Environmental temperature variation affects brain protein expression and cognitive abilities in adult zebrafish (Danio rerio): A proteomic and behavioural study.

Water temperature is an important environmental parameter influencing the distribution and the health of fishes and it plays a central role in ectothermic animals. The aim of this study is to determine the effects of environmental temperature on the brain proteome and the behavioural responses in zebrafish, a widely used animal model for environmental "omics" studies. Adult specimens of wild-type zebrafish were kept at 18 °C, 34 °C and 26 °C (control) for 21 days. Proteomic data revealed that several proteins involved in cytoskeletal organization, mitochondrial regulation and energy metabolism are differently regulated at the extreme temperatures. In particular, the expression of proteins associated to synapses and neurotransmitter release is down-regulated at 18 °C and 34 °C. In both thermal conditions, fish exhibited a reduced interest for the novel environment and an impairment of cognitive abilities during Y-Maze behavioural tests. The observed pathways of protein expression are possibly associated to functional alterations of the synaptic transmission that may result in cognitive functions impairment at central nervous system level as those revealed by behavioural tests. This study indicates that temperature variations can elicit biochemical changes that may affect fish health and behaviour. This combined approach provides insights into mechanisms supporting thermal acclimation and plasticity in fishes. SIGNIFICANCE: Environmental temperature variation may impact on all levels of biological life. Understanding the impact of thermal variation on the nervous system and animal behaviour is of primary importance since the results obtained can be applied from the ecological to the biomedical fields

Brain Proteome and Behavioural Analysis in Wild Type, BDNF+/− and BDNF−/− Adult Zebrafish (Danio rerio) Exposed to Two Different Temperatures

Experimental evidence suggests that environmental stress conditions can alter the expression of BDNF and that the expression of this neurotrophin influences behavioural responses in mammalian models. It has been recently demonstrated that exposure to 34 degrees C for 21 days alters the brain proteome and behaviour in zebrafish. The aim of this work was to investigate the role of BDNF in the nervous system of adult zebrafish under control and heat treatment conditions. For this purpose, zebrafish from three different genotypes (wild type, heterozygous BDNF+/- and knock out BDNF-/-) were kept for 21 days at 26 degrees C or 34 degrees C and then euthanized for brain molecular analyses or subjected to behavioural tests (Y-maze test, novel tank test, light and dark test, social preference test, mirror biting test) for assessing behavioural aspects such as boldness, anxiety, social preference, aggressive behaviour, interest for the novel environment and exploration. qRT-PCR analysis showed the reduction of gene expression of BDNF and its receptors after heat treatment in wild type zebrafish. Moreover, proteomic analysis and behavioural tests showed genotype- and temperature-dependent effects on brain proteome and behavioural responding. Overall, the absent expression of BDNF in KO alters (1) the brain proteome by reducing the expression of proteins involved in synapse functioning and neurotransmitter-mediated transduction; (2) the behaviour, which can be interpreted as bolder and less anxious and (3) the cellular and behavioural response to thermal treatment

Proteomic dissection of nanotopography-sensitive mechanotransductive signaling hubs that foster neuronal differentiation in PC12 cells

Neuronal cells are competent in precisely sensing nanotopographical features of their microenvironment. The perceived microenvironmental information will be \ue2\u80\u9cinterpreted\ue2\u80\u9d by mechanotransductive processes and impacts on neuronal functioning and differentiation. Attempts to influence neuronal differentiation by engineering substrates that mimic appropriate extracellular matrix (ECM) topographies are hampered by the fact that profound details of mechanosensing/-transduction complexity remain elusive. Introducing omics methods into these biomaterial approaches has the potential to provide a deeper insight into the molecular processes and signaling cascades underlying mechanosensing/-transduction but their exigence in cellular material is often opposed by technical limitations of major substrate top-down fabrication methods. Supersonic cluster beam deposition (SCBD) allows instead the bottom-up fabrication of nanostructured substrates over large areas characterized by a quantitatively controllable ECM-like nanoroughness that has been recently shown to foster neuron differentiation and maturation. Exploiting this capacity of SCBD, we challenged mechanosensing/-transduction and differentiative behavior of neuron-like PC12 cells with diverse nanotopographies and/or changes of their biomechanical status, and analyzed their phosphoproteomic profiles in these settings. Versatile proteins that can be associated to significant processes along the mechanotransductive signal sequence, i.e., cell/cell interaction, glycocalyx and ECM, membrane/f-actin linkage and integrin activation, cell/substrate interaction, integrin adhesion complex, actomyosin organization/cellular mechanics, nuclear organization, and transcriptional regulation, were affected. The phosphoproteomic data suggested furthermore an involvement of ILK, mTOR, Wnt, and calciumsignaling in these nanotopography- and/or cell mechanics-related processes. Altogether, potential nanotopography-sensitive mechanotransductive signaling hubs participating in neuronal differentiation were dissected

Effect of thermal stress on Danio rerio behaviour: a proteomic study to understand the molecular mechanism.

Environmental temperature variations affect many properties and functions of biomolecules and structural components at the cellular level, and influence the animal physiology and behavior at the organism level. It is well accepted that the biochemical changes induced by temperature stress are attributed to gene expression modulations. The aim of this work was to understand the molecular mechanism and the behavioural responses correlated to temperature stress in zebrafish (Danio rerio), which is a widely used animal model for environmental genomics researches. Adult specimens of wild type zebrafish were kept at three different temperatures: 18°C, 34°C and 26°C (used as a control) for 21 days and then subjected to behavioural tests using a YMaze task to evaluate the response to novelty and the spatial memory. Proteomic analysis were carried out on brains to evaluate the thermal effect at the central nervous system level. Briefly, for each temperature 9 brains were lysed and after reduction and derivatization, the proteins were digested with trypsin. LC-ESI MS/MS analysis was performed on a Dionex UltiMate 3000 HPLC. The eluate was electrosprayed into an LTQ Orbitrap Velos. Four technical replicate analyses of each sample were performed. Mass spectra were analyzed using MaxQuant and Peak Studio software. Finally, the bioinformatic analysis was carried out by DAVID and PANTHER softwares to evaluate enriched categories filtered for biological processes, molecular function (MF), cellular component (CC) and pathways involved. Preliminary results suggest that thermal stress at the cellular level influences the CC organization, biogenesis, structural morphogenesis and MF and at the organism level affects the interest for the new environment and the spatial memor

EFFECT OF THERMAL STRESS ON DANIO RERIO BEHAVIOUR: A PROTEOMIC STUDY TO UNDERSTAND THE MOLECULAR MECHANISM

Environmental temperature variations affect many properties and functions of biomolecules and structural components at the cellular level, and influence the animal physiology and behavior at the organism level. It is well accepted that the biochemical changes induced by temperature stress are attributed to gene expression modulations. The aim of this work was to understand the molecular mechanism and the behavioural responses correlated to temperature stress in zebrafish (Danio rerio), which is a widely used animal model for environmental genomics researches. Adult specimens of wild type zebrafish were kept at three different temperatures: 18\ub0C, 34\ub0C and 26\ub0C (used as a control) for 21 days and then subjected to behavioural tests using a YMaze task to evaluate the response to novelty and the spatial memory. Proteomic analysis were carried out on brains to evaluate the thermal effect at the central nervous system level. Briefly, for each temperature 9 brains were lysed and after reduction and derivatization, the proteins were digested with trypsin. LC-ESI MS/MS analysis was performed on a Dionex UltiMate 3000 HPLC. The eluate was electrosprayed into an LTQ Orbitrap Velos. Four technical replicate analyses of each sample were performed. Mass spectra were analyzed using MaxQuant and Peak Studio software. Finally, the bioinformatic analysis was carried out by DAVID and PANTHER softwares to evaluate enriched categories filtered for biological processes, molecular function (MF), cellular component (CC) and pathways involved. Preliminary results suggest that thermal stress at the cellular level influences the CC organization, biogenesis, structural morphogenesis and MF and at the organism level affects the interest for the new environment and the spatial memory

Environmental temperature variation affects brain protein expression and cognitive abilities in adult zebrafish (Danio rerio): A proteomic and behavioural study

Water temperature is an important environmental parameter influencing the distribution and the health of fishes and it plays a central role in ectothermic animals. The aim of this study is to determine the effects of environmental temperature on the brain proteome and the behavioural responses in zebrafish, a widely used animal model for environmental “omics” studies. Adult specimens of wild-type zebrafish were kept at 18 °C, 34 °C and 26 °C (control) for 21 days. Proteomic data revealed that several proteins involved in cytoskeletal organization, mitochondrial regulation and energy metabolism are differently regulated at the extreme temperatures. In particular, the expression of proteins associated to synapses and neurotransmitter release is down-regulated at 18 °C and 34 °C. In both thermal conditions, fish exhibited a reduced interest for the novel environment and an impairment of cognitive abilities during Y-Maze behavioural tests. The observed pathways of protein expression are possibly associated to functional alterations of the synaptic transmission that may result in cognitive functions impairment at central nervous system level as those revealed by behavioural tests. This study indicates that temperature variations can elicit biochemical changes that may affect fish health and behaviour. This combined approach provides insights into mechanisms supporting thermal acclimation and plasticity in fishes

Proteins and mirnas in feline renal amyloid deposits

Amyloidosis is a group of diseases occurring in humans and animals, due to the deposition of misfolding proteins in different organs. In humans, deposits were characterized using proteomics and more recently the role of miRNAs in the pathogenesis was proposed. In Abyssinian cat the main target is the kidney. Little is known on the mechanisms underlying the disease. The aim of this study is to profile proteins and miRNAs in healthy and affected Abyssinian cats, to evaluate their differential expression and clarify the pathogenesis mechanisms. Formalin-fixed paraffin-embedded kidney slices were collected from 7 affected and 5 healthy Abyssinians and used for proteomic and miRNAs analyzes. Peptides were analyzed with an LTQ-Orbitrap Velos mass spectrometer (MS). MS spectra were searched against the F.catus NCBI sequence database (release31.01.2017) by MaxQuant. Bioinformatic analysis was performed with DAVID and Panther softwares. MiRNAs were sequenced on the Illumina NextSeq500 platform. MiRDeep2 was used to map reads on the genome vs9.0, identify putative miRNAs, quantify their expression and identify homologous. Filtered proteins and miRNs statistically different were identified using a student t-test and a moderate t-test respectively (p-value 64.05). Part of the proteins was exclusively detected in the affected (n.175), part in the healthy (n.47) and part were common to the two groups (n.160). A fraction of the latter resulted upregulated (n.16) or down regulated (n.18) in affected compared to the healthy cats (p-value 64.05). Annotation and functional grouping suggested an effect on extracellular matrix and macromolecular complex subunit organization. MiRNA analysis detected 341 representatives, 22 differentially expressed between affected and healthy (p<.05). Six miRNAs out of 22 (four with a P-value<.009) are known to be involved in Alzheimer Disease. Interestingly, miR-26a-5p (P-value 0.120) is involved in the human immunoglobulin light chain amyloidosis onset. This study identified different miRNA and protein renal compositions in Abyssinian affected and healthy cats. The pathways involving these molecules are under investigation, providing new insights for the pathogenesis understanding