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In a patient with long-standing idiopathic hypereosinophilia with no apparent organ damage we measured serum eosinophil cationic protein (ECP) and eosinophil protein X (EPX) titers, activated circulating eosinophil rates (by means of monoclonal antibodies EG1 and EG2), and the release of ECP and EPX in vitro by leukocytes at different cultures stages in order to detect possible functional abnormalities associated with hypereosinophilia. Our patient had elevated serum levels of both ECP and EPX, together with a high EG2 count, which would suggest eosinophil activation. However, serum levels of ECP and EPX were not significantly high in relation to the total number of eosinophil cells, although they were more numerous than in healthy controls. Moreover, the release of intracytoplasmic basic proteins by the patient's eosinophils was poor even after in vitro stimulation. Since hypereosinophilic syndrome (HES) with organ damage can appear as long as 8-9 years after the presence of a hypereosinophilic state, the absolutely benign nature of our patient's condition still cannot be defined. Thus, there is the possibility it could be slow-onset or smoldering HES.

Hibernation has been proposed as a tool for human space travel. In recent years, a procedure to induce a metabolic state known as &ldquo, synthetic torpor&rdquo, 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.

Each of the wake-sleep states is characterized by specific changes in autonomic activity and bodily functions. The goal of such changes is not always clear. During non-rapid eye movement (NREM) sleep, the autonomic outflow and the activity of the endocrine system, the respiratory system, the cardiovascular system, and the thermoregulatory system seem to be directed at increasing energy saving. During rapid eye movement (REM) sleep, the goal of the specific autonomic and regulatory changes is unclear, since a large instability of autonomic activity and cardiorespiratory function is observed in concomitance with thermoregulatory changes, which are apparently non-functional to thermal homeostasis. Reciprocally, the activation of thermoregulatory responses under thermal challenges interferes with sleep occurrence. Such a double-edged and reciprocal interaction between sleep and thermoregulation may be favored by the fact that the central network controlling sleep overlaps in several parts with the central network controlling thermoregulation. The understanding of the central mechanism behind the interaction between sleep and thermoregulation may help to understand the functionality of thermoregulatory sleep-related changes and, ultimately, the function(s) of sleep. © 2021 American Physiological Society. Compr Physiol 11:1591-1604, 2021.

Counting cells in fluorescent microscopy is a tedious, time-consuming task that researchers have to accomplish to assess the effects of different experimental conditions on biological structures of interest. Although such objects are generally easy to identify, the process of manually annotating cells is sometimes subject to fatigue errors and suffers from arbitrariness due to the operator’s interpretation of the borderline cases. We propose a Deep Learning approach that exploits a fully-convolutional network in a binary segmentation fashion to localize the objects of interest. Counts are then retrieved as the number of detected items. Specifically, we introduce a Unet-like architecture, cell ResUnet (c-ResUnet), and compare its performance against 3 similar architectures. In addition, we evaluate through ablation studies the impact of two design choices, (i) artifacts oversampling and (ii) weight maps that penalize the errors on cells boundaries increasingly with overcrowding. In summary, the c-ResUnet outperforms the competitors with respect to both detection and counting metrics (respectively, $$F_1$$ F 1 score = 0.81 and MAE = 3.09). Also, the introduction of weight maps contribute to enhance performances, especially in presence of clumping cells, artifacts and confounding biological structures. Posterior qualitative assessment by domain experts corroborates previous results, suggesting human-level performance inasmuch even erroneous predictions seem to fall within the limits of operator interpretation. Finally, we release the pre-trained model and the annotated dataset to foster research in this and related fields.

Monitoring of bioelectric signals in peripheral sympathetic nerves of small animal models is crucial to gain understanding of how the autonomic nervous system controls specific body functions related to disease states. Advances in minimally-invasive electrodes for such recordings in chronic conditions rely on electrode materials that show low-impedance ionic/electronic interfaces and elastic mechanical properties compliant with the soft and fragile nerve strands. Here we report a highly stretchable low-impedance electrode realized by microcracked gold films as metallic conductors covered with stretchable conducting polymer composite to facilitate ion-to-electron exchange. The conducting polymer composite based on poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) obtains its adhesive, low-impedance properties by controlling thickness, plasticizer content and deposition conditions. Atomic Force Microscopy measurements under strain show that the optimized conducting polymer coating is compliant with the micro-crack mechanics of the underlying Au-layer, necessary to absorb the tensile deformation when the electrodes are stretched. We demonstrate functionality of the stretchable electrodes by performing high quality recordings of renal sympathetic nerve activity under chronic conditions in rats.

In mammals, torpor/hibernation is a state that is characterized by an active reduction in metabolic rate followed by a progressive decrease in body temperature. Torpor was successfully mimicked in non-hibernators by inhibiting the activity of neurons within the brainstem region of the Raphe Pallidus, or by activating the adenosine A1 receptors in the brain. This state, called synthetic torpor, may be exploited for many medical applications, and for space exploration, providing many benefits for biological adaptation to the space environment, among which an enhanced protection from cosmic rays. As regards the use of synthetic torpor in space, to fully evaluate the degree of physiological advantage provided by this state, it is strongly advisable to move from Earth-based experiments to 'in the field' tests, possibly on board the International Space Station.

Prader–Willi syndrome (PWS) is a neurodevelopmental disorder that is characterized by metabolic alteration and sleep abnormalities mostly related to rapid eye movement (REM) sleep disturbances. The disease is caused by genomic imprinting defects that are inherited through the paternal line. Among the genes located in the PWS region on chromosome 15 (15q11-q13), small nucleolar RNA 116 (Snord116) has been previously associated with intrusions of REM sleep into wakefulness in humans and mice. Here, we further explore sleep regulation of PWS by reporting a study with PWScrm+/p− mouse line, which carries a paternal deletion of Snord116. We focused our study on both macrostructural electrophysiological components of sleep, distributed among REMs and nonrapid eye movements. Of note, here, we study a novel electroencephalography (EEG) graphoelements of sleep for mouse studies, the well-known spindles. EEG biomarkers are often linked to the functional properties of cortical neurons and can be instrumental in translational studies. Thus, to better understand specific properties, we isolated and characterized the intrinsic activity of cortical neurons using in vitro microelectrode array. Our results confirm that the loss of Snord116 gene in mice influences specific properties of REM sleep, such as theta rhythms and, for the first time, the organization of REM episodes throughout sleep–wake cycles. Moreover, the analysis of sleep spindles present novel specific phenotype in PWS mice, indicating that a new catalog of sleep biomarkers can be informative in preclinical studies of PWS.

Under conditions of scarce food availability and cool ambient temperature, the mouse (Mus Musculus) enters into torpor, a state of transient metabolic suppression mediated in part by the autonomic nervous system. Hypothalamic orexins are involved in the coordination of behaviors and autonomic function. We tested whether orexins are necessary for the coordinated changes in physiological variables, which underlie torpor and represent its physiological signature. We performed simultaneous measurements of brain temperature, electroencephalographic, and electromyographic activity allowing objective assessment of wake-sleep behavior, and cardiovascular, respiratory, and metabolic variables in orexin knockout mice (ORX-KO) and wild-type mice (WT) during torpor bouts elicited by caloric restriction and mild cold stress. We found that torpor bouts in WT are characterized by an exquisitely coordinated physiological signature. The characteristics of torpor bouts in terms of duration and rate of change of brain temperature and electromyographic activity at torpor entrance and exit did not differ significantly between ORX-KO and WT, and neither did the cardiovascular, respiratory, and metabolic characteristics of torpor. ORX-KO and WT also had similar wake-sleep state changes associated with torpor bouts, with the exception of a significantly higher rapid-eye movement sleep time in ORX-KO at torpor entrance. Our results demonstrate that orexins are not necessary either for the normal physiological adaptations occurring during torpor in mice or for their coordination, suggesting that mechanisms different from orexin peptide signaling may be involved in the regulation and the coordination of these physiological responses.

The integrated activity of the autonomic nervous system (ANS) with the somatomotor and the neuroendocrine systems allows an animal to maintain internal homoeostasis and regulate its interaction with the external environment. Significant changes in ANS activity occur on passing from wakefulness to sleep, and this pattern of autonomic activation is profoundly different even between NREMS and REMS. While, during NREMS, the ANS works to maintain body homeostasis in accordance with the reduced metabolic needs of this state, during REMS ANS activity is greatly variable, and not visibly oriented to the maintenance of the stability of physiological variables. Sleep derangements determine an alteration of autonomic activity that can also extend to waking, favoring the development of pathological conditions. In this review, we summarize new insights into the impact of sleep derangements on ANS control of immune, respiratory and cardiovascular function, and into the role of ANS in thermoregulation and energy expenditure.

Tau protein is of primary importance for many physiological processes in neurons, where it affects the dynamics of the microtubule system. When hyperphosphorylated (PP-Tau), Tau monomers detach from microtubules and tend to aggregate firstly in oligomers, and then in neurofibrillary tangles, as it occurs in a group of neurodegenerative disorders named thauopathies. A hypothermia-related accumulation of PP-Tau, which is quickly reversed after the return to normothermia, has been shown to occur in the brain of hibernators during torpor. Since, recently, in our lab, a hypothermic torpor-like condition (synthetic torpor, ST) was pharmacologically induced in the rat, a non-hibernator, the aim of the present work was to assess whether ST can lead to a reversible PP-Tau accumulation in the rat brain. PP-Tau was immunohistochemically assessed by staining for AT8 (phosphorylated Tau) and Tau-1 (non-phosphorylated Tau) in 19 brain structures, which were chosen mostly due to their involvement in the regulation of autonomic and cognitive functions in relation to behavioral states. During ST, AT8 staining was strongly expressed throughout the brain, while Tau-1 staining was reduced compared to control conditions. During the following recovery period, AT8 staining progressively reduced close to zero after 6 h from ST. However, Tau-1 staining remained low even after 38 h from ST. Thus, overall, these results show that ST induced an accumulation of PP-Tau that was, apparently, only partially reversed to normal during the recovery period. While the accumulation of PP-Tau may only depend on the physicochemical characteristics of the enzymes regulating Tau phosphorylation, the reverse process of dephosphorylation should be actively regulated, also in non-hibernators. In conclusion, in this work a reversible and widespread PP-Tau accumulation has been induced through a procedure that leads a non-hibernator to a degree of reversible hypothermia, which is comparable to that observed in hibernators. Therefore, the physiological mechanism involved in this process can sustain an adaptive neuronal response to extreme conditions, which may however lead to neurodegeneration when particular intensities and durations are exceeded.

The Raphe Pallidus (RPa) is a region of the brainstem that was shown to modulate the sympathetic outflow to many tissues and organs involved in thermoregulation and energy expenditure. In rodents, the pharmacological activation of RPa neurons was shown to increase the activity of the brown adipose tissue, heart rate, and expired CO2, whereas their inhibition was shown to induce cutaneous vasodilation and a state of hypothermia that, when prolonged, leads to a state resembling torpor referred to as synthetic torpor. If translatable to humans, this synthetic torpor-inducing procedure would be advantageous in many clinical settings. A first step to explore such translatability, has been to verify whether the neurons within the RPa play the same role described for rodents in a larger mammal such as the pig. In the present study, we show that the physiological responses inducible by the pharmacological stimulation of RPa neurons are very similar to those observed in rodents. Injection of the GABAA agonist GABAzine in the RPa induced an increase in heart rate (from 99 to 174bpm), systolic (from 87 to 170mmHg) and diastolic (from 51 to 98mmHg) arterial pressure, and end-tidal CO2 (from 49 to 62 mmHg). All these changes were reversed by the injection in the same area of the GABAA agonist muscimol. These results support the possibility for RPa neurons to be a key target in the research for a safe and effective procedure for the induction of synthetic torpor in humans.

Torpor is a peculiar mammalian behaviour, characterized by the active reduction of metabolic rate, followed by a drop in body temperature. To enter torpor, the activation of all thermogenic organs that could potentially defend body temperature must be prevented. Most of these organs, such as the brown adipose tissue, are controlled by the key thermoregulatory region of the Raphe Pallidus (RPa). Currently, it is not known which brain areas mediate the entrance into torpor. To identify these areas, the expression of the early gene c-Fos at torpor onset was assessed in different brain regions in mice injected with a retrograde tracer (Cholera Toxin subunit b, CTb) into the RPa region. The results show a network of hypothalamic neurons that are specifically activated at torpor onset and a direct torpor-specific projection from the Dorsomedial Hypothalamus to the RPa that could putatively mediate the suppression of thermogenesis during torpor.

Background Scoring of wake–sleep states by trained investigators is a time-consuming task in many sleep experiments. We aimed to validate SCOPRISM, a new open-source algorithm for sleep scoring based on automatic graphical clustering of epoch distribution. Methods We recorded sleep and blood pressure signals of 36 orexin-deficient, 7 leptin knock-out, and 43 wild-type control mice in the PRISM laboratory. Additional groups of mice ( n = 14) and rats ( n = 6) recorded in independent labs were used to validate the algorithm across laboratories. Results The overall accuracy, specificity and sensitivity values of SCOPRISM (97%, 95%, and 94%, respectively) on PRISM lab data were similar to those calculated between human scorers (98%, 98%, and 94%, respectively). Using SCOPRISM, we replicated the main sleep and sleep-dependent cardiovascular findings of our previous studies. Finally, the cross-laboratory analyses showed that the SCOPRISM algorithm performed well on mouse and rat data. Comparison with existing methods SCOPRISM performed similarly or even better than recently reported algorithms. SCOPRISM is a very simple algorithm, extensively (cross)validated and with the possibility to evaluate its efficacy following a quick and easy visual flow chart. Conclusions We validated SCOPRISM, a new, automated and open-source algorithm for sleep scoring on a large population of mice, including different mutant strains and on subgroups of mice and rats recorded by independent labs. This algorithm should help accelerate basic research on sleep and integrative physiology in rodents.

Therapeutic hypothermia is today used in several clinical settings, among them the gut related diseases that are influenced by ischemia/reperfusion injury. This perspective paved the way to the study of hibernation physiology, in natural hibernators, highlighting an unexpected importance of the gut microbial ecosystem in hibernation and torpor. In natural hibernators, intestinal microbes adaptively reorganize their structural configuration during torpor, and maintain a mutualistic configuration regardless of long periods of fasting and cold temperatures. This allows the gut microbiome to provide the host with metabolites, which are essential to keep the host immunological and metabolic homeostasis during hibernation. The emerging role of the gut microbiota in the hibernation process suggests the importance of maintaining a mutualistic gut microbiota configuration in the application of therapeutic hypothermia as well as in the development of new strategy such as the use of synthetic torpor in humans. The possible utilization of tailored probiotics to mold the gut ecosystem during therapeutic hypothermia can also be taken into consideration as new therapeutic strategy.

Background The independent prognostic impact of diabetes mellitus ( DM ) and prediabetes mellitus (pre‐ DM ) on survival outcomes in patients with chronic heart failure has been investigated in observational registries and randomized, clinical trials, but the results have been often inconclusive or conflicting. We examined the independent prognostic impact of DM and pre‐ DM on survival outcomes in the GISSI ‐HF (Gruppo Italiano per lo Studio della Sopravvivenza nella Insufficienza Cardiaca‐Heart Failure) trial. Methods and Results We assessed the risk of all‐cause death and the composite of all‐cause death or cardiovascular hospitalization over a median follow‐up period of 3.9 years among the 6935 chronic heart failure participants of the GISSI ‐ HF trial, who were stratified by presence of DM (n=2852), pre‐ DM (n=2013), and non‐ DM (n=2070) at baseline. Compared with non‐ DM patients, those with DM had remarkably higher incidence rates of all‐cause death (34.5% versus 24.6%) and the composite end point (63.6% versus 54.7%). Conversely, both event rates were similar between non‐ DM patients and those with pre‐ DM . Cox regression analysis showed that DM , but not pre‐ DM , was associated with an increased risk of all‐cause death (adjusted hazard ratio, 1.43; 95% CI , 1.28–1.60) and of the composite end point (adjusted hazard ratio, 1.23; 95% CI , 1.13–1.32), independently of established risk factors. In the DM subgroup, higher hemoglobin A1c was also independently associated with increased risk of both study outcomes (all‐cause death: adjusted hazard ratio, 1.21; 95% CI , 1.02–1.43; and composite end point: adjusted hazard ratio, 1.14; 95% CI , 1.01–1.29, respectively). Conclusions Presence of DM was independently associated with poor long‐term survival outcomes in patients with chronic heart failure. Clinical Trial Registration URL : http://www.clinicaltrials.gov . Unique identifier: NCT 00336336.

In recent years, sleep medicine has evolved into a full-grown discipline, featuring a multidisciplinary approach to diagnosis and treatment of patients with sleep disorders. Sleep medicine cuts across the boundaries of different conventional disciplines and is therefore open to medical and non-medical professionals with different specialty backgrounds. The aim of the current paper is to introduce a qualification for those professionals whose main occupation is to practice sleep medicine in the setting of a sleep medicine centre. The drafting of guidelines dealing with requirements for such qualification was entrusted to a task force by the European Sleep Research Society. The guidelines are the result of a progressive consensus procedure in which standards were defined for education, training, and evaluation. The final step along this pathway is a theoretical and practical examination, providing proof of proficiency in the field of sleep medicine. This paper describes the object of specific competences, the scope of sleep medicine, and the qualification procedures that pertain to three professional categories: medical specialists, non-medical professionals with a university master degree (such as psychologists and biologists), and nurses and technologists. Indices of preceding practical experience and theoretical knowledge are presented in Appendices 1 and 2. These guidelines are a European standard. They may be adapted in the future according to new scientific insights. National certification programs that comply with these guidelines may be subject to homologation by the ESRS.

In the albino rat, a REM sleep (REMS) onset can be induced with a high probability and a short latency when the light is suddenly turned off (dark pulse, DP) during non-REM sleep (NREMS). The aim of this study was to investigate to what extent DP delivery could overcome the integrative thermoregulatory mechanisms that depress REMS occurrence during exposure to low ambient temperature (Ta). To this aim, the efficiency of a non-rhythmical repetitive DP (3 min each) delivery during the first 6-h Light period of a 12h:12h Light-Dark cycle in inducing REMS was studied in the rat, through the analysis of electroencephalogram (EEG), electrocardiogram (EKG), hypothalamic temperature (Thy), and motor activity at different Tas. The results showed that DP delivery triggers a transition from NREMS to REMS comparable to that which occurs spontaneously. However, the efficiency of DP delivery in inducing REMS was reduced during cold exposure to an extent comparable with that observed in spontaneous REMS occurrence. Such impairment was associated with low Delta activity and high sympathetic tone when DPs were delivered. Repetitive DP administration increased REMS amount during the delivery period and a subsequent negative REMS rebound was observed. In conclusion, DP delivery did not overcome the integrative thermoregulatory mechanisms that depress REMS in the cold. These results underline the crucial physiological meaning of the mutual exclusion of thermoregulatory activation and REMS occurrence, and support the hypothesis that the suspension of the central control of body temperature is a prerequisite for REMS occurrence

The effects of a single intraperitoneal administration of lithium, a drug used to prevent the recurrence of mania in bipolar disorders, were determined in the rat by studying changes in: (i) the wake-sleep cycle; (ii) autonomic parameters (hypothalamic and tail temperature, heart rate); (iii) the capacity to accumulate cAMP and IP(3) in the preoptic-anterior hypothalamic region (PO-AH) and in the cerebral cortex (CC) under an hypoxic stimulation at normal laboratory and at low ambient temperature (T(a)). In the immediate hours following the injection, lithium induced: (i) a significant reduction in REM sleep; (ii) a non-significant reduction in the delta power density of the EEG in NREM sleep; (iii) a significant decrease in the concentration of cAMP in PO-AH at normal laboratory T(a); (iv) a significant increase of IP(3) concentration in CC following exposure to low T(a). The earliest and most sensitive effects of lithium appear to be those concerning sleep. These changes are concomitant with biochemical effects that, in spite of a systemic administration of the substance, may be differentiated according to the second messenger involved, the brain region and the ambient condition.

a

Serotonin [5-hydroxytryptamine (5-HT)] plays an inhibitory role in rapid-eye-movement (REM) sleep although the exact mechanism(s) and site(s) of action are not known. It is commonly assumed that 5-HT exerts its influence on REM sleep via input from the dorsal raphe nucleus (DRN) directly onto cholinergic neurons involved in the generation of REM sleep. 5-HT 2 receptor sites have been found on cholinergic neurons in the laterodorsal tegmental nucleus (LDT) and pedunculopontine tegmental nucleus (PPT). We locally microinjected the 5-HT 2 agonist DOI ((±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl) and the 5-HT 2 antagonist, ketanserin, in LDT in rats to determine whether these receptor sites are involved in the regulation of behavioral states. DOI and ketanserin primarily affected REM sleep, by significantly decreasing or increasing, respectively, the number, but not the duration, of REM sleep episodes. DOI specifically decreased the occurrence of clusters of REM sleep episodes appearing at intervals less than or equal to 3 min (sequential episodes) without affecting single episodes separated by more than 3 min. An opposite effect of ketanserin on REM sleep clusters, although not statistically significant, was observed.

Body homeostasis and sleep homeostasis may both rely on the complex integrative activity carried out by the hypothalamus. Thus, the three main wake-sleep (WS) states (i.e. wakefulness, NREM sleep, and REM sleep) may be better understood if the different cardio-respiratory and metabolic parameters, which are under the integrated control of the autonomic and the endocrine systems, are studied during sleep monitoring. According to this view, many physiological events can be considered as an expression of the activity that physiological regulations should perform in order to cope with the need to fulfill body and sleep homeostasis. This review is aimed at making an assessment of data showing the existence of a physiological interplay between body homeostasis and sleep homeostasis, starting from the spontaneous changes observed in the somatic and autonomic activity during sleep, through evidence showing the deep changes occurring in the central integration of bodily functions during the different WS states, to the changes in the WS states observed when body homeostasis is challenged by the external environment and when the return to normal ambient conditions allows sleep homeo- stasis to run without apparent physiological restrictions. The data summarized in this review suggest that an approach to the dichotomy between NREM and REM sleep based on physiological regulations may offer a framework within which observations that a traditional behavioral approach may overlook can be interpreted. The study of the interplay between body and sleep homeostasis appears, therefore, to be a way to understand the function of complex organisms beyond that of the specific regulations.

Thermoregulatory responses to temperature changes are not operant during REM sleep (REMS), but fully operant in non-REM sleep and wakefulness. The specificity of the relationship between REMS and the impairment of thermoregulation was tested by eliciting the reflex release of Thyrotropin Releasing Hormone (TRH), which is integrated at hypothalamic level. By inducing the sequential secretion of Thyroid Stimulating Hormone (TSH) and Thyroid Hormone, TRH intervenes in the regulation of obligatory and non-shivering thermogenesis. Experiments were performed on male albino rats implanted with epidural electrodes for EEG recording and 2 silver-copper wire thermodes, bilaterally placed in the preoptic-hypothalamic area (POA) and connected to small thermoelectric heat pumps driven by a low-voltage high current DC power supply. In preliminary experiments, a thermistor was added in order to measure hypothalamic temperature. The activation of TRH hypophysiotropic neurons by the thermode cooling of POA was indirectly assessed, in conditions in which thermoregulation was either fully operant (wakefulness) or not operant (REMS), by a radioimmunoassay determination of plasmatic levels of TSH. Different POA cooling were performed for 120 s or 40 s at current intensities of 80 mA and 125 mA, respectively. At both current intensities, POA cooling elicited, with respect to control values (no cooling current), a significant increase in plasmatic TSH levels in wakefulness, but not during REMS. These results confirm the inactivation of POA thermal sensitivity during REMS and show, for the first time, that this inactivation concerns also the fundamental endocrine control of non-shivering thermogenesis.

The possibility of inducing a suspended animation state similar to natural torpor would be greatly beneficial in medical science, since it would avoid the adverse consequence of the powerful autonomic activation evoked by external cooling. Previous attempts to systemically inhibit metabolism were successful in mice, but practically ineffective in nonhibernators. Here we show that the selective pharmacological inhibition of key neurons in the central pathways for thermoregulatory cold defense is sufficient to induce a suspended animation state, resembling natural torpor, in a nonhibernator. In rats kept at an ambient temperature of 15°C and under continuous darkness, the prolonged inhibition (6 h) of the rostral ventromedial medulla, a key area of the central nervous pathways for thermoregulatory cold defense, by means of repeated microinjections (100 nl) of the GABAAagonist muscimol (1 mm), induced the following: (1) a massive cutaneous vasodilation; (2) drastic drops in deep brain temperature (reaching a nadir of 22.44 ± 0.74°C), heart rate (from 440 ± 13 to 207 ± 12 bpm), and electroencephalography (EEG) power; (3) a modest decrease in mean arterial pressure; and (4) a progressive shift of the EEG power spectrum toward slow frequencies. After the hypothermic bout, all animals showed a massive increase in NREM sleep Delta power, similarly to that occurring in natural torpor. No behavioral abnormalities were observed in the days following the treatment. Our results strengthen the potential role of the CNS in the induction of hibernation/torpor, since CNS-driven changes in organ physiology have been shown to be sufficient to induce and maintain a suspended animation state.

Capitoli 2-4. Fisiologia della cellula nervosa e muscolare. Fisiologia del siistema nervoso centrale. Fisiologia del sistema nervoso autonomo

This Chapter is aimed to provide a general overview of the hypothalamus and its functions. The main features of hypothalamic functional neuroanatomy will be addressed. This will lay the ground to discuss the role of hypothalamic neural pathways in different body functions, such as the regulation of wake-sleep behavior, body temperature, body fluid osmolarity, energy balance, and the cardiovascular system. In this picture, the role of hypothalamic hypocretin neurons in orchestrating behavioral and autonomic responses to environmental challenges will stand out. Knowledge of such integrative physiologic role of hypocretin neurons is a prerequisite to understand the pathophysiology of patients with narcolepsy-cataplexy, in whom hypocretin neurons are functionally lost.

In different species, rapid eye movement sleep (REMS) is characterized by a thermoregulatory impairment. It has been postulated that this impairment depends on a general insufficiency in the hypothalamic integration of autonomic function. This study aims to test this hypothesis by assessing the hypothalamic regulation of body fluid osmolality during the different wake-sleep states in the rat. Arginine-vasopressin (AVP) plasma levels were determined following intracerebroventricular (ICV) infusions of artificial cerebrospinal fluid (aCSF), either isotonic or made hypertonic by the addition of NaCl at three different concentrations (125, 250 and 500 mM). Animals were implanted with a cannula within a lateral cerebral ventricle for ICV infusions and with electrodes for the recording of the electroencephalogram. ICV infusions were made in different animals during Wake, REMS or non-REM sleep (NREMS). The results show that ICV infusion of hypertonic aCSF during REMS induced an increase in AVP plasma levels that was not different from that observed during either Wake or NREMS. These results suggest that the thermoregulatory impairment that characterizes REMS does not depend on a general impairment in the hypothalamic control of body homeostasis.

Putative sympathetic premotor neurons controlling cutaneous vasomotion are contained within the rostral ventromedial medulla (RVMM) between levels corresponding, rostrally, to the rostral portion of the nucleus of the facial nerve (RVMM(fn)) and, caudally, to the rostral pole of the inferior olive (RVMM(io)). Cutaneous vasoconstrictor premotor neurons in the RVMM(fn) play a major role in mediating thermoregulatory changes in cutaneous vasomotion that regulate heat loss. To determine the role of neurons in the RVMM(io) in regulating cutaneous blood flow, we examined the changes in the tail and paw skin temperature of free-behaving rats following chemically-evoked changes in the activity of neurons in the RVMM(io). Microinjection of the GABA A agonist, muscimol, within either the RVMM(fn) or the RVMM(io) induced a massive peripheral vasodilation; microinjection of the GABA A antagonist bicuculline methiodide within the RVMM(fn) reversed the increase in cutaneous blood flow induced by warm exposure and, unexpectedly, disinhibition of RVMM(io) neurons produced a rapid cutaneous vasodilation. We conclude that the tonically-active neurons driving cutaneous vasoconstriction, likely sympathetic premotor neurons previously described in the RVMM(fn), are also located in the RVMM(io). However, in the RVMM(io), these are accompanied by a population of neurons that receives a tonically-active GABAergic inhibition in the conscious animal and that promotes a cutaneous vasodilation upon relief of this inhibition. Whether the vasodilator neurons located in the RVMM(io) play a role in thermoregulation remains to be determined.

Recent research proposes that putative sympathetic premotor neurons controlling cutaneous vasomotion are located within the rostral ventromedial medulla (RVMM), an area delimited rostrally by the nucleus of the facial nerve (RVMM(fn)) and caudally by the rostral pole of the inferior olive (RVMM(io)). Data from several experiments have shown that cutaneous blood flow is increased by the inhibition and reduced by the activation of neurons within the RVMM(fn), but no data are available on the role of the RVMM(io) on cutaneous vasomotion. To clarify the role of RVMM(fn) and RVMM(io) neurons in controlling cutaneous vasomotion, microinjection of the GABAA agonist muscimol and GABAA antagonist bicuculline methiodide were performed within the RVMM(fn) or the RVMM(io). A massive peripheral vasodilatation was evoked by muscimol injection within both the two areas, while disinhibition of either RVMM(fn) or RVMM(io) neurons produced, respectively, a reversal of the increase in cutaneous blood flow induced by warm exposure and an unexpected rapid cutaneous vasodilatation. The results show that two intermingled sets of neurons are located within the RVMM: a vasoconstrictory and thermoregulatory set, which is tonically active, is located in both the RVMM(io) and the RVMM(fn), while a vasodilatatory and non-thermoregulatory set, which is tonically inhibited, is located in the RVMM(io).

Studies conducted on anaesthetized animals have shown that neurons within the rostral ventromedial medulla (RVMM) can modulate the cardiac sympathetic outflow. Disinhibition of RVMM neurons by microinjection of the GABA-A antagonist bicuculline methiodide induces a strong increase in heart rate (HR), but inhibition of the same neurons by microinjection of the GABA-A agonist muscimol does not produce clear effects on HR. However, general anaesthesia affects sympathetic outflow and does not permit a evaluation of heart rate variability (HRV) measure. In the present study the role of RVMM neurons in mediating tonic cardiac sympathetic outflow has been assessed in free behaving rats. HRV has been evaluated in six free behaving rats before and after microinjection of muscimol (1mM) within the RVMM. HRV was analyzed within both the time domain (R-R mean; R-R standard deviation (STD-RR); root mean square successive difference (RMSSD)) and frequency domain (High (HF, 0.6–2.4 Hz) and Low Frequency (LF, 0.06–0.6 Hz) bands). Effects of injections were separated into six temporal blocks according to the dynamics of changes in core temperature following RVMM inhibition. At the end of the experiment, animals the site of injection was histologically verified. The power of the LF band, expression of cardiovascular sympathetic drive, was clearly reduced following RVMM neurons inhibition, suggesting that RVMM neurons participate in maintaining tonic cardiac sympathetic outflow.