How to obtain an integrated picture of the molecular networks involved in adaptation to microgravity in different biological systems?

Periodically, the European Space Agency (ESA) updates scientific roadmaps in consultation with the scientific community. The ESA SciSpacE Science Community White Paper (SSCWP) 9, "Biology in Space and Analogue Environments", focusses in 5 main topic areas, aiming to address key community-identified knowledge gaps in Space Biology. Here we present one of the identified topic areas, which is also an unanswered question of life science research in Space: "How to Obtain an Integrated Picture of the Molecular Networks Involved in Adaptation to Microgravity in Different Biological Systems?" The manuscript reports the main gaps of knowledge which have been identified by the community in the above topic area as well as the approach the community indicates to address the gaps not yet bridged. Moreover, the relevance that these research activities might have for the space exploration programs and also for application in industrial and technological fields on Earth is briefly discussed.

General and species-specific recommendations for minimal requirements for the use of cephalopods in scientific research.

Here we list species-specific recommendations for housing, care and management of cephalopod molluscs employed for research purposes with the aim of contributing to the standardization of minimum requirements for establishments, care and accommodation of these animals in compliance with the principles stated in Directive 2010/63/EU. Maximizing their psychophysical welfare was our priority. General recommendations on water surface area, water depth and tank shape here reported represent the outcome of the combined action of the analysis of the available literature and an expertise-based consensus reached - under the aegis of the COST Action FA1301 - among researchers working with the most commonly used cephalopod species in Europe. Information on water supply and quality, environmental conditions, stocking density, feeding and handling are also provided. Through this work we wish to set the stage for a more fertile ground of evidence-based approaches on cephalopod laboratory maintenance, thus facilitating standardization and replicability of research outcomes across laboratories, at the same time maximizing the welfare of these animals.

Circulating and Salivary NGF and BDNF Levels in SARS-CoV-2 Infection: Potential Predictor Biomarkers of COVID-19 Disease-Preliminary Data.

COVID-19 continues to afflict the global population, causing several pathological diseases and exacerbating co-morbidities due to SARS-CoV-2's high mutation. Recent interest has been devoted to some neuronal manifestations and to increased levels of Nerve Growth Factor (NGF) and Brain-derived Neurotrophic Factor (BDNF) in the bloodstream during SARS-CoV-2 infection, neurotrophins that are well-known for their multifactorial actions on neuro-immune-endocrine and visual functions. Nineteen (19) patients were enrolled in this monocentric prospective study and subjected to anamnesis and biosamples collection (saliva and blood) at hospitalization (acute phase) and 6 months later (remission phase). NGF and BDNF were quantified by ELISA, and biochemical data were related to biostrumental measurements. Increased NGF and BDNF levels were quantified in saliva and serum during the acute phase of SARS-CoV-2 infection (hospitalized patients), and reduced levels were observed in the next 6 months (remission phase), never matching the baseline values. Salivary and circulating data would suggest the possibility of considering sera and saliva as useful matrices for quickly screening neurotrophins, in addition to SARS-CoV2 antigens and RNA. Overall, the findings described herein highlight the importance of NGF and BDNF as dynamic biomarkers for monitoring disease and reinforces the possibility of using saliva and sera for quick, non-invasive COVID-19 screening.

When Nerve Growth Factor Met Behavior.

Since its first characterization in the early 1950s, the role of the polypeptidic nerve growth factor (NGF) in controlling behavior remained elusive. Since the mid-1980s, we undertook a series of experiments aimed at elucidating the biological role(s) played by neurotrophins, particularly NGF, in adult rodents. At the beginning, we concentrated on the submandibular salivary gland of the male mouse, which was known to store massive amount of NGF. We found that under specific stress conditions, the salivary NGF is released in the bloodstream: intermale fighting between isolated males was the first reported context in which salivary NGF was released, thus providing a physiological significance for its presence in the adult, territorial males. We also found that dominant males release less NGF than subordinates and provided a loop-type model which includes intermale social confrontation, adrenal gland size, and functional status, corticosterone release, a model resulting in likelihood to be stabilized in a "dominant" or a "subordinate" social status. A variety of social anxiety contexts of mammals, humans included, has been described since then, and further studies carried out on humans showed that NGF is released in the bloodstream of parachutists at their first skydiving experience and in the case of ranking high on the Passionate Love Scale (amour fou). Ethological data from lab rodents helped in understanding NGF function in subtly controlling social "status" of male mice: the considerations about the interplay among neurobiological, physiological, and behavioral factors in structuring the dominant vs subordinate phenotypes may well apply to other vertebrate species, specifically addressing the underlying role of neurotrophins in relating behavior and brain neuroplasticity.

Relaxation Response in Stressed Volunteers: Psychometric Tests and Neurotrophin Changes in Biological Fluids.

Background: To evaluate the beneficial effects of relaxation response (RR) training in adult stressed subjects by evaluating the psychometric response recorded at relaxation session. Cortisol as well as nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) mediators were quantified in both saliva and tears, and their levels were related to each other and to the psychometric response. Methods: Stressed subjects (n = 23; 10M/13F; age range 21-53 years old) were voluntarily enrolled in the study. RR training sessions were carried out for 2 months, 1 day per week, at the same time (3-5 p.m.). Two different psychological questionnaires, the Perceived Stress Scale-10 (PSS-10) and the Beck Depression Inventory - Short Form (BDI-SF) and Ocular Surface Disease Index (OSDI) tests, were administered before each session. Saliva and tears were sampled for cortisol (EIA), NGF (ELISA), and BDNF (ELISA) quantifications. Questionnaires' data were analyzed and compared to biochemical ones. Results: All subjects reported beneficial effects from training. RR significantly reduced the psychological stress indexes (p = 0.039 for PSS-10 and p = 0.001 for BDI-SF). Specifically, RR training lowered the perception of Perceived Helplessness (items 1, 3, 10; p < 0.05) in PSS-10 and increased the Perceived Self-Efficacy (p < 0.05). OSDI score was in the normal range (0-25). Biochemically, a decrease in cortisol, a trend to a decrease in NGF, and an increase in BDNF levels were observed in saliva samples after RR treatment. Furthermore, a trend to a decrease in NGF and an increase in BDNF were quantified in tear samples. A correlation between PSS-10 total score and saliva NGF variation (%) as well as between BDI-SF total score and BDNF tear levels were also observed. Conclusion: RR training appeared useful to lowering psychological, mental, and physical stress, as supported by both psychological total and single scores. The finding on biochemical levels of BDNF in saliva and tears are sustained by previous studies while those of NGF require further investigation. Overall, these data on a small population highlight the potential use of RR training and potential neurotrophic changes in biological fluids, in stressed volunteers.

Effects of low-dose methylcyclopentadienyl manganese tricarbonyl-derived manganese on the development of diencephalic dopaminergic neurons in zebrafish.

Fuel additive methylcyclopentadienyl manganese tricarbonyl (MMT) is counted as an organic manganese (Mn)-derived compound. The toxic effects of Mn (alone and complexed) on dopaminergic (DA) neurotransmission have been investigated in both cellular and animal models. However, the impact of environmentally relevant Mn exposure on DA neurodevelopment is rather poorly understood. In the present study, the MMT dose of 100 µM (about 5 mg Mn/L) caused up-regulation of DA-related genes in association with cell body swelling and increase in the number of DA neurons of the ventral diencephalon subpopulation DC2. Furthermore, our analysis identified significant brain Mn bioaccumulation and enhancement of total dopamine levels in association with locomotor hyperactivity. Although DA levels were restored at adulthood, we observed a deficit in the acquisition and consolidation of memory. Collectively, these findings suggest that developmental exposure to low-level MMT-derived Mn is responsible for the selective alteration of diencephalic DA neurons and with long-lasting effects on fish explorative behaviour in adulthood.

Review: Environmental impact on ocular surface disorders: Possible epigenetic mechanism modulation and potential biomarkers.

Throughout life, external and the internal environments interact in normal development and tissue homeostasis as well as in onset of disease. Epigenetic modifications occur in response to environmental changes and play a fundamental role in controlling gene expression without modification of the DNA base sequence. Aging, inflammation, drugs, infections and ultraviolet exposure may have profound effects on epigenetic modifications and trigger susceptibility to diseases. Increasing evidence has demonstrated that epigenetic mechanisms play a key role in regulating the physiopathology of the ocular surface. The evaluation of epigenetic factors in ocular disease would lead to further investigation regarding the potential use of therapeutic targets and diagnostic biomarkers. This review examines specific epigenetic and biochemical mechanisms that may occur in the ocular surface microenvironment. Potential correlation between epigenetic factors and targets should be considered in future approaches to prevention and/or treatment of specific ocular surface disorders.

The Oral Health-Related Quality of Life in State Institutionalized Older Adults in Malta.

PURPOSE: The objective of this study was to determine the oral health-related quality of life in state institutionalized older adults and correlate it to their oral health status. MATERIALS AND METHODS: A group of 278 older adults (average age: 83.5±6.5 years) from 9 state institutions in Malta was randomly selected. Participants were clinically examined and answered three questionnaires (Oral Health Impact Profile [OHIP-14], Geriatric Oral Health Assessment Index [GOHAI], and Denture Satisfaction). RESULTS: Oral health-related quality of life measurements were significantly associated with pocket depth (OHIP-14 and GOHAI, P<.05); decayed, missing, or filled teeth (OHIP-14 and GOHAI, P<.05); carious teeth (OHIP-14 and GOHAI, P<.05); number of missing teeth (OHIP-14 and GOHAI, P<.05); and maxillary and mandibular dentures (OHIP-14 and GOHAI, P<.0001). Denture satisfaction was associated with denture age (P=.010) and types of prostheses (P<.05). CONCLUSION: Institutionalized older adults have high levels of oral disease. Although oral health-related quality of life is generally good, those participants with a poorer oral health-related quality of life had significant associations with poor oral health. However, the extent varies with the individual's perception of good health.

The oral health of institutionalized older adults in Malta.

PURPOSE: Studies on the oral health status of institutionalized older adults are less prevalent than those of community-dwelling older adults, as institutionalized older adults tend to be frailer. Poor oral health in older adults has a negative impact on the quality of life and self-confidence of older people and potentially poses a financial burden on both the older adult and society in general. The objective of this study was to assess and compare the oral health status of state institutionalized older adults in Malta with that of their European counterparts. MATERIALS AND METHODS: A total of 278 older adults with an average age of 83.6 ± 6.5 years from nine state institutions in Malta were randomly selected. Participants were clinically examined for caries, periodontal disease, oral mucosal lesions, and prosthetic status. RESULTS: The oral health status of state institutionalized older adults was poor, with a very low routine dental attendance (29.3%) and, consequently, a high level of treatment need (44.4% of dentate individuals required extractions and 42.1% of dentate individuals required restorations). Only 2% of dentate individuals had healthy periodontal tissues with no clinical attachment loss over 4 mm. Edentulism among state institutionalized older adults stood at 41%. CONCLUSIONS: Institutionalized older adults from Malta have a poor oral health status comparable to institutionalized older adults from Europe in general, which poses fiscal and cultural challenges that need to be addressed by the dental community.

Changes in stress hormones and metabolism during a 105-day simulated Mars mission.

BACKGROUND: The Mars-105 project was aimed at simulating crew's activities, workload, and communication during a mission to Mars, evaluating the homeostatic adaptations to prolonged confinement and cohabitation. METHODS: Fasting plasma glucose (FPG) and insulin, C-peptide, leptin, cortisol, and NGF and BDNF plasma levels were monitored in six healthy nonsmoking male subjects taking part in a 105-d Mars mission simulation. Samples were collected from each subject before (0 wk), during (2.5 wk; 5 wk; 10 wk; 15 wk), and after confinement (+1 wk). RESULTS: Confinement resulted in impaired glucometabolic parameters, since FPG increased during the first 5 wk (baseline: 85.2 ± 10.8 mg · dl⁻¹; 2.5 wk: 98.4 ± 4.7 mg · dl⁻¹; 5 wk: 92.5 ± 6.0 mg · dl⁻¹) and insulin dropped at 2.5 wk (baseline: 14.4 ± 4.8 mU · L⁻¹; 2.5 wk: 7.7 ± 2.1 mU · L⁻¹), subsequently returning to baseline values. HOMA-IR paralleled plasma insulin, dropping to 1.8 ± 0.5 at 2.5 wk (baseline: 3.0 ± 1.2). At all time-points tested, plasma leptin levels were decreased (baseline: 4.4 ± 3.3 ng · dl⁻¹; 2.5 wk: 1.6 ± 1.2 ng · dl⁻¹; 5 wk: 1.3 ± 0.8 ng · dl⁻¹; 10 wk: 1.5 ± 1.1 ng · dl⁻¹; 15 wk:1.7 ± 0.8 ng · dl⁻¹), whereas cortisol levels were increased (baseline: 10.8 ± 4.9 ng · dl⁻¹; 2.5 wk: 16.8 ± 3.5 ng · dl⁻¹; 5 wk: 18.1 ± 7.6 ng · dl⁻¹; 10 wk: 18.1 ± 8.3 ng · dl⁻¹; 15 wk:14.2 ± 4.4 ng · dl⁻¹), resulting in a negative correlation between these hormones. BDNF levels increased only at 5 and 10 wk (baseline: 67.1 ± 36.0 pg · ml⁻¹; 5 wk: 164 ± 54 pg · ml⁻¹; and 10 wk: 110.2 ± 28.9 pg · ml⁻¹). DISCUSSION: The data obtained with the Mars-105 experiment suggest that environmental stress has a strong impact upon metabolic and stress response, indicating the need for further studies and the implementation of specific countermeasures.

Behavioural response of European starlings exposed to video playback of conspecific flocks: effect of social context and predator threat.

We studied the behavioural response of European starlings to a socially mediated predation event. Adult starlings were exposed to either a video of a flock attacked by a peregrine falcon or a video of a flock not under attack. We investigated whether the social condition affected the anti-predator response under the hypothesis that in such a gregarious species singletons would increase their anti-predator behaviour more than individuals in groups, to compensate for potential increased risk. The video of the flock under attack caused an increase in immobility and vigilance, more marked in singletons, both during and after the exposure. The video of the non-threatened flock caused an increase in activity levels, especially during the exposure. Furthermore, we observed a marked increase in comfort activities in singletons as well as in social interactions and vocalizations in mini-flocks. Only birds in mini-flocks vocalized, which may be explained by an audience effect, a process of social cognition mediated by the social context, and not only by the stimulus. The results are in line with previous field studies, which showed that isolated starlings are exposed to a higher risk of predation compared to individuals in flocks.

Development of a Maltese version of oral health-associated questionnaires: OHIP-14, GOHAI, and the Denture Satisfaction Questionnaire.

PURPOSE: To show the reliability of the Maltese translations of OHIP-14, GOHAI, and the Denture Satisfaction Questionnaire, define the reliability of the responses, and determine the correlation between OHIP-14 and GOHAI. MATERIALS AND METHODS: The items of the three questionnaires (OHIP-14, GOHAI, and Denture Satisfaction) were translated into Maltese and back into English to compare with the original version. Specific sampling of a population well versed in Maltese and English was carried out to obtain a sample of respondents for each questionnaire. Data were gathered through self-administered questionnaires: first administering the Maltese version and following with the English version 1 week later. RESULTS: Participation rates were high (98%). Cronbach's alpha for all three questionnaires was high (> 0.7), indicating satisfactory test-retest reliability of the instruments. Similarly, the Spearman correlation coefficients for both the English and Maltese versions of OHIP-14 and GOHAI were good (> 0.6). CONCLUSION: The Maltese versions of OHIP-14, GOHAI, and the Denture Satisfaction Questionnaire can be safely used as a valid alternative to the English versions in studies of patients who are limited in linguistic proficiency.

Evaluation of gene, protein and neurotrophin expression in the brain of mice exposed to space environment for 91 days.

Effects of 3-month exposure to microgravity environment on the expression of genes and proteins in mouse brain were studied. Moreover, responses of neurobiological parameters, nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF), were also evaluated in the cerebellum, hippocampus, cortex, and adrenal glands. Spaceflight-related changes in gene and protein expression were observed. Biological processes of the up-regulated genes were related to the immune response, metabolic process, and/or inflammatory response. Changes of cellular components involving in microsome and vesicular fraction were also noted. Molecular function categories were related to various enzyme activities. The biological processes in the down-regulated genes were related to various metabolic and catabolic processes. Cellular components were related to cytoplasm and mitochondrion. The down-regulated molecular functions were related to catalytic and oxidoreductase activities. Up-regulation of 28 proteins was seen following spaceflight vs. those in ground control. These proteins were related to mitochondrial metabolism, synthesis and hydrolysis of ATP, calcium/calmodulin metabolism, nervous system, and transport of proteins and/or amino acids. Down-regulated proteins were related to mitochondrial metabolism. Expression of NGF in hippocampus, cortex, and adrenal gland of wild type animal tended to decrease following spaceflight. As for pleiotrophin transgenic mice, spaceflight-related reduction of NGF occurred only in adrenal gland. Consistent trends between various portions of brain and adrenal gland were not observed in the responses of BDNF to spaceflight. Although exposure to real microgravity influenced the expression of a number of genes and proteins in the brain that have been shown to be involved in a wide spectrum of biological function, it is still unclear how the functional properties of brain were influenced by 3-month exposure to microgravity.

The Mice Drawer System (MDS) experiment and the space endurance record-breaking mice.

The Italian Space Agency, in line with its scientific strategies and the National Utilization Plan for the International Space Station (ISS), contracted Thales Alenia Space Italia to design and build a spaceflight payload for rodent research on ISS: the Mice Drawer System (MDS). The payload, to be integrated inside the Space Shuttle middeck during transportation and inside the Express Rack in the ISS during experiment execution, was designed to function autonomously for more than 3 months and to involve crew only for maintenance activities. In its first mission, three wild type (Wt) and three transgenic male mice over-expressing pleiotrophin under the control of a bone-specific promoter (PTN-Tg) were housed in the MDS. At the time of launch, animals were 2-months old. MDS reached the ISS on board of Shuttle Discovery Flight 17A/STS-128 on August 28(th), 2009. MDS returned to Earth on November 27(th), 2009 with Shuttle Atlantis Flight ULF3/STS-129 after 91 days, performing the longest permanence of mice in space. Unfortunately, during the MDS mission, one PTN-Tg and two Wt mice died due to health status or payload-related reasons. The remaining mice showed a normal behavior throughout the experiment and appeared in excellent health conditions at landing. During the experiment, the mice health conditions and their water and food consumption were daily checked. Upon landing mice were sacrificed, blood parameters measured and tissues dissected for subsequent analysis. To obtain as much information as possible on microgravity-induced tissue modifications, we organized a Tissue Sharing Program: 20 research groups from 6 countries participated. In order to distinguish between possible effects of the MDS housing conditions and effects due to the near-zero gravity environment, a ground replica of the flight experiment was performed at the University of Genova. Control tissues were collected also from mice maintained on Earth in standard vivarium cages.

Early exposure to low doses of atrazine affects behavior in juvenile and adult CD1 mice.

Environmental exposure to endocrine disrupting chemicals is receiving increasing attention, with particular regard to distinct periods of development where neuroendocrine circuitries are critical for shaping the mammalian brain. Atrazine (ATZ), a widely used herbicide, has been reported to affect steroid hormones and interfere with pathways critical for sex-specific physiological and behavioral development. Aim of the present study was to evaluate effects of perinatal exposure to environmentally relevant subtoxic doses of ATZ, on neurobehavioral development in mice and investigate possible alterations in steroid hormone metabolism. Neurobehavioral development of female and male mice delivered from CD1 dams, and daily exposed from Gestational Day 14 until Postnatal Day 21 (PND 21) to 1 or 100 µg/kg bw ATZ, was investigated. Specifically, locomotor and exploratory activity, social interactions and cognitive performance were evaluated at PND 16, 31 and 60, respectively. Moreover, general toxicity clinical signs, testicular parameters, rate of testosterone metabolism and aromatase activity in F1 male liver were analyzed at adulthood. Changes in exploratory profile and in affiliative/investigative behavior were observed, revealing a feminization of behavioral profile in ATZ-exposed males. Alteration in learning performance at adulthood was also evident. A limited decreased sperm count and concentration, as well as some slight impairment in hepatic testosterone metabolism and in aromatase activity (slightly but not significantly decreased) were observed in both low and high dose exposed animals. In conclusion developmental exposure to non-toxic, environmentally relevant doses of ATZ can produce subtle functional alterations, detectable in juvenile rodents by a detailed behavioral analysis. Behavioral disturbances appeared mainly related with neurodevelopmental disorder affecting the social domain and the emotional/affective repertoire, although further research is needed to elucidate the mechanism through which the effects are induced.

More than antioxidant: N-acetyl-L-cysteine in a murine model of endometriosis.

N-acetyl-L-cysteine exerts a complex action on endometrial cells, involving regulation of gene expression and protein activity and location, all converging into a decreased proliferation and a switch toward a differentiating, less invasive, and less inflammatory phenotype. Also considering the lack of undesired side effects, including unaffected fertility potential, this suggests a beneficial use of NAC in endometriosis clinical treatment.

Bird populations as sentinels of endocrine disrupting chemicals.

Exposure to endocrine disrupting chemicals (EDCs) is a widespread phenomenon in nature. Although the mechanisms of action of EDCs are actively studied, the consequences of endocrine disruption (ED) at the population level and the adaptations evolved to cope with chronic EDC exposure have been overlooked. Birds probably represent the animal taxon most successfully adapted to synanthropic life. Hence, birds share with humans a similar pattern of exposure to xenobiotics. In this article, we review case studies on patterns of behaviour that deviate from the expectation in bird species exposed to EDCs. We provide behavioural and ecological parameters to be used as endpoints of ED; methodological requirements and caveats based on species-specific life-history traits, behavioural repertoires, developmental styles, and possibility of captive breeding; a list of species that could be used as sentinels to assess the quality of man-made environment.

Birth spacing in the mouse communal nest shapes adult emotional and social behavior.

The interactions with the mother and with peers are among the most relevant early environmental factors shaping adult brain function and behavior. In order to investigate the role of these factors, we exploited a novel early manipulation, the Communal Nest (CN), consisting in a single nest where three mothers give birth, keep their pups and share care-giving behavior from birth to weaning. In particular, we reared CD-1 swiss mice in three different CN conditions, each one characterized by a different interval between the three deliveries (Birth Spacing) of 3, 5 or 7 days (respectively, CN+/-3, CN+/-5, CN+/-7). Length of birth spacing affected maternal behavior, CN+/-7 pups receiving the highest levels. At adulthood, mice reared in the different conditions showed differences in emotional response and social skills. In the plus maze test, short birth spacing was found to be associated with enhanced emotionality, CN+/-3 mice showing highest levels of anxiety-like responses in the plus maze compared to the other two CN groups. In the social interaction test, the strategies to achieve dominance differed among the three groups. While CN+/-3 mice appeared to have a more aggressive strategy, displaying high levels of attack behavior in the first encounter, CN+/-5 and CN+/-7 mice displayed a more affiliative strategy based on social investigation. Overall, these findings show that birth spacing shapes the early mouse social environment and, in turn, affects the development of social skills and emotional responses.

A mouse model of neurobehavioural response to altered gravity conditions: an ontogenetical study.

To determine the influence of gravity during critical periods of development is important in the perspective of long-term spaceflight and exploration, data coming from this kind of studies providing insight into basical biological phenomena underlying the development of the nervous system and its plasticity. Aim of the present study was to evaluate neurobehavioural responses to hypergravity exposure in CD-1 mice at different stage of development. Early adolescent (postnatal day 28, PND 28), adolescent (PND 42) and young-adult (PND 60) male and female mice were exposed to acute 2g rotational-generated hypergravity. Motion sickness index and behavioural performances pre, during and after rotation were recorded, and long-lasting effects on exploratory behaviour (hole-board test) and emotional/anxiety-like responses (plus-maze test) were investigated. Furthermore, in order to correlate behavioural changes with alterations in central levels of neurotrophins, brain amounts of Nerve Growth Factor (NGF) and Brain Derived Neurotrophic Factor (BDNF) were also assessed on PND 90, following a re-exposure to hypergravity. Age and sex differences were observed, females being more vulnerable than males to motion sickness, and susceptibility to hypergravity increasing with age of exposure. Moreover, mice showed a general reduction in spontaneous activity during the rotation, while recovery time after rotation became progressively longer with increasing age of the experimental subjects. Long-term effects on exploratory behaviour and emotional/anxiety-like response were also observed, behavioural profiles mainly changing in those animals experiencing hypergravity as young-adults. Finally, major changes in brain levels of NGF and BDNF were detected in mice firstly exposed as young-adults.

Early social enrichment affects responsiveness to different social cues in female mice.

Communal nesting (CN), an early social enrichment procedure in which multiple females rear the offspring in a single nest, increases maternal care levels received by offspring and interaction with peers. It has been shown that male mice reared under CN conditions show increased social competence and propensity to social interactions at adulthood. In the present study we investigated long-term behavioural effects of CN on female offspring. Mouse pups were reared under two different experimental conditions: standard nesting (SN, where single mother rears her pups) and CN (three females rearing their pups in a single nest). At adulthood CN and SN virgin females underwent three different behavioural tests: (i) maternal induction following presentation of foster pups; (ii) social recognition test in which ultrasound vocalizations (USVs) and social investigation behaviour emitted by a resident female in the presence of a female partner were recorded; (iii) zero-maze to analyze anxiety profiles. CN females showed (i) decreased licking response in the maternal induction test accompanied by an increased sniffing response; (ii) decreased of social interest towards a novel partner (during the Retest Different phase), and decreased USV emission rate in the social recognition test; CN and SN females did not differ in the emotional responses measured in the zero-maze apparatus. As a whole these data suggest that CN rearing render female mice less reactive to social novelty.