Stressful experiences are linked to anxiety disorders in humans. Similar effects are observed in rodent models, where anxiety is often measured in classic conflict tests such as the open-field test. Spontaneous rearing behavior, in which rodents stand on their hind legs to explore, can also be observed in this test yet is often ignored. We define two forms of rearing, supported rearing (in which the animal rears against the walls of the arena) and unsupported rearing (in which the animal rears without contacting the walls of the arena). Using an automated open-field test, we show that both rearing behaviors appear to be strongly context dependent and show clear sex differences, with females rearing less than males. We show that unsupported rearing is sensitive to acute stress, and is reduced under more averse testing conditions. Repeated testing and handling procedures lead to changes in several parameters over varying test sessions, yet unsupported rearing appears to be rather stable within a given animal. Rearing behaviors could therefore provide an additional measure of anxiety in rodents relevant for behavioral studies, as they appear to be highly sensitive to context and may be used in repeated testing designs.

The locus coeruleus (LC) is a region in the brainstem that produces noradrenaline and is involved in both normal and pathological brain function. Pupillometry, the measurement of pupil diameter, provides a powerful readout of LC activity in rodents, primates and humans. The protocol detailed here describes a miniaturized setup that can screen LC activity in rodents in real-time and can be established within 1-2 d. Using low-cost Raspberry Pi computers and cameras, the complete custom-built system costs only 300 euros, is compatible with stereotaxic surgery frames and seamlessly integrates into complex experimental setups. Tools for pupil tracking and a user-friendly Pupillometry App allow quantification, analysis and visualization of pupil size. Pupillometry can discriminate between different, physiologically relevant firing patterns of the LC and can accurately report LC activation as measured by noradrenaline turnover. Pupillometry provides a rapid, non-invasive readout that can be used to verify accurate placement of electrodes/fibers in vivo, thus allowing decisions about the inclusion/exclusion of individual animals before experiments begin.

30Psychiatric diseases have a strong heritable component known to not be restricted to 31 DNA sequence-based genetic inheritance alone but to also involve epigenetic factors in 32 germ cells 1,2 . Initial evidence suggested that sperm RNA is causally linked 2,3 to the 33 transmission of symptoms induced by traumatic experiences. Here we show that 34 alterations in long RNA in sperm contribute to the inheritance of specific trauma 35 symptoms. Injection of long RNA fraction from sperm of males exposed to postnatal 36 trauma recapitulates the effects on food intake, glucose response to insulin and risk-37 taking in adulthood whereas the small RNA fraction alters body weight and behavioral 38 despair. Alterations in long RNA are maintained after fertilization, suggesting a direct 39 link between sperm and embryo RNA. 40 41 42 43 Adverse experiences can have long-lasting transgenerational effects on mental and 46 physical health, and often increase disease risk 4,5 . Traumatic stress in early life in 47 particular, can induce pathologies like psychosis, depression and metabolic 48 dysfunctions in adulthood across generations 6 . To examine the biological factors 49 involved, we recapitulated heritable behavioural and metabolic effects of postnatal 50 trauma across several generations using a previously established model of 51 unpredictable maternal separation combined with unpredictable maternal stress 52(MSUS) in the mouse, that shows symptoms through up to three generations ( Fig.1) 2,7-53 13 . We have shown that such postnatal trauma alters small RNA in sperm and that 54 injection of total sperm RNA from exposed male mice into naïve fertilized oocytes elicits 55 symptoms reminiscent of those observed in natural offspring of exposed fathers 2 . Other 56 studies have demonstrated that adult stress 14,15 and environmental insults like altered 57 diet or vinclozolin exposure, or positive factors such as exercise or environmental 58 enrichment can affect small RNA in sperm 16-21 and somatic tissues 22 in the offspring.59Recently, tRNA fragments and their modifications were also found to be affected by 60 nutritional insult, and unmodified or modified sperm small RNA injected into fertilized 61 oocytes could mimic metabolic changes resulting from altered parental diet in the 62 progeny 18,23,24 . These studies therefore suggest that small RNA in sperm can be 63 carrier of heritable information. Here we sought to determine whether long RNA in 64 sperm also contributes to the transmission of the effects of previous exposure. 66 Materials and Methods 68Mice. C57BL/6J mice were housed in a temperature and humidity-controlled facility 69 under a reverse light-dark cycle, and food and water were provided ad libitum. 70 Experimental procedures were performed during the animals' active cycle. All71 experiments were approved by the cantonal veterinary office, Zurich (license 55/12 then 72 57/15).73 74 MSUS paradigm. C57BL/6J primiparous females and males were mated at 2.5-3 75 months of age. Randomly selected dams and litt...

AbstracttreeclimbR is for analyzing hierarchical trees of entities, such as phylogenies or cell types, at different resolutions. It proposes multiple candidates that capture the latent signal and pinpoints branches or leaves that contain features of interest, in a data-driven way. It outperforms currently available methods on synthetic data, and we highlight the approach on various applications, including microbiome and microRNA surveys as well as single-cell cytometry and RNA-seq datasets. With the emergence of various multi-resolution genomic datasets, treeclimbR provides a thorough inspection on entities across resolutions and gives additional flexibility to uncover biological associations.