Ultrasonic vocalisations during rapid eye movement sleep in the rat

Rats are known to use a 22-kHz ultrasonic vocalisation as a distress call to warn of danger to other members of their group. We monitored 22-kHz ultrasonic vocalisation emissions in rats (lean and obese) as part of a sleep deprivation study to detect the eventual presence of stress during the procedure. Unexpectedly, we detected ultrasonic vocalisation emission during rapid eye movement (REM) sleep, but not during non-REM (NREM) sleep, in all the rats. The event occurs during the expiratory phase and can take place singularly or as a train. No difference was detected in the number or duration of these events in lean versus obese rats, during the light versus the dark period, and after sleep deprivation. As far as we know, this is the first report showing that rats can vocalise during REM sleep

Synthetic torpor triggers a regulated mechanism in the rat brain, favoring the reversibility of Tau protein hyperphosphorylation

Introduction: Hyperphosphorylated Tau protein (PPTau) is the hallmark of tauopathic neurodegeneration. During "synthetic torpor" (ST), a transient hypothermic state which can be induced in rats by the local pharmacological inhibition of the Raphe Pallidus, a reversible brain Tau hyperphosphorylation occurs. The aim of the present study was to elucidate the - as yet unknown - molecular mechanisms underlying this process, at both a cellular and systemic level.Methods: Different phosphorylated forms of Tau and the main cellular factors involved in Tau phospho-regulation were assessed by western blot in the parietal cortex and hippocampus of rats induced in ST, at either the hypothermic nadir or after the recovery of euthermia. Pro- and anti-apoptotic markers, as well as different systemic factors which are involved in natural torpor, were also assessed. Finally, the degree of microglia activation was determined through morphometry.Results: Overall, the results show that ST triggers a regulated biochemical process which can dam PPTau formation and favor its reversibility starting, unexpectedly for a non-hibernator, from the hypothermic nadir. In particular, at the nadir, the glycogen synthase kinase-beta was largely inhibited in both regions, the melatonin plasma levels were significantly increased and the antiapoptotic factor Akt was significantly activated in the hippocampus early after, while a transient neuroinflammation was observed during the recovery period.Discussion: Together, the present data suggest that ST can trigger a previously undescribed latent and regulated physiological process, that is able to cope with brain PPTau formation

Molecular changes induced by a centrally-induced state of synthetic torpor in multiple organs:a new strategy for radioprotection.

Torpor is a successful survival strategy displayed by several mammalian species to cope with harsh environmental conditions. A complex interplay of ambient, genetic and circadian stimuli acts centrally to induce a severe suppression of metabolic rate, usually followed by an apparently undefended reduction of body temperature. Some animals, such as marmots, are able to maintain this physiological state for months (hibernation), during which torpor bouts are periodically interrupted by short interbouts of normothermia (arousals). Interestingly, torpor adaptations have been shown to be associated with a large resistance towards stressors, such as radiation: indeed, if irradiated during torpor, hibernators can tolerate higher doses of radiation, showing an increased survival rate. New insights for radiotherapy and long-term space exploration could arise from the induction of torpor in non-hibernators, like humans. The present research project is centered on synthetic torpor (ST), a hypometabolic/hypothermic condition induced in a non-hibernator, the rat, through the pharmacological inhibition of the Raphe Pallidus, a key brainstem area controlling thermogenic effectors. By exploiting this procedure, this thesis aimed at: i) providing a multiorgan description of the functional cellular adaptations to ST; ii) exploring the possibility, and the underpinning molecular mechanisms, of enhanced radioresistance induced by ST. To achieve these aims, transcriptional and histological analysis have been performed in multiple organs of synthetic torpid rats and normothermic rats, either exposed or not exposed to 3 Gy total body of X-rays. The results showed that: i) similarly to natural torpor, ST induction leads to the activation of survival and stress resistance responses, which allow the organs to successfully adapt to the new homeostasis; ii) ST provides tissue protection against radiation damage, probably mainly through the cellular adaptations constitutively induced by ST, even though the triggering of specific responses when the animal is irradiated during hypothermia might play a role

Sleep deprivation soon after recovery from synthetic torpor enhances tau protein dephosphorylation in the rat brain

Neuronal Tau protein hyperphosphorylation (PPtau) is a hallmark of tauopathic neurodegeneration. However, a reversible brain PPtau occurs in mammals during either natural or "synthetic" torpor (ST), a transient deep hypothermic state that can be pharmacologically induced in rats. Since in both conditions a high sleep pressure builds up during the regaining of euthermia, the aim of this work was to assess the possible role of post-ST sleep in PPtau dephosphorylation. Male rats were studied at the hypothermic nadir of ST, and 3-6 h after the recovery of euthermia, after either normal sleep (NS) or total sleep deprivation (SD). The effects of SD were studied by assessing: (i) deep brain temperature (Tb); (ii) immunofluorescent staining for AT8 (phosphorylated Tau) and Tau-1 (non-phosphorylated Tau), assessed in 19 brain structures; (iii) different phosphorylated forms of Tau and the main cellular factors involved in Tau phospho-regulation, including pro- and anti-apoptotic markers, assessed through western blot in the parietal cortex and hippocampus; (iv) systemic factors which are involved in natural torpor; (v) microglia activation state, by considering morphometric variations. Unexpectedly, the reversibility of PPtau was more efficient in SD than in NS animals, and was concomitant with a higher Tb, higher melatonin plasma levels, and a higher frequency of the microglia resting phenotype. Since the reversibility of ST-induced PPtau was previously shown to be driven by a latent physiological molecular mechanism triggered by deep hypothermia, short-term SD soon after the regaining of euthermia seems to boost the possible neuroprotective effects of this mechanism

Reversible Tau Phosphorylation Induced by Synthetic Torpor in the Spinal Cord of the Rat

Tau is a key protein in neurons, where it affects the dynamicsof the microtubulesystem. The hyperphosphorylation of Tau (PP-Tau) commonlyleads to the formationof neurofibrillary tangles, as it occurs in tauopathies, a group of neurodegenerativediseases, including Alzheimer’s. Hypothermia-related accumulation of PP-Tau has beendescribed in hibernators and during synthetic torpor (ST),a torpor-like condition that hasbeen induced in rats, a non-hibernating species. Remarkably, in ST PP-Tau is reversibleand Tau de-phosphorylates within a few hours following the torpor bout, apparentlynot evolving into pathology. These observations have been limited to the brain, but inanimal models of tauopathies, PP-Tau accumulation also appears to occur in the spinalcord (SpCo). The aim of the present work was to assess whetherST leads to PP-Tauaccumulation in the SpCo and whether this process is reversible. Immunofluorescence(IF) for AT8 (to assess PP-Tau) and Tau-1 (non-phosphorylated Tau) was carried out onSpCo coronal sections. AT8-IF was clearly expressed in the dorsal horns (DH) duringST, while in the ventral horns (VH) no staining was observed.The AT8-IF completelydisappeared after 6h from the return to euthermia. Tau-1-IFdisappeared in both DH andVH during ST, returning to normal levels during recovery. Toshed light on the cellularprocess underlying the PP-Tau pattern observed, the inhibited form of the glycogen-synthase kinase 3β(the main kinase acting on Tau) was assessed using IF: VH (i.e., inmotor neurons) were highly stained mainly during ST, while in DH there was no staining.Since tauopathies are also related to neuroinflammation, microglia activation was alsoassessed through morphometric analyses, but no ST-inducedmicroglia activation wasfound in the SpCo. Taken together, the present results show that, in the DH of SpCo, STinduces a reversible accumulation of PP-Tau. Since during ST there is no motor activity,the lack of AT8-IF in VH may result from an activity-related process at a cellular level. Thus,ST demonstrates a newly-described physiological mechanism that is able to resolve theaccumulation of PP-Tau and apparently avoid the neurodegenerative outcome.Keywords: hypothermia, hibernation, microglia, tauopathies, GSK3β, motor neurons, adaptive respons

Western-blot results from Synthetic-torpor experiments conducted in 2019-2020

This dataset contains the original results for the western-blot (WB) determinations from experiments carried out in the "Physiological regulations in the wake-sleep cycle" lab, at the Department of Biomedical and Neuromotor Sciences - University of Bologna, Italy. The WB procedure, the bands acquisitions and their intensity quantifications were conducted at the “Centre for Applied Biomedical Research – CRBA” - University of Bologna, St. Orsola Hospital, Italy

Fluorescent Neuronal Cells v2: multi-task, multi-format annotations for deep learning in microscopy

Abstract Fluorescent Neuronal Cells v2 is a collection of fluorescence microscopy images and the corresponding ground-truth annotations, designed to foster innovative research in the domains of Life Sciences and Deep Learning. This dataset encompasses three image collections wherein rodent neuronal cell nuclei and cytoplasm are stained with diverse markers to highlight their anatomical or functional characteristics. Specifically, we release 1874 high-resolution images alongside 750 corresponding ground-truth annotations for several learning tasks, including semantic segmentation, object detection and counting. The contribution is two-fold. First, thanks to the variety of annotations and their accessible formats, we anticipate our work will facilitate methodological advancements in computer vision approaches for segmentation, detection, feature extraction, unsupervised and self-supervised learning, transfer learning, and related areas. Second, by enabling extensive exploration and benchmarking, we hope Fluorescent Neuronal Cells v2 will catalyze breakthroughs in fluorescence microscopy analysis and promote cutting-edge discoveries in life sciences

Hibernation and Radioprotection: Gene Expression in the Liver and Testicle of Rats Irradiated under Synthetic Torpor

Hibernation has been proposed as a tool for human space travel. In recent years, a procedure to induce a metabolic state known as "synthetic torpor" in non-hibernating mammals was successfully developed. Synthetic torpor may not only be an efficient method to spare resources and reduce psychological problems in long-term exploratory-class missions, but may also represent a countermeasure against cosmic rays. Here we show the preliminary results from an experiment in rats exposed to ionizing radiation in normothermic conditions or synthetic torpor. Animals were irradiated with 3 Gy X-rays and organs were collected 4 h after exposure. Histological analysis of liver and testicle showed a reduced toxicity in animals irradiated in torpor compared to controls irradiated at normal temperature and metabolic activity. The expression of ataxia telangiectasia mutated (ATM) in the liver was significantly downregulated in the group of animal in synthetic torpor. In the testicle, more genes involved in the DNA damage signaling were downregulated during synthetic torpor. These data show for the first time that synthetic torpor is a radioprotector in non-hibernators, similarly to natural torpor in hibernating animals. Synthetic torpor can be an effective strategy to protect humans during long term space exploration of the solar system