Increased intracellular pH at the macula densa activates nNOS during tubuloglomerular feedback.
Increased intracellular pH at the macula densa activates nNOS during tubuloglomerular feedback.
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DOI：
10.1111/j.1523-1755.2005.00282.x
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发表时间：
2005-05
影响因子：
19.6
通讯作者：
Ruisheng Liu;O. Carretero;Yilin Ren;J. Garvin
中科院分区：
文献类型：
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作者：
Ruisheng Liu;O. Carretero;Yilin Ren;J. Garvin
背景致密斑感觉到管状液中氯化钠浓度的增加，并通过称为管球反馈（TGF）的过程增加传入小动脉张力。一氧化氮（NO）的生产由致密斑神经元型一氧化氮合酶（nNOS）的增强，通过增加在致密斑管腔中的NaCl，和NO，从而形成抑制TGF。用阿米洛利阻断顶端Na（+）/H（+）交换增加TGF并模拟nNOS抑制的作用。我们假设增加致密斑内腔中的NaCl会升高致密斑细胞内pH（pH（i））并激活nNOS。方法取肾动脉升支和部分远端小管，行显微解剖和灌流。将致密斑灌注液从低（10 mmol/L）改变为高NaCl溶液（80 mmol/L）以模拟诱导TGF的条件。两种溶液的渗透压摩尔浓度均为180 mOsm，因此改变溶液不会改变细胞体积。结果当灌流液由低到高时，致密斑pH（i）由7.0 ± 0.5增加到7.8 ± 0.6（P < 0.05; N= 5）。当加入阿米洛利以抑制Na（+）/H（+）交换时，TGF过程中pH（i）的增加被阻断（N= 5）。增加鲁米那NaCl（N= 5）后，NO敏感染料的荧光强度增加了28.8 +/- 4.1%，表明NO产生增加。在Na（+）/H（+）交换抑制剂amiloride或nNOS抑制剂7-NI的存在下，通过将致密斑灌流液从低到高切换诱导的NO增加被钝化。为了研究pH（i）的变化是否能直接改变NO的产生，我们使用了一种K（+）/H（+）离子载体尼日利亚菌素来平衡腔内和细胞内的pH。当在低NaCl溶液中存在10（-5）mol/L尼日利亚菌素时，当致密斑的pH从7.3升高到7.8时，在致密斑中的γ-D-2的荧光增加了17.9 ± 1.3%（P < 0.01; N= 5）。在7-NI存在下，由升高pH（i）诱导的NO增加被阻断（N= 5）。结论TGF-β 1诱导致密斑pH（i）升高，pH（i）升高激活nNOS。
BACKGROUND The macula densa senses increasing NaCl concentrations in tubular fluid and increases afferent arteriole tone by a process known as tubuloglomerular feedback (TGF). Nitric oxide (NO) production by macula densa neuronal nitric oxide synthase (nNOS) is enhanced by increasing NaCl in the macula densa lumen, and the NO thus formed inhibits TGF. Blocking apical Na(+)/H(+) exchange with amiloride augments TGF and mimics the effect of nNOS inhibition. We hypothesized that increasing NaCl in the macula densa lumen raises macula densa intracellular pH (pH(i)) and activates nNOS. METHODS The thick ascending limb and a portion of the distal tubule with intact macula densa plaque adherent to the glomerulus were microdissected and perfused. Macula densa perfusate was changed from a low (10 mmol/L) to high NaCl solution (80 mmol/L) to mimic the conditions that induce TGF. Osmolality of both solutions was 180 mOsm, so that changing the solutions did not alter cell volume. RESULTS Macula densa pH(i) increased significantly from 7.0 +/- 0.5 to 7.8 +/- 0.6 when the perfusate was changed from low to high (P < 0.05; N= 5). When amiloride was added to inhibit Na(+)/H(+) exchange, the increase in pH(i) during TGF was blocked (N= 5). Fluorescence intensity of DAF-2, an NO-sensitive dye, increased by 28.8 +/- 4.1% after increasing luminal NaCl (N= 5), indicating an increase in NO production. In the presence of the Na(+)/H(+) exchanger inhibitor amiloride or the nNOS inhibitor 7-NI, the increase in NO induced by switching the macula densa perfusate from low to high was blunted. To study whether changes in pH(i) can directly alter NO production, we used nigericin, a K(+)/H(+) ionophore, to equilibrate luminal and intracellular pH. When macula densa pH was raised from 7.3 to 7.8 in the presence of 10(-5) mol/L nigericin in the low NaCl solution, fluorescence of DAF-2 in the macula densa increased by 17.9 +/- 1.3% (P < 0.01; N= 5). In the presence of 7-NI, the increase in NO induced by raising pH(i) was blocked (N= 5). CONCLUSION We concluded that macula densa pH(i) increases during TGF, and this increase in pH(i) activates nNos.