Mechanisms mediating osmoreception: Sustained response in the tilapia cell model

渗透调节是复杂生物体生命的基础。控制生命过程的分子的结构和功能由弱作用力(疏水相互作用和氢键)维持，因此对其离子和渗透环境的微小变化很敏感。为此，生物体投入相当大的能量来控制细胞内和细胞外的液体。例如，在鱼类中，渗透调节通常消耗总新陈代谢产出的25%-50%。除此之外，渗透平衡是一种脆弱的平衡，通过神经内分泌阵列的主要部分的持续相互作用来维持。事实上，临床医学中的一个长期困难的挑战是调节重症患者的盐和水平衡。考虑到渗透调节的成本和重要性，具有讽刺意味的是，人们对脊椎动物监测和控制盐分和水分平衡的机制知之甚少。然而，更仔细的观察揭示了障碍：渗透感受细胞和组织典型的复杂结构和排列。在几乎所有的情况下，人们不能同时测量细胞大小和渗透调节输出(激素释放)的变化，因为细胞可以明确地被区分为渗透感受细胞。在硬骨鱼中，催乳素(PRL)是一种重要的渗透压调节激素，维持淡水中的水矿物质平衡，其分泌直接受细胞外渗透压的控制。与这些作用相一致的是，淡水中的罗非鱼血浆催乳素水平升高，当细胞外渗透压下降时，催乳素的释放受到刺激。我们之前的NSF支持的研究表明，细胞外渗透压的下降导致水被动流入PRL细胞，从而在几分钟内增加细胞体积。这通过拉伸激活的通道启动细胞外钙离子的内流，产生细胞内钙[钙]i升高，从而刺激催乳素的释放。这一快速反应在体外持续刺激下延长，并在低渗介质中观察到24小时内PRL释放增加。另一方面，在高渗条件下，PRL的释放减少，反映了其对盐水渗透调节的抵消作用。然而，影响PRL持续释放和基因表达的渗透调节的信号转导机制尚不清楚。事实上，人们对任何渗透调节内分泌系统中传递急性或慢性渗透信息的细胞机制知之甚少。本工作旨在利用罗非鱼PRL细胞模型来表征长期暴露在环境渗透压生理变化中引起PRL释放和基因表达持续变化的细胞机制。这些研究将为调节渗透调节激素释放和产生的持续变化的细胞信号机制提供新的见解，这是目前其他模型系统无法获得的。这项工作将涉及从高中到研究生的多个层次的学生，包括几个代表性不足的少数民族，他们将在一个重要的研究领域接受培训，这将对细胞生理学产生影响。

Osmoregulation is fundamental to life in complex organisms. The structure and function of molecules that control the processes of life are maintained by weak forces (hydrophobic interactions and hydrogen bonds) and thus are sensitive to small changes in their ionic and osmotic environment. For this reason, organisms invest considerable energy in controlling intracellular and extracellular fluids. In fish, for example, osmoregulation typically consumes 25-50% of total metabolic output. Beyond this, osmotic equilibrium is a fragile balance that is maintained through the continuous interplay of a major portion of the neuroendocrine array. Indeed, a persistently difficult challenge in clinical medicine is the regulation of salt and water balance in seriously ill patients. In view of the cost and importance of osmoregulation, it may seem ironic that the mechanisms that monitor and control salt and water balance in vertebrates are so poorly understood. Closer attention, however, reveals the impediment: the typically complex structure and arrangement of osmoreceptive cells and tissues. In virtually all cases, one cannot simultaneously measure changes in cell size and the osmoregulatory output (hormone release) in a cell that can be distinguished specifically as osmoreceptive cells. In teleost fish, prolactin (PRL) is an essential osmoregulatory hormone that maintains hydromineral balance in fresh water and whose secretion is controlled directly by extracellular osmolality. Consistent with these actions, plasma PRL is elevated in the tilapia, Oreochromis mossambicus, a euryhaline teleost, in fresh water and PRL release is stimulated when extracellular osmolality declines. Our previous NSF-supported studies demonstrated that a fall in extracellular osmolality leads to the passive influx of water into the PRL cell that increases cell volume within minutes. This initiates an influx of extracellular Ca2+ through stretch-activated channels, producing a rise in intracellular calcium [Ca2+]i which stimulates PRL release. This rapid response is extended under sustained stimulation in vitro with increased PRL release being observed for 24 h in hyposmotic medium. On the other hand, PRL release is reduced under hyperosmotic conditions, reflecting its counteraction to salt water osmoregulation. However, signal transduction mechanisms that underlie the osmotic control of sustained PRL release and gene expression are not known. In fact, very little is known about the cellular mechanisms that transduce either acute or chronic osmotic information in any osmoregulatory endocrine system. The present work is aimed at characterizing cellular mechanisms by which extended exposure to physiological changes in ambient osmolality evoke sustained changes in PRL release and gene expression using the tilapia PRL cell model. These studies will provide new insight into the cell-signaling mechanisms that mediate the sustained alteration of the release and production of osmoregulatory hormones that at present cannot be obtained with other model systems. This work will involve students at many levels from high school to graduate students, including several underrepresented minorities, who will be trained in an important area of research, which will have an impact in cell physiology.