Neonatal Proximal Tubule Acidification

描述(申请人提供)：新生儿比成人更容易出现液体和电解质紊乱，部分原因是肾脏不成熟。结果表明，新生儿近端小管氯化钠和碳酸氢钠的转运速度和特点与成人有显著差异。近端小管对氯化钠和碳酸氢钠的重吸收在很大程度上依赖于管腔Na+/H+交换器，在成年人中是NHE3。新生儿尽管有大量的Na+/H+交换活性，但几乎没有顶膜NHE3。我们提供的证据表明，新生儿的Na+/H+亚型与成人不同。我们发现，与主要表达NHE3和少量NHE8的成人相比，新生儿近端小管的刷状缘膜表达丰富的NHE8和少量的NHE3。然而，我们对NHE8的功能特征、它的调控以及是什么导致NHE8在出生后发育过程中向NHE3的异构体切换知之甚少。这项建议的目的是研究NHE8在体外和体内的功能。我们将在全细胞贴片结构上使用高度敏感的pH选择电极技术，在那里我们可以完全控制细胞外和细胞内环境，以表征NHE8的功能。我们将使用一种研究肾脏发育的新动物模型-肾上腺切除-甲状腺功能减退大鼠来研究出生后发育过程中导致NHE8亚型转换的因素，在该模型中，近端小管Na+/H+交换和NHE3的成熟几乎停止在新生水平。我们已经证明，糖皮质激素和甲状腺激素会增加NHE3的水平。切除肾上腺-甲状腺功能减退的大鼠将允许我们测试这一假设，即新生儿相对甲状腺功能低下、糖皮质激素缺乏状态阻止NHE3的表达，使另一种具有不同调节特性的异构体成为新生儿近端小管酸化所必需的。我们将研究NHE8是否被糖皮质激素和甲状腺激素以相反的方向调节，从而导致成人的下调。最后，我们现在有了一只NHE8缺失的小鼠，它将使我们能够检查这种发育中的NHE亚型在肾脏酸化中的相对功能重要性，以及它在新生儿和成人酸中毒反应中的相对重要性。 与公共卫生相关：新生儿更容易因腹泻等可能危及生命的疾病而发生酸中毒和容量耗竭。这种对酸中毒的易感性在很大程度上是因为新生儿的肾脏不太发达，在某些情况下，使用不同于成年人的转运蛋白进行电解质运输。这项提议将描述一种可能在新生儿肾脏酸化和盐分转运中发挥重要作用的新生儿转运蛋白。

DESCRIPTION (provided by applicant): Neonates are more prone to fluid and electrolyte disorders than adults due, in part, to the immature kidney. We have shown that there are marked differences between the rate and characteristics of neonatal proximal tubule NaCl and NaHCO3 transport than that in the adult. Proximal tubule reabsorption of NaCl and NaHCO3 is dependent in large part on the luminal Na+/H+ exchanger, which in adults is NHE3. Neonates have almost no apical membrane NHE3 despite having a significant amount of Na+/H+ exchanger activity. We provide evidence that the neonate has a different Na+/H+ isoform than the adult. We show that the brush border membrane of the neonatal proximal tubule expresses abundant NHE8 and little NHE3, compared to the adult that primarily expresses NHE3 and little NHE8. However, we know very little about the functional characteristics of NHE8, its regulation and what causes the isoform switch during postnatal development to NHE3. The aims of this proposal are to characterize the function of NHE8 in vitro and in vivo. We will use a highly sensitive technique of pH selective electrodes on the whole-cell patch configuration where we have total control of the extra and intracellular milieu to characterize the function of NHE8. We will examine the factors responsible for the isoform switch from NHE8 during postnatal development using a novel animal model to study renal development, the adrenalectomized-hypothyroid rat, where the maturation of proximal tubule Na+/H+ exchange and NHE3 is nearly arrested at the neonatal level. We have shown that NHE3 levels are increased by the glucocorticoids and thyroid hormone. The adrenalectomized-hypothyroid rat will allow us to test the hypothesis that the relative hypothyroid, glucocorticoid deficient state of the neonate prevents the expression of NHE3 making another isoform with different regulatory properties necessary for neonatal proximal tubule acidification. We will examine if NHE8 is regulated in the opposite direction by glucocorticoids and thyroid hormone resulting in down regulation in the adult. Finally, we now have a NHE8 null mouse that will allow us to examine the relative functional importance of this developmental NHE isoform in renal acidification and its relative importance in response to acidosis in both neonates and adults. PUBLIC HEALTH RELEVANCE: Neonates are more prone to develop acidosis and volume depletion from diseases such as diarrhea that can be life threatening. This susceptibility to acidosis is in large part due to the fact that the newborn kidney is less well developed and in some cases uses different transport proteins for electrolyte transport than adults. This proposal will characterize a neonatal transporter that likely plays an important role in neonatal kidney acidification and salt transport.