A cross-sectional observational study for ethno-geographical disparities in sleep quality, brain morphometry and cognition (a SOLACE study) in Indians residing in India, and South Asians and Europeans residing in the UK - a study protocol.

Introduction: As individuals age, their sleep patterns change, and sleep disturbances can increase the risk of dementia. Poor sleep quality can be a risk factor for mild cognitive impairment (MCI) and dementia. Epidemiological studies show a connection between sleep quality and cognitive changes, with brain imaging revealing grey matter volume reduction and amyloid beta accumulation in Alzheimer's disease. However, most research has focused on Europeans, with little attention to other ethnic groups. Methods: This is a cross sectional study comparing effects across countries and ethnicities. Group 1 (n = 193) will be Indians residing in India (new participant recruitment), Group 2 will be South Asians residing in UK and group 3 will be Europeans residing in the UK. For group 2 and 3 (n = 193), data already collected by UK-based Southall and Brent REvisited (SABRE) tri-ethnic study will be used. For group 1, Pittsburgh Sleep Quality Index questionnaire (PSQI) will be used for assessment of sleep quality, Indian Council of Medical Research (Neurocognitive ToolBox) (ICMR-NCTB) for cognition testing and a 3 T MRI cerebral scan for brain morphometry. The data will be compared to sleep, cognitive function and brain MRI parameters from SABRE. Discussion: Racial and ethnic differences can impact the relationships of cognitive function, sleep quality and brain structure in older adults. Earlier studies have highlighted higher prevalence of poor sleep among black individuals compared to white individuals. Genetic or epigenetic mechanisms may contribute to these variations. Socio-cultural and environmental factors, such as neighbourhood, migration, lifestyle, stress and perceived discrimination may influence sleep patterns. The aim of the study is to examine the ethnogeographic variations in sleep quality, cognitive performance and brain morphometry among Indians living in India, and South Asians and Europeans residing in the UK.

A novel methodology to demonstrate vestibulo-ocular reflex using caloric stimulation in undergraduate physiology laboratory.

The study aims to develop a novel methodology to demonstrate the vestibulo-ocular reflex (VOR) and nystagmus by caloric stimulation in an undergraduate medical physiology laboratory. The experimental setup involved two sets of electrodes: one set positioned laterally to both eyes, and another set positioned vertically over either the right or left eye. The caloric method is used to stimulate ears, which involves irrigation of warm (44°C) and cold (30°C) water into the ears while maintaining a temperature difference of approximately ±7°C from the body temperature. The changes in chorioretinal potential were calibrated to angular displacement by a two-point calibration method, and angular velocity was derived after taking the first-time derivative. The results obtained from the digital data acquisition system were compared to the traditional instrument used in our Otorhinolaryngology Department [Interacoustics Videonystagmography (VNG) System for hospitals, medical grade] for the normal subject's data. No significant differences in angular velocity were noted (P > 0.05). The cold stimuli elicit a more pronounced VOR compared to the warm stimuli. It has been consistently observed that the onset of nystagmus occurs approximately 20 s after irrigation, reaching its peak intensity between 45 and 90 s, and gradually diminishing until it ceases after approximately 200 s. Our developed methodology enables the recording and quantification of nystagmus using easily accessible equipment. This study serves the goal of visualizing the physiological process of VOR and thereby fulfills the goal of an effective teaching tool for demonstrating to undergraduate medical students.NEW & NOTEWORTHY We developed a novel methodology to demonstrate and visualize the most common and important physiological phenomenon like the vestibulo-ocular reflex as a teaching module for undergraduate students.

Introducing virtual classrooms for undergraduate physiology teaching during the COVID-19 pandemic: acceptance by students and subjective impact on learning.

The COVID-19 pandemic and worldwide lockdowns brought major changes in education systems. There was a sudden obligatory shift toward utilization of digital resources for teaching and learning purposes. Medical education, specifically physiology teaching, comprises hands-on training in the laboratory. It is challenging to offer a course like physiology in a virtual format. The objective of this study was to assess the effectiveness and influence of virtual classroom technology on online physiology education in a sample size of 83 first-year MBBS undergraduates. A questionnaire comprising questions related to technology accessibility and utilization, comprehensibility and effectiveness of instructions, faculty proficiency, and learning outcomes was administered to the group. The responses were collected and analyzed. Validation through principal components and factor analysis showed that online teaching is not very effective and has a limited application in the physiology education of undergraduate MBBS students. Our study also revealed that virtual physiology teaching of undergraduate medical students during the COVID-19 pandemic had a moderate level of effectiveness.NEW & NOTEWORTHY In the present qualitative study, we have conducted and validated an online physiology teaching platform at a medical college to continue medical education during the peak times of the COVID-19 pandemic and prolonged lockdowns. Furthermore, we have evaluated the effectiveness of online physiology teaching through multidimensional feedback from undergraduate MBBS students. It is experimental evidence of inadequate sustainability, moderate efficacy, limited application, and poor first-hand experience gained by the students in virtual physiology teaching in a preclinical and clinical setting.

Rac1 controls cell turnover and reversibility of the involution process in postpartum mammary glands.

Cell turnover in adult tissues is essential for maintaining tissue homeostasis over a life span and for inducing the morphological changes associated with the reproductive cycle. However, the underlying mechanisms that coordinate the balance of cell death and proliferation remain unsolved. Using the mammary gland, we have discovered that Rac1 acts as a nexus to control cell turnover. Postlactational tissue regression is characterised by the death of milk secreting alveoli, but the process is reversible within the first 48 h if feeding recommences. In mice lacking epithelial Rac1, alveolar regression was delayed. This defect did not result from failed cell death but rather increased cell turnover. Fitter progenitor cells inappropriately divided, regenerating the alveoli, but cell death also concomitantly accelerated. We discovered that progenitor cell hyperproliferation was linked to nonautonomous effects of Rac1 deletion on the macrophageal niche with heightened inflammation. Moreover, loss of Rac1 impaired cell death with autophagy but switched the cell death route to apoptosis. Finally, mammary gland reversibility failed in the absence of Rac1 as the alveoli failed to recommence lactation upon resuckling.

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

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

Surveillance of Zika and Dengue viruses in field-collected Aedes aegypti mosquitoes from different states of India.

The emergence of Zika created a panic situation worldwide and burdened the public health system of the affected countries. In India, Zika outbreak occurred in 2017 and in July 2021 one laboratory-confirmed case was also reported in Kerala state. The study aimed to assess any positivity in field-collected mosquitoes for ZIKV and DENV during 2016-2021. Mosquito samples were received from 4 Zika affected citites, whereas samples were collected from Delhi. Out of 2346 pools, 4 pools for ZIKV, and 23 pools for DENV were found positive. ZIKV positive pools were from Shastri Nagar, Jaipur (2 pools), Rajpur Hotel, Jaipur (1 pool) and Kanpur (1 pool). Ten localities of Delhi were found positive for DENV. It was observed that the maximum positivity rate for both viruses were found in September-October. The active role of the vector in local transmission of ZIKV warrants regular surveillance and monitoring of Aedes vectors in India.

How Losing Sleep Following Vaccination May Weaken the Response to SARS-CoV Vaccines.

With the emergence of new SARS-CoV-2 variants, a close analysis of factors that affect the efficacy of the vaccine in different groups is a must. It is important to elucidate the role of clinical, behavioral and host factors on modulation of immunogenicity of the SARS-CoV-2 vaccine. Data from other vaccines have shown that duration and efficiency of sleep affect the immunogenicity of the vaccine. There is a need for identification of circadian influence and sleep on SARS-CoV-2 vaccine using validated immune correlates of protection. We propose that sleep acts as a natural adjuvant by promoting the immunological synapse formation between the antigen presenting cells and CD4+ T cells thereby leading to activation of cellular and humoral immunity in SARS-CoV-2 infection. Thus, apart from the titer of neutralizing antibodies, the cellular immunity including CD4+ T cells and memory T cells must be assessed to clearly demarcate the long-term effect of sleep duration and efficiency on vaccine immunogenicity.

Mediodorsal thalamus lesion increases paradoxical sleep in rats.

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

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

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

Changes in sleep-wake cycle after microinjection of agonist and antagonist of endocannabinoid receptors at the medial septum of rats.

The role of medial septum in the genesis of slow-wave sleep and the inhibition of rapid eye movement sleep has been established using neurotoxic lesion and chemical stimulation of the medial septum. Intracerebroventricular injection of endocannabinoids (anandamide) decreases wake and increases slow-wave and rapid eye movement sleep in rats. Central cannabinoid (CB1) receptors are localized in the rat medial septum; however, the role of cannabinoid receptors at the medial septum on the regulation of sleep-wakefulness in rats lacks evidence. In this study, we have examined the changes in sleep architecture of 21 male Wistar rats, divided into three groups. Initially, 6 rats were used for dose standardization. Subsequently, one group (n = 6) was microinjected with CB1 receptor agonist, R-(+)-WIN 55,212-2 mesylate salt, the second group (n = 6) received microinjection of CB1 receptor antagonist LY 320,135, and the third group (n = 5) was microinjected with the vehicle, DMSO at the medial septum using stereotaxy. The sleep-wake cycle was recorded using electroencephalogram, electro-oculogram, and electromyogram. Microinjection of CB1 receptor agonist at the medial septum decreased slow-wave sleep and increased total sleep time. The increase in total sleep time was due to an increased percentage of rapid eye movement sleep. After the third and fourth hour of CB1 receptor antagonist microinjection at the medial septum, slow-wave sleep decreased when compared to vehicle injection, while rapid eye movement sleep decreased compared to baseline. We conclude that the endocannabinoid system at the septal nucleus acts through CB1 receptors to increase rapid eye movement sleep in rats.

First report on the transmission of Zika virus by Aedes (Stegomyia) aegypti (L.) (Diptera: Culicidae) during the 2018 Zika outbreak in India.

In a recent outbreak of Zika virus (ZIKV) infection in Jaipur city (Rajasthan, India), a total of 159 cases were reported in September 2018. In order to identify vector responsible for Zika transmission, mosquitoes were collected from houses with reported Zika cases and nearby houses. A total of 108 pools containing 522 mosquitoes were tested for presence of ZIKV using RT-PCR and Real Time RT-PCR. We detected presence of ZIKV in three pools of Aedes (Stegomyia) aegypti (L.), out of a total of 79 pools with 383 Ae. aegypti through RT-PCR as well as real-time RT-PCR. The presence of ZIKV in Ae. aegypti was further confirmed by DNA sequencing of the partial envelope region of ZIKV. Homology search of DNA sequence revealed highest identity (100%) with a ZIKV isolate from human from the study area which support the role of Ae. aegypti acting as a ZIKV vector. All other mosquitoes (Aedes vittatus and Culex quinquefasciatus) were negative for ZIKV. None of the F1 generation mosquito pools (276 mosquitoes in 43 pools) were found positive. This is the first report of presence of ZIKV in Ae. aegypti from the Indian subcontinent.

ß1 integrin is a sensor of blood flow direction.

Endothelial cell (EC) sensing of fluid shear stress direction is a critical determinant of vascular health and disease. Unidirectional flow induces EC alignment and vascular homeostasis, whereas bidirectional flow has pathophysiological effects. ECs express several mechanoreceptors that respond to flow, but the mechanism for sensing shear stress direction is poorly understood. We determined, by using in vitro flow systems and magnetic tweezers, that ß1 integrin is a key sensor of force direction because it is activated by unidirectional, but not bidirectional, shearing forces. ß1 integrin activation by unidirectional force was amplified in ECs that were pre-sheared in the same direction, indicating that alignment and ß1 integrin activity has a feedforward interaction, which is a hallmark of system stability. En face staining and EC-specific genetic deletion studies in the murine aorta revealed that ß1 integrin is activated and is essential for EC alignment at sites of unidirectional flow but is not activated at sites of bidirectional flow. In summary, ß1 integrin sensing of unidirectional force is a key mechanism for decoding blood flow mechanics to promote vascular homeostasis.This article has an associated First Person interview with the first author of the paper.

Hijacking a Morphogenesis Proteinase for Cancer Cell Invasion.

A long-standing question in biology is whether cancer cells exploit developmental processes to invade the surrounding stroma. In this issue of Developmental Cell, Feinberg et al. (2018) identify the matrix metalloproteinase Mmp14 as a key driver of mammary invasion common to both development and cancer but expose an underlying twist.

Role of An. culicifacies as a vector of malaria in changing ecological scenario of Northeastern states of India.

BACKGROUND & OBJECTIVES: Malaria has become endemic and subject of concern in most part of the India especially Northeastern states of India. Surveys before 2000 revealed that Anopheles minimus was major vector responsible for transmission of malaria in this region followed by An. dirus and An. fluviatilis. However, recent studies indicate replacement of An. minimus vector by An. culicifacies due to different ecological changes and change in landuse pattern etc. The objective of present study was to explore the vectorial role of An. culicifacies in transmission of malaria in four malaria endemic states, viz. Assam, Meghalaya, Manipur and Sikkim of India. METHODS: The seven surveys were conducted in 176 selected villages belonging to eight districts of the four states in both pre-monsoon (March-April) and post-monsoon (September-October) seasons from 2010 to 2013. However, in 2011 surveys could not be carried out due to public inconvenience in pre-monsoon season. For vectorial role of all vector species collected, ELISA and PCR were assayed. RESULTS: A total of 19,173 specimens belonging to 30 anopheline species were collected, out of which 4315 belonged to four established vector species. In total, 4183 specimens were processed through ELISA, out of which 236 specimens were found positive for circumsporozoite (CS) protein. Further, infectivity was confirmed by PCR in 35 samples, of which 12 samples were found positive for Plasmodium falciparum and three for P. vivax. Out of 12 Plasmodium falciparum positive samples, nine samples were positive for An. culicifacies, two for An. fluviatilis and one for An. minimus. While out of three Plasmodium vivax positive samples, two samples were positive for An. dirus and one sample was positive for An. culicifacies. INTERPRETATION & CONCLUSION: Anopheles culicifacies replaced the An. minimus, the vector of malaria in Northeastern states of India, as it was found to be highly infected with malaria parasite as compared to An. minimus by ELISA and PCR analysis, and thus playing a major role in malaria transmission in this region. The ecological changes like deforestation, development of irrigation channels and change in landuse pattern, have helped in evolution of An. culicifacies in the study area. Therefore, modified vector control strategies are required on urgent basis.

Rac1 Controls Both the Secretory Function of the Mammary Gland and Its Remodeling for Successive Gestations.

An important feature of the mammary gland is its ability to undergo repeated morphological changes during each reproductive cycle with profound tissue expansion in pregnancy and regression in involution. However, the mechanisms that determine the tissue's cyclic regenerative capacity remain elusive. We have now discovered that Cre-Lox ablation of Rac1 in mammary epithelia causes gross enlargement of the epithelial tree and defective alveolar regeneration in a second pregnancy. Architectural defects arise because loss of Rac1 disrupts clearance in involution following the first lactation. We show that Rac1 is crucial for mammary alveolar epithelia to switch from secretion to a phagocytic mode and rapidly remove dying neighbors. Moreover, Rac1 restricts the extrusion of dying cells into the lumen, thus promoting their eradication by live phagocytic neighbors while within the epithelium. Without Rac1, residual milk and cell corpses flood the ductal network, causing gross dilation, chronic inflammation, and defective future regeneration.

An integrin-ILK-microtubule network orients cell polarity and lumen formation in glandular epithelium.

The extracellular matrix has a crucial role in determining the spatial orientation of epithelial polarity and the formation of lumens in glandular tissues; however, the underlying mechanisms remain elusive. By using Cre­Lox deletion we show that ß1 integrins are required for normal mammary gland morphogenesis and lumen formation, both in vivo and in a three-dimensional primary culture model in which epithelial cells directly contact a basement membrane. Downstream of basement membrane ß1 integrins, Rac1 is not involved; however, ILK is needed to polarize microtubule plus ends at the basolateral membrane and disrupting each of these components prevents lumen formation. The integrin­microtubule axis is necessary for the endocytic removal of apical proteins from the basement-membrane­cell interface and for internal Golgi positioning. We propose that this integrin signalling network controls the delivery of apical components to the correct surface and thereby governs the orientation of polarity and development of lumens.

Specific ß-containing integrins exert differential control on proliferation and two-dimensional collective cell migration in mammary epithelial cells.

Understanding how cell cycle is regulated in normal mammary epithelia is essential for deciphering defects of breast cancer and therefore for developing new therapies. Signals provided by both the extracellular matrix and growth factors are essential for epithelial cell proliferation. However, the mechanisms by which adhesion controls cell cycle in normal epithelia are poorly established. In this study, we describe the consequences of removing the ß1-integrin gene from primary cultures of mammary epithelial cells in situ, using CreER. Upon ß1-integrin gene deletion, the cells were unable to progress efficiently through S-phase, but were still able to undergo collective two-dimensional migration. These responses are explained by the presence of ß3-integrin in ß1-integrin-null cells, indicating that integrins containing different ß-subunits exert differential control on mammary epithelial proliferation and migration. ß1-Integrin deletion did not inhibit growth factor signaling to Erk or prevent the recruitment of core adhesome components to focal adhesions. Instead the S-phase arrest resulted from defective Rac activation and Erk translocation to the nucleus. Rac inhibition prevented Erk translocation and blocked proliferation. Activated Rac1 rescued the proliferation defect in ß1-integrin-depleted cells, indicating that this GTPase is essential in propagating proliferative ß1-integrin signals. These results show that ß1-integrins promote cell cycle in mammary epithelial cells, whereas ß3-integrins are involved in migration.

Molecular dissection of integrin signalling proteins in the control of mammary epithelial development and differentiation.

Cell-matrix adhesion is essential for the development and tissue-specific functions of epithelia. For example, in the mammary gland, beta1-integrin is necessary for the normal development of alveoli and for the activation of endocrine signalling pathways that determine cellular differentiation. However, the adhesion complex proteins linking integrins with downstream effectors of hormonal signalling pathways are not known. To understand the mechanisms involved in connecting adhesion with this aspect of cell phenotype, we examined the involvement of two proximal beta1-integrin signalling intermediates, integrin-linked kinase (ILK) and focal adhesion kinase (FAK). By employing genetic analysis using the Cre-LoxP system, we provide evidence that ILK, but not FAK, has a key role in lactogenesis in vivo and in the differentiation of cultured luminal epithelial cells. Conditional deletion of ILK both in vivo and in primary cell cultures resulted in defective differentiation, by preventing phosphorylation and nuclear translocation of STAT5, a transcription factor required for lactation. Expression of an activated RAC (RAS-related C3 botulinum substrate) in ILK-null acini restored the lactation defect, indicating that RAC1 provides a mechanistic link between the integrin/ILK adhesion complex and the differentiation pathway. Thus, we have determined that ILK is an essential downstream component of integrin signalling involved in differentiation, and have identified a high degree of specificity within the integrin-based adhesome that links cell-matrix interactions with the tissue-specific function of epithelia.