Endosomal pH regulation in neurons: Roles of Na+/H+ exchangers

内吞作用是一种通过内吞体内膜结合细胞器内化膜成分和受体相关配体的机制。当配体解离时，大量受体通过“循环内小体”返回质膜。虽然已知内小体的酸性管腔pH在配体从受体解离及其降解中起重要作用，但我们对内小体再循环pH调节的了解有限，尤其是在神经元中。我们研究的长期目标是确定神经细胞内体pH的生物学作用。直到最近，研究人员才开始意识到，与非神经元细胞相比，神经元中的内小体在功能上更加多样化，蛋白质和脂类通过这些专门的内小体被分选到轴突和树突。然而，神经细胞中内体多样性是如何建立的仍不清楚。我们假设神经元比非神经元细胞含有更多的酸性内小体，这为选择性靶向轴突和树突的受体提供了基础。因此，内体pH可能通过神经递质及其受体增加对神经元通讯的新的机械洞察力。*我们将通过两种互补的方法来研究内体pH在神经元中的生物学作用，一种主要是生化的，另一种是在整个生物体水平上的。以Obj的身份。1，我们将研究内体pH在神经元分化所需的神经元信号和受体靶向中的作用。原代培养的大鼠神经元将用于我们实验室已经使用的生化分析，但需要进行一些修改。以Obj的身份。2，我们将利用果蝇基因敲除品系来探索神经元循环内小体的pH变化对动物行为的影响。果蝇之所以被使用，是因为强大的基因技术和敏感的行为测试。将讨论内体pH调节剂及其结合伙伴的生化相互作用的生理学意义。我们预计，我们的研究将极大地促进对神经元内体功能的基本理解，并具有跨认知科学的普遍适用性。我们在该领域的经验、UBC强大的合作环境、接触到优秀的学员以及真正令人兴奋的初步结果使我们能够成功地开展我们的研究计划。我相信，通过这项研究，我们将通过获取关键基础知识以及通过我们的计划培养熟练和高度上进的未来科学家，为加拿大社会做出贡献。我们的研究成果将发表在同行评议的期刊上，并在国际会议上发表。**

Endocytosis is a mechanism that internalizes membrane components and receptor-associated ligands through "endosomes", membrane-bound organelles. Upon dissociation of ligands, a large population of receptors returns to the plasma membrane via "recycling endosomes". While it is known that the acidic luminal pH of endosomes plays important roles in ligand-dissociation from the receptor and its degradation, our knowledge of recycling endosomal pH regulation, especially in neurons, is limited. The long-term goal of our research is to define biological roles of endosomal pH in neuronal cells. It is only very recently that researchers have begun to realize that endosomes are functionally more diverse in neurons than in non-neuronal cells, and proteins and lipids are sorted to axons and dendrites through those specialized endosomes. However, how the endosomal diversity is established in neuronal cells remains unknown. We hypothesize that neurons contain more acidic endosomes than non-neuronal cells, which provides a basis for preferential targeting of selected receptors to axons and dendrites. Thus, endosomal pH may add a new mechanistic insight into neuronal communication through the neurotransmitter and its receptor.*** We are going to investigate biological roles of endosomal pH in neurons by using two complementary approaches, one primarily biochemical, the second at the level of the whole organism. As Obj. 1, we will examine the role of endosomal pH in neuronal signaling and receptor targeting that are required for neuronal differentiation. Rat primary cultured neurons will be used for biochemical analyses already used in our laboratory with some modifications. As Obj. 2, we will explore the impact of pH changes in neuronal recycling endosomes on animal behavior using knock-out strains of fruit flies. Fruit flies are used because of the availability of powerful genetic techniques and sensitive behavioral tests. The physiological significance of biochemical interactions of endosomal pH-regulators and their binding partners will be addressed. We anticipate that our research will substantially advance basic understanding of endosomal function in neurons, with general applicability across cognitive science. Our experience in the field, the strong collaborative environment at UBC, access to outstanding trainees, and truly exciting preliminary results uniquely enable us to carry out our research program successfully. I am confident that through this research we will contribute to the Canadian society both through the acquisition of critical base knowledge as well as by training skilled and highly motivated future scientists through our program. The outcome of our research will be published in peer reviewed journals and presented at international conferences.**