Compartmentalization of no in the Juxtaglomerular Apparatus (JGA)

Abstract

23 We previously reported that NO generated by the afferent arteriolar (Af-Art) endothelium caused dilatation and that NO produced by nNOS in the macula densa (MD) blunts tubuloglomerular feedback (TGF) via a cGMP-dependent mechanism. Here we hypothesized that a) NO is compartmentalized in the JGA, with endothelial NO only acting on vascular smooth muscle cells and MD NO acting within the MD, and b) all effects of MD NO are mediated by cGMP. We microperfused the Af-Art (at 60 mm Hg) with MD attached. MDs were perfused with solutions containing either 6 mM Na and 3 mM Cl (low NaCl) or 80 mM Na and 77 mM Cl (high NaCl). We first studied the effect of the soluble guanylate cyclase inhibitor LY83583 (1 μM) on TGF when added to the MD perfusate. Increasing the NaCl concentration in the MD perfusate decreased Af-Art diameter by 3.2 ± 0.5 μm. Adding LY83583 to the MD increased TGF response to 6.3 ± 1.1 μm (p<0.031 vs control; n=6). Inhibiting MD guanylate cyclase did not alter the effect of acetylcholine on the preconstricted Af-Art. Next we studied the effect of NO produced by the Af-Art endothelium on TGF. During the control period, the TGF response was 3.0 ± 0.9 μm. After LY83583 was added to the lumen of the Af-Art, TGF was 2.3 ± 0.8 μm (n.s.; n=6). This concentration completely blocked acetylcholine-induced dilatation. Removal of the Af-Art endothelium with an antibody against factor VIII-related antigen and complement did not alter TGF response. Before removing the endothelium, Af-Art diameter decreased from 18.6 ± 1.4 to 16.1 ± 1.1 μm; after removal, Af-Art diameter decreased from 19.1 ± 1.4 to 15.9 ± 1.1 μm (n=6). To determine whether the effect of NO in the MD is entirely mediated by cGMP, TGF was studied after giving a) LY83583 or b) LY83583 plus 7NI. In the presence of LY83583 alone, the TGF response was 3.8 ± 0.8 μm. In the presence of LY83583 plus 7NI, the TGF response was 4.2 ± 1.6 μm (n.s.; n=6). We concluded that: 1) NO is compartmentalized in the JGA, and therefore MD NO acts upon the MD itself rather than diffusing to the Af-Art; and 2) all of the effects of NO in the MD are mediated by cGMP.