Physiological and environmental variables, or covariates, can account for an important portion of the variability\r\nobserved in behavioural/physiological results from different laboratories even when using the same type of animals\r\nand phenotyping procedures. We present the results of a behavioural study with a sample of 1456 genetically\r\nheterogeneous N/Nih-HS rats, including males and females, which are part of a larger genome-wide fine-mapping\r\nQTL (Quantitative Trait Loci) study. N/Nih-HS rats have been derived from 8 inbred strains and provide very small\r\ndistance between genetic recombinations, which makes them a unique tool for fine-mapping QTL studies. The\r\nbehavioural test battery comprised the elevated zero-maze test for anxiety, novel-cage (open-field like) activity,\r\ntwo-way active avoidance acquisition (related to conditioned anxiety) and context-conditioned freezing (i.e.\r\nclassically conditioned fear). Using factorial analyses of variance (ANOVAs) we aimed to analyse sex differences in\r\nanxiety and fear in this N/Nih-HS rat sample, as well as to assess the effects of (and interactions with) other\r\nindependent factors, such as batch, season, coat colour and experimenter. Body weight was taken as a quantitative\r\ncovariate and analysed by covariance analysis (ANCOVA). Obliquely-rotated factor analyses were also performed\r\nseparately for each sex, in order to evaluate associations among the most relevant variables from each behavioural\r\ntest and the common dimensions (i.e. factors) underlying the different behavioural responses. ANOVA analyses\r\nshowed a consistent pattern of sex effects, with females showing less signs of anxiety and fear than males across\r\nall tests. There were also significant main effects of batch, season, colour and experimenter on almost all\r\nbehavioural variables, as well as ââ?¬Å?sex Ã?â?? batchââ?¬Â, ââ?¬Å?sex Ã?â?? seasonââ?¬Â and ââ?¬Å?sex Ã?â?? experimenterââ?¬Â interactions. Body weight\r\nshowed significant effects in the ANCOVAs of most behavioural measures, but sex effects were still present in spite\r\nof (and after controlling for) these ââ?¬Å?body weightââ?¬Â effects. Factor analyses of relevant variables from each test\r\nshowed a two-fold factor structure in both sexes, with the first factor mainly representing anxiety and conditioned\r\nfear in males, while in females the first factor was dominated by loadings of activity measures. Thus, besides\r\nshowing consistent sex differences in anxiety-, fear- and activity-related responses in N/Nih-HS rats, the present\r\nstudy shows that femalesââ?¬â?¢ behaviour is predominantly influenced by activity while males are more influenced by\r\nanxiety. Moreover, the results point out that, besides ââ?¬Å?sexââ?¬Â effects, physiological variables such as colour and body weight, and environmental factors as batch/season or ââ?¬Å?experimenterââ?¬Â, have to be taken into account in both\r\nbehavioural and quantitative genetic studies because of their demonstrated influences on phenotypic outcomes.
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