Increased glomerular filtration rate (GFR), also known as glomerular hyperfiltration, is a common observation in the early stage of diabetes and is considered a key risk factor for the later development of diabetic kidney disease (DKD). Although the pathophysiological mechanisms of glomerular hyperfiltration in diabetes have not been fully understood, the tubuloglomerular feedback (TGF) has been implicated. In particular, the “tubular hypothesis” is now widely recognized. It describes that proximal tubular reabsorption of sodium and glucose via the sodium- glucose cotransporter 2 (SGLT2) is upregulated in the diabetic kidney, thereby reducing the NaCl delivery to the macula densa, which decreases the TGF activity and increases the single-nephron GFR in an effort to normalize NaCl excretion (SGLT2-NaCl-TGF pathway). We have recently identified a new and complementing mechanism that increases GFR in the diabetic kidney: the increased tubular glucose load at the macula densa is sensed through the apical sodium-glucose cotransporter 1 (SGLT1) and enhances the neuronal nitric oxide synthase (NOS1)-derived nitric oxide (NO) generation, which blunts TGF responsiveness and likewise increases the single-nephron GFR (SGLT1-NOS1-TGF pathway). SGLT2 inhibition is currently changing the perspectives of antidiabetic therapy with significant renoprotective effects. The benefits of SGLT2 inhibition in the diabetic kidney have been proposed to be largely independent of its blood glucose-lowering effect and mainly based on the central role of this transporter in the “tubular hypothesis” of diabetic glomerular hyperfiltration and nephropathy. In accordance with the “tubular hypothesis”, inhibition of SGLT2 increases the macula densa NaCl delivery and reduces diabetic hyperfiltration and thereby protects the diabetic kidney. However, inhibition of SGLT2 also increases glucose delivery to the macula densa, which is expected to decrease the TGF responsiveness through the macula densa SGLT1-NOS1-TGF pathway, and thereby limit both the GFR-lowering effect of SGLT2 inhibition and its protective effects on kidney. Therefore, in the present proposal, we will examine the effects of SGLT2 inhibition on the tubular glucose load at the macula densa, the macula densa NOS1/NO levels, and TGF responsiveness in diabetic mice, and then determine the significance of the macula densa SGLT1-NOS1 pathway in the SGLT2 inhibition-induced alterations in TGF and GFR by using mouse models with SGLT1 knockout or macula densa-specific NOS1 knockout; moreover, we will compare the effects of dual SGLT1/SGLT2 inhibition and selective SGLT2 inhibition on TGF activity, GFR and kidney injury in diabetic mice, and further determine the significance of the TGF mechanism in these effects by using a mouse model that lacks TGF due to macula densa-specific NKCC2 knockout. Successful completion of this proposal will establish the opposing role of the macula densa SGLT1-NOS1-TGF pathway in the protective e...