Investigating the role of Eph receptors and their molecular partners in anxiogenesis

Psychological stress leads to the enhancement of anxiety and can trigger a variety of psychiatric disorders. The mechanisms by which stress regulates anxiety are unclear. Eph receptors and their ligands, Ephrins, are attractive candidates to consider due to their multifaceted functions and high expression in the limbic system. Eph/Ephrins have been shown to regulate both functional and structural neuronal plasticity as well as hippocampus-dependent behaviour in mice. Thus, the aim of this study was to investigate the roles of Eph/Ephrins in the hippocampus and the amygdala and their interaction with molecular partners upon stress. First, I found that plasmin, a stress-related protease, cleaves EphA4 with high specificity. Mass spectrometry and bioinformatic analyses revealed the cleavage site is located within the fibronectin-like repeats of EphA4. EphA4, highly expressed in the hippocampus, interacts with EphrinB2. Following stress their interaction increases, as does the expression of EphrinB2. Studies in mice in which EphrinB2 was conditionally deleted in forebrain neurons demonstrated that EphrinB2 signalling is critical to the formation of anxiety-like behaviour. Furthermore, EphrinB2 mediates stress-related potentiation of contextual fear conditioning. These findings implicate the EphrinB2/EphA4/plasmin pathway as a new player in hippocampal regulation of anxiety. Second, I found that in the amygdala an extracellular serine protease, neuropsin, cleaves EphB2 shortly after stress. This molecular event alters EphB2 membrane expression and promotes a dynamic interaction of EphB2 with NMDA receptors. Consistent with the role of EphB2 in the stress response, bilateral amygdala infusion of anti-EphB2 antibody before stress prevents the development of stress-induced anxiety. The anxiolytic phenotype of neuropsin-deficient mice is rescued by the infusion of neuropsin into the amygdala before stress, confirming the effect of neuropsin is acute and not developmental, and pointing to the amygdala as the locus of the neuropsin’s effect. Taken together these findings implicate the acute, stress related cleavage of EphB2 by neuropsin in the amygdala as a key event in the development of anxiety-related behaviour.