Role of Neuroligin-2 and its Interacting Inhibitory Synapse Organizers in the Circuits of Fear Learning and Anxiety

Abstract

Neuroligin-2 (Nlgn2) is a synaptic adhesion molecule that promotes the maturation and function of inhibitory synapses. Lack of Nlgn2 in mice produced a reduction in inhibitory synaptic transmission in many brain regions. However, at the level of behavioral circuits, Nlgn2 knockout mice (Nlgn2 KO) exhibited phenotypes that were specifically related to the function of emotional circuits, such as anxiety-like behavior and cognitive impairments involving aversive emotional valence. Aiming to understand how Nlgn2 regulates the function and plasticity of emotional circuits, I characterized its role in fear learning using the Pavlovian fear conditioning (FC) paradigm. I observed a behavioral impairment measured at the level of short and long-term fear memory retrieval in Nlgn2 KO mice that was specific to auditory, but not contextual FC. Using immediate-early gene expression assay to probe neuronal activation during FC retrieval, I detected dysregulated activation in Nlgn2 KO mice in brain regions that are involved in fear memory processing such as the lateral amygdala (LA), the medial prefrontal cortex (mPFC) and the auditory cortex (AuC). In these regions, a lack of activation in response to FC was detected in Nlgn2 KO compared to WT mice, in line with impaired plasticity required for fear memory formation. Using cell-type specific gene knockout approach, I found that the FC deficit in Nlgn2 KO was caused by lack of Nlgn2 from specific inhibitory neurons subtypes rather than excitatory neurons. Particularly, deletion of Nlgn2 from vasoactive intestinal peptide (VIP) expressing inhibitory neurons, but not from Ca+2/Calmodulin-dependent protein kinase-II (CAMKII) expressing excitatory neurons, recapitulated the FC deficit in Nlgn2 KO. This result was also supported by learning-dependent overactivation of VIP interneurons (INs) in the LA during FC retrieval. While the reduction in fear learning in a mouse model with increased anxiety-like behavior is intriguing, I found that these two behaviors are controlled by effects of Nlgn2 on different cell types. Unlike what I observed in fear learning, deletion of Nlgn2 from excitatory CAMKII neurons fully recapitulated the anxiety-like behavior of Nlgn2 KO mice in the open field test. In contrast, mice lacking Nlgn2 from specific inhibitory neurons showed normal exploratory behavior using the same test. Moreover, the modulation of Nlgn2 function by its interaction partner IgSF9b was also distinct between fear learning and anxiety. Particularly, Nlgn2-IgSF9b double deletion exacerbated the fear learning deficit in Nlgn2 KO mice, in contrast to the normalization of anxiety-like behavior reported previously in the double KO mice. Taken together, my data reveals the complexity by which inhibitory synapse organizers regulate brain function in a circuit specific and a cell-type specific manner.