Effects of fear learning on early olfactory processing

描述（由申请人提供）：高阶（Li et al. 2008; Edeline & Weinberger 1991; Gdalyahu et al. 2012）和早期（Fletcher 2012）加工中心的感觉系统生理都可以通过情绪学习进行修改。感觉病理实际上是许多情感障碍的一个共同因素，从创伤后应激障碍的完全幻觉（Mueser & Butler 1987; Freeman & Fowler 2009）到注意力处理的更微妙的变化（Desseilles等人2009；Adenauer等人2010），这些变化可能会使传入的感觉信息偏向于焦虑障碍的威胁预测刺激（Beck & Clark 1997; Buckley 2000）。焦虑障碍通常在动物模型中使用联想恐惧条件反射范式进行研究，在该模型中，动物学习到中性感觉刺激，如气味（CS+），预测厌恶刺激的发生，如脚震。这些模型为研究感觉加工在情绪学习中的作用提供了一个独特的机会，在初步研究中，我们发现鉴别嗅觉恐惧条件反射会导致CS+诱发的嗅觉感觉神经元（嗅觉感觉神经元是大脑的主要（第一）感觉输入）的神经递质释放的惊人增强。这种增强可能有助于以后检测CS+或将注意力集中到类似的感觉刺激上，或者，也可能是适应不良并导致焦虑。为了进一步研究早期感觉加工与焦虑样状态之间的关系，本研究将对经历区别嗅觉恐惧条件反射的小鼠进行纵向体内生理和形态光学成像程序。在所有成熟的OSN轴突末端表达synaptophorin （spH）胞吐指示器的个体小鼠中，在基线、恐惧学习后和灭绝学习后，将通过植入背侧嗅球上方的双侧颅窗观察气味诱发的OSN向嗅球肾小球的突触输出（Bozza et al. 2004）。研究人员将在恐惧条件反射前后观察OSN轴突末端加载钙敏感染料的小鼠突触前嗅觉锁定Ca2+信号，以评估情绪学习如何改变CS+反应性OSN的气味诱发反应动力学。将对生理成像实验中确定的肾小球进行体内双光子形态计量学分析，以评估OSN肾小球神经支配的潜在结构可塑性（Jones et al. 2008）。我们的初步数据表明，CS+的增强表征是由嗅球回路的变化介导的，嗅球回路对传入的感觉信息进行了限制（即，情绪学习可能导致大门打开）。为了在经历恐惧条件反射的小鼠体内验证这一点，我们将通过药理操作突触前调节OSN活动的小叶内回路，电刺激嗅觉神经，并直接观察突触前抑制OSN轴突末端神经递质释放的gaba能中间神经元的气味诱发活动（在gad65能中间神经元中表达遗传编码的Ca2+指示剂GCamP3的小鼠中）。

DESCRIPTION (provided by applicant): Sensory system physiology in both higher order (Li et al. 2008; Edeline & Weinberger 1991; Gdalyahu et al. 2012) and early (Fletcher 2012) processing centers can be modified by emotional learning. Sensory pathologies are in fact a common element of many affective disorders, ranging from outright hallucinations in post traumatic stress disorder (Mueser & Butler 1987; Freeman & Fowler 2009) to more subtle changes in attentional processing (Desseilles et al. 2009; Adenauer et al. 2010) which may bias incoming sensory information to threat-predictive stimuli in anxiety disorders (Beck & Clark 1997; Buckley 2000). Anxiety disorders are routinely studied in animal models using associative fear conditioning paradigms, in which an animal learns that a neutral sensory stimulus, such as an odor (the CS+), predicts the occurrence of an aversive stimulus, such as a footshock. Such models provide a unique opportunity to investigate the role of sensory processing in emotional learning, and in preliminary studies we found that discriminative olfactory fear conditioning causes a surprising enhancement of CS+-evoked neurotransmitter release from olfactory sensory neurons (OSNs), the primary (first) sensory inputs to the brain. This enhancement may be beneficial in facilitating later detection of the CS+ or focusing attention towards similar sensory stimuli, or alternatively, may be maladaptive and contribute to anxiety. To further investigate the associations between early sensory processing and anxiety-like states, this study will perform longitudinal in vivo physiological and morphometric optical imaging procedures in mice that undergo discriminative olfactory fear conditioning. Odorant-evoked OSN synaptic output to olfactory bulb glomeruli will be visualized through a bilateral cranial window implanted above the dorsal olfactory bulbs at baseline, after fear learning, and again after extinction learning in individual mice expressing the synaptopHluorin (spH) exocytosis indicator in all mature OSN axon terminals (Bozza et al. 2004). Presynaptic sniff-locked Ca2+ signals in mice whose OSN axon terminals have been loaded with calcium-sensitive dye will be visualized before and after fear conditioning to assess how odorant-evoked response dynamics of CS+-responsive OSNs are modified by emotional learning. In vivo two-photon-based morphometric analyses will be performed on glomeruli identified during physiological imaging experiments to assess potential structural plasticity in OSN glomerular innervation (Jones et al. 2008). Our preliminary data suggest that the enhanced representation of the CS+ is mediated by changes in the olfactory bulb circuitry that gates incoming sensory information (i.e., emotional learning may cause the gate to open). To test this in vivo in mice that undergo fear conditioning, we will pharmacologically manipulate the intraglomerular circuit that presynaptically modulates OSN activity, electrically stimulate the olfactory nerve, and directly visualize odor- evoked activity n GABAergic interneurons that presynaptically inhibit neurotransmitter release from OSN axon terminals (in mice expressing the genetically-encoded Ca2+ indicator GCamP3 in GAD65-ergic interneurons).