was calculated from various frequency bands in different time bins. EEG data were grouped by areas, namely, central, frontal, prefrontal, parietal, temporal, and occipital in time bins. Sleep diary parameters were also compared for pre-post-yoga nidra training. Results: After 2 weeks of yoga nidra practice, awake was scored throughout the session (n = 26). PSD results (mean difference in dB between different time bins; P value) showed significant changes. When compared to pre-yoga nidra, there was an increase in delta power in the central area (1.953; P = 0.033) and a decrease in the prefrontal area (2.713; P = 0.041) during yoga nidra. Sleep diary showed improvement in sleep duration (P = 0.0001), efficiency (P = 0.0005), quality (P = 0.0005), and total wake duration (P = 0.00005) after 2 weeks of practice. Interpretations and Conclusions: Yoga nidra practice in novices is electrophysiologically an awake state with signs of slow waves locally, often referred to as local sleep. Clinical Trial: Clinical Trial Registry of India, http://www.ctri.nic.in/Clinicaltrials/pmaindet2.php? trialid = 6253, 2013/05/003682.

Yoga nidra practice shows improvement in sleep in patients with chronic insomnia: A randomized controlled trial.

Background Yoga nidra is practised by sages for sleep. The practice is simple to use and has been clearly laid out, but its role in the treatment of chronic insomnia has not been well studied. Methods In this randomized parallel-design study conducted during 2012-16, we enrolled 41 patients with chronic insomnia to receive conventional intervention of cognitive behavioural therapy for insomnia (n=20) or yoga nidra (n=21). Outcome measures were both subjective using a sleep diary and objective using polysomnography (PSG). Salivary cortisol levels were also measured. PSG was done before the intervention in all patients and repeated only in those who volunteered for the same. Results Both interventions showed an improvement in subjective total sleep time (TST), sleep efficiency, wake after sleep onset, reduction in total wake duration and enhancement in subjective sleep quality. Objectively, both the interventions improved TST and total wake duration and increased N1% of TST. Yoga nidra showed marked improvement in N2% and N3% in TST. Salivary cortisol reduced statistically significantly after yoga nidra (p=0.041). Conclusion Improvement of N3 sleep, total wake duration and subjective sleep quality occurred following yoga nidra practice. Yoga nidra practice can be used for treatment of chronic insomnia after supervised practice sessions.

Mediodorsal thalamus lesion increases paradoxical sleep in rats.

INTRODUCTION: The mediodorsal thalamic nucleus has extensive connections with prefrontal cortex, which is considered as seat of cognition. It also receives connections from sleep-wakefulness regulating areas in the brainstem and hypothalamus. Decreased volume and degeneration of mediodorsal thalamic nuclei have been reported in schizophrenia and fatal familial insomnia, respectively. In both conditions, the sleep is abnormal. OBJECTIVE: To study the role of mediodorsal thalamic nuclei in sleep wakefulness in rats. MATERIAL AND METHODS: Neurotoxic lesion of mediodorsal thalamic nuclei with ibotenic acid was performed in adult male Wistar rats and sleep wakefulness was recorded. The recordings were taken on 2nd, 7th and 14th days after lesion and compared with the baseline recordings. In order to study the diurnal changes, lesion recordings were of 24h duration. We also performed L-glutamate excitation of mediodorsal thalamic nuclei in another set of animals. After L-glutamate microinjection, sleep wakefulness was recorded for 4h. The recordings were obtained in a digital acquisition system (BSL 4.0 MP 36, Biopac Systems, Inc., USA). RESULTS: In the present investigation, ibotenic acid lesion of mediodorsal thalamic nuclei reduced the wakefulness and increased paradoxical sleep, which contradicts the reports from earlier lesion studies in cats. Glutamate excitation of mediodorsal thalamic nuclei produced prolonged wakefulness. DISCUSSION: The results suggest that the mediodorsal thalamic nuclei augments arousal in the ascending reticular wake promoting pathways in contrast to the earlier reports that mediodorsal thalamic nucleus is involved in generation of slow wave sleep. The present study adds another evidence for the role of thalamus in sleep-wake regulation.

Muscle temperature is least altered during total sleep deprivation in rats.

It has often been said that the brain is mostly benefitted from sleep. To understand the importance of sleep, extensive studies on other organs are too required. One such unexplored area is the understanding of muscle physiology during the sleep-wake cycle. Changes in muscle tone with different sleep phases are evident from the rapid eye movement sleep muscle atonia. There is variation in brain and body temperature during sleep stages, the brain temperature being higher during rapid eye movement sleep than slow-wave sleep. However, the change in muscle temperature with different sleep stages is not known. In this study, we have implanted pre-calibrated K-type thermocouples in the hypothalamus and the dorsal nuchal muscle, and a peritoneal transmitter to monitor the hypothalamic, muscle, and body temperature respectively in rats during 24 h sleep-wake cycle. The changes in muscle, body, and hypothalamic temperature during total sleep deprivation were also monitored. During normal sleep-wake stages, the temperature in the decreasing order was that of the hypothalamus, body, and muscle. Total sleep deprivation by gentle handling caused a significant increase in hypothalamic and body temperature, while there was least change in the muscle temperature. The circadian rhythm of the hypothalamic and body temperature in the sleep-deprived rats was disrupted, while the same was preserved in the muscle temperature. The results of our study show that muscle atonia during rapid eye movement sleep is a physiologically regulated thermally quiescent muscle state offering a conducive environment for muscle rest and repair.

Acute sleep deprivation elevates brain and body temperature in rats.

Available sleep deprivation studies lack data on simultaneous changes in hypothalamic, cortical and body temperature during sleep deprivation and recovery. Ten adult male Wistar rats chronically implanted with electroencephalogram, electro-oculogram and electromyogram electrodes for recording sleep were used in this study. Hypothalamic and cortical temperatures were measured by pre-implanted thermocouples. A radio transmitter (TA10TAF-40, DSI USA) was implanted intraperitoneally to measure body temperature. All the temperatures were measured simultaneously at 15-s intervals during baseline conditions, sleep deprivation and recovery sleep. Sleep deprivation was carried out for 24 hr by the gentle handling method; however, sleep and temperature were only recorded during the first 12 hr of deprivation. During sleep deprivation the body, hypothalamic and cortical temperatures increased significantly as compared to baseline. During recovery sleep, body and cortical temperature recovered earlier than the hypothalamic temperature. Hypothalamic temperature remained higher than the baseline values throughout 12 hr of recovery sleep. In the recovery sleep, cortical temperature decreased immediately and reached near baseline by 4 hr. We observed a quicker return of cortical temperature towards control temperature during recovery sleep compared with hypothalamic and body temperature. The results of the present study show that acute sleep deprivation results in a rise in both cortical and hypothalamic temperature, along with body temperature. A rise in cortical temperature may be a contributing factor for cognitive dysfunction resulting from sleep deprivation.

Guidelines of the Indian Society for Sleep Research (ISSR) for Practice of Sleep Medicine during COVID-19.

BACKGROUND: Sleep services are assigned a non-essential status during COVID-19. The American Academy of Sleep Medicine strongly urges sleep clinicians to continue postponing non-urgent care until a later date, if such a recommendation is made by state officials due to local conditions. At the same time, one cannot ignore the fact that sleep is important for people's health and wellbeing. Therefore, to protect the health of the population, it is essential to find ways and means to continue the practice of sleep medicine even during the COVID-19 pandemic. METHOD: Social environment and work ethics in sleep clinics and sleep laboratories in Asia, Africa, and Latin America are different from those in the US. Under these circumstances, the Indian Society for Sleep Research (ISSR) created a task force to develop guidelines for the practice of sleep medicine, not only for the Indian environment but also for other countries that are affected by the COVID-19 pandemic. The task force examined documents regarding practice of sleep medicine and associated specialities  during COVID-19 by various professional organizations and governmental authorities. The recommendations were examined for their applicability. Wherever gaps were identified, consensus was reached keeping in view the available evidences. OUTCOME AND RECOMMENDATIONS: The emphasis of the guidelines is on avoiding doctor to patient contact during the pandemic. Teleconsultation and other modes of audio-visuals can be used as modes for medical practice during the COVID-19 pandemic. However, in addition to the patient, the presence of a family member, or a reliable informant is recommended. Patients of most sleep disorders can be provided tele-aftercare service. ISSR guidelines also give a list of medications allowed to be prescribed during the first and the follow-up teleconsultation. Hospitals and clinics are slowly opening in India and many other countries. As sleep services resume operations, there is a need to find innovative ways to reduce contact with COVID-19 patients, follow personal protection guidelines, as well as social distancing. This article does discuss strategies for the safe conduct of Level 1 sleep studies. Home sleep testing, which had greater acceptance during the last few years, should be given more attention during the COVID-19 period. Once the decision to reopen the sleep laboratory and resume operations is made, the safety of the patients and office staff should become the major priority. The ISSR recommendation is to postpone and reschedule in-laboratory positive pressure therapy, but it mentions the considerations to be followed in emergency situations. At the same time, high clinical risk patients may be diagnosed on the basis of clinical findings, and without performing polysomnography or home sleep testing. However, at some point, there is a need to reinitiate the in-lab testing. In addition, daily assessment of the COVID-19 situation in the community, along with a review of the situation with local public health and the state health department is advised.

Cortex senses environmental temperature earlier than the hypothalamus in awake rats.

Simultaneous and direct recording of temperature from the body, hypothalamus, and cortex in animals exposed to acute thermal challenges lack evidence. This study was conducted to assess the usual concept, that brain temperature is rather stable when animals are exposed to different ambient temperatures. In this study, we report the characteristic changes in the body, hypothalamic, and cortical temperature, when the rats were acutely exposed to cold (6 °C) and hot (36 °C) ambient temperature. The results of our study show that the body temperature is robustly regulated while hypothalamic and cortical temperatures vary on challenges to ambient cold (6 °C) and warm (36 °C) exposure in awake rats. The onset of response was observed quickest in the cortex, indicating that the cortical thermal sensing may relay intracranial thermal input to the hypothalamus for the regulation of body temperature within narrow limits. The present findings contradict earlier evidence, which stated that the brain does not participate in thermal sensing.

Participation of preoptic area TRPV4 ion channel in regulation of body temperature.

Transient receptor potential vanilloid 4 (TRPV4) ion channel is a non-selective cation channel and its role in cutaneous thermosensation is emerging. It is expressed in many areas of the brain including the preoptic area (POA)/anterior hypothalamus which is the key neural site for thermoregulation. The present study was conducted to find out the role of TRPV4 ion channel in the POA in thermoregulation. Rats preimplanted with guide cannulae with indwelling styli 2.0mm above the POA received TRPV4 agonist/antagonist/isotonic saline injections bilaterally in the POA using an injector cannula in three separate groups of six rats each. Body temperature (Tb) was recorded telemetrically by preimplanted radio transmitter in the peritoneal cavity. The injection of TRPV4 agonist (GSK1016790A) in the POA decreased Tb while its antagonist (RN1734) increased Tb. Immunohistochemical localization showed presence of TRPV4 ion channel in the POA. The results of the present study suggest that TRPV4 ion channels in the POA may play an important role in thermoregulation.

Effects of NMDA and non-NMDA ionotropic glutamate receptors in the medial preoptic area on body temperature in awake rats.

Glutamate when microinjected at the medial preoptic area (mPOA) influences brain temperature (Tbr) and body temperature (Tb) in rats. Glutamate and its various receptors are present at the mPOA. The aim of this study was to identify the contribution of each of the ionotropic glutamatergic receptors at the mPOA on changes in Tbr and Tb in freely moving rats. Adult male Wistar rats (n=40) were implanted with bilateral guide cannula with indwelling styli above the mPOA. A telemetric transmitter was implanted at the peritoneum to record Tb and locomotor activity (LMA). A precalibrated thermocouple wire implanted near the hypothalamus was used to assess Tbr. Specific agonist for each ionotropic glutamate receptor was microinjected into the mPOA and its effects on temperature and LMA were measured in the rats. The rats were also microinjected with the respective ionotropic receptor antagonists, 15min prior to the microinjection of each agonist. Amongst amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), N-methyl-d-aspartate (NMDA) and kainic acid, AMPA increased Tb and LMA when injected at the mPOA. Specific antagonists for AMPA receptors was able to attenuate this increase (p<0.005). Pharmacological blockade of NMDA was able to lower Tbr only. Microinjection of kainic acid and its antagonist had no effect on the variables. The finding of the study suggests that activation of the AMPA receptors at the mPOA, leads to the rise in body temperature.

Intragastric administration of glutamate increases REM sleep in rats.

Monosodium glutamate, a umami taste substance is commonly used flavor enhancer. The effect of intragastric administration of 1.5 ml of 0.12M monosodium glutamate on sleep-wake was studied in 10 adult male Wistar rats. Sleep-wake parameters were recorded through chronically implanted electroencephalogram, electrooculogram and electromyogram electrodes using a digital recording system (BIOPAC system Inc. BSL PRO 36, USA). The sleep-wake was recorded for 6h after the intragastric administration of either glutamate or saline. Sleep-wake stages were analyzed as wake, slow wave sleep and REM sleep. Compared to saline, intragastric administration of glutamate significantly increased REM sleep duration and episode frequency. REM sleep duration was increased in all the three 2h bins, 10:00-12:00 h (p=0.037), 12:00-14:00 h (p=0.037) and 14:00-16:00 h (p=0.007). The slow wave sleep and total sleep time were not affected. It is concluded that intragastric glutamate administration increases REM sleep.

Intracerebroventricular injection of orexin-2 receptor antagonist promotes REM sleep.

Orexins are important regulators of sleep-wakefulness (S-W). Rats were intracerebroventricularly (ICV) administered selective orexin-2 receptor antagonist, TCS-OX2-29. There was decreased wakefulness due to a decrease in the average wake episode duration and an increased REM sleep due to an increase in the number of REM episodes, indicating an inhibitory role of the central orexin-2 receptors on REM sleep generation.

Basal forebrain thermoregulatory mechanism modulates auto-regulated sleep.

Regulation of body temperature and sleep are two physiological mechanisms that are vital for our survival. Interestingly neural structures implicated in both these functions are common. These areas include the medial preoptic area (POA), the lateral POA, the ventrolateral POA, the median preoptic nucleus, and the medial septum, which form part of the basal forebrain (BF). When given a choice, rats prefer to stay at an ambient temperature of 27°C, though the maximum sleep was observed when they were placed at 30°C. Ambient temperature around 27°C should be considered as the thermoneutral temperature for rats in all sleep studies. At this temperature the diurnal oscillations of sleep and body temperature are properly expressed. The warm sensitive neurons of the POA mediate the increase in sleep at 30°C. Promotion of sleep during the rise in ambient temperature from 27 to 30°C, serve a thermoregulatory function. Autonomous thermoregulatory changes in core body temperature and skin temperature could act as an input signal to modulate neuronal activity in sleep-promoting brain areas. The studies presented here show that the neurons of the BF play a key role in regulating sleep. BF thermoregulatory system is a part of the global homeostatic sleep regulatory mechanism, which is auto-regulated.

Glutamate microinjection in the medial septum of rats decreases paradoxical sleep and increases slow wave sleep.

The role of the medial septum in suppressing paradoxical sleep and promoting slow wave sleep was suggested on the basis of neurotoxic lesion studies. However, these conclusions need to be substantiated with further experiments, including chemical stimulation studies. In this report, the medial septum was stimulated in adult male rats by microinjection of L-glutamate. Sleep-wakefulness was electrophysiologically recorded, through chronically implanted electrodes, for 2 h before the injection and 4 h after the injection. There was a decrease in paradoxical sleep during the first hour and an increase in slow wave sleep during the second hour after the injection. The present findings not only supported the lesion studies but also showed that the major role of the medial septum is to suppress paradoxical sleep.

Hypothalamic temperature: a key regulator in homeostatic restoration of sleep during chronic cold exposure in rats.

Exposure to cold ambient temperature (Ta) affects sleep-wake (S-W) state. The vigilance states on the other hand influence thermal status of the animals. Simultaneous recording of body temperature (Tb) with S-W is crucial to understand the homeostatic relationship between the two. In the present study we recorded both Tb and hypothalamic temperature (Thy) along with S-W, during acute and chronic exposure to mild cold (Ta). Electrooculogram (EOG), electroencephalogram (EEG) and electromyogram (EMG) electrodes were chronically implanted in rats to assess S-W. A thermocouple, near the preoptic area, and radio transmitter in the peritoneum, were implanted, to record Thy and Tb respectively. After three days of baseline recordings of S-W, Thy and Tb at Ta of 26 dergrees C, the rats were exposed to mild cold Ta (18 degrees C) for 28 days. All the parameters were recorded during cold exposure and also for five days after the termination of cold exposure. On the first day of cold exposure there was a decrease in slow wave sleep and paradoxical sleep, but they were restored by the 21st day of continued exposure. The Thy remained decreased throughout the cold exposure. Though the Tb showed a slight decrease on the first day of cold exposure, there was no appreciable change during the subsequent days. The Thy came back to near pre exposure level on termination of cod exposure. The decrease in Thy during mild cold exposure would have triggered cold defense mechanisms. Increase in wakefulness during acute cold exposure and non-shivering thermogenesis during chronic cold exposure are probably responsible for the maintenance of Tb. Decrease in Thy is probably the key trigger for initiating thermoregulatory measures to maintain Tb and homeostatic restoration of sleep.

Warm sensitive neurons of the preoptic area regulate ambient temperature related changes in sleep in the rat.

Warm sensitive neurons (WSN) play a major role not only in body temperature regulation, but also in sleep regulation. The present study was undertaken to investigate the role of WSN of the preoptic area (POA) in mediating the ambient temperature (T(amb)) related changes in sleep. The effect of T(amb) changes on sleep and body temperature was studied in rats before and after destruction of WSN of the POA by local intracerebral injection of capsaicin. Though the rats preferred 27 degrees C T(amb), they slept maximum at 30 degrees C. After destruction of WSN of the POA, slow wave sleep (SWS) peak was brought down to 27 degrees C, which was the preferred T(amb) of the rats. This indicates that WSN of the POA mediate the increase in SWS, at temperatures higher than preferred T(amb). On the other hand, in WSN destroyed rats, rapid eye movement (REM) sleep was maximum at 33 degrees C. It suggests that the REM sleep generation is under inhibitory control of the WSN of the POA. The study supports several earlier reports that the neurons of the POA play a key role in coordinating sleep and body temperature regulation.

Glutamate microinjection at the medial preoptic area enhances slow wave sleep in rats.

A large body of evidence has established the role of the medial preoptic area (mPOA) in regulation of slow wave sleep (SWS). Although the mPOA neurons contain excitatory neurotransmitter glutamate, its role in sleep-wakefulness is not known. In the present study microinjection of monosodium glutamate (40, 80 and 120 ng) into the mPOA augmented SWS. Earlier reports have shown enhancement of paradoxical sleep by glutamate in other brain areas.

Lysine and arginine reduce the effects of cerebral ischemic insults and inhibit glutamate-induced neuronal activity in rats.

Intravenous administration of arginine was shown to be protective against cerebral ischemic insults via nitric oxide production and possibly via additional mechanisms. The present study aimed at evaluating the neuroprotective effects of oral administration of lysine (a basic amino acid), arginine, and their combination on ischemic insults (cerebral edema and infarction) and hemispheric brain swelling induced by transient middle cerebral artery occlusion/reperfusion in rats. Magnetic resonance imaging and 2,3,5-triphenyltetrazolium chloride staining were performed 2 days after ischemia induction. In control animals, the major edematous areas were observed in the cerebral cortex and striatum. The volumes associated with cortical edema were significantly reduced by lysine (2.0 g/kg), arginine (0.6 g/kg), or their combined administration (0.6 g/kg each). Protective effects of these amino acids on infarction were comparable to the inhibitory effects on edema formation. Interestingly, these amino acids, even at low dose (0.6 g/kg), were effective to reduce hemispheric brain swelling. Additionally, the effects of in vivo microiontophoretic (juxtaneuronal) applications of these amino acids on glutamate-evoked neuronal activity in the ventromedial hypothalamus were investigated in awake rats. Glutamate-induced neuronal activity was robustly inhibited by microiontophoretic applications of lysine or arginine onto neuronal membranes. Taken together, our results demonstrate the neuroprotective effects of oral ingestion of lysine and arginine against ischemic insults (cerebral edema and infarction), especially in the cerebral cortex, and suggest that suppression of glutamate-induced neuronal activity might be the primary mechanism associated with these neuroprotective effects.

Glycine enhances glutamate-induced excitation in ventromedial hypothalamic neurons in awake rats.

The finding that glycine potentiates N-methyl-D-aspartate (NMDA) receptor-mediated responses, has tremendously changed our understanding of glutamatergic synaptic transmission in the brain. Although the phenomenon has been confirmed in number of preparations, it is yet to be demonstrated in awake animals. Further, the controversy that glycine binding sites of NMDA receptor are saturated in vivo or not, can be best verified in awake animals. Here, we have demonstrated that glycine enhanced glutamate-induced neuronal discharges in the ventromedial nucleus of the hypothalamus of awake behaving rats using microiontophoresis technique, suggesting that the glycine binding sites of NMDA receptor are not saturated under physiological conditions.