Lmo4 in the Basolateral Complex of the Amygdala Modulates Fear Learning

Pavlovian fear conditioning is an associative learning paradigm in which mice learn to associate a neutral conditioned stimulus with an aversive unconditioned stimulus. In this study, we demonstrate a novel role for the transcriptional regulator Lmo4 in fear learning. LMO4 is predominantly expressed in pyramidal projection neurons of the basolateral complex of the amygdala (BLC). Mice heterozygous for a genetrap insertion in the Lmo4 locus (Lmo4gt/+), which express 50% less Lmo4 than their wild type (WT) counterparts display enhanced freezing to both the context and the cue in which they received the aversive stimulus. Small-hairpin RNA-mediated knockdown of Lmo4 in the BLC, but not the dentate gyrus region of the hippocampus recapitulated this enhanced conditioning phenotype, suggesting an adult- and brain region-specific role for Lmo4 in fear learning. Immunohistochemical analyses revealed an increase in the number of c-Fos positive puncta in the BLC of Lmo4gt/+ mice in comparison to their WT counterparts after fear conditioning. Lastly, we measured anxiety-like behavior in Lmo4gt/+ mice and in mice with BLC-specific downregulation of Lmo4 using the elevated plus maze, open field, and light/dark box tests. Global or BLC-specific knockdown of Lmo4 did not significantly affect anxiety-like behavior. These results suggest a selective role for LMO4 in the BLC in modulating learned but not unlearned fear.

巴甫洛夫恐惧条件化（Pavlovian fear conditioning）是一类经典的联想学习范式，在此范式中，小鼠能够建立中性条件刺激与厌恶性非条件刺激之间的关联。本研究揭示了转录调控因子Lmo4在恐惧学习中的全新功能。Lmo4主要在杏仁核基底外侧复合体（basolateral complex of the amygdala, BLC）的锥体投射神经元中表达。携带Lmo4基因座基因陷阱插入的杂合小鼠（Lmo4gt/+），其Lmo4表达水平较野生型（wild type, WT）小鼠降低50%；这类小鼠在对应刺激的环境与线索中均表现出增强的僵立行为。通过短发夹RNA（small-hairpin RNA, shRNA）在BLC中敲低Lmo4，而非海马齿状回区域，可重现这一增强的条件化表型，这提示Lmo4在恐惧学习中发挥成年阶段及脑区特异性的调控作用。免疫组织化学分析显示，恐惧条件化后，Lmo4gt/+小鼠BLC内的c-Fos阳性颗粒数量显著高于野生型小鼠。最后，本研究通过高架十字迷宫、旷场实验及明暗箱实验，检测了Lmo4gt/+小鼠以及BLC特异性Lmo4敲低小鼠的类焦虑行为。结果表明，全身性或BLC特异性的Lmo4敲低并未对类焦虑行为产生显著影响。综上，上述结果提示LMO4在BLC中选择性调控习得性恐惧，而非非习得性恐惧。