Net K Secretion in Thick Ascending Limb of Mice on Low Na High K Diet

低钠高钾饮食的小鼠粗升肢的净钾分泌

• 批准号: 9049067
• 负责人: Bangchen Wang
• 金额: $ 3.17万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-17 至 2020-09-16
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9049067
• 关键词: Adrenalectomy; Affect; Aldosterone; Angiotensin II; Angiotensin II Receptor; Antihypertensive Agents; Apical; Back; Bumetanide; Cardiovascular Diseases; Cells; Chronic Kidney Failure; Diet; Distal; Diuresis; Diuretics; Drug usage; Effectiveness; Enzyme Inhibitor Drugs; Enzyme Inhibitors; Excretory function; Furosemide; Hypertension; Hypokalemia; Intake; Ion Transport; Kidney; Knockout Mice; Limb structure; Measures; Mediating; Mediator of activation protein; Mediterranean Diet; Membrane; Micropuncture; Mus; Na(+)-K(+)-Exchanging ATPase; Nephrons; Patch-Clamp Techniques; Patients; Potassium; Potassium Channel; Recycling; Regulation; Renin-Angiotensin-Aldosterone System; Role; Spectrometry; Structure of ascending limb of Henle' s loop; Thick; Time; Vegetarian diet; Western Blotting; apical membrane; basolateral membrane; dietary salt; driving force; hyperkalemia; inhibitor/antagonist; interstitial; prevent; public health relevance; receptor; salt intake; sodium-potassium chloride cotransporter 2 protein; tool; uptake; wasting

 DESCRIPTION (provided by applicant): Hyperkalemia is a common condition in chronic kidney disease (CKD) patients because of decreased ability of kidney to excrete K as well as the use of inhibitors of renin-angiotensin-aldosterone system. Paleolithic and Mediterranean diets have low Na, high K contents and are well known to be protective against cardiovascular diseases and CKD. However, it remains unclear how the kidneys handle such a high K load in the context of low Na intake. This is crucial to choosing effective diuretics and preventing the drastic consequences of hypokalemia and hyperkalemia for patients on these diets. For instance, our preliminary findings in this proposal suggest that the K-wasting loop diuretics may become K-sparing and decrease K excretion on low Na high K diet. In the thick ascending limb (TAL), Na+, K+, and Cl- are reabsorbed via Na+-K+-2Cl- cotransporter (NKCC2), and most of the K+ is recycled back into lumen by apical Renal Outer Medullary K channel (ROMK) to facilitate NaCl reabsorption by NKCC2. It has been long understood that the high interstitial K concentration induced by high K diet would inhibit the predominant basolateral K channels, leading to membrane depolarization and inhibition of Cl- exit, and thus inhibit NaCl reabsorption via NKCC2 in the TAL. However, our lab has recently discovered that in mice on a low Na, high K diet (LNaHK), there is a considerable furosemide-sensitive Na+ reabsorption as well as a net K+ secretion in the TAL. Contrary to the traditional view, we have also found that the NKCC2 expression is increased on LNaHK diet. We hypothesize that on LNaHK diet, apical ROMK is upregulated and becomes the predominant K channel which is not affected by high interstitial K+ concentration. NKCC2 is upregulated to supply Na+ to the basolateral Na+-K+ ATPase and bring in more K+ to be secreted. Unlike previous studies, we now have more advanced tools to further investigate the ion transport in TAL such as knockout mice. The hypothesis of this proposal is that the high levels of aldosterone (aldo) and angiotensin II (Ang II) induced by LNaHK diet increase NKCC2 and ROMK activities in the apical membrane of the TAL, thereby causing a net K+ secretion. Two specific aims will investigate the mechanisms that regulate net K secretion in TAL: Specific Aim 1. Determine the role and regulation of NKCC2 in the net K+ secretion in TAL. Specific Aim 2: Determine the role and regulation of ROMK in the net K+ secretion in TAL. We will determine the expression of NKCC2 and ROMK in TAL using western blot and real- time PCR in mice on LNaHK compared to control diet. We will measure the functional activity of NKCC2 and ROMK in TAL using micropuncture, atomic absorptive spectrometry, and patch clamp techniques. We will also perform adrenalectomy (ADX) and use aldosterone (aldo) replacement and inhibitors of renin-angiotensin- aldosterone system to elucidate the regulation of NKCC2 by aldo and Ang II. By fulfilling this proposal, we will have a better understanding of the renal K handling in people on the cardioprotective low Na high K diets.

 描述（由适用提供）：高钾血症是慢性肾脏疾病（CKD）患者的常见疾病，因为肾脏对极端K的能力降低以及使用肾素 - 血管紧张素 - 醛固酮系统的抑制剂。旧石器时代和地中海饮食的Na含量很低，K含量很高，并且众所周知可以保护心血管疾病和CKD。但是，尚不清楚肾脏在低Na摄入量的情况下如何处理如此高的K负载。这对于选择有效的利尿剂和防止患者对这些饮食中患者的低钾血症和高钾血症的剧烈后果至关重要。例如，我们在该提案中的初步发现表明，低Na高k饮食中的K浪费循环利尿剂可能会变成k的k左右，并减少k的排泄物。在较厚的上升肢（TAL）中，Na+，K+和Cl-通过Na+-k+-k+-2Cl-共转运蛋白（NKCC2）重新吸收，并且大多数K+通过顶端肾外内髓质K通道（ROMK）恢复为腔，以促进NACL通过NKCC2促进NACL的NACLIPTINP。长期以来，人们已经理解，高k饮食诱导的高间隙K浓度会抑制主要的基底外侧K通道，从而导致膜沉积和抑制cl-出口，从而抑制NACL通过TAL中NKCC2通过NKCC2的吸收。但是，我们的实验室最近发现，在低Na，高K饮食（LNAHK）的小鼠中，有一个考虑速尿敏感的Na+重吸收以及TAL中的K+分泌。与传统观点相反，我们还发现lnahk饮食中的NKCC2表达增加。我们假设在lnahk饮食上，顶端romk已更新，并成为不受高间隙K+浓度影响的主要K通道。 NKCC2已更新以向基底外侧Na+ -K+ ATPase提供NA+，并引入更多的K+分泌。与以前的研究不同，我们现在拥有更多的高级工具来进一步研究TAL中的离子运输，例如敲除小鼠。该提议的假设是，lnahk饮食诱导的高水平的醛固酮（Aldo）和血管紧张素II（ANG II）增加了TAL的顶膜中的NKCC2和ROMK活动，从而导致净K+分泌。两个具体的目的将调查在TAL中调节净k分泌的机制：特定目的1。确定NKCC2在TAL净K+分泌中的作用和调节。与对照饮食相比，我们将使用Western blot和实时PCR在TAL中确定NKCC2和ROMK在TAL中的表达。我们将使用微型函数，原子吸收光谱和斑块夹技术在TAL中测量NKCC2和ROMK的功能活性。我们还将进行肾上腺切除术（ADX）并使用醛固酮（ALDO）的替代品和肾素 - 血管紧张素 - 甲求化酮系统的抑制剂，以通过Aldo和Ang II阐明NKCC2的调节。通过履行这一建议，我们将更好地了解心脏保护低Na高K饮食中人们的肾脏K处理。