Electric Studies of Excitation, Secretion & Contraction

描述（由申请人提供）：松果体是大脑中的一个内分泌器官，主要受交感神经系统释放的去甲肾上腺素调节。它是我们生物钟系统的一部分，将夜间荷尔蒙褪黑激素广播到全身。松果体细胞的夜间状态伴随着相对于白天状态的基因表达的广泛变化。许多表达的基因是由姐妹谱系光感受器表达的，除了哺乳动物的松果体细胞不具有光敏性。我们的总体假设是松果体细胞在两种非常不同的电生理兴奋性之间切换 有助于调节褪黑激素的分泌以及昼夜节律的夹带和维持的状态。使用生物物理技术，培养的大鼠松果体细胞的昼夜状态将在其离子通道补体，钙信号动力学，受体激活，第二信使水平和分泌机制方面进行对比。通过在适当的神经递质中预孵育培养的原代细胞来实现状态变化。将从大鼠松果体中分离细胞，置于细胞培养物中，用去甲肾上腺素处理以模拟夜晚，并在显微镜下使用全细胞千兆密封记录、钙敏感染料光度测定、神经递质电流测定法和HPLC以及使用第二信使的各种活细胞指示剂等技术进行研究。夜间松果体细胞被假设为具有与光感受器相关的电兴奋性，并且白天松果体细胞被假设为相对静止。的假设，5-羟色胺分泌的松果体细胞的量子胞吐，而褪黑激素和N-乙酰5-羟色胺分泌的疏水扩散也将被测试。了解松果体的节律性分泌机制将对治疗睡眠障碍，短日照的季节性情感反应以及因时差和轮班工作而引起的注意力丧失做出重要贡献。

DESCRIPTION (provided by applicant): The pineal gland is an endocrine organ in the brain that is primarily regulated by noradrenaline released by the sympathetic nervous system. It is the part of our circadian clock system that broadcasts the night hormone, melatonin, to all of the body. The night state of pinealocytes is accompanied by extensive changes in gene expression relative to the day state. Many expressed genes are those expressed by a sister lineage, photoreceptors, except that mammalian pinealocytes are not photosensitive. Our overall hypothesis is that pinealocytes switch between two very different electrophysiological excitability states that contribute to regulated secretion of melatonin and to entrainment and maintenance of circadian rhythms. Using biophysical techniques, the night and day states of cultured rat pinealocytes will be contrasted in terms of their ion channel complement, calcium signal dynamics, receptor activation, second messenger levels, and secretory mechanisms. The state changes will be achieved by preincubating cultured primary cells in appropriate neurotransmitters. Cells will be dissociated from rat pineal glands, placed in cell culture, treate with norepinephrine to mimic night, and studied under the microscope with techniques such as whole-cell gigaseal recording, photometry of calcium-sensitive dyes, amperometry and HPLC of neurotransmitters, and use of various live-cell indicators for second messengers. Night pinealocytes are hypothesized to have electrical excitability related to that of photoreceptors, and day pinealocytes are hypothesized to be relatively quiescent. The hypothesis that serotonin is secreted from pinealocytes by quantal exocytosis whereas melatonin and N-acetyl serotonin are secreted by hydrophobic diffusion will also be tested. Understanding the rhythmic secretory mechanisms of the pineal will make an important contribution towards treating sleep disorders, seasonal affective responses to short days, and loss of attention due to jet lag and shift work.