Molecular and cellular analysis of accessory olfactory circuits in mice

在小鼠中，犁鼻器（VNO）是用于检测信息素的主要感觉器官，信息素是 影响社会行为，包括：领土，性识别和产妇护理。VNO可以分为 分为不同性质的顶层和底层。VNO感觉神经元（VSNs）在顶层 表达G蛋白偶联受体（GPCR）的Vmn 1 r家族成员，而基础VSN表达 GPCR的Vmn 2 r家族的成员。Vmn 2 r属于GPCR的相同家族，其包括 代谢型谷氨酸、GABAB和味觉受体。基底VSN不服从每个神经元一个受体的原则 这一规则在嗅觉系统中的大多数其他感觉神经元中起作用。Vmn 2 r系列由四个 指定为A、B、C和D类。A类、B类和D类的成员彼此关系更密切 而不是C类的七个成员（Vmn 2 r1 -7）。每个基底VSN表达一个Vmn 2 rC和一个Vmn 2 rC。 Vmn2rABD。因此，7种C类受体在VNO中广泛表达，而ABD类受体在VNO中广泛表达。 像其他嗅觉受体一样，这种受体的表达很少。的功能意义 Vmn 2 rs的共表达还不清楚。我们假设Vmn 2 rCs和Vmn 2 rABDs形成 异二聚体，并且这种相互作用改变了基础VSN的功能特性。Vmn 2 rCs可能会影响 Vmn 2 rABD的亚细胞定位、其配体结合和信号传导特性。 为了检验这些假设，我们设计了一个多管齐下的策略，包括分子和 生化研究、神经解剖学检查和行为分析。我们已经生成了一条鼠标线 使用CRISPR/Cas9介导的染色体工程携带Vmn 2 r1 -7基因簇的缺失 然后是Cre重组。此外，我们产生了一系列针对所有Vmn 2 rC的特异性抗体， 针对特定的C类受体，以及针对Vmn 2 rAs的进化枝。有了这些新试剂，我们可以 解决这些假设。我们的研究将揭示基础VSNs功能的机制。 更广泛地说，这些实验将揭示嗅觉系统如何介导社会行为， 对物种的生存至关重要。到目前为止，只有少数Vmn 2 r配体已被确定。既然我们 预测Vmn 2 rC簇的缺失将影响动物正确响应的能力。 Vmn 2 rABD配体，我们已经产生的簇敲除线将是非常宝贵的评估新的 vmn 2 r配体和它们的行为重要性，因为这些配体被鉴定。这样，我们的研究将 加深我们对以多种方式控制社会行为的理解。最后，我们的实验将 为GPCR家族功能的分子机制提供了额外的见解， vmn 2 rs属于。这些受体中的一些与人类疾病有关。这个项目将为塔里克 坎诺尼尔，最近布朗毕业于代表性不足的少数民族，一个独特的培训机会，将 为他申请研究生院继续教育奠定了坚实的基础。

In mice, the vomeronasal organ (VNO) is a main sensory organ for detecting pheromones, chemicals that affect social behaviors including: territoriality, sexual recognition and maternal care. The VNO can be divided into apical and basal layers that differ in their properties. VNO sensory neurons (VSNs) in the apical layer express members of the Vmn1r family of G protein coupled receptors (GPCRs), while basal VSNs express members of the Vmn2r family of GPCRs. Vmn2rs belong to the same family of GPCRs that includes the metabotropic glutamate, GABAB, and taste receptors. Basal VSNs do not obey the one receptor per neuron rule that is operative in most other sensory neurons in the olfactory system. The Vmn2r family consists of four classes designated A, B, C and D. Members of classes A, B and D are more closely related to one another than to the seven members of class C (Vmn2r1-7). Each basal VSN expresses one Vmn2rC and one Vmn2rABD. Consequently, The seven class C receptors are broadly expressed in the VNO while class ABD receptors are sparsely expressed, like the rest of the olfactory receptors. The functional significance of the coexpression of Vmn2rs is not well understood. We hypothesize that Vmn2rCs and Vmn2rABDs form heterodimers and that this interaction alters the functional properties of the basal VSNs. Vmn2rCs might affect the subcellular localization of Vmn2rABDs, their ligand binding and signaling properties. To examine these hypotheses, we have devised a multipronged strategy encompassing molecular and biochemical studies, neuroanatomical examination, and behavioral analysis. We have generated a mouse line carrying a deletion of the Vmn2r1-7 gene cluster using CRISPR/Cas9-mediated chromosome engineering followed by Cre recombination. Further, we generated a battery of specific antibodies against all Vmn2rCs, against specific class C receptors, and against a clade of Vmn2rAs. With these new reagents, we are poised to address these hypotheses. Our studies will reveal the mechanisms underlying the function of basal VSNs. More broadly, these experiments will shed light on how the olfactory system mediates social behaviors that are critical for the survival of the species. Thus far, only a handful of Vmn2r ligands have been identified. Since we predict that the deletion of the Vmn2rC cluster will affect the ability of the animals to properly respond to Vmn2rABD ligands, the cluster knockout line that we have generated will be invaluable for evaluating new Vmn2r ligands and their behavioral importance, as these ligands are identified. In this manner, our studies will deepen our understanding of the control of social behavior in multiple ways. Finally, our experiments will provide additional insight into the molecular mechanisms underlying the function of the GPCR family to which Vmn2rs belong. Some of these receptors have relevance to human disease. This project will offer Tariq Cannonier, a recent Brown graduate from an under-represented minority, a unique training opportunity that will provide him a solid foundation for applying to graduate school for his continuing education.