Photoperiod-driven concurrent changes in hypothalamic and brainstem transcription of sleep and immune genes in migratory redheaded bunting.

The molecular regulation of sleep in avian migrants is still obscure. We thus investigated this in migratory redheaded buntings, where four life-history states (LHS; i.e. non-migratory, pre-migratory, migratory and refractory states) were induced. There was increased night-time activity (i.e. Zugunruhe) during the migratory state with reduced daytime activity. The recordings of the sleep-wake cycle in buntings showed increased night-time active wakefulness coupled with drastically reduced front and back sleep during migratory phase. Interestingly, we found the buntings to feed and drink even after lights-off during migration. Gene expression studies revealed increased hypothalamic expression of glucocorticoid receptor (nr3c1), and pro-inflammatory cytokines (il1b and il6) in pre-migratory and migratory states, respectively, whereas in brainstem Ca2+/calmodulin-dependent protein kinase 2 (camk2) was upregulated during the migratory state. This suggested a heightened pro-inflammatory state during migration which is a feature of chronic sleep loss, and a possible role of Ca2+ signalling in promoting wakefulness. In both the hypothalamus and brainstem, the expression of melatonin receptors (mel1a and mel1b) was increased in the pre-migratory state, and growth hormone-releasing hormone (ghrh, known to induce sleep) was reduced during the migratory state. The current results demonstrate key molecules involved in the regulation of sleep-wake cycle across LHS in migratory songbirds.

Photoperiodic modulation of melatonin receptor and immune genes in migratory redheaded bunting.

The transcriptional regulation of innate immune function across annual life history states (LHS) remains obscure in avian migrants. We, therefore, investigated this in a migratory passerine songbird, redheaded bunting (Emberiza bruniceps), which exhibits long-distance vernal migration from India to Central Asia. We exposed the birds (N = 10) to differential photoperiodic conditions to induce a non-migratory (NM), pre-migratory (PM), migratory (MIG), and refractory (REF) state, and performed gene expression assays of melatonin receptors (MEL1A and MEL1B), and innate immunity-linked genes (IL1B, IL6, TLR4, and NFKB) in spleen and blood. We found a significant reduction in splenic mass and volume, and a parallel increase in fat accumulation, and testicular growth in birds under migratory state. The gene expression assay revealed an upregulation of MEL1A and MEL1B mRNA levels in both the tissues in MIG. Additionally, we found a nocturnal increase of splenic IL1B expression, and IL1B, IL6, and TLR4 expression in the blood. The mRNA expression of melatonin receptors and proinflammatory cytokine showed a positive correlation. These results suggest that melatonin relays the photoperiodic signal to peripheral immune organs, which shows LHS-dependent changes in mRNA expression of immune genes.

Hypothalamic plasticity in response to changes in photoperiod and food quality: An adaptation to support pre-migratory fattening in songbirds?

In latitudinal avian migrants, increasing photoperiods induce fat deposition and body mass increase, and subsequent night-time migratory restlessness in captive birds, but the underlying mechanisms remain poorly understood. We hypothesized that an enhanced hypothalamic neuronal plasticity was associated with the photostimulated spring migration phenotype. We tested this idea in adult migratory red-headed buntings (Emberiza bruniceps), as compared with resident Indian weaverbirds (Ploceus philippinus). Birds were exposed to a stimulatory long photoperiod (14L:10D, LP), while controls were kept on a short photoperiod (10L:14D, SP). Under both photoperiods, one half of birds also received a high calorie, protein- and fat-rich diet (SP-R, LP-R) while the other half stayed on the normal diet (SP-N, LP-N). Thirty days later, as expected, the LP had induced multiple changes in the behaviour and physiology in migratory buntings. Photostimulated buntings also developed a preference for the rich food diet. Most interestingly, the LP and the rich diet, both separately and in association, increased neurogenesis in the mediobasal hypothalamus (MBH), as measured by an increased number of cells immunoreactive for doublecortin (DCX), a marker of recently born neurons, in buntings, but not weaverbirds. This neurogenesis was associated with an increased density of fibres immunoreactive for the orexigenic neuropeptide Y (NPY). This hypothalamic plasticity observed in a migratory, but not in a non-migratory, species in response to photoperiod and food quality might represent an adaptation to the pre-migratory fattening, as required to support the extensive energy expenses that incur during the migratory flight.

Ambient temperature affects multiple drivers of physiology and behaviour: adaptation for timely departure of obligate spring migrants.

We investigated the role of ambient temperature in departure from wintering areas of migratory black-headed buntings in spring. Birds transferred at 22 and 35°C to long days were compared with one another and with controls held on short days for indices of readiness to migrate (Zugunruhe, fattening, mass gain), levels of testosterone and gonadal recrudescence. Temperature affected the development of migratory behaviour and physiology: buntings under long days at 35°C, compared with those at 22°C, showed altered migratory behaviour (daily activity and Zugunruhe onset), and enhanced muscle growth and plasma testosterone levels, but showed no effect on testis growth. Temperature was perceived at both peripheral and central levels, and affected multiple molecular drivers culminating into the migratory phenotype. This was evidenced by post-mortem comparison of the expression of 13 genes with known functions in the skin (temperature-sensitive TRP channels: trpv4 and trpm8), hypothalamus and/or midbrain (migration-linked genes: th, ddc, adcyap1 and vps13a) and flight muscles (muscle growth associated genes: ar, srd5a3, pvalb, mtor, myod, mstn and hif1a). In photostimulated birds, the expression of trpv4 in skin, th in the hypothalamus and midbrain, and srd5a3, ar, pvalb and mtor in flight muscle, in parallel with testosterone levels, was greater at 35°C than at 22°C. These results demonstrate the role of ambient temperature in development of the spring migration phenotype, and suggest that transcriptional responsiveness to temperature is a component of the overall adaptive strategy in latitudinal songbird migrants for timely departure from wintering areas in spring.

Neuromorphometric changes associated with photostimulated migratory phenotype in the Palaearctic-Indian male redheaded bunting.

Neural substrates, including brain areas, differential gene expression and neuroendocrine basis, of migration are known. However, very little is known about structural changes in the brain that underlie the development and cessation of migration in long-distance avian migrants. Towards this, we investigated neuromorphological changes in the higher-order associative areas in male redheaded bunting (Emberiza bruniceps), which is a Palaearctic-Indian night migrant with wintering grounds in India. Photosensitive birds (8L:16D; SD) were exposed to stimulatory long days (16L:8D; LD), with controls retained on non-stimulatory short days. LD birds depicted shifts to, and sustained night-time activity as recorded by actograms. LD birds demonstrated increased body mass, fat deposition and testicular volume in keeping with the migratory phenotype. When LD birds had exhibited 10.0 ± 2.4 cycles of Zugunruhe (intense nighttime activity in captives, akin to night migratory flight in the wild), bird brains were fixed by transcardial perfusion, and changes in the neuronal morphometry of pallial, sub-pallial and hypothalamic brain regions studied using rapid Golgi technique with modifications, as used and validated in our laboratory. There were significant differences in both area and perimeter of soma in the visual hyperpallium apicale implicated in migratory orientation and the neuroendocrine control region for timing of migration, the mediobasal hypothalamus. We attribute these neuromorphometric changes in the soma area and perimeter to the photostimulated changes associated with the development of migration and reproductive phenotypes in redheaded buntings. It is suggested that changes in the neuronal plasticity in brain control regions parallel photoperiod-induced physiological responses.

Development of vernal migration in redheaded buntings: concurrent behavioral, physiological and neural changes under stimulatory photoperiods.

We investigated changes in behavior, physiology and selected brain regions during the development of vernal migration and reproduction phenotypes in migratory redheaded buntings. We monitored 24 h activity-rest pattern and measured food intake, fat deposition, and body mass of buntings exposed for 12 weeks to short (SP, 8L : 16D) and long (LP, 13L : 11D) photoperiods at 22 ± 2 °C temperature. Under LP, not SP, buntings exhibited a photostimulated spring migration phenotype (hyperphagia, fat deposition and body mass gain). However, there were sex differences in the development of vernal migration, as shown by faster and earlier induction of Zugunruhe (nocturnal migratory restlessness) in males than in females. In the next experiment, increasing photoperiods over 12 weeks following the vernal equinox induced behavioural and physiological changes associated with vernal migration phenotypes in both male and female buntings, but in a sex-dependent manner. In a subsequent experiment over 8 weeks corresponding to the spring migration period we found an increased expression of CART, not NPY, in INc, and decreased expression of GnRH-I in POA in the brain by week 6 of the observation under increasing photoperiods. There was also an increased expression of doublecortin (a marker of neuronal incorporation) in the olfactory bulb and song control nuclei (Area X and HVC, higher vocal centre) in male birds. These results demonstrate changes in the brain peptides and neuronal recruitment along with changes in the behaviour and physiology, and give insights into the concurrent photoperiodic induction of the seasonal response at multiple levels in migratory songbirds.

Difference in control between spring and autumn migration in birds: insight from seasonal changes in hypothalamic gene expression in captive buntings.

We hypothesized differences in molecular strategies for similar journeys that migrants undertake to reproduce in spring and to overwinter in autumn. We tested this in redheaded buntings (Emberiza bruniceps) photoinduced into spring and autumn migratory states, with winter and summer non-migratory states as controls. Compared with controls, buntings fattened, gained weight and showed Zugunruhe (nocturnal migratory restlessness) in the migratory state. Spring migration was associated with greater fat and body mass, and higher intensity of Zugunruhe, compared with autumn migration. Circulating corticosterone levels were higher in spring, while T3 levels were higher in autumn. Hypothalamic expression of thyroid hormone-responsive (dio2, dio3), light-responsive (per2, cry1, adcyap1) and th (tyrosine hydroxylase, involved in dopamine biosynthesis) genes showed significant changes with transition from non-migratory to the migratory state. There were significantly higher mRNA expressions in autumn, except for higher th levels in the spring. Furthermore, the expression patterns of dnmt3a (not dnmt3b) and tet2 genes suggested an epigenetic difference between the non-migrant and migrant periods, and the spring and autumn migrant periods. These results demonstrate for the first time seasonal transition in hypothalamic gene expressions, and suggest differences in regulatory strategies at the transcriptional level for spring and autumn migrations in songbirds.

Changes in brain peptides associated with reproduction and energy homeostasis in photosensitive and photorefractory migratory redheaded buntings.

Present study examined the expression of brain peptides associated with the reproduction and energy homeostasis (GnRH/GnIH, NPY/VIP), and assessed their possible functional association in the photosensitive (non-breeding, pre-breeding), photostimulated (breeding) and photorefractory (post-breeding) migratory redheaded buntings (Emberiza bruniceps), using double-labeled immunohistochemistry. Particularly, we measured immunoreactive (-ir) cell numbers, per cent cell area and cell optical density (OD) in the preoptic area (GnRH-I), midbrain (GnRH-II), paraventricular nucleus (GnIH), dorsomedial hypothalamus, DMH and infundibular complex, INc (NPY and VIP), and lateral septal organ (VIP) of buntings kept under natural photoperiods at the wintering latitude (26°55'N). There was a significant seasonal difference in GnRH-I, not GnRH-II, with reduced -ir cells in the photosensitive and photorefractory buntings, and notably with increased cell OD between the refractory and non-breeding states with no increase in testis size. Also, increased cell OD of GnIH neurons in non-breeding state indicated its role in the maintenance of small testes during the post-refractory period. Overall, seasonal changes in GnRH-I and GnIH were found consistent with their suggested roles in reproductive regulation of absolute photorefractory birds. Further, there was a significant seasonal change in cell OD of NPY neurons in DMH, not the INc. In contrast, VIP immunoreactivity was seasonally altered, with a significantly higher VIP-ir cells in breeding than the pre-breeding state. Finally, close proximity between perikarya with fibres suggested functional interactions between the GnRH and GnIH, and NPY and VIP. Thus, seasonal plasticity of brain peptides is perhaps the part of neural regulation of seasonal reproduction and associated energy homeostasis in migratory songbirds.

Nocturnal melatonin levels decode daily light environment and reflect seasonal states in night-migratory blackheaded bunting (Emberiza melanocephala).

We proposed two perhaps overlapping hypotheses. Hypothesis 1 examined whether daily light information was transduced by a change in the pattern of daily melatonin secretion. Hypothesis 2 tested whether the melatonin amplitude peak was contingent upon seasonal states. To test these hypotheses, we performed three experiments on night migratory blackheaded buntings (Emberiza melanocephala). The first two experiments measured plasma melatonin levels in buntings exposed to light-dark (LD) cycles, with white and/or blue (450 nm) and red (640 nm) light periods differing by about 10-12 fold in the level of illuminance. In birds exposed to a 12 h day (white light) at 0.66 W m(-2) (dim) and 7.85 W m(-2) (bright) light intensities (experiment 1), night melatonin levels were significantly affected by the end of 8-week exposure in the dim, not bright, day with buntings showing a bimodal nocturnal melatonin peak. Similarly, in birds on 13 h days in white, blue and red light periods at 0.028 W m(-2) (dim) and 0.28 (bright) W m(-2) intensities (experiment 2), mid night melatonin levels were significantly higher in red than in the blue or white light periods after 4.5 weeks of exposure. The third experiment measured plasma melatonin levels in buntings that were held under natural light conditions (NDL, 27°N; experiment 3A), or exposed to LD cycles (experiment 3B). There were slightly more elevated melatonin levels early at night during the spring (photosensitive) than during the autumn (photorefractory) migration season, without a difference in nocturnal melatonin peak between these two times providing almost similar light hours. Similarly, there was no difference in mid night melatonin levels between photosensitive and photorefractory buntings subjected to a skeleton photoperiod (1L : 11D : 1L : 11D), although melatonin levels were higher in the first than in the second 11 h dark phase in photosensitive birds. Overall, these results show that (i) nocturnal melatonin levels decode the intensity and wavelength of the daily light environment, and (ii) the daily melatonin secretion pattern subtly reflects seasonal states in the migratory blackheaded bunting.

Molecular basis of photoperiodic control of reproductive cycle in a subtropical songbird, the Indian weaver bird (Ploceus philippinus).

Less is known about genetic basis of photoperiodic regulation of reproductive cycle in subtropical birds. This study measured the expression levels of DIO2, DIO3, GnRH, and GnIH genes in Indian weaver birds subjected to short days (8h light:16h darkness, 8L:16D) and long days (16L:8D) for 48weeks. Whereas small, reproductively inactive testes were maintained under short days, weaver birds underwent testis recrudescence - regression cycle under long days. Relative expression levels of DIO2, DIO3, GnRH and GnIH genes were quantified by the real-time PCR (qPCR) in hypothalamus of birds (n=4) sampled at the beginning of the experiments, and after 10 and 48weeks of short and long day exposures. These sample times represented photosensitive unstimulated (day 0), and under long days the recrudescence (photostimulated, after 10weeks) and regression (photorefractory, after 48weeks) testicular phases. Birds under short days served as controls. The expression pattern of these genes corresponded with testicular phases. High and low GnRH and DIO2 levels were found in birds with large and small testes, respectively. By-and-large the converse was true for GnIH and DIO3 expression levels. Thus, after 10weeks of exposure, there was a significant difference in the mRNA levels between short and long day birds, with small and large testes, respectively. The results also suggest for a possible rapid switching between DIO2 and DIO3 and GnRH and GnIH expressions during testis maturation - regression cycle in Indian weaver birds.

Adaptation of oxidative phosphorylation to photoperiod-induced seasonal metabolic states in migratory songbirds.

Eukaryotic cells produce chemical energy in the form of ATP by oxidative phosphorylation of metabolic fuels via a series of enzyme mediated biochemical reactions. We propose that the rates of these reactions are altered, as per energy needs of the seasonal metabolic states in avian migrants. To investigate this, blackheaded buntings were photoperiodically induced with non-migratory, premigratory, migratory and post-migratory phenotypes. High plasma levels of free fatty acids, citrate (an intermediate that begins the TCA cycle) and malate dehydrogenase (mdh, an enzyme involved at the end of the TCA cycle) confirmed increased availability of metabolic reserves and substrates to the TCA cycle during the premigratory and migratory states, respectively. Further, daily expression pattern of genes coding for enzymes involved in the oxidative decarboxylation of pyruvate to acetyl-CoA (pdc and pdk) and oxidative phosphorylation in the TCA cycle (cs, odgh, sdhd and mdh) was monitored in the hypothalamus and liver. Reciprocal relationship between pdc and pdk expressions conformed with the altered requirements of acetyl-CoA for the TCA cycle in different metabolic states. Except for pdk, all genes had a daily expression pattern, with high mRNA expression during the day in the premigratory/migratory phenotypes, and at night (cs, odhg, sdhd and mdh) in the nonmigratory phenotype. Differences in mRNA expression patterns of pdc, sdhd and mdh, but not of pdk, cs and odgh, between the hypothalamus and liver indicated a tissue dependent metabolism in buntings. These results suggest the adaptation of oxidative phosphorylation pathway(s) at gene levels to the seasonal alternations in metabolism in migratory songbirds.

Intermittent Food Absence Motivates Reallocation of Locomotion and Feeding in Spotted Munia (Lonchura punctulata).

BACKGROUND: Daily feeding and locomotion are interrelated behaviours. The time spent in feeding and rate of food intake depends on food availability. In low food condition, the birds would show intense movement (locomotion) for a longer time throughout the day however during abundant food supply they may chose higher activity and food intake in the morning and evening only. In the present study we hypothesized that in Spotted Munia (Lonchura punctulata), intermittent food availability during day would reallocate their interrelated behaviors, the feeding (food intake) and locomotor activity patterns. METHODS: Two groups of birds (N = 6 each) were kept individually in activity cages under 12L:12D. Group 1 (Control; C) had ad libitum food but group 2 (Treatment; T) had food for 6 hours only (2 h presence followed by 2 h absence; 2P:2A) during 12 hour light period. In the first week, group 2 received food with 'lights on' (TI; ZT 0-2, 4-6 and 8-10; where ZT 0= zeitgeber time 0, time of lights ON). In the following week, the food was given 2 hours after 'lights on' (TII; ZT 2-4, 6-8, 10-12). The food intake and locomotor activity under each condition were observed. RESULTS: The results showed that locomotor activity was induced during food deprivation and suppressed during food availability. Also the food deprivation led to increased food intake. CONCLUSION: Our results suggest that intermittent food availability/deprivation reallocates the locomotor activity and food intake in Spotted Munia.

Circadian rhythms are not involved in the regulation of circannual reproductive cycles in a sub-tropical bird, the spotted munia.

Circannual rhythms regulate seasonal reproduction in many vertebrates. The present study investigated whether circannual reproductive phenotypes (rhythms in growth of gonads and molt) were generated independently of the circadian clocks in the subtropical non-photoperiodic spotted munia (Lonchura punctulata). Birds were subjected to light:dark (LD) cycles with identical light but varying dark hours, such that the period of LD cycle (T) equaled 16 h (T16; 12 h L:4 h D), 21 h (T21; 12 h L:9 h D), 24 h (T24; 12 h L:12 h D) and 27 h (T27; 12 h L:15 h D), or to continuous light (LL, 24 h L:0 h D) at ~18°C. During the ~21 month exposure, munia underwent at least two cycles of gonadal development and molt; changes in body mass were not rhythmic. This was similar to the occurrence of annual cycles in reproduction and molt observed in wild birds. A greater asynchrony between circannual cycles of gonad development and molt indicated their independent regulation. Females showed reproductive rhythms with similar circannual periods, whilst in males, circannual periods measured between peak gonadal size were longer in T21 and T24 than in T16 or T27. This suggested a sex-dependent timing of annual reproduction in the spotted munia. Also, food availability periods may not influence the circannual timing of reproduction, as shown by the results on the rhythm in gonadal growth and regression in munia under T-photocycles and LL that provided differential light (feeding) hours. Further, a short-term experiment revealed that activity-rest patterns in munia were synchronized with T-photocycles, but were arrhythmic under LL. We conclude that circadian rhythms are not involved in the timing of the annual reproductive cycle in the spotted munia.

Photoperiodic effects on activity behaviour in the spiny eel (Macrognathus pancalus).

The study focused on the characteristics of circadian locomotor activity in the spiny eel, M. pancalus, kept under different photoperiodic conditions. Two experiments were conducted. Experiment 1 tested the light intensity dependent effect on circadian rhythmicity of the locomotor activity in spiny eel. Three groups of fish were entrained to 12L:12D conditions for 10 days. Thereafter, they were released to constant conditions for 15 days as indicated below: group 1-DD (0 lux), group 2- LL(dim) (-1 lux) and group 3-LL(bright) (-500 lux). The locomotor activity of the fish, housed singly in an aquarium, was recorded continuously with infrared sensors connected to a computer. More than 90% activity of the eels was confined to the dark hours suggesting nocturnal habit. Under constant conditions, the activity in 7/9 fish in group 1, 4/8 in group 2 and 3/8 in group 3, started free running with a mean circadian period of 24.48 +/- 0.17 h, 23.21 +/- 0.47 h and 25.54 +/- 1.13 h in respective groups. Remaining fish in each group became arrhythmic. This suggests that spiny eel can be synchronised to LD cycle and under constant conditions they free run with a circadian period. However, their activity under LL is light intensity dependent; higher the intensity, more disruption in circadian locomotor activity. Experiment 2 was conducted to study the effect of decreasing night length (increasing photoperiod) on circadian locomotor activity. The fish were sequentially exposed to 16D (8L:16D), 12D (12L:12D), 8D (16L:8D), 4D (20L:4D) and 2D (22L:2D) for 10 days in each condition, thereafter, they were released in constant dark (DD= 0lux). The results showed that the duration of night length affects both, the amplitude and duration of locomotor activity. It can be concluded that the spiny eels are nocturnal and that their locomotor activity is under the circadian control and may be influenced by the photoperiod.

Light wavelength dependent circadian and seasonal responses in blackheaded bunting.

Animals in the wild are exposed to daily variations in sun light, viz. duration, intensity and spectrum. Photosensitive blackheaded buntings (Emberiza melanocephala) were exposed to photoperiods differing in the length of light period, wavelengths and intensity. The effects of such light changes were measured on locomotor activity rhythm as well as seasonal responses like development of migratory restlessness: Zugunruhe, body mass and gonadal growth. The results show that the buntings are differentially responsive to light wavelengths and intensities and are indicative of a phase-dependent action of light on the circadian photoperiodic system. These birds seem to use changes in the light variables of the solar environment to regulate their circadian and seasonal responses.

Gut dysbiosis impacts the immune system and promotes prostate cancer.

The gut microbiota is a system of microorganisms in the human gastrointestinal (GI) system, consisting of trillions of microorganisms residing in epithelial surfaces of the body. Gut microbiota are exposed to various external and internal factors and form a unique gut-associated immunity maintained through a balancing act among diverse groups of microorganisms. The role of microbiota in dysbiosis of the gut in aiding prostate cancer development has created an urgency for extending research toward comprehension and preventative measures. The gut microbiota varies among persons based on diet, race, genetic background, and geographic location. Bacteriome, mainly, has been linked to GI complications, metabolism, weight gain, and high blood sugar. Studies have shown that manipulating the microbiome (bacteriome, virome, and mycobiome) through the dietary intake of phytochemicals positively influences physical and emotional health, preventing and delaying diseases caused by microbiota. In this review, we discuss the wealth of knowledge about the GI tract and factors associated with dysbiosis-mediated compromised gut immunity. This review also focuses on the relationship of dysbiosis to prostate cancer, the impact of microbial metabolites short-chain fatty acids (SCFAs) on host health, and the phytochemicals improving health while inhibiting prostate cancer.

The presence/absence of conspecifics modulates the circadian locomotor activity and body mass in spotted munia (Lonchura punctulata).

Biological clocks regulate the behavior and physiology of animals by tracking the local time using diverse time cues. Social cues are relevant in studying the behavior of gregarious animals, but these cues have not been widely studied in birds. Temporal information for circadian timekeeping is socially communicated through visual, physical, olfactory, and auditory means. We examined the efficacy of pulsatile social interactions on locomotor activity and its associated characteristics such as distribution profile of rest and activity, total counts, activity duration, phase shift in activity onset, and circadian periodicity in spotted munia. Besides, we analyzed the effect of such social interactions on their body mass. Spotted munia exhibited phase shift in the onset of activity when subjected to social isolation, but these cues could not affect their circadian periodicity. In Pair as well as in Group, social isolation led to increased activity and activity duration, and decreased body mass in guests relative to the host bird. Our results suggest that the circadian rhythm of locomotor activity in spotted munia is quite sensitive to socialization and isolation, and isolation is detrimental for the birds. Consistent with these observations, the decline in body mass revealed the physiological consequences of social isolation on spotted munia.

Evaluation of social jetlag and chronotypes in Indian school-going adolescents.

Background: School-going students' school time acts as a zeitgeber during the weekdays, and in the weekends, due to the absence of the zeitgeber, they show free run and wake up late in the morning. This may result in poor health hazards in adolescents. Objective: The aim of this study was to assess the prevalence of social jetlag along with the distribution of their chronotype in school-going adolescents. Materials and Methods: This is a cross-sectional study conducted between May and June 2019 with the help of the MCTQ (Munich Chronotype Questionnaire) and MEQ (Morningness and Eveningness Questionnaire) in the adolescent population (N = 1154). Statistical Analysis: The study was performed using the Mann-Whitney U test to compare continuous variables of sleep parameters between workdays and free days. The significance was taken at P < 0.05. Results: The results showed that out of the total population, 81% were morning type and 67% of adolescents were socially jetlagged. There was a significant delay in the time to bed on free days by 1.88 hours and a significant delay in the wake-up time by 2.46 hours. The total sleep duration was also significantly higher on free days by 1.88 hours in the population. Conclusion: This is our first study in the Indian population to show that morning-type individuals showed irregular sleep timings and were more socially jetlagged. This needs to be focused on by the school authorities and government to keep a flexible school timing for the students on the basis of their chronotype for their better academic performance.

Automated qualitative batch measurement of lipid droplets in the liver of bird using ImageJ.

Oil red O staining is effective in detection and quantification of neutral lipid droplets in tissues such as the liver. However, converting images into testable data using ImageJ can be time-consuming and is prone to inaccuracy or bias. We describe a protocol for automated qualitative measurement of lipid droplets in the bird liver using a batch processing macro script. We explain steps for extracting tissue, cryosectioning, staining, and imaging, followed by script generation for quantification of lipid in the images.

Surviving high temperatures: a case study of the spotted munia (Lonchura punctulata).

Every year, a combination of summer with extreme weather events such as "heatwaves" affects the life of organisms on earth. Previous studies on humans, rodents, and some birds signify the impact of heat stress on their survival and existence. Over the past four decades, the frequency of heatwaves has increased because of global warming. Therefore, we performed a longitudinal study on a resident bird species, the spotted munia (Lonchura punctulata) by simulating a heatwave-like condition. We were interested in understanding how a Passeriformes native to a sub-tropical country deals with heatwave-like conditions. Initially, the birds were subjected to room temperature (25 ± 2 °C; T1) for 10 days, followed by a simulated heatwave-like condition (42 ± 1 °C; T2) for 7 days and again back to room temperature (25 ± 2 °C; RT1) for the next 7 days. To elucidate how birds cope with simulated heatwave conditions, we examined different behavioral and physiological parameters. We found that although heat stress significantly reduced total activity counts and food intake but, the body mass, blood glucose, and hemoglobin levels remained unaffected by any of the temperature conditions. Furthermore, HSP70 and biochemical markers of liver injuries such as ALP, AST, ALT, bilirubin direct, and bilirubin total were found elevated in response to the simulated heatwave-like condition, whereas uric acid and triglyceride were reduced. Creatinine and total protein levels were unaffected by the heatwave. The post heatwave treatment resulted in a rebound of the behavioral and physiological responses, but the recovered responses were not equivalent to the pre-heatwave levels (T1 conditions). Thus, the present study demonstrates heatwave-associated behavioral and physiological changes in a resident passerine finch which has tremendous physiological flexibility.