Fig. 2.

ENaC and the ET_B receptor are necessary for shear stress-mediated nitrite production. Shear stress (low = 3 dyn/cm² or high = 30 dyn/cm²) or static conditions (0 dyn/cm²) applied to mouse inner medullary CD segment 3 cells (mIMCD-3) for 1 h. A: pretreatment (30 min) of cells with the ENaC inhibitor benzamil (0.2 μM) blunted the shear stress-mediated increase in mIMCD-3 nitrite under high shear stress (n = 4–8, P = 0.025). Benzamil did not prevent the shear stress-mediated increase in nitrite under low shear stress, and there was no significant effect under static conditions (n = 4–8, P > 0.5). *Represents a significant increase compared with the corresponding static condition. †Represents a significant difference from the corresponding low shear stress group. ‡Represents significant difference between vehicle and benzamil groups with high shear. B: pretreatment for 30 min with the ET_B antagonist BQ788 (1 μM), but not the ET_A antagonist BQ123 (1 μM), reduced nitrite production in static conditions and blunted the high shear stress increase in nitrite production. ET_B receptor blockade and inhibition of nitric oxide synthase (NOS)1 with 1 μM VNIO abolished the shear stress-mediated increase in nitrite (n = 5, *P < 0.05 compared with vehicle static, †P < 0.05 compared with corresponding vehicle). C: NOS inhibitors were given for 30 min before sample collection. Inhibition of NOS2 by 100 nM 1400W had no significant effect on static or high shear stress-mediated nitrite production; however, NOS1 inhibition by 1 μM VNIO significantly blunted static and shear stress-mediated NO production. Inhibition of total NOS by 1 mM l-NAME significantly reduced nitrite production in static and high shear conditions (n = 5–11, P < 0.05). *Represents a significant increase compared with the corresponding static condition. †Represents a significant difference from the corresponding vehicle group. D: % contribution of NOS1, NOS2, and NOS3 on nitrite production under static and shear stress conditions.