A quantitative trait locus on chromosome 1 modulates intermale aggression in mice.

Aggression between male conspecifics is a complex social behavior that is likely modulated by multiple gene variants. In this study, the BXD recombinant inbred mouse strains (RIS) were used to map quantitative trait loci (QTLs) underlying behaviors associated with intermale aggression. Four hundred and fifty-seven males from 55 strains (including the parentals) were observed at an age of 13 ± 1 week in a resident-intruder test following 10 days of isolation. Attack latency was measured directly within a 10-minute time period and the test was repeated 24 hours later. The variables we analyzed were the proportion of attacking males in a given strain as well as the attack latency (on days 1 and 2, and both days combined). On day 1, 29% of males attacked, and this increased to 37% on day 2. Large strain differences were obtained for all measures of aggression, indicating substantial heritability (intraclass correlations 0.10-0.18). We identified a significant QTL on chromosome (Chr) 1 and suggestive QTLs on mouse Chrs 1 and 12 for both attack and latency variables. The significant Chr 1 locus maps to a gene-sparse region between 82 and 88.5 Mb with the C57BL/6J allele increasing aggression and explaining about 18% of the variance. The most likely candidate gene modulating this trait is Htr2b which encodes the serotonin 2B receptor and has been implicated in aggressive and impulsive behavior in mice, humans and other species.

Functional implications of decreases in neurogenesis following chronic mild stress in mice.

Numerous data from human and animal studies suggest that hippocampal plasticity might be a key element in depression. However, the connection remains loose at best and further data are needed. Human studies are of necessity limited, but animal models can help providing further insight. Unpredictable chronic mild stress (UCMS) is a commonly used model because it mimics depression-like phenotypes satisfactorily. Its rationale is based on the underlying stress-induced difficulties found in many depressed patients. We therefore studied learning and hippocampal neurogenesis in mice from three different inbred strains subjected to UCMS. Learning was assessed in different hippocampus-dependent and independent tasks. The rate of survival of newly generated brain cells was determined in behaviorally-naive animals. Results demonstrated a dramatic reduction of surviving new brain cells in both the hippocampus and the subventricular zone of UCMS-treated animals. This reduction was observed both for neurons and for other cells of the hippocampus. Behavioral data demonstrated an impairment of hippocampus-dependent learning, whereas hippocampus-independent learning was spared. However, the specific results were strongly dependent on strain and sex so that there does not appear to be a direct causative relationship between the deficits in neurogenesis and behavior.

A quantitative-genetic analysis of hippocampal variation in the mouse.

This report analyses the genetic underpinnings of the proportions of the hippocampal terminal fields in the mouse at the midseptotemporal level. We used 5 inbred strains and all possible F(1) crosses between them (diallel cross). Broad heritabilities ranged from 11 to 53%. Additive genetic variation was present for all phenotypes analyzed. Directional dominance was found for the relative size of the suprapyramidal mossy fiber terminal field only. For the stratum lacunosum-moleculare, ambidirectional dominance emerged. These findings suggest that, in evolutionary history, directional selection has operated for a proportionally large suprapyramidal terminal field. For all other hippocampal variables (viz. the relative sizes for the strata oriens, pyramidale, radiatum, lacunosum-moleculare, CA4, intra- and infrapyramidal mossy fiber terminal field and the absolute size of the regio inferior) past stabilizing selection was inferred.

QTL and systems genetics analysis of mouse grooming and behavioral responses to novelty in an open field.

The open field is a classic test used to assess exploratory behavior, anxiety and locomotor activity in rodents. Here, we mapped quantitative trait loci (QTLs) underlying behaviors displayed in an open field, using a panel of 53 BXD recombinant inbred mouse strains with deep replication (10 per strain and sex). The use of these strains permits the integration and comparison of data obtained in different laboratories, and also offers the possibility to study trait covariance by exploiting powerful bioinformatics tools and resources. We quantified behavioral traits during 20-min test sessions including (1) percent time spent and distance traveled near the wall (thigmotaxis), (2) leaning against the wall, (3) rearing, (4) jumping, (5) grooming duration, (6) grooming frequency, (7) locomotion and (8) defecation. All traits exhibit moderate heritability making them amenable to genetic analysis. We identified a significant QTL on chromosome M.m. 4 at approximately 104 Mb that modulates grooming duration in both males and females (likelihood ratio statistic values of approximately 18, explaining 25% and 14% of the variance, respectively) and a suggestive QTL modulating locomotion that maps to the same locus. Bioinformatic analysis indicates Disabled 1 (Dab1, a key protein in the reelin signaling pathway) as a particularly strong candidate gene modulating these behaviors. We also found 2 highly suggestive QTLs for a sex by strain interaction for grooming duration on chromosomes 13 and 17. In addition, we identified a pairwise epistatic interaction between loci on chromosomes 12 at 36-37 Mb and 14 at 34-36 Mb that influences rearing frequency in males.

Systems genetic analysis of hippocampal neuroanatomy and spatial learning in mice.

Variation in hippocampal neuroanatomy correlates well with spatial learning ability in mice. Here, we have studied both hippocampal neuroanatomy and behavior in 53 isogenic BXD recombinant strains derived from C57BL/6J and DBA/2J parents. A combination of experimental, neuroinformatic and systems genetics methods was used to test the genetic bases of variation and covariation among traits. Data were collected on seven hippocampal subregions in CA3 and CA4 after testing spatial memory in an eight-arm radial maze task. Quantitative trait loci were identified for hippocampal structure, including the areas of the intra- and infrapyramidal mossy fibers (IIPMFs), stratum radiatum and stratum pyramidale, and for a spatial learning parameter, error rate. We identified multiple loci and gene variants linked to either structural differences or behavior. Gpc4 and Tenm2 are strong candidate genes that may modulate IIPMF areas. Analysis of gene expression networks and trait correlations highlight several processes influencing morphometrical variation and spatial learning.

A note on the effect of within-strain sample sizes on QTL mapping in recombinant inbred strain studies.

This note explores the effect of within-strain sample sizes on the correlations between a phenotype and a molecular-genetic marker in a battery of inbred strains. It is shown that the maximum correlation possible between a molecular marker and a behavioral or neuronal phenotype equals the additive-genetic correlation. How close the strain correlation will approach the additive-genetic correlation depends only on heritability and within-strain sample sizes. The equations derived can be used to optimize designs of studies attempting to localize Quantitative Trait Loci utilizing Recombinant Inbred Strains, provided information about the heritability of the character under study is available.

Hippocampal mossy fibers and radial-maze learning in the mouse: a correlation with spatial working memory but not with non-spatial reference memory.

One hundred and eight male mice from nine different inbred strains were tested for two aspects of learning in an eight-arm radial maze. In the first experimental arrangement of the maze, measuring spatial working memory, clear strain differences were found on the fifth day of training. Furthermore, this type of learning showed a high positive correlation with the size of the intra- and infrapyramidal hippocampal mossy fiber terminal field as revealed with Timm's staining. In the second experiment, in which non-spatial reference memory was tested, significant strain differences were found for the learning variables, but there were no significant covariations with the sizes of the intra- and infrapyramidal mossy fiber terminal fields. These results, combined with previous data, suggest that heritable variations of the hippocampal intra- and infrapyramidal mossy fiber projection influence processes determining spatial learning capabilities in mice.

Genetic dissection of mouse exploratory behaviour.

A large variety of apparatus and procedures are being employed to measure mouse exploratory behaviour. Definitions of what constitutes exploration also vary widely. The present article reviews two studies whose results permit a genetic dissection of behaviour displayed in an open-field situation. The results agree that factors representing exploration and stress/fear underlie this type of behaviour. Both factors appear to be linked to neuroanatomical variation in the sizes of the hippocampal intra- and infrapyramidal mossy fibre terminal fields. Multivariate analysis of genetic correlations may render important insights into the structure of behaviour and its relations with neuroanatomical and neurophysiological systems.

Spatial and non-spatial spontaneous alternation and hippocampal mossy fibre distribution in nine inbred mouse strains.

Ten male mice from each one of nine inbred strains were tested for spontaneous alternation in a T-maze, which was placed in a spatially richly structured room. Each test consisted of two trials, the first choice to be made was forced, the second one free. By turning the maze 180 degrees between choices during 8 of the 16 tests, 3 variables could be measured: total alternation, spatial alternation, and non-spatial alternation. After Timm's staining, the sizes of the intra- and infrapyramidal mossy fibre terminal fields (iip-MF) were measured. Significant strain differences were found for all variables, but none of the behavioural variables correlated with hippocampal variation. These results disagree with earlier experiments, where large correlations between the iip-MF and spatial working memory were found. Some hypotheses to explain this discrepancy are presented.

Correlations between radial-maze learning and structural variations of septum and hippocampus in rodents.

Large, but non-pathological, individual differences in neuroanatomy of the brain exist in rodents, which have been shown to covary with behavioral traits. In the present review, we explore the relationship between variations in the extent of the intra- and infrapyramidal mossy fiber projection of the hippocampus and spatial and non-spatial learning capacities in mice and rats. Preliminary data concerning anatomical variation in the septo-hippocampal cholinergic system and its consequences for individual behavior are also presented. We conclude that the hippocampal intra- and infrapyramidal mossy fiber projection is intimately involved in the regulation of spatial, but not of non-spatial learning capabilities. Although lesion studies have shown that a well-functioning cholinergic system is a prerequisite for performance in spatial learning tasks, our preliminary data suggest that individual differences in the cholinergic system do not explain individual differences in learning.

Hippocampal mossy fiber distribution covaries with open-field habituation in the mouse.

We studied the sizes of the hippocampal intra- and infrapyramidal mossy fiber (iip-MF) terminal fields and habituation to a new environment (open-field) in 25 genetically different groups of mice. Based on previous findings and theoretical considerations, a positive relationship between the size of the iip-MF terminal fields and the extent of behavioral change between two subsequent exposures to the open-field was expected. In fact, such a relationship was revealed by a factor analysis. Our results indicate that mice possessing large iip-MF terminal fields are more efficient in the processing of spatial information.

Further phenotypical characterisation of two substrains of C57BL/6J inbred mice differing by a spontaneous single-gene mutation.

Males from two substrains of C57BL/6J mice, which have been found to differ for open-field exploration, radial-maze learning, and the sizes of their hippocampal intra- and infra-pyramidal mossy fibre (IIPMF) terminal fields, were compared for offensive aggression, thermoregulatory nest-building, and their behaviour in the Light-Dark choice test. The substrain with the smaller IIPMF showed higher aggression and more nest-building behavior than the one with the larger IIPMF, whereas only tentative differences were found in the Light Dark choice test. These findings confirm and expand on previously found genetic links between the IIPMF and behaviours in mice. These substrains provide a powerful tool to localise the gene involved and subsequently investigate the pathway leading from gene to behaviour.

No correlations between spatial and non-spatial reference memory in a T-maze task and hippocampal mossy fibre distribution in the mouse.

Two reference memory tasks were tested in a T-maze, which was placed in a spatially richly structured environment and turned 180 degrees between trials following a semi-random schedule. Male mice from 9 different inbred strains were either trained always to go to the same place (spatial task) or always to make the same turn (non-spatial task). Animals were subsequently processed for Timm's stain and the sizes of their intra- and infrahippocampal mossy fibre terminal fields (ipp-MF) were measured. Significant strain differences were found for this variable and in both learning tasks, but learning and hippocampal variation did not correlate. This disagrees with earlier findings in a radial maze, where significant correlations between the iipMF and spatial reference memory were obtained. Two hypotheses are brought forward to explain this discrepancy. First, in radial mazes (multiple choices) different memory capabilities might be used than in T-mazes (only two choices). Second, a considerable amount of stress appeared to be present in our subjects, possibly induced by the large size of the T-maze. This might have interfered negatively with acquisition. Further experiments will be needed to test these hypotheses.

Behavioural and neuroanatomical divergence between two sublines of C57BL/6J inbred mice.

Male mice of strains C57BL/6J and DBA/2J differ reproducibly in a number of behaviours displayed in an open-field. In particular, C57BL/6J mice show a higher rearing-up frequency than do DBA/2J animals. Recently, a marked drop occurred in the frequency with which this behaviour is displayed by the C57BL/6J//Nmg (N) subline, that has been separated from the original C57BL/6J line over 62 generations. Comparison of our animals with the C57BL/6J//Kun(K) subline, separated from the Jackson parent over at least 40 generations, showed a significant strain difference for rearing. Since both a positive additive-genetic correlation between rearing and the size of the intra- and infrapyramidal mossy fibre terminal field (iipMF) and a correlated response in the size of the iipMF to selection for rearing have been found previously, we expected to find smaller iipMF in N, as compared with K. After processing for Timm's stain, this predicted difference was indeed found. Skin grafting demonstrated that the two sublines were still completely histocompatible, excluding a possible genetic contamination of N. This provides very strong support for the hypothesis that both the behavioural and the neuroanatomical differences between these sublines are caused by a single spontaneous mutation in the N line and strengthens the idea of a functional relationship between the structural and the behavioural variable.

Behavioral responses to novelty and structural variation of the hippocampus in mice. II. Multivariate genetic analysis.

On the basis of results from lesion studies in rodents, covariations are expected to exist between naturally-occurring heritable variations in hippocampal morphology and exploratory behavior elicited by novel surroundings. For this reason, we set up a full diallel cross between five inbred mouse strains and analyzed the behavioral and the hippocampal anatomical variation in male animals from this cross. Employing a bivariate extension of the diallel-cross analysis, estimates were obtained for the phenotypical, environmental, and genetical correlations between the phenotypes studied. A factor analysis performed on the matrix of additive-genetic correlations revealed that variations in the size of the intra- and infrapyramidal mossy fiber terminal fields (iip-MF) are negatively related to open-field exploration and novelty-induced fear. These results indicate that having larger iip-MF projections promotes the collection and processing of information about a novel environment, entailing lower levels of exploration and fear.

Behavioral responses to novelty and structural variation of the hippocampus in mice. I. Quantitative-genetic analysis of behavior in the open-field.

As a first step towards a multivariate quantitative-genetic analysis of covariations between heritable variation in hippocampal structure and mouse behavior, a univariate analysis of the genetic architecture of behavioral responses to novelty is presented. For several components of exploratory behavior considerable amounts of genetic variation were found and an evolutionary history of stabilizing selection for intermediate levels of exploration was inferred. Comparison of these results with those from a previous study indicated that even a relatively small diallel cross, involving 4-5 inbred strains, may provide useful genetic information on a specific sample of animals. Larger numbers of strains are needed to provide precise estimates of genetic parameters in a population.

A genetic-correlational study of hippocampal structural variation and variation in exploratory activities of mice.

Our previous work provided evidence that hippocampal opioid peptides form an important neurochemical substrate underlying the gene-dependent exploratory behavior of mice. A prominent hippocampal opioid is dynorphin B, which resides in the mossy fibers exclusively. In order to seek support for causal relationships between dynorphinergic hippocampal mechanisms and exploration, a quantitative-genetic method was chosen. For this purpose, mice from the inbred strains C57BL/6, DBA/2, BLN, and CPB-K were used. Their hippocampal mossy fiber projections were visualized by means of immunohistochemistry, using a highly specific anti-dynorphin B antiserum. The additive-genetic correlations that were estimated suggest pleiotropic gene effects on locomotion, rearing-up, wall-leaning, and several intra- and infrapyramidal mossy fiber (iipMF) variables. Long iipMF, in particular, were found to be associated with high exploratory activity.

A genetic-correlational study of hippocampal neurochemical variation and variation in exploratory activities of mice.

Previously, we have demonstrated that hippocampal mossy fibers, containing the opioid peptide dynorphin B, are functionally connected with the gene-dependent exploratory behavior of mice. In order to seek further evidence of causal relationships between dynorphin B action and exploration, a quantitative-genetic method was chosen. For this purpose, mice from the inbred strains C57BL/6, DBA/2, BLN, and CPB-K were used. By means of radioimmunoassay, the hippocampal level of dynorphin B was monitored in mice that had been exposed to environmental novelty, as compared to naive animals. Clear evidence was obtained that novelty induces the release of hippocampal dynorphin B. Furthermore, low tissue content was found to be causally connected with high exploratory scores.

Neuroanatomical divergence between two substrains of C57BL/6J inbred mice entails differential radial-maze learning.

Compared to the parental strain C57BL/6J, male mice from the mutated substrain C57BL/6JNmg show smaller hippocampal intra- and infrapyramidal mossy fiber projections and a correlated inability to master a simple spatial radial-maze task. Possibly, these two substrains differ for only one single gene, making them a valuable model to investigate the physiological pathways leading from genotype to neurobehavioral phenotype.

Radial-maze performance and structural variation of the hippocampus in mice: a correlation with mossy fibre distribution.

Twenty-four male mice, belonging to 8 different inbred strains, were tested in an 8-arm radial maze. Clear strain differences were found for performance on the third day of training, which correlated very strongly with the size of the hippocampal intra- and infrapy ramidal mossy fibre (iip-MF) terminal fields. These results, combined with those from earlier experiments, indicate that genetic variations of the iip-MF projection influence processes that determine behavioural abilities of mice.