R. V. Kozhemyakina

Genetic Architecture of Tameness in a Rat Model of Animal Domestication

A common feature of domestic animals is tameness—i.e., they tolerate and are unafraid of human presence and handling. To gain insight into the genetic basis of tameness and aggression, we studied an intercross between two lines of rats (Rattus norvegicus) selected over >60 generations for increased tameness and increased aggression against humans, respectively. We measured 45 traits, including tameness and aggression, anxiety-related traits, organ weights, and levels of serum components in >700 rats from an intercross population. Using 201 genetic markers, we identified two significant quantitative trait loci (QTL) for tameness. These loci overlap with QTL for adrenal gland weight and for anxiety-related traits and are part of a five-locus epistatic network influencing tameness. An additional QTL influences the occurrence of white coat spots, but shows no significant effect on tameness. The loci described here are important starting points for finding the genes that cause tameness in these rats and potentially in domestic animals in general.

Genetic influences on brain gene expression in rats selected for tameness and aggression

Interindividual differences in many behaviors are partly due to genetic differences, but the identification of the genes and variants that influence behavior remains challenging. Here, we studied an F2 intercross of two outbred lines of rats selected for tame and aggressive behavior toward humans for >64 generations. By using a mapping approach that is able to identify genetic loci segregating within the lines, we identified four times more loci influencing tameness and aggression than by an approach that assumes fixation of causative alleles, suggesting that many causative loci were not driven to fixation by the selection. We used RNA sequencing in 150 F2 animals to identify hundreds of loci that influence brain gene expression. Several of these loci colocalize with tameness loci and may reflect the same genetic variants. Through analyses of correlations between allele effects on behavior and gene expression, differential expression between the tame and aggressive rat selection lines, and correlations between gene expression and tameness in F2 animals, we identify the genes Gltscr2, Lgi4, Zfp40, and Slc17a7 as candidate contributors to the strikingly different behavior of the tame and aggressive animals.

Functional Characteristics of Serotonin 5-HT2A and 5-HT2C Receptors in the Brain and the Expression of the 5-HT2A and 5-HT2C Receptor Genes in Aggressive and Non-Aggressive Rats

The functional activity of serotonin 5-HT2A and 5-HT2C receptors and the expression of the genes encoding them were studied in Norway rats bred for 60 generations for the presence and absence of high levels of stress-evoked aggression to humans. There were no significant differences in the levels of 5-HT2A receptor mRNA in the midbrain, frontal cortex, and hippocampus and the extents of head twitching evoked by the 5-HT2A agonist DOI in rats with and without genetically determined high levels of aggression. Administration of the selective 5-HT2C agonist MK-212 weakened reflex startle in response to an acoustic signal (the acoustic startle response) in non-aggressive animals but had no significant effects on the response in aggressive animals. Increases in the level of 5-HT2C receptor mRNA were seen in the frontal cortex and hippocampus in non-aggressive rats as compared with aggressive animals. Increases in the expression of the 5-HT2C receptor gene and the functional state of 5-HT2C receptors were seen in the brains of non-aggressive rats, without any changes in the 5-HT2A receptor mRNA level or receptor sensitivity; this is evidence for the involvement of 5-HT2C receptors in the mechanisms inhibiting fear-evoked aggressive behavior.

Serotonin 5-HT1A receptor in infancy-onset aggression: Comparison with genetically defined aggression in adult rats

Antisocial aggressive behavior in adolescents represents growing clinical and social problem. Previously the implication of 5-HT1A receptor in the regulation of fear-induced aggression was shown. Here, the involvement of 5-HT1A receptor in infancy-onset genetically defined aggression was studied on Norway rats selectively bred for high level or for the lack of aggression toward man. The aggressive behavior and the expression of 5-HT1A receptor gene, 5-HT1A receptor density and functional activity were determined in infant (15-day-old) and adult rats. Considerable differences in aggressive response to man between infant rats of aggressive (A) and nonaggressive (NA) strains were found. In contrast to infant NA rats, A infants elicited marked aggressive response to handling, although its expression was less than in adult A rats. 5-HT1A receptor agonist 8-OH-DPAT (0.2 and 0.5mg/kg) decreased aggressive behavior in both A infant and adult rats. The desensitization of 5-HT1A receptors in the brain of A infant and adult rats was revealed. In contrast to decreased 5-HT1A gene expression in the midbrain of A adult rats, the 5-HT1A gene expression in the midbrain of infant rats did not differ between A and NA strains. There was no difference in 5-HT1A receptor density in infant rats. The data showed (1) the implication of 5-HT1A receptor in genetically defined infancy-onset fear-induced aggression, (2) the desensitization of 5-HT1A receptors as essential factor in infancy-onset aggression, and (3) the increased complexity of 5-HT-ergic control of aggressive behavior in adult rats with the involvement of 5-HT1A gene and the density of 5-HT1A receptors.

Brain-derived neurotrophic factor (BDNF) and its precursor (proBDNF) in genetically defined fear-induced aggression.

The brain-derived neurotrophic factor (BDNF), its precursor (proBDNF) and BDNF mRNA levels were studied in the brain of wild rats selectively bred for more than 70 generations for either high level or for the lack of affective aggressiveness towards man. Significant increase of BDNF mRNA level in the frontal cortex and increase of BDNF level in the hippocampus of aggressive rats was revealed. In the midbrain and hippocampus of aggressive rats proBDNF level was increased, whereas BDNF/proBDNF ratio was reduced suggesting the prevalence and increased influence of proBDNF in highly aggressive rats. In the frontal cortex, proBDNF level in aggressive rats was decreased. Thus, considerable structure-specific differences in BDNF and proBDNF levels as well as in BDNF gene expression between highly aggressive and nonaggressive rats were shown. The data suggested the implication of BDNF and its precursor proBDNF in the mechanism of aggressiveness and in the creation of either aggressive or nonaggressive phenotype.

The effects of prolonged selection for behavior on the stress response and activity of the reproductive system of male grey mice ( Rattus norvegicus )

Two Norway rat (Rattus norvegicus) populations contrasting in behavior have been raised at the Institute of Cytology and Genetics, Novosibirsk, Russia with long-term selection for the absence or enhancement of an aggressive response towards humans. They are designated as tame and aggressive, respectively. In this work we investigated the effects of the selection on behavior, stress responsiveness, and function of testes in males of the 78th generation. It is shown that the difference between the strains in their response to humans remains the same as in previous generations. However, the differences in stress response and rate of maturation contradict earlier data. For the first time, we note a higher glucocorticoid-mediated response to restriction stress and retarded sexual maturation in tame rats compared to aggressive ones, according to morphometric indices of gonads and testosterone levels. It is conceivable that the change in selection effects is determined by the disjunction of the directions of selection for behavior and the modification of the stress response. This study is the first to characterize males recently (six or seven generations of propagation in captivity) caught in the wild with regard to the indices under consideration and used as a control group. Wild rats have the highest stress response and rate of sexual maturation as compared to those selected.

Immune reactivity in rats selected for the enhancement or elimination of aggressiveness towards humans

This study analyzes immune reactivity in two lines of rats selected for the enhancement or elimination of aggressiveness toward humans. Compared to nonaggressive line, aggressive rats showed increased blood ratio of CD4(+) and CD8(+)T lymphocytes, monocyte chemoattractant protein (MCP)-1 level both before and after immunization with sheep red blood cells (SRBC), enhanced IgM-immune response, as well as decreased level of interleukin (IL)-1α before immunization. However, antigen administration produced IL-1α increase in aggressive rats and its decrease in nonaggressive rats compared to non-immunized rats of the same lines. In addition, line-dependent alterations of T lymphocyte distribution in response to immune activation have been found only in the spleen. It is suggested that genetic differences in aggressive behavior may contribute to differences in immune function.

[Expression of apoptosis genes in the brain of rats with genetically defined fear-induced aggression].

The programmed cell death (or apoptosis) plays an important role both in developing and mature brains. Multiple data indicate the involvement of processes of apoptosis in mechanisms of different psychopathologies. At the same time, nothing is known about the role of apoptosis in the regulation of genetically defined aggression. In the present work, the expression of the genes that encode main pro- and antiapoptotic BAX and BCL-XL proteins, as well as caspase 3 (the main effector of apoptosis), in different brain structures of rats that were selected on a high aggression towards human (or its absence) was studied. A significant increase in the expression of the gene encoding caspase 3 was detected in the hypothalamus. This was accompanied by a significant decrease in the expression of proapoptotic Bax gene in the hippocampus and increase in mRNA level of antiapoptotic Bcl-xl gene in the raphe nuclei area of midbrain in highly aggressive rats. An increase in the ratio Bcl-xl: Bax was found in the midbrain and amygdala; a trend towards an increase in the ratio was also found in hippocampus of aggressive animals compared to tame animals. Thus, we demonstrated that genetically defined fear-induced aggression is associated with significant changes in the genetic control of apoptosis in the brain. It is assumed that an increase in the Bcl-xl gene expression (accompanied by a decrease in the Bax gene expression) can indicate an increase in the threshold of neuronal apoptosis in highly aggressive rats.

5-HT1A receptor gene silencers Freud-1 and Freud-2 are differently expressed in the brain of rats with genetically determined high level of fear-induced aggression or its absence.

Serotonin 5-HT1A receptor is known to play a crucial role in the mechanisms of genetically defined aggression. In its turn, 5-HT1A receptor functional state is under control of multiple factors. Among others, transcriptional factors Freud-1 and Freud-2 are known to be involved in the repression of 5-HT1A receptor gene expression. However, implication of these factors in the regulation of behavior is unclear. Here, we investigated the expression of 5-HT1A receptor and silencers Freud-1 and Freud-2 in the brain of rats selectively bred for 85 generations for either high level of fear-induced aggression or its absence. It was shown that Freud-1 and Freud-2 levels were different in aggressive and nonaggressive animals. Freud-1 protein level was decreased in the hippocampus, whereas Freud-2 protein level was increased in the frontal cortex of highly aggressive rats. There no differences in 5-HT1A receptor gene expression were found in the brains of highly aggressive and nonaggressive rats. However, 5-HT1A receptor protein level was decreased in the midbrain and increased in the hippocampus of highly aggressive rats. These data showed the involvement of Freud-1 and Freud-2 in the regulation of genetically defined fear-induced aggression. However, these silencers do not affect transcription of the 5-HT1A receptor gene in the investigated rats. Our data indicate the implication of posttranscriptional rather than transcriptional regulation of 5-HT1A receptor functional state in the mechanisms of genetically determined aggressive behavior. On the other hand, the implication of other transcriptional regulators for 5-HT1A receptor gene in the mechanisms of genetically defined aggression could be suggested.

The influence of social environment in early life on the behavior, stress response, and reproductive system of adult male Norway rats selected for different attitudes to humans

The influence of social disturbance in early life on behavior, response of blood corticosterone level to restraint stress, and endocrine and morphometric indices of the testes was studied in 2-month Norway rat males from three populations: not selected for behavior (unselected), selected for against aggression to humans (tame), and selected for increased aggression to humans (aggressive). The experimental social disturbance included early weaning, daily replacement of cagemates from days 19 to 25, and subsequent housing in twos till the age of 2months. The social disturbance increased the latent period of aggressive behavior in the social interaction test in unselected males and reduced relative testis weights in comparison to the corresponding control groups. In addition, experimental unselected rats had smaller diameters of seminiferous tubules and lower blood testosterone levels. In the experimental group, tame rats had lower basal corticosterone levels, and aggressive animals had lower hormone levels after restraint stress in comparison to the control. The results suggest that the selection in two directions for attitude to humans modifies the response of male rats to social disturbance in early life. In this regard, the selected rat populations may be viewed as a model for investigation of (1) neuroendocrinal mechanisms responsible for the manifestation of aggression and (2) interaction of the hypothalamic-pituitary-adrenal and hypothalamic-pituitary-gonadal systems in stress.