Multiple studies suggest that interaction between the higher brain structures like cortex and hippocampus on one side, and a so called defensive circuitry (which includes amygdala, hypothalamus and periaqueductal grey), on the other side, determine fear/defensive behavior. Moreover, changes in such interaction may lead to mental illness.? ? We hypothesized that Brain Derived Neurotrophic Factor (BDNF) modulates such interaction by regulating plasticity in synapses between the higher brain structures and lower defensive circuit. To test this hypothesis and to identify mechanisms regulating fear/defensive responses, we study behavioral and related neuronal changes in mice with the conditional knockout of BDNF.? ? The goals of this study include:? 1) Identification of possible changes in fear/defensive behaviors in BDNF KO mice? 2) Determining molecular and neuronal mechanisms underlying such changes.? ? 1) Identification of possible changes in fear/defensive behaviors in BDNF KO mice? In the previous years we found that mice lacking BDNF in the CA3 area of hippocampus are abnormally aggressive towards their cage partners. During the last fiscal year, we characterized this home cage aggression by documenting frequency and age of its onset between paired littermates of either same or different genotypes. By the age 60 days, 90 % of paired knockouts, 60 % of pairs between wild type and knockout and only 10 % of the wild type pairs show aggressive behavior. In all pairs between knockout and wild type mice, knockout animal was the aggressor.? ? We are testing several behavioral hypotheses which might explain the home cage fighting phenotype: 1) increase in territorial aggression, 2) impaired social recognition memory leading, 3) failure to establish social hierarchy.? ? To test territorial aggression we used intruder test. Surprisingly, testing of isolated animals revealed no differences in territorial aggression between genotypes. This result suggests that aggressive behavior in BDNF knockouts is of a social origin due to possible abnormality in interactions between animals. It might originate from loss of social memory leading to a perception of another animal as an intruder. We excluded this possibility by confirming normal social recognition of ovariectomized female mice. To determine whether sensory cues from another animal were triggering heightened aggression in BDNF knockouts, we housed animals in cages with perforated partition that does not allow any fighting, but permits olfactory, visual and some tactile contact between neighboring animals. Such housing, in comparison with completely isolated conditions, did not alter aggressiveness of the control animals, but increased it in the knockouts, indicating that their excessive aggressive behavior is triggered by social experience. We continue investigation of specific cues triggering aggression and possible effects of BDNF knockout on formation of social hierarchy by looking at development of dominant/submissive behaviors.? ? 2) Finding neuronal mechanisms of increased aggressiveness in BDNF knockout mice? The major goal of our study is identification of neuronal circuits responsible for increase aggression. We are testing two potential hypotheses that may explain increased aggression: the lateral septum hypothesis and monoamine hypothesis. ? ? Since the knockout of BDNF is mostly restricted to hippocampal area CA3, we are looking at possible changes in connections between CA3 and other brain areas known to control aggression. The only such area that is directly connected to CA3 is lateral septum, which is known to suppress aggressive behaviors and which receives abundant projections from CA3. Thus our working hypothesis is that lack of BDNF secretion from CA3 cells might influence function of neurons in lateral septum that normally suppresses aggression. In the past year we found a reduction in BDNF level in the lateral septum of BDNF CA3restricted knockout mice. This finding indicates that BDNF is delivered form CA3 to lateral septum where it may act locally and suppress aggression. We were developing tool for testing this idea. We initiated collaboration with Dr. Reichardt lab in UCSF on production of mice allowing conditional knockout of BDNF receptor TrkB. In order to locate BDNF target responsible for control of aggressive behavior, we will ablate BDNF receptor TrkB in two candidate areas, lateral septum and hippocampal area CA3, using conditional knockout. To delete TrkB from lateral septum, we developed high titer lentivirus expressing Cre-recombinase that will be injected in the lateral septum of mice generated in Reichardts lab. To delete TrkB in hippocampal area CA3, we are using CA3-specific Cre-line generated by Dr. Nakazawa.? ? The monoamine hypothesis is based on previous studies showing that BDNF may influence levels of serotonin that, when decreased, may enhance aggressive behavior. To determine whether serotonin levels are altered in BDNF knockout mice, during the last fiscal year, we established a microdialysis/HPLC system for measuring levels of mono-amines in freely moving animals. Currently experiments on measuring levels of serotonin are on the way.