REGULATION OF THE CHOLINERGIC COTRANSPORTER--MOLECULAR ASPECTS

高亲和力的胆碱转运系统被认为是支持胆碱能传递的重要因素。这种运输者对钠(ChCoT)有强制性的要求，并与之协同运输。这种联合转运蛋白的一个重要特征是，转运蛋白的活动似乎可以调节神经元的活动。因此，ACh的调节在生理上具有重要意义。ChCoT的这种生理调节的确切分子机制还不是很清楚。我们提出了一个三部分的假说来解释ChCoT的这种生理调节。我们提出的假设的第一部分是，胆碱在胆碱能突触的转运速度的变化反映了质膜中活跃的ChCoT数量的平行变化。假设的第二部分是，ChCoT的数量是由含有ChCoT的载体囊泡的调节和构成循环决定的，ChCoT是末端(质膜)膜，是由含有ChCoT的载体囊泡在胞浆和质膜之间的胞液隔间的调节和构成循环决定的，受细胞内信使的影响。为了验证这三个部分的假设，我们将实现以下一组具体目标。1.建立循环ChCoT的囊泡性质，并表征参与该循环的分子机制。2.确定蛋白激酶C(PKC)参与ChCoT囊泡循环的分子机制。3.分离、纯化和鉴定作为可招募ChCoT池的细胞质来源的载体囊泡。

A high affinity choline transport system has been characterized as an essential element in supporting of cholinergic transmission. This transporter has a obligatory requirement for, and co-transports sodium (ChCoT). An important feature of this co-transporter is that the transport activity appears to adjust neuronal activity. The adjustment of ACh to be met and thus is physiologically highly significant. The precise molecular mechanisms responsible for this physiological regulation of the ChCoT are not well understood. We propose a three part hypothesis to explain this physiological regulation of the ChCoT. The first part of our proposed hypothesis is that the changes in the transport rates of choline at the cholinergic synapse reflect parallel changes in the number of active ChCoTs in the plasma membrane. The second part of the hypothesis is that the number of ChCoTs is the terminal (plasma) membrane in determined by the regulated and constitutive cycling of carrier vesicles containing ChCoTs is the terminal (plasma) membrane in determined by the regulated and constitutive cycling of carrier vesicles containing ChCoTs between a cytosolic compartment between the cytosol and the plasma membrane is under the influence of intracellular messengers. To test this three part hypothesis we will accomplish the following set of Specific Aims. 1. Establish the vesicular nature of the cycling ChCoT and characterize the molecular mechanisms involved in this cycling. 2. Determine the molecular mechanism of protein kinase C (PKC) involvement in the cycling of ChCoT vesicles. 3. Isolate,, purify and initiate the characterization of the carrier vesicle that serves as the cytoplasmic source of the recruitable pool of ChCoTs.