Low-dose L-NAME induces salt sensitivity associated with sustained increased blood volume and sodium-chloride cotransporter activity in rodents.

To investigate the cause of salt sensitivity in a normotensive animal model, we treated rats with a low-dose of the nitric oxide synthase inhibitor, L-NAME, that does not elevate blood pressure per se or induce kidney fibrosis. A high salt diet increased the circulating blood volume both in L-NAME-treated and nontreated animals for the first 24 hours. Thereafter, the blood volume increase persisted only in the L-NAME-treated rats. Blood pressure was higher in the L-NAME-treated group from the start of high salt diet exposure. Within the first 24 hours of salt loading, the L-NAME treated animals failed to show vasodilation and maintained high systemic vascular resistance in response to blood volume expansion. After four weeks on the high salt diet, the slope of the pressure-natriuresis curve was blunted in the L-NAME-treated group. An increase in natriuresis was observed after treatment with hydrochlorothiazide, but not amiloride, a change observed in parallel with increased phosphorylated sodium-chloride cotransporter (NCC). In contrast, a change in blood pressure was not observed in L-NAME-treated NCC-deficient mice fed a high salt diet. Moreover, direct L-NAME-induced NCC activation was demonstrated in cells of the mouse distal convoluted tubule. The vasodilatator, sodium nitroprusside, downregulated phosphorylated NCC expression. The effect of L-NAME on phosphorylated NCC was blocked by both the SPAK inhibitor STOCK2S-26016 and the superoxide dismutase mimetic TEMPO which also attenuated salt-induced hypertension. These results suggest that the initiation of salt sensitivity in normotensive rodents could be due to hyporeactivity of the vasculature and that maintaining blood pressure could result in a high circulating volume due to inappropriate NCC activity in the low-dose L-NAME model. Thus, even slightly impaired nitric oxide production may be important in salt sensitivity regulation in healthy rodents.

A rapid method for measuring serum oxidized albumin in a rat model of proteinuria and hypertension.

Oxidative stress is a risk for and cause of various disease, however, measurements of oxidative stress are either time-consuming or non-specific. Here, we established a rapid method of using high performance liquid chromatography (HPLC) to measure serum oxidized albumin in a rat model. We optimized HPLC conditions for rat oxidized albumin. To validate our method, three-week-old male Sprague-Dawley rats were uninephrectomized and treated normal diet, high salt diet or high salt diet with Tempol, a superoxide dismutase (SOD) mimetic. After 4 weeks of treatment, we analyzed serum oxidized albumin. The main findings are listed as below. (i) Our method of oxidized albumin measurement only takes 16 minutes, with an intra-day and inter-day deviation within 1% and a detection limit concentration of 6.4 mg/ml. (ii) Oxidized albumin levels were significantly higher in the high salt diet group than in the normal salt diet group, and this effect was reversed by Tempol. (iii) Oxidized albumin levels also correlated with urinary protein and 8-isoprostane levels. In conclusion, we have established a simple method for evaluating rat serum oxidized albumin using HPLC. Our method is rapid and has an advantage over conventional methods and may be useful for animal models of oxidative stress.