The circadian molecular clock in the suprachiasmatic nucleus is necessary but not sufficient for fear entrainment.

We show that nocturnal aversive stimuli presented to mice while they are eating and drinking outside of their safe nest can entrain circadian behaviors, leading to a shift toward daytime activity. We also show that the canonical molecular circadian clock is necessary for fear entrainment and that an intact molecular clockwork in the suprachiasmatic nucleus, the site of the central circadian pacemaker, is necessary but not sufficient to sustain fear entrainment of circadian rhythms. Our results demonstrate that entrainment of a circadian clock by cyclic fearful stimuli can lead to severely mistimed circadian behavior that persists even after the aversive stimulus is removed. Together, our findings support the interpretation that circadian and sleep symptoms associated with fear and anxiety disorders are, in part, the output of a fear-entrained clock, and provide a mechanistic insight into this clock.

[EXCESS WEIGHT AND SOCIODEMOGRAPHIC RISK FACTORS FOR SARS-COV-2 INFECTION IN ISRAEL].

BACKGROUND: Studies to date have demonstrated an increased prevalence of obesity and low socioeconomic status (SES( among people with confirmed cases of COVID-19, and low SES has been linked to obesity. OBJECTIVES: Our goal was to better understand the important relationship between body weight and sociodemographic variables affecting the COVID-19 burden. METHODS: We conducted a cross-sectional study of subjects presenting to Israel's largest emergency department and their odds for positive SARS-CoV-2 virus PCR testing during the first wave of the pandemic. RESULTS: We found that as BMI rises, as compared to normal weight, it is associated with increasing odds for testing positive, independently of age, gender, SES and population density (BMI 25-29.9 kg/m2: OR = 1.42, 95% CI 1.07 - 1.90; BMI 30-34.9 kg/m2: OR = 1.50, 95% CI 1.06 - 2.11; BMI ≥ 35 kg/m2: OR = 1.61, 95% CI 1.02 - 2.46). Furthermore, male gender, low SES and high population density are also associated with excess risk for positive test results independently of body weight. CONCLUSIONS: Understanding these risk factors for infection and how they might interplay can help the medical community develop approaches to protect at-risk groups from infection and severe disease secondary to seasonal and pandemic viral infections.

The circadian molecular clock in the suprachiasmatic nucleus is necessary but not sufficient for fear entrainment in the mouse.

Nocturnal aversive stimuli presented to mice during eating and drinking outside of their safe nest can entrain circadian behaviors, leading to a shift toward daytime activity. We show that the canonical molecular circadian clock is necessary for fear entrainment and that an intact molecular clockwork in the suprachiasmatic nucleus (SCN), the site of the central circadian pacemaker, is necessary but not sufficient to sustain fear entrainment of circadian rhythms. Our results demonstrate that entrainment of a circadian clock by cyclic fearful stimuli can lead to severely mistimed circadian behavior that persists even after the aversive stimulus is removed. Together, our results support the interpretation that circadian and sleep symptoms associated with fear and anxiety disorders may represent the output of a fear-entrained clock. One-Sentence Summary: Cyclic fearful stimuli can entrain circadian rhythms in mice, and the molecular clock within the central circadian pacemaker is necessary but not sufficient for fear-entrainment.

Habitat aridity as a determinant of the trade-off between water conservation and evaporative heat loss in bats.

The maintenance of water balance in arid environments might represent a formidable challenge for Chiroptera, since they have high surface-to-volume ratios. In deserts, bats conserve water, for example, using daily torpor, but they also might experience episodic heat bouts, when they may need to increase total evaporative water loss (TEWL) to thermoregulate. We hypothesized that in bats, habitat aridity and its variability determine a trade-off between water conservation and thermoregulation via evaporative means. To test this hypothesis, we collated data from the literature of 22 species of bats on TEWL, body temperature and resting metabolic rate, in torpor and euthermy. We also collected data on ambient temperature (Ta) and precipitation of the locations where bats were captured, calculated an aridity index, and built an index of variability of the environment. After correcting for phylogeny, we found that, as aridity and variability of the environment increased, bats had lower values of TEWL, but the rate at which TEWL increases with Ta was higher, supporting our hypothesis. These results suggest that at high Ta there is a trade-off between water conservation and evaporative heat loss in bats. The evolution of physiological mechanisms that allow water conservation and tolerance to conditions of high Ta without access to free water might thus be crucial to explain the distribution of desert bats.

Lipid composition of the stratum corneum in different regions of the body of Kuhl's pipistrelle from the Negev Desert, Israel.

The most superficial epidermal layer in endotherms is the stratum corneum (SC), which is composed of dead corneocytes embedded in a lipid matrix with free fatty acids, cholesterol, ceramides, and cerebrosides; the lipid composition of the SC determines its permeability to water vapor. Lipids that are more polar, have longer hydrocarbon chains, and are less bulky are often packed in more ordered phase states to slow cutaneous evaporative water loss (CEWL); these lipids also resist transitions to more disordered phases at high ambient temperatures (Ta). In bats, wing and tail membranes (wing patagia and tail uropatagium, respectively) allow powered flight, but increase surface area, and hence CEWL, with implications for survival in arid environments. We captured Pipistrellus kuhlii from an arid habitat and measured the lipid composition of the SC of the plagiopatagium in the wing, the uropatagium, and the non-membranous region (NMR) of the body using thin layer chromatography and reversed phase high performance liquid chromatography coupled with atmospheric pressure photoionization mass spectrometry. The patagia contained more cholesterol and shorter-chained ceramides, and fewer cerebrosides than the NMR, indicating that the lipid phase transition temperature in the patagia is lower than in the NMR. Thus, at moderate Ta the lipids in the SC in all body regions will remain in an ordered phase state, allowing water conservation; but as Ta increases, the lipids in the SC of the patagia will more easily transition into a disordered phase, resulting in increased CEWL from the patagia facilitating efficient heat dissipation in hot environments.

Managing Obesity in Lockdown: Survey of Health Behaviors and Telemedicine.

Since the outbreak of COVID-19, billions of people have gone into lockdown, facing pandemic related challenges that engender weight gain, especially in the obese. We report the results of an online survey, conducted during Israel's first quarantine, of 279 adults treated in hospital-based obesity clinics with counseling, medications, surgery, endoscopic procedures, or any combination of these for weight loss. In this study, we assessed the association between changes in dietary and lifestyle habits and body weight, and the benefits of receiving weight management care remotely through telemedicine during lockdown. Compared to patients not receiving obesity care via telemedicine, patients receiving this care were more likely to lose weight (OR, 2.79; p = 0.042) and also to increase participation in exercise (OR, 2.4; p = 0.022). While 40% of respondents reported consuming more sweet or salty processed snacks and 33% reported less vegetables and fruits, 65% reported more homemade foods. At the same time, 40% of respondents reported a reduction in exercise and 52% reported a decline in mood. Alterations in these eating patterns, as well as in exercise habits and mood, were significantly associated with weight changes. This study highlights that lockdown affects health behaviors associated with weight change, and advocates for the use of telemedicine to provide ongoing obesity care during future quarantines in order to promote weight loss and prevent weight gain.

Spatial and Temporal Dynamics of Mal Secco Disease Spread in Lemon Orchards in Israel.

Mal Secco is a severe disease of citrus in which the fungus Plenodomus tracheiphilus (formerly Phoma tracheiphila) penetrates the vascular system of the host. In this study, we characterized the spatial dynamics of the disease in seven lemon orchards. A representative block of trees from each orchard was evaluated monthly during 3 consecutive years. In addition, scouts assessed disease severity in 75 orchards from three different geographical regions and tested for association between disease severity and measures of orchard management, environmental factors, cultural practices, and cultivar type. We assessed disease incidence and characteristics of spatial patterns using Ripley's K function and fitted logistic regression models for different neighboring tree structures followed by model selection methods to provide insight into the spatial and temporal dynamics of disease progress. We found different rates of disease spread in different orchards, which are most likely the result of differences in orchard management practices or less likely the result of differences in climatic conditions. There was an indication that agricultural tools contribute to spread of the disease within rows of trees. The results confirm that the lemon cultivar Interdonato is less susceptible compared with other citrus cultivars, and they suggest that the density of urban terrain surrounding each orchard is positively correlated with the severity of the disease. In contrast to our expectations, no correlation was found between the density of lemon orchards surrounding an orchard and the severity of the disease within it, which corroborates previous findings regarding the limited distribution of the disease.

Circadian regulation of sleep in a pre-clinical model of Dravet syndrome: dynamics of sleep stage and siesta re-entrainment.

STUDY OBJECTIVES: Sleep disturbances are common co-morbidities of epileptic disorders. Dravet syndrome (DS) is an intractable epilepsy accompanied by disturbed sleep. While there is evidence that daily sleep timing is disrupted in DS, the difficulty of chronically recording polysomnographic sleep from patients has left our understanding of the effect of DS on circadian sleep regulation incomplete. We aim to characterize circadian sleep regulation in a mouse model of DS. METHODS: Here we exploit long-term electrocorticographic recordings of sleep in a mouse model of DS in which one copy of the Scn1a gene is deleted. This model both genocopies and phenocopies the disease in humans. We test the hypothesis that the deletion of Scn1a in DS mice is associated with impaired circadian regulation of sleep. RESULTS: We find that DS mice show impairments in circadian sleep regulation, including a fragmented rhythm of non-rapid eye movement (NREM) sleep and an elongated circadian period of sleep. Next, we characterize re-entrainment of sleep stages and siesta following jet lag in the mouse. Strikingly, we find that re-entrainment of sleep following jet lag is normal in DS mice, in contrast to previous demonstrations of slowed re-entrainment of wheel-running activity. Finally, we report that DS mice are more likely to have an absent or altered daily "siesta". CONCLUSIONS: Our findings support the hypothesis that the circadian regulation of sleep is altered in DS and highlight the value of long-term chronic polysomnographic recording in studying the role of the circadian clock on sleep/wake cycles in pre-clinical models of disease.

Kisspeptin Neurons in the Arcuate Nucleus of the Hypothalamus Orchestrate Circadian Rhythms and Metabolism.

Successful reproduction in female mammals is precisely timed and must be able to withstand the metabolic demand of pregnancy and lactation. We show that kisspeptin-expressing neurons in the arcuate hypothalamus (Kiss1ARH) of female mice control the daily timing of food intake, along with the circadian regulation of locomotor activity, sleep, and core body temperature. Toxin-induced silencing of Kiss1ARH neurons shifts wakefulness and food consumption to the light phase and induces weight gain. Toxin-silenced mice are less physically active and have attenuated temperature rhythms. Because the rhythm of the master clock in the suprachiasmatic nucleus (SCN) appears to be intact, we hypothesize that Kiss1ARH neurons signal to neurons downstream of the master clock to modulate the output of the SCN. We conclude that, in addition to their well-established role in regulating fertility, Kiss1ARH neurons are a critical component of the hypothalamic circadian oscillator network that times overt rhythms of physiology and behavior.

Sleepmore in Seattle: Later school start times are associated with more sleep and better performance in high school students.

Most teenagers are chronically sleep deprived. One strategy proposed to lengthen adolescent sleep is to delay secondary school start times. This would allow students to wake up later without shifting their bedtime, which is biologically determined by the circadian clock, resulting in a net increase in sleep. So far, there is no objective quantitative data showing that a single intervention such as delaying the school start time significantly increases daily sleep. The Seattle School District delayed the secondary school start time by nearly an hour. We carried out a pre-/post-research study and show that there was an increase in the daily median sleep duration of 34 min, associated with a 4.5% increase in the median grades of the students and an improvement in attendance.

The Dorsal Medial Habenula Minimally Impacts Circadian Regulation of Locomotor Activity and Sleep.

In nocturnal rodents, voluntary wheel-running activity (WRA) represents a self-reinforcing behavior. We have previously demonstrated that WRA is markedly reduced in mice with a region-specific deletion of the transcription factor Pou4f1 (Brn3a), which leads to an ablation of the dorsal medial habenula (dMHb). The decrease in WRA in these dMHb-lesioned (dMHbCKO) mice suggests that the dMHb constitutes a critical center for conveying reinforcement by exercise. However, WRA also represents a prominent output of the circadian system, and the possibility remains that the dMHb is a source of input to the master circadian pacemaker located in the suprachiasmatic nucleus (SCN) of the hypothalamus. To test this hypothesis, we assessed the integrity of the circadian system in dMHbCKO mice. Here we show that the developmental lesion of the dMHb reduces WRA under both a light-dark cycle and constant darkness, increases the circadian period of WRA, but has no effect on the circadian amplitude or period of home cage activity or the daily amplitude of sleep stages, suggesting that the lengthening of period is a result of the decreased WRA in the mutant mice. Polysomnographic sleep recordings show that dMHbCKO mice have an overall unaltered daily amplitude of sleep stages but have fragmented sleep and an overall increase in total rapid eye movement (REM) sleep. Photoresponsiveness is intact in dMHbCKO mice, but compared with control animals, they reentrain faster to a 6-h abrupt phase delay protocol. Circadian changes in WRA of dMHbCKO mice do not appear to emerge within the central pacemaker, as circadian expression of the clock genes Per1 and Per2 within the SCN is normal. We do find some evidence for fragmented sleep and an overall increase in total REM sleep, supporting a model in which the dMHb is part of the neural circuitry encoding motivation and involved in the manifestation of some of the symptoms of depression.

The cutaneous lipid composition of bat wing and tail membranes: a case of convergent evolution with birds.

The water vapour permeability barrier of mammals and birds resides in the stratum corneum (SC), the outermost layer of the epidermis. The molar ratio and molecular arrangement of lipid classes in the SC determine the integrity of this barrier. Increased chain length and polarity of ceramides, the most abundant lipid class in mammalian SC, contribute to tighter packing and thus to reduced cutaneous evaporative water loss (CEWL). However, tighter lipid packing also causes low SC hydration, making it brittle, whereas high hydration softens the skin at the cost of increasing CEWL. Cerebrosides are not present in the mammalian SC; their pathological accumulation occurs in Gaucher's disease, which leads to a dramatic increase in CEWL. However, cerebrosides occur normally in the SC of birds. We tested the hypothesis that cerebrosides are also present in the SC of bats, because they are probably necessary to confer pliability to the skin, a quality needed for flight. We examined the SC lipid composition of four sympatric bat species and found that, as in birds, their SC has substantial cerebroside contents, not associated with a pathological state, indicating convergent evolution between bats and birds.

Differential effects of photoperiod length on depression- and anxiety-like behavior in female and male diurnal spiny mice.

The relationships between biological rhythms and affective disorders are known but their underlying biology not clear. There is difficulty in studying circadian rhythms in humans and appropriate animal models are hard to identify or develop. Some studies show that diurnal rodents can be advantageous model animals for the study of interactions between biological rhythms and affective disorders but previous studies did not include females whereas in humans there are sex differences in affective disorders. The present study tested the effects of short photoperiods in both males and females of the diurnal golden spiny mouse (Acomys russatus). Adult, female and male spiny mice were housed in either neutral photoperiod (12:12 light/dark; NP), or short photoperiod (5:19 light/dark; SP) conditions. After 3weeks acclimatization, animals were tested for spontaneous activity in an open field, elevated plus maze (EPM), sweet solution preference (SSP) and the forced swim test (FST). Both sexes responded to the SP, but while SP males showed increased anxiety-like behavior in the EPM and depression-like behavior in the FST, females showed increased activity, reduced anxiety-like behavior in the EPM, depression-like response in the SSP and no effect in the FST. Differences between sexes were previously demonstrated in behavioral tests that followed a variety of manipulations, and were usually explained in the context of sex hormones. Yet, the current results cannot be compared with previous data from diurnal rodents and further testing of females from other diurnal rodents are needed to explore whether these differences are a general phenomenon or possibly unique to golden spiny mice.

Circadian Forced Desynchrony of the Master Clock Leads to Phenotypic Manifestation of Depression in Rats.

In mammals, a master circadian clock within the suprachiasmatic nucleus (SCN) of the hypothalamus maintains the phase coherence among a wide array of behavioral and physiological circadian rhythms. Affective disorders are typically associated with disruption of this fine-tuned "internal synchronization," but whether this internal misalignment is part of the physiopathology of mood disorders is not clear. To date, depressive-like behavior in animal models has been induced by methods that fail to specifically target the SCN regulation of internal synchronization as the mode to generate depression. In the rat, exposure to a 22-h light-dark cycle (LD22) leads to the uncoupling of two distinct populations of neuronal oscillators within the SCN. This genetically, neurally, and pharmacologically intact animal model represents a unique opportunity to assess the effect of a systematic challenge to the central circadian pacemaker on phenotypic manifestations of mood disorders. We show that LD22 circadian forced desynchrony in rats induces depressive-like phenotypes including anhedonia, sexual dysfunction, and increased immobility in the forced swim test (FST), as well as changes in the levels and turnover rates of monoamines within the prefrontal cortex. Desynchronized rats show increased FST immobility during the dark (active) phase but decreased immobility during the light (rest) phase, suggesting a decrease in the amplitude of the normal daily oscillation in this behavioral manifestation of depression. Our results support the notion that the prolonged internal misalignment of circadian rhythms induced by environmental challenge to the central circadian pacemaker may constitute part of the etiology of depression.

Metabolic rate, evaporative water loss and thermoregulatory state in four species of bats in the Negev desert.

Life in deserts is challenging for bats because of their relatively high energy and water requirements; nevertheless bats thrive in desert environments. We postulated that bats from desert environments have lower metabolic rates (MR) and total evaporative water loss (TEWL) than their mesic counterparts. To test this idea, we measured MR and TEWL of four species of bats, which inhabit the Negev desert in Israel, one species mainly restricted to hyper-arid deserts (Otonycteris hemprichii), two species from semi-desert areas (Eptesicus bottae and Plecotus christii), and one widespread species (Pipistrellus kuhlii). We also measured separately, in the same individuals, the two components of TEWL, respiratory water loss (RWL) and cutaneous evaporative water loss (CEWL), using a mask. In all the species, MR and TEWL were significantly reduced during torpor, the latter being a consequence of reductions in both RWL and CEWL. Then, we evaluated whether MR and TEWL in bats differ according to their geographic distributions, and whether those rates change with Ta and the use of torpor. We did not find significant differences in MR among species, but we found that TEWL was lowest in the species restricted to desert habitats, intermediate in the semi-desert dwelling species, and highest in the widespread species, perhaps a consequence of adaptation to life in deserts. Our results were supported by a subsequent analysis of data collected from the literature on rates of TEWL for 35 bat species from desert and mesic habitats.

Ambient temperature and nutritional stress influence fatty acid composition of structural and fuel lipids in Japanese quail (Coturnix japonica) tissues.

In birds, fatty acids (FA) serve as the primary metabolic fuel during exercise and fasting, and their composition affects metabolic rate and thus energy requirements. To ascertain the relationship between FAs and metabolic rate, a distinction should be made between structural and fuel lipids. Indeed, increased unsaturation of structural lipid FAs brings about increased cell metabolism, and changes in the FA composition of fuel lipids affects metabolic rate through selective mobilization and increasing availability of specific FAs. We examined the effects of acclimation to a low ambient temperature (Ta: 12.7±3.0°C) and nutritional status (fed or unfed) on the FA composition of four tissues in Japanese quail, Coturnix japonica. Differentiating between neutral (triglycerides) and polar (phospholipids) lipids, we tested the hypothesis that both acclimation to low Ta and nutritional status modify FA composition of triglycerides and phospholipids. We found that both factors affect FA composition of triglycerides, but not the composition of phospholipids. We also found changes in liver triacylglyceride FA composition in the low-Ta acclimated quail, namely, the two FAs that differed, oleic acid (18:1) and arachidonic acid (20:4), were associated with thermoregulation. In addition, the FAs that changed with nutritional status were all reported to be involved in regulation of glucose metabolism, and thus we suggest that they also play a role in the response to fasting.

Waking to drink: rates of evaporative water loss determine arousal frequency in hibernating bats.

Bats hibernate to cope with low ambient temperatures (T(a)) and low food availability during winter. However, hibernation is frequently interrupted by arousals, when bats increase body temperature (T(b)) and metabolic rate (MR) to normothermic levels. Arousals account for more than 85% of a bat's winter energy expenditure. This has been associated with variation in T(b), T(a) or both, leading to a single testable prediction, i.e. that torpor bout length (TBL) is negatively correlated with T(a) and T(b). T(a) and T(b) were both found to be correlated with TBL, but correlations alone cannot establish a causal link between arousal and T(b) or T(a). Because hydration state has also been implicated in arousals from hibernation, we hypothesized that water loss during hibernation creates the need in bats to arouse to drink. We measured TBL of bats (Pipistrellus kuhlii) at the same T(a) but under different conditions of humidity, and found an inverse relationship between TBL and total evaporative water loss, independent of metabolic rate, which directly supports the hypothesis that hydration state is a cue to arousal in bats.

The relationship between cutaneous water loss and thermoregulatory state in Kuhl's pipistrelle Pipistrellus kuhlii, a Vespertillionid bat.

Total evaporative water loss is the sum of respiratory water loss (RWL) and cutaneous water loss (CWL) and constitutes the main avenue of water loss in bats. Because bats fly and have large surface-to-volume ratios, they potentially have high rates of RWL and CWL. Most species of small insectivorous bats have the ability to reduce their body temperature (T(b)) at rest, which substantially reduces energy expenditure and water loss. We hypothesized that bats reduce evaporative water loss during bouts of deep hypothermia (torpor) by decreasing RWL and CWL. We measured T(b), RWL, CWL, and resting metabolic rate (RMR) in Kuhl's pipistrelle Pipistrellus kuhlii, a small insectivorous bat. In support of our hypothesis, we found that RWL decreased with decreasing RMR. We found that CWL was lower in torpid individuals than in normothermic bats; however, bats in deep torpor had similar or higher CWL than bats in shallow torpor, suggesting that they exert a less effective physiological control over CWL when in deep torpor. Because insectivorous bats spend most of their lives in torpor or hibernation, the regulation of CWL in different heterothermic states has relevant ecological and evolutionary consequences.

Dietary fatty acid composition influences tissue lipid profiles and regulation of body temperature in Japanese quail.

Many avian species reduce their body temperature (T(b)) to conserve energy during periods of inactivity, and we recently characterized how ambient temperature (T(a)) and nutritional stress interact with one another to influence physiologically controlled hypothermic responses in Japanese quail (Coturnix japonica). In the present study, we examined how the fatty acid (FA) composition of the diet influences the FA composition of phospholipids in major organs and how these affect controlled hypothermic responses and metabolic rates in fasted birds. For 5 weeks prior to fasting, quail were fed a standard diet and gavaged each morning with 0.7 ml of water (control), or a vegetable oil comprising saturated fatty acids (SFA; coconut oil), or unsaturated fatty acids (UFA; canola oil). Birds were then fasted for 4 days at a T(a) of 15°C. We found that, while fasting, both photophase and scotophase T(b) decreased significantly more in the SFA treatment group than in the control group; apparently the former down-regulated their T(b) set point. This deeper hypothermic response was correlated with changes in the phospholipid composition of the skeletal muscle and liver, which contained significantly more oleic acid (18:1) and less arachidonic acid (20:4), respectively. Our data imply that these two FAs may be associated with thermoregulation.