Interactions of Human Organic Anion Transporters with Diuretics

Abstract

The tubular secretion of diuretics in the proximal tubule has been shown to be critical for the action of drugs. To elucidate the molecular mechanisms for the tubular excretion of diuretics, we have elucidated the interactions of human organic anion transporters (hOATs) with diuretics using cells stably expressing hOATs. Diuretics tested were thiazides, including chlorothiazide, cyclothiazide, hydrochlorothiazide, and trichlormethiazide; loop diuretics, including bumetanide, ethacrynic acid, and furosemide; and carbonic anhydrase inhibitors, including acetazolamide and methazolamide. These diuretics inhibited organic anion uptake mediated by hOAT1, hOAT2, hOAT3, and hOAT4 in a competitive manner. hOAT1 exhibited the highest affinity interactions for thiazides, whereas hOAT3 did those for loop diuretics. hOAT1, hOAT3, and hOAT4 but not hOAT2, mediated the uptake of bumetanide. hOAT3 and hOAT4, but not hOAT1 mediated the efflux of bumetanide. hOAT1 and hOAT3, but not hOAT2 and hOAT4 mediated the uptake of furosemide. In conclusion, it was suggested that hOAT1 may play an important role in the basolateral uptake of thiazides, and hOAT3 in the uptake of loop diuretics. In addition, it was also suggested that bumetanide taken up by hOAT3 and/or hOAT1 is excreted into the urine by hOAT4.

Overlapping in vitro and in vivo specificities of the organic anion transporters OAT1 and OAT3 for loop and thiazide diuretics

“…Furosemide and bendroflumethiazide inhibited tracer uptake with IC 50 values for mOat1 of 8 Ϯ 1 and 8 Ϯ 3 M, respectively (Fig. 1A), which were comparable with previously reported K i values for rat and human OAT1 (13,36). The IC 50 values for furosemide and bendroflumethiazide in mOat3 were 2.8 Ϯ 0.1 and 21 Ϯ 3 M, respectively (Fig.…”

“…The IC 50 values for furosemide and bendroflumethiazide in mOat3 were 2.8 Ϯ 0.1 and 21 Ϯ 3 M, respectively (Fig. 1B), also comparable with values previously obtained for human OAT3 in one prior study (13). Thus both furosemide and bendroflumethiazide inhibited tracer uptake by mOat1 and mOat3 in a concentration-dependent manner, consistent with their being substrates for these transporters.…”

“…Previous expression studies in Xenopus oocytes with rat OAT1 (36) or in mouse S2 cell lines with human OAT1 and human OAT3 (13) had indicated that, in vitro, both OAT1 and OAT3 can transport thiazides as well as loop diuretics, but the in vivo relevance of these transport mechanisms remained unclear. Because of the tremendous clinical importance of diuretic handling by the kidney, it is important to examine transport in vivo.…”

Multidrug Resistance–Associated Protein 4 Is Involved in the Urinary Excretion of Hydrochlorothiazide and Furosemide

“…Organic anion transporter 1 (OAT1/SLC22A6) and OAT3 (SLC22A8) have broad substrate specificities, and kinetic analyses using rat kidney slices have suggested that Oat1 mainly is responsible for the renal uptake of hydrophilic and small organic anions, whereas Oat3 is responsible for the uptake of more bulky organic anions (8 -10). Diuretics interact with OAT1 and OAT3, and, particularly, loop diuretics such as furosemide and bumetanide are substrates of OAT (11,12). Recently, Oat1 knockout mice were generated, and it was demonstrated that Oat1 was responsible for the renal uptake and pharmacologic action of furosemide (13).…”

Development and Validation of Fluorescence-Based and Automated Patch Clamp–Based Functional Assays for the Inward Rectifier Potassium Channel Kir4.1

“…[10][11][12] Loop and thiazide diuretics must first be secreted by renal proximal tubule cells before reaching their sites of action on the apical membrane. [13][14][15][16] Interactions between diuretics and other secreted drugs, including antibiotics, NSAIDs, antivirals, as well as organic acid in the setting of renal failure, can limit their secretion and natriuretic effects. Kir4.1/5.1 inhibitors that act directly on the basolateral membrane should circumvent this limitation.…”