A high-salt diet stimulates thick ascending limb eNOS expression by raising medullary osmolality and increasing release of endothelin-1

<jats:p>A high-salt diet increases renal endothelin (ET) production and thick ascending limb (THAL) endothelial nitric oxide synthase (eNOS) expression. ET stimulates THAL eNOS expression via ET<jats:sub>B</jats:sub>receptors. The tonicity of the renal medulla is highly variable, and hyperosmolality stimulates ET-1 synthesis by endothelial cells. We hypothesized that a high-salt diet raises medullary osmolality, increases ET release by the THAL, and thus enhances eNOS expression. Seven days of high salt (1% NaCl in drinking water) increased eNOS expression in THALs by 125 ± 31%. High salt increased outer medullary osmolality from 362 ± 13 to 423 ± 6 mosmol/kgH<jats:sub>2</jats:sub>O ( P < 0.05). Bosentan, a dual-ET receptor antagonist, blocked the increase in THAL eNOS expression caused by high salt (2.66 ± 0.44 absorbance units with bosentan vs. 5.15 ± 0.67 for vehicle; P < 0.05). Conscious systolic blood pressure did not differ between the two groups. In primary cultures of medullary THALs, raising osmolality from 300 to 350 and 400 mosmol/kgH<jats:sub>2</jats:sub>O using NaCl increased eNOS expression by 39 ± 11% ( P < 0.05) and 71 ± 16%, respectively ( P < 0.05). In primary cultures of THALs, raising osmolality from 300 to 400 mosmol/kgH<jats:sub>2</jats:sub>O for 1 h increased ET-1 release from 62 ± 7 to 113 ± 2 pg/mg protein ( P < 0.05). BQ-788, an ET<jats:sub>B</jats:sub>receptor antagonist (1 μM), blocked the stimulatory effect of 400 mosmol/kgH<jats:sub>2</jats:sub>O on eNOS expression (70 ± 13% vs. −5 ± 10%; paired difference, 74 ± 15%; P < 0.05). BQ-788 alone had no significant effect. We concluded that high salt stimulates THAL eNOS expression by increasing outer medullary osmolality, ET-1 release by the THAL and ET<jats:sub>B</jats:sub>receptor activation. This may be an important regulatory mechanism of THAL NaCl absorption when dietary salt intake is increased.</jats:p>