8-Aminoguanine and Its Actions on Renal Excretory Function

Abstract

Background: The endogenous purine 8-aminoguanine induces diuresis/natriuresis/glucosuria by inhibiting PNPase (purine nucleoside phosphorylase); however, mechanistic details are unknown. Methods: Here, we further explored in rats 8-aminoguanine’s effects on renal excretory function by combining studies using intravenous 8-aminoguanine, intrarenal artery infusions of PNPase substrates (inosine and guanosine), renal microdialysis, mass spectrometry, selective adenosine receptor ligands, adenosine receptor knockout rats, laser doppler blood flow analysis, cultured renal microvascular smooth muscle cells, HEK293 cells expressing A 2B receptors and homogeneous time resolved fluorescence assay for adenylyl cyclase activity. Results: Intravenous 8-aminoguanine caused diuresis/natriuresis/glucosuria and increased renal microdialysate levels of inosine and guanosine. Intrarenal inosine, but not guanosine, exerted diuretic/natriuretic/glucosuric effects. In 8-aminoguanine-pretreated rats, intrarenal inosine did not induce additional diuresis/natriuresis/glucosuria. 8-Aminoguanine did not induce diuresis/natriuresis/glucosuria in A 2B -receptor knockout rats, yet did so in A 1 - and A 2A -receptor knockout rats. Inosine’s effects on renal excretory function were abolished in A 2B knockout rats. Intrarenal BAY 60-6583 (A 2B agonist) induced diuresis/natriuresis/glucosuria and increased medullary blood flow. 8-Aminoguanine increased medullary blood flow, a response blocked by pharmacological inhibition of A 2B , but not A 2A , receptors. In HEK293 cells expressing A 2B receptors, inosine activated adenylyl cyclase, and this was abolished by MRS 1754 (A 2B antagonist). In renal microvascular smooth muscle cells, 8-aminoguanine and forodesine (PNPase inhibitor) increased inosine and 3’,5’-cAMP; however, in cells from A 2B knockout rats, 8-aminoguanine and forodesine did not augment 3’,5’-cAMP yet increased inosine. Conclusions: 8-Aminoguanine induces diuresis/natriuresis/glucosuria by increasing renal interstitial levels of inosine which, via A 2B receptor activation, increases renal excretory function, perhaps in part by increasing medullary blood flow.