Diabetes mellitus is the most common and rapidly growing cause of end-stage renal disease in developed countries. A classic hallmark of early diabetes mellitus includes activation of the renin-angiotensin system (RAS), which may lead to hypertension and renal tissue injury, but the mechanism of RAS activation is elusive. Here we identified a paracrine signaling pathway in the kidney in which high levels of glucose directly triggered the release of the prohypertensive hormone renin. The signaling cascade involved the local accumulation of succinate and activation of the kidney-specific G protein-coupled metabolic receptor, GPR91, in the glomerular endothelium as observed in rat, mouse, and rabbit kidney sections. Elements of signal transduction included endothelial Ca 2+ , the production of NO and prostaglandin (PGE 2 ), and their paracrine actions on adjacent renin-producing cells. This GPR91 signaling cascade may serve to modulate kidney function and help remove metabolic waste products through renal hyperfiltration, and it could also link metabolic diseases, such as diabetes, or metabolic syndrome with RAS overactivation, systemic hypertension, and organ injury.

Abstract-In addition to the juxtaglomerular apparatus, renin is also synthesized in renal tubular epithelium, including the collecting duct (CD). Angiotensin (Ang) II differentially regulates the synthesis of juxtaglomerular (inhibition) and CD (stimulation) renin. Because diabetes mellitus, a disease with high intrarenal renin-Ang system and Ang II activity, is characterized by high prorenin levels, we hypothesized that the CD is the major source of prorenin in diabetes. Renin granular content was visualized using in vivo multiphoton microscopy of the kidney in diabetic Munich-Wistar rats. Diabetes caused a 3.5-fold increase in CD renin, in contrast to less pronounced juxtaglomerular changes. Ang II type 1 receptor blockade with Olmesartan reduced CD renin to control levels but significantly increased juxtaglomerular renin. Using a fluorogenic renin assay, the prorenin component of CD renin content was measured by assessing the difference in enzymatic activity of medullary homogenates before and after trypsin activation of prorenin. Trypsinization caused no change in control renin activity but a 5-fold increase in diabetes. Studies on a CD cell line (M1) showed a 22-fold increase in renin activity after trypsinization and a further 35-fold increase with Ang II treatment. Therefore, prorenin significantly contributes to baseline CD renin. Diabetes, possibly via Ang II, greatly stimulates CD prorenin and causes hyperplasia of renin-producing connecting segments. These novel findings suggest that, in a rat model of diabetes, prorenin content and release from the CD may be more important than the juxtaglomerular apparatus in contrast to the existing paradigm. (Hypertension. 2008;51:1597-1604.)

Multiphoton fluorescence microscopy offers the advantages of deep optical sectioning of living tissue with minimal phototoxicity and high optical resolution. More importantly, dynamic processes and multiple functions of an intact organ can be visualized in real time using noninvasive methods, and quantified. These studies aimed to extend existing methods of multiphoton fluorescence imaging to directly observe and quantify basic physiological parameters of the kidney including glomerular filtration rate (GFR) and permeability, blood flow, urinary concentration/dilution, renin content and release, as well as more integrated and complex functions like the tubuloglomerular feedback (TGF)-mediated oscillations in glomerular filtration and tubular flow. Streptozotocin-induced diabetes significantly increased single-nephron GFR (SNGFR) from 32.4 +/- 0.4 to 59.5 +/- 2.5 nl/min and glomerular permeability to a 70-kDa fluorophore approximately eightfold. The loop diuretic furosemide 2-fold diluted and increased approximately 10-fold the volume of distal tubular fluid, while also causing the release of 20% of juxtaglomerular renin content. Significantly higher speeds of individual red blood cells were measured in intraglomerular capillaries (16.7 +/- 0.4 mm/s) compared with peritubular vessels (4.7 +/- 0.2 mm/s). Regular periods of glomerular contraction-relaxation were observed, resulting in oscillations of filtration and tubular flow rate. Oscillations in proximal and distal tubular flow showed similar cycle times ( approximately 45 s) to glomerular filtration, with a delay of approximately 5-10 and 25-30 s, respectively. These innovative technologies provide the most complex, immediate, and dynamic portrayal of renal function, clearly depicting the components and mechanisms involved in normal physiology and pathophysiology.

Macula densa (MD) cells of the juxtaglomerular apparatus (JGA) are salt sensors and generate paracrine signals that control renal blood flow, glomerular filtration, and release of the prohypertensive hormone renin. We hypothesized that the recently identified succinate receptor GPR91 is present in MD cells and regulates renin release. Using immunohistochemistry, we identified GPR91 in the apical plasma membrane of MD cells. Treatment of MD cells with succinate activated mitogen-activated protein kinases (MAPKs; p38 and extracellular signal-regulated kinases 1/2) and cyclooxygenase 2 (COX-2) and induced the synthesis and release of prostaglandin E 2 , a potent vasodilator and classic paracrine mediator of renin release. Using microperfused JGA and real-time confocal fluorescence imaging of quinacrinelabeled renin granules, we detected significant renin release in response to tubular succinate (EC 50 350 M). Genetic deletion of GPR91 (GPR91 Ϫ/Ϫ mice) or pharmacologic inhibition of MAPK or COX-2 blocked succinate-induced renin release. Streptozotocin-induced diabetes caused GPR91-dependent upregulation of renal cortical phospho-p38, extracellular signal-regulated kinases 1/2, COX-2, and renin content. Salt depletion for 1 wk increased plasma renin activity seven-fold in wild-type mice but only 3.4-fold in GPR91 Ϫ/Ϫ mice. In summary, MD cells can sense alterations in local tissue metabolism via accumulation of tubular succinate and GPR91 signaling, which involves the activation of MAPKs, COX-2, and the release of prostaglandin E 2 . This mechanism may be integral in the regulation of renin release and activation of the renin-angiotensin system in health and disease.

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