Tubuloglomerular Feedback Response Research Articles

Purinergic Receptor Antagonists: A Complementary Treatment for Hypertension.

The treatment of hypertension has improved in the last century; attention has been directed to restoring several altered pathophysiological mechanisms. However, regardless of the current treatments, it is difficult to control blood pressure. Uncontrolled hypertension is responsible for several cardiovascular complications, such as chronic renal failure, which is frequently observed in hypertensive patients. Therefore, new approaches that may improve the control of arterial blood pressure should be considered to prevent serious cardiovascular disorders. The contribution of purinergic receptors has been acknowledged in the pathophysiology of hypertension; this review describes the participation of these receptors in the alteration of kidney function in hypertension. Elevated interstitial ATP concentrations are essential for the activation of renal purinergic receptors; this becomes a fundamental pathway that leads to the development and maintenance of hypertension. High ATP levels modify essential mechanisms implicated in the long-term control of blood pressure, such as pressure natriuresis, the autoregulation of the glomerular filtration rate and renal blood flow, and tubuloglomerular feedback responses. Any alteration in these mechanisms decreases sodium excretion. ATP stimulates the release of vasoactive substances, causes renal function to decline, and induces tubulointerstitial damage. At the same time, a deleterious interaction involving angiotensin II and purinergic receptors leads to the deterioration of renal function.

The Role of Macula Densa Nitric Oxide Synthase 1 Beta Splice Variant in Modulating Tubuloglomerular Feedback.

Abnormalities in renal electrolyte and water excretion may result in inappropriate salt and water retention, which facilitates the development and maintenance of hypertension, as well as acid-base and electrolyte disorders. A key mechanism by which the kidney regulates renal hemodynamics and electrolyte excretion is via tubuloglomerular feedback (TGF), an intrarenal negative feedback between tubules and arterioles. TGF is initiated by an increase of NaCl delivery at the macula densa cells. The increased NaCl activates luminal Na-K-2Cl cotransporter (NKCC2) of the macula densa cells, which leads to activation of several intracellular processes followed by the production of paracrine signals that ultimately result in a constriction of the afferent arteriole and a tonic inhibition of single nephron glomerular filtration rate. Neuronal nitric oxide (NOS1) is highly expressed in the macula densa. NOS1β is the major splice variant and accounts for most of NO generation by the macula densa, which inhibits TGF response. Macula densa NOS1β-mediated modulation of TGF responses plays an essential role in control of sodium excretion, volume and electrolyte hemostasis, and blood pressure. In this article, we describe the mechanisms that regulate macula densa-derived NO and their effect on TGF response in physiologic and pathologic conditions. © 2023 American Physiological Society. Compr Physiol 13:4215-4229, 2023.

Changes in Proximal Tubular Reabsorption Modulate Microvascular Regulation via the TGF System.

This review paper considers the consequences of modulating tubular reabsorption proximal to the macula densa by sodium–glucose co-transporter 2 (SGLT2) inhibitors, acetazolamide, and furosemide in states of glomerular hyperfiltration. SGLT2 inhibitors improve renal function in early and advanced diabetic nephropathy by decreasing the glomerular filtration rate (GFR), presumably by activating the tubuloglomerular feedback (TGF) mechanism. Central in this paper is that the renoprotective effects of SGLT2 inhibitors in diabetic nephropathy can only be partially explained by TGF activation, and there are alternative explanations. The sustained activation of TGF leans on two prerequisites: no or only partial adaptation should occur in reabsorption proximal to macula densa, and no or only partial adaptation should occur in the TGF response. The main proximal tubular and loop of Henle sodium transporters are sodium–hydrogen exchanger 3 (NHE3), SGLT2, and the Na-K-2Cl co-transporter (NKCC2). SGLT2 inhibitors, acetazolamide, and furosemide are the most important compounds; inhibiting these transporters would decrease sodium reabsorption upstream of the macula densa and increase TGF activity. This could directly or indirectly affect TGF responsiveness, which could oppose sustained TGF activation. Only SGLT2 inhibitors can sustainably activate the TGF as there is only partial compensation in tubular reabsorption and TGF response. SGLT2 inhibitors have been shown to preserve GFR in both early and advanced diabetic nephropathy. Other than for early diabetic nephropathy, a solid physiological basis for these effects in advanced nephropathy is lacking. In addition, TGF has hardly been studied in humans, and therefore this role of TGF remains elusive. This review also considers alternative explanations for the renoprotective effects of SGLT2 inhibitors in diabetic patients such as the enhancement of microvascular network function. Furthermore, combination use of SGLT2 inhibitors and angiotensin-converting enzyme inhibitors (ACEi) or angiotensin receptor blockers (ARBs). in diabetes can decrease inflammatory pathways, improve renal oxygenation, and delay the progression of diabetic nephropathy.

Macula Densa Intracellular Alkalinization Activates NOS1β but Suppresses NOS1α during Tubuloglomerular Feedback

Increased NaCl at the macula densa, the same stimulus of the tubuloglomerular feedback (TGF), also induces intracellular alkalinization (pH: from 7.4 to 7.8) and enhances nitric oxide (NO) generation. Neuronal NO synthase (NOS1) is the primary isoform of NOS expressed in the macula densa and its activity is strongly pH‐dependent. Nevertheless, the impact of pH on the catalytic capacity of NOS1 remains controversial with inconsistent observations. A potential explanation for this discrepancy may be due to the presence of different splice variants of NOS1 (NOS1α and NOS1β). Therefore, we hypothesized that the macula densa intracellular alkalinization during TGF activates NOS1β but suppresses NOS1α, which modulates the NO production and thus the TGF responsiveness. To test this hypothesis, we measured the NOS activity of purified NOS1α and NOS1β proteins in various pH environments using the Griess method. The pH‐dependent NO production curves of NOS1α and NOS1β were different. The optimal pH was ~6.5 for NOS1α, but ~8.5 for NOS1β. The maximum activities of NOS1α and NOS1β were similar at their optimal pH of 6.5 and 8.5, respectively. Then, we measured and compared the renal cortical NOS1 expression, macula densa NO generation, TGF response in vitro and in vivo, renal vascular resistance (RVR), and mean arterial pressure (MAP) in the Sprague Dawley (SD) rats and the spontaneously hypertensive rats (SHRs). While the total NOS1 expression was not significantly different between SD rats and SHRs, the alternative splicing patterns were contradictory. The ratio of NOS1α/NOS1β was greater in SHRs than SD rats (2.94±0.48 vs 0.36±0.07; n=6, p<0.01 vs SD), which was associated with the reduced NO production (21.6±4.5 vs 38.3±5.6 units/min; n=8, p<0.01 vs SD), enhanced TGF in vitro (4.9±0.4 vs 3.5±0.5 µm; n=10, p<0.05 vs SD) and in vivo (12.8±2.1 vs 9.1±1.7 mmHg; n=9, p<0.01 vs SD), increased RVR (23.9±1.3 vs 17.8±1.5 mmHg.ml‐1.min‐1.g kidney wt‐1; n=6, p<0.05 vs SD), and elevated MAP (127.3±6.6 vs 101.7±4.2 mmHg; n=6, p<0.05 vs SD). We also measured and compared the NOS1 expression in the renal biopsy (cortex) tissues of Caucasians (CAs) and African Americans (AAs). There was no significant difference in the total NOS1 expression between CAs and AAs, whereas the ratio of NOS1α/NOS1β was greater in AAs than CAs (3.53±1.64 vs 0.43±0.31; n=10‐11, p<0.01 vs CAs), which was associated with the higher serum creatinine level (1.25±0.08 vs 1.03±0.12 mg/dl; n=10‐11, p<0.05 vs CAs). Lastly, we determined the effects of pH on the macula densa NO generation as well as the TGF responsiveness in SD rats and SHRs. When the pH of the tubular perfusate was switched from 7.4 to 8.0, the macula densa NO production increased by 29.5±6.9% (from 39.7±4.5 to 51.4±5.7 units/min) in SD rats, whereas it decreased by 26.2±4.3% (from 22.5±3.4 to 16.6±4.2 units/min) in SHRs (n=7, p<0.05 vs 7.4); the TGF response in vitro decreased by 24.3±5.4% (from 3.7±0.4 to 2.8±0.6 µm) in SD rats, while it increased by 19.1±3.9% (from 4.7±0.4 to 5.6±0.5 µm) in SHRs (n=7, p<0.05 vs 7.4). In conclusion, the present study demonstrates that the macula densa intracellular alkalinization during TGF activates NOS1β but suppresses NOS1α, which regulates the NO generation, the TGF response and potentially the renal hemodynamics.

First Author Spotlight.

AnnouncementFirst Author SpotlightPublished Online:28 Mar 2022https://doi.org/10.1152/ajprenal.2022.322.4.AUMoreSectionsPDF (372 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations HANNAH M. COSTELLODepartment of Physiology and Functional Genomics, University of Florida, Gainesville, FloridaFigure 1.Hannah CostelloDownload figureDownload PowerPointHannah Costello is a postdoctoral fellow in Dr. Michelle Gumz’s laboratory at the University of Florida in the Department of Physiology and Functional Genomics. She completed her British Heart Foundation-funded PhD in 2021 at the University of Edinburgh. Her research interest focuses on the role of the adrenal and kidney clock in blood pressure regulation. In her spare time, she enjoys traveling, hiking, and reading [Photo: Fig. 1].https://doi.org/10.1152/ajprenal.00385.2021LAUREN G. DOUMADepartment of Physiology and Functional Genomics, University of Florida, Gainesville, FloridaFigure 2.Lauren DoumaDownload figureDownload PowerPointLauren Douma is a Research Assistant Professor at the University of Florida in the Department of Biochemistry and Molecular Biology. She completed her PhD in Genetics and Genomics at the University of Florida. In 2017, she began her postdoctoral fellowship with Dr. Michelle Gumz and since then has been studying the role of the kidney molecular circadian clock in blood pressure regulation. Additionally, she teaches undergraduate- and graduate-level biochemistry and molecular biology courses. She enjoys running, baking, and reading comic books in her free time [Photo: Fig. 2].https://doi.org/10.1152/ajprenal.00385.2021RADMILA MICANOVICDivision of Nephrology and Hypertension, Indiana University School of Medicine, Indianapolis, IndianaFigure 3.Dr. Radmila MicanovicDownload figureDownload PowerPointDr. Radmila Micanovic is an Assistant Research Professor of Medicine at the Indiana University School of Medicine in the Division of Nephrology and Hypertension. She is a researcher and laboratory manager in Dr. Tarek El-Ashkar’s laboratory, a Principal Investigator of the same Division. Prior to joining the Indiana University School of Medicine, she spent 20 yr working in the pharmaceutical industry in drug discovery. She earned her PhD in Biochemistry from the University of Wyoming and completed her postdoctoral studies at the former Roche Institute of Molecular Biology. Her BS degree in Molecular Biology and Physiology was completed in her home country, at the University of Belgrade, Republic of Serbia. Her current work with Dr. El-Achkar focuses on the role of Tamm-Horsfall protein in renal physiology and in the pathophysiology of acute kidney injury. Away from the laboratory, she enjoys great literature, movies, and European TV shows [Photo: Fig. 3].https://doi.org/10.1152/ajprenal.00322.2021SUMIT R. MONUDivision of Hypertension and Vascular Research, Department of Internal Medicine, Henry Ford Hospital, Detroit, MichiganFigure 4.Sumit R. MonuDownload figureDownload PowerPointSumit R. Monu is currently an Instructor-II in the Division of Hypertension and Vascular Research (Internal Medicine) at Henry Ford Hospital and an affiliate Assistant Professor at Wayne State University. He completed his Veterinary Medicine degree (DVM) from Madras Veterinary College, India, and did his PhD at the University of Toledo School of Medicine. Currently, he is working on unraveling the role/mechanism of altered renal hemodynamics in kidney damage commonly seen in obesity, diabetes, and hypertension. Away from the laboratory, he enjoys traveling along with his wife, Mani, and playing with his son, Neil, and is fond of great food and music [Photo: Fig. 4].https://doi.org/10.1152/ajprenal.00307.2021This article has no references to display. Download PDF Back to Top Next FiguresReferencesRelatedInformation Related ArticlesKidney-specific KO of the circadian clock protein PER1 alters renal Na+ handling, aldosterone levels, and kidney/adrenal gene expression 15 Mar 2022American Journal of Physiology-Renal PhysiologyThe kidney releases a nonpolymerizing form of uromodulin in the urine and circulation that retains the external hydrophobic patch domain 15 Mar 2022American Journal of Physiology-Renal PhysiologyDecreased tubuloglomerular feedback response in high-fat diet-induced obesity 15 Mar 2022American Journal of Physiology-Renal Physiology More from this issue > Volume 322Issue 4April 2022Pages i-ii Crossmark Copyright & PermissionsCopyright © 2022 the American Physiological Society.https://doi.org/10.1152/ajprenal.2022.322.4.AUPubMed35344383History Published online 28 March 2022 Published in print 1 April 2022 Metrics

Decreased tubuloglomerular feedback response in high-fat diet-induced obesity.

Obesity increases the risk of renal damage, but the mechanisms are not clear. Normally, kidneys autoregulate to keep the glomerular capillary pressure (PGC), renal blood flow, and glomerular filtration rate in a steady state. However, in obesity, higher PGC, renal blood flow, and glomerular filtration rate are noted. Together, these may lead to glomerular damage. PGC is controlled mainly by afferent arteriole resistance, which, in turn, is regulated by tubuloglomerular feedback (TGF), a vasoconstrictor mechanism. High fat-induced obesity causes renal damage, and this may be related to increased PGC. However, there are no studies as to whether high-fat diet (HFD)-induced obesity affects TGF. We hypothesized that TGF would be attenuated in obesity caused by HFD feeding (60% fat) in Sprague-Dawley rats. Sprague-Dawley rats fed a normal-fat diet (NFD; 12% fat) served as the control. We studied 4 and 16 wk of HFD feeding using in vivo renal micropuncture of individual rat nephrons. We did not observe significant differences in body weight, TGF response, and mean arterial pressure at 4 wk of HFD feeding, but after 16 wk of HFD, rats were heavier and hypertensive. The maximal TGF response was smaller in HFD-fed rats than in NFD-fed rats, indicating an attenuation of TGF in HFD-induced obesity. Baseline PGC was higher in HFD-fed rats than in NFD-fed rats and was associated with higher glomerulosclerosis. We conclude that attenuated TGF and higher PGC along with hypertension in HFD-fed obese Sprague-Dawley rats could explain the higher propensity of glomerular damage observed in obesity.NEW & NOTEWORTHY Reduced tubuloglomerular feedback, higher glomerular capillary pressure, and hypertension in combination may explain the higher glomerular damage observed in high-fat diet-induced obesity.

Role of the macula densa sodium glucose cotransporter type 1-neuronal nitric oxide synthase-tubuloglomerular feedback pathway in diabetic hyperfiltration

An increase of glomerular filtration rate (GFR) is a common observation in early diabetes and is considered a key risk factor for subsequent kidney injury. However, the mechanisms underlying diabetic hyperfiltration have not been fully clarified. Here, we tested the hypothesis that macula densa neuronal nitric oxide synthase (NOS1) is upregulated via sodium glucose cotransporter type 1 (SGLT1) in diabetes, which then inhibits tubuloglomerular feedback (TGF) promoting glomerular hyperfiltration. Therefore, we examined changes in cortical NOS1 expression and phosphorylation, nitric oxide production in the macula densa, TGF response, and GFR during the early stage of insulin-deficient (Akita) diabetes in wild-type and macula densa-specific NOS1 knockout mice. A set of sophisticated techniques including microperfusion of juxtaglomerular apparatus invitro, micropuncture of kidney tubules invivo, and clearance kinetics of plasma fluorescent-sinistrin were employed. Complementary studies tested the role of SGLT1 in SGLT1 knockout mice and explored NOS1 expression and phosphorylation in kidney biopsies of cadaveric donors. Diabetic mice had upregulated macula densa NOS1, inhibited TGF and elevated GFR. Macula densa-selective NOS1 knockout attenuated the diabetes-induced TGF inhibition and GFR elevation. Additionally, deletion of SGLT1 prevented the upregulation of macula densa NOS1 and attenuated inhibition of TGF in diabetic mice. Furthermore, the expression and phosphorylation levels of NOS1 were increased in cadaveric kidneys of diabetics and positively correlated with blood glucose as well as estimated GFR in the donors. Thus, our findings demonstrate that the macula densa SGLT1-NOS1-TGF pathway plays a crucial role in the control of GFR in diabetes.

Knockout of Macula Densa Neuronal Nitric Oxide Synthase Increases Blood Pressure in db/db Mice.

[Figure: see text].

Macula Densa NOS1β Modulates Renal Hemodynamics and Blood Pressure during Pregnancy: Role in Gestational Hypertension.

Regulation of renal hemodynamics and BP via tubuloglomerular feedback (TGF) may be an important adaptive mechanism during pregnancy. Because the β-splice variant of nitric oxide synthase 1 (NOS1β) in the macula densa is a primary modulator of TGF, we evaluated its role in normal pregnancy and gestational hypertension in a mouse model. We hypothesized that pregnancy upregulates NOS1β in the macula densa, thus blunting TGF, allowing the GFR to increase and BP to decrease. We used sophisticated techniques, including microperfusion of juxtaglomerular apparatus in vitro, micropuncture of renal tubules in vivo, clearance kinetics of plasma FITC-sinistrin, and radiotelemetry BP monitoring, to determine the effects of normal pregnancy or reduced uterine perfusion pressure (RUPP) on macula densa NOS1β/NO levels, TGF responsiveness, GFR, and BP in wild-type and macula densa-specific NOS1 knockout (MD-NOS1KO) mice. Macula densa NOS1β was upregulated during pregnancy, resulting in blunted TGF, increased GFR, and decreased BP. These pregnancy-induced changes in TGF and GFR were largely diminished, with a significant rise in BP, in MD-NOS1KO mice. In addition, RUPP resulted in a downregulation in macula densa NOS1β, enhanced TGF, decreased GFR, and hypertension. The superimposition of RUPP into MD-NOS1KO mice only caused a modest further alteration in TGF and its associated changes in GFR and BP. Finally, in African green monkeys, renal cortical NOS1β expression increased in normotensive pregnancies, but decreased in spontaneous gestational hypertensive pregnancies. Macula densa NOS1β plays a critical role in the control of renal hemodynamics and BP during pregnancy.

A Novel Model of Renal Autoregulation Demonstrates Dynamic Modulatory Interactions between TGF and Myogenic Mechanisms

The afferent arteriole regulates glomerular pressure and flow by altering its diameter in response to changes in afferent arteriole wall tension and glomerular filtration via the myogenic and tubuloglomerular feedback (TGF) mechanisms, respectively. Previous studies have indicated that TGF modulates myogenic mechanism sensitivity (Walker III, Matthew, et al. Amer. J. of Physiology-Renal 279.5 (2000): F858-F865), however the degree to which these two mechanisms interact in controlling afferent arteriole smooth muscle cell (SMC) tone remains unclear. We hypothesized that TGF signals modulate the magnitude of the myogenic response to changes in wall tension such that the TGF and myogenic mechanisms act synergistically to control afferent arteriole SMC tone. Accordingly, we developed a mathematical model of renal autoregulation that would allow for testing of differing models of convergence of the TGF and myogenic signals. We considered a “separated” model, in which the TGF and myogenic signals acted additively and independently to influence SMC tone; and an “integrated” model, in which TGF signals modulated the myogenic mechanism sensitivity in influencing SMC tone. Afferent arteriole SMC tone was translated to afferent arteriole wall tension which was used to change the diameter over time. We parameterized the afferent arteriole model according to data previously obtained using the juxtamedullary nephron preparation (Takenaka, Tsuneo, et al. Amer. J. of Physiology-Renal Physiology 267.5 (1994): F879-F887). We combined our afferent arteriole model with a model of glomerular filtration previously developed by us and a model of solute concentration dynamics on the length of the thick ascending limb of the Loop of Henle to simulate autoregulatory responses to a rapid change in perfusion pressure. By nature of the time lag of the tubule model in updating macula densa solute concentration in response to a change in SNGFR, the TGF response was delayed whereas the myogenic response was assumed to be instantaneous. We subjected each model to a step increase in perfusion pressure and compared the models’ responses to that of a model of furosemide administration which lacked TGF signals in calculating SMC tone. We found that the integrated model reached a lower minimum diameter in a shorter time duration after the pressure increase. The slope of the change in diameter immediately after the pressure increase was largest for the integrated model at 0.16 mm/s, while the separated model at 0.11 mm/s did not compete with the model lacking TGF at 0.13 mm/s. These results corroborate previous experimental findings that the speed of the myogenic constrictive response is increased with increasing signals from the macula densa, and that the myogenic mechanism is blunted in the absence of TGF input. Experiments with our novel mathematical model of renal autoregulation support the concept that the myogenic and TGF mechanisms act synergistically to control SMC tone and support the concept that the TGF mechanism modulates the sensitivity of the myogenic response.

Effects of SGLT2 inhibitor and dietary NaCl on glomerular hemodynamics assessed by micropuncture in diabetic rats.

Inhibitors of the main proximal tubular Na-glucose cotransporter (SGLT2) mitigate diabetic glomerular hyperfiltration and have been approved by the United States Food and Drug Administration for slowing the progression of diabetic kidney disease. It has been proposed that SGLT2 inhibitors improve hard renal outcomes by reducing glomerular capillary pressure (PGC) via a tubuloglomerular feedback (TGF) response to a decrease in proximal reabsorption (Jprox). However, the effect of SGLT2 inhibition on PGC has not been measured. Here, we studied the effects of acute SGLT2 blockade (ertugliflozin) on Jprox and glomerular hemodynamics in two-period micropuncture experiments using streptozotocin-induced diabetic rats fed high- or low-NaCl diets. PGC was measured by direct capillary puncture or computed from tubular stop-flow pressure (PSF). TGF is intact while measuring PGC directly but rendered inoperative when measuring PSF. Acute SGLT2 inhibitor reduced Jprox by ∼30%, reduced PGC by 5-8 mmHg, and reduced glomerular filtration rate (GFR) by ∼25% (all P < 0.0001) but had no effect on PSF. The decrease in PGC was larger with the low-NaCl diet (8 vs. 5 mmHg, P = 0.04) where PGC was higher to begin with (54 vs. 50 mmHg, P = 0.003). Greater decreases in PGC corresponded, unexpectedly, to lesser decreases in GFR (P = 0.04). In conclusion, these results confirm expectations that PGC would decline in response to acute SGLT2 inhibition and that a functioning TGF system is required for this. We infer a contribution of postglomerular vasorelaxation to the TGF responses where decreases in PGC were large and decreases in GFR were small.NEW & NOTEWORTHY It has been theorized that Na-glucose cotransporter (SGLT2) blockade slows progression of diabetic kidney disease by reducing physical strain on the glomerulus. This is the first direct measurement of intraglomerular pressure during SGLT2 blockade. Findings confirmed that SGLT2 blockade does reduce glomerular capillary pressure, that this is mediated through tubuloglomerular feedback, and that the tubuloglomerular feedback response to SGLT2 blockade involves preglomerular vasoconstriction and postglomerular vasorelaxation.

Genetic susceptibility of hypertension-induced kidney disease.

Hypertension is the second leading cause of end‐stage renal disease (ESRD) after diabetes mellitus. The significant differences in the incidence of hypertensive ESRD between different patient populations worldwide and patients with and without family history indicate that genetic determinants play an important role in the onset and progression of this disease. Recent studies have identified genetic variants and pathways that may contribute to the alteration of renal function. Mechanisms involved include affecting renal hemodynamics (the myogenic and tubuloglomerular feedback responses); increasing the production of reactive oxygen species in the tubules; altering immune cell function; changing the number, structure, and function of podocytes that directly cause glomerular damage. Studies with hypertensive animal models using substitution mapping and gene knockout strategies have identified multiple candidate genes associated with the development of hypertension and subsequent renal injury. Genome‐wide association studies have implicated genetic variants in UMOD, MYH9, APOL‐1, SHROOM3, RAB38, and DAB2 have a higher risk for ESRD in hypertensive patients. These findings provide genetic evidence of potential novel targets for drug development and gene therapy to design individualized treatment of hypertension and related renal injury.

Roles of Insulin Receptor Substrates (IRS) in renal function and renal hemodynamics.

We have reported previously that renal hemodynamic abnormalities exist in the prediabetic stage of type II diabetic rats. At this prediabetic stage these rats have hyperinsulinemia, insulin resistance and metabolic syndrome. It is well known that insulin resistance is frequently associated with renal abnormalities, but the mechanism underlying this association has remained speculative. Although insulin is known to modify renal hemodynamics, little is known about the roles of insulin receptor substrates (IRS1, IRS2) in the renal actions of insulin. To address this issue, the effects of insulin on renal function and renal hemodynamics were investigated in C57BL/6 (WT: wild type), insulin receptor substrate 1- knockout (IRS1–/–), and IRS2-knockout (IRS2–/–) mice. IRS2–/–mice had elevated glucose level as expected. 24-h urine collections and serum creatinine revealed that creatinine clearance did not significantly differ between these groups. Albuminuria was found in IRS1–/–and IRS2–/–groups. We examined the effects on the IRS during the administration of Losartan, which is widely used for diabetic nephropathy. After the administration of Losartan the IRS displayed improved renal hemodynamics. Moreover, the subjects were also given Pioglitazone, which improves insulin resistance. Losartan significantly reduced albuminuria in both groups. Pioglitazone also showed similar results. We assessed the autoregulatory responses of the total renal blood flow (RBF), the superficial (SBF) and the deep renal cortical blood flow (DBF) with stepwise reductions of renal perfusion pressure (RPP), which was induced by a manual clamp on the abdominal aorta. During the clamp induced reductions of the RPP by 10 to 20mm HG, RBF, SBF and the DBF fell significantly more in the IRS1 and IRS2 than in the WT mice. Furthermore micropuncture studies showded that compared to the WT tubuloglomerular feedback (TGF) responses of the stop flow pressure (Psf) were reduced in both the IRS1 -/- and IRS2 -/-. The results of the IRS1 and IRS2 mice displayed the pressence of hemodynamic abnormalities. Losartan and Pioglitazone have shown the potential to improve these abnormalities. In conclusion the results indicate that IRS plays a major role in the stimulation of renal functions and renal hemodynamics in type type II diabetes.

A new mechanism for the sex differences in angiotensin II-induced hypertension: the role of macula densa NOS1β-mediated tubuloglomerular feedback.

Females are protected against the development of angiotensin II (ANG II)-induced hypertension compared with males, but the mechanisms have not been completely elucidated. In the present study, we hypothesized that the effect of ANG II on the macula densa nitric oxide (NO) synthase 1β (NOS1β)-mediated tubuloglomerular feedback (TGF) mechanism is different between males and females, thereby contributing to the sexual dimorphism of ANG II-induced hypertension. We used microperfusion, micropuncture, clearance of FITC-inulin, and radio telemetry to examine the sex differences in the changes of macula densa NOS1β expression and activity, TGF response, natriuresis, and blood pressure (BP) after a 2-wk ANG II infusion in wild-type and macula densa-specific NOS1 knockout mice. In wild-type mice, ANG II induced higher expression of macula densa NOS1β, greater NO generation by the macula densa, and a lower TGF response in vitro and in vivo in females than in males; the increases of glomerular filtration rate, urine flow rate, and Na+ excretion in response to an acute volume expansion were significantly greater and the BP responses to ANG II were significantly less in females than in males. In contrast, these sex differences in the effects of ANG II on TGF, natriuretic response, and BP were largely diminished in knockout mice. In addition, tissue culture of human kidney biopsies (renal cortex) with ANG II resulted in a greater increase in NOS1β expression in females than in males. In conclusion, macula densa NOS1β-mediated TGF is a novel and important mechanism for the sex differences in ANG II-induced hypertension.

Kidney alkalization increases macula densa NOS1β expression and improves transplanted renal graft function

Ischemia‐reperfusion injury (IRI) is a major hurdle for the survival of transplanted grafts. Currently, there are no effective strategies for prevention and treatment of IRI during kidney transplantation (KTX). Neuronal NO synthase beta (NOS1β) is the primary splice variant of NOS1 in macula densa (MD) and contributes to most of the MD‐NO generation, which blunts tubuloglomerular feedback (TGF) response and increase GFR, which is one of the important factors in evaluation of renal function. While TGF response modulated by NOS1β has been demonstrated to play an essential role in the long‐term control of hemodynamics, the role of NOS1 in the MD in the prevention of IRI and protection of transplanted graft function remains unknown.KTX was performed in C57BL/6 mice. The expression of NOS1 and graft function were evaluated. To test the response of NOS1 to the tubular pH, the mice were given bicarbonate or NH4Cl for two weeks. Blood, urine pH and expression of NOS1 were measured. KTX were performed after the treatment. Graft injury and function were evaluated again. To further investigate the role of the NOS1 in the protection of the graft function, we repeated the above experiments in kidney specific NOS1 knockout (KO) mice and compared the results with C57BL/6 mice.The control recipients showed moderate graft injury with plasma creatinine (Pcr) of 0.68 ± 0.13 mg/dl and reduced graft function indicated by 23% decrease in GFR at 5 days after KTX. NOS1β expression decreased by 60%. TGF response was measured by micropuncture which was 4.1 ± 0.3 mmHg for sham‐operated mice and 7.2 ± 0.9 mmHg for KTX mice. These data indicated that decreased NOS1β expression may contribute to the decrease of GFR and deterioration in graft function by enhanced TGF response. KO recipients showed more severe graft injury with the Pcr of 1.07±0.14mg/dl which demonstrated that deletion of NOS1 from macula densa significantly aggravated renal graft injury.The pH‐NOS activity response curve showed that NOS1 was sensitive to tubular pH with the optimal pH of 8.0. Bicarbonate treatment increased NOS1 expression in macula densa by 65% while NH4Cl intake decreased the NOS1 expression by 40%.KTX were performed in C57BL/6 and KO mice after two weeks treatment of bicarbonate or NH4Cl. Pcr levels were about 30% and 100% lower in bicarbonate‐treated group than H2O and NH4Cl groups, respectively, indicating that bicarbonate treatment significantly alleviated graft injury. However, there were no significant differences in the graft injury for the KO mice among different groups.In summary, NOS1β expression in the macula densa decreases after KTX, which enhances TGF responsiveness and contributes to the decrease in GFR and decline of graft function. Rescue of NOS1β in the macula densa by renal alkalization decreases kidney injury and improves transplanted graft functions.

Antihypertensive and Renal Mechanisms of SGLT2 (Sodium-Glucose Linked Transporter 2) Inhibitors.

Empaglifolzin, canagliflozin, and dapagliflozin are SGLT2 (sodium-glucose linked transporter type 2) inhibitors for treatment of type 2 diabetes mellitus that also reduce blood pressure, mortality, and cardiovascular disease and slow the loss of glomerular filtration rate. SGLT2 inhibitors inhibit the coupled reabsorption of sodium and glucose from the proximal tubules, thereby increasing renal glucose and sodium excretion, but they have more widespread renal effects, including inhibition of the sodium:proton exchanger. They increase the delivery of sodium to the loop of Henle and can thereby activate the tubuloglomerular feedback response to correct glomerular hyperfiltration. There are multiple potential mechanisms whereby these drugs lower blood pressure and preserve kidney function that are the focus of this review.

Clinical Implications

Clinical Implications

New Mechanism for the Sex Differences in Salt-Sensitive Hypertension: The Role of Macula Densa NOS1β-Mediated Tubuloglomerular Feedback.

Females are relatively resistant to salt-sensitive hypertension than males, but the mechanisms are not completely elucidated. We recently demonstrated a decisive role of macula densa neuronal NOS1β (nitric oxide synthase β)-mediated tubuloglomerular feedback (TGF) in the long-term control of glomerular filtration rate, sodium excretion, and blood pressure. In the present study, we hypothesized that the macula densa NOS1β-mediated TGF mechanism is different between male and female, thereby contributing to the sexual dimorphism of salt-sensitive hypertension. We used microperfusion, micropuncture, clearance of fluorescein isothiocyanate-inulin, and radio telemetry to examine the sex differences in the changes of macula densa NOS1β expression and activity, TGF response, natriuresis, and blood pressure after salt loading in wild-type and macula densa-specific NOS1 knockout mice. In wild-type mice, a high-salt diet induced greater increases in macula densa NOS1β expression and phosphorylation at Ser 1417, greater nitric oxide generation by the macula densa, and more inhibition in TGF response in vitro and in vivo in females than in males. Additionally, the increases of glomerular filtration rate, urine flow rate, and sodium excretion in response to an acute volume expansion were significantly greater in females than in males. The blood pressure responses to angiotensin II plus a high-salt diet were significantly less in females than in males. In contrast, these sex differences in TGF, natriuretic response, and blood pressure were largely diminished in knockout mice. In conclusion, macula densa NOS1β-mediated TGF is a novel and important mechanism for the sex differences in salt-sensitive hypertension.

Aging Impairs Renal Autoregulation in Mice.

Impaired renal autoregulation permits more transmission of disturbance in systemic blood pressure, which initiates barotrauma in intrarenal microvasculatures such as glomerular and tubulointerstitial capillaries, contributing to the development of kidney damage and deterioration in renal function, especially under the conditions with high blood pressure. Although it has been postulated that autoregulatory efficiency is attenuated in the aging kidney, direct evidence remains lacking. In the present study, we measured the autoregulation of renal blood flow, myogenic response of afferent arteriole (Af-Art), tubuloglomerular feedback in vivo with micropuncture, as well as tubuloglomerular feedback in vitro in isolated perfused juxtaglomerular apparatus in young and aged C57BL/6 mice. We found that renal blood flow was not significantly changed in response to a defined elevation of renal arterial pressure in young mice but significantly increased in aged mice. Additionally, myogenic response of Af-Art measured by microperfusion with a stepwise increase in perfusion pressure was significantly blunted in the aging kidney, which is associated with the attenuation of intraluminal pressure-induced intracellular calcium increases, as well as the reduced expression of integrin α5 (Itga5) in Af-Art. Moreover, both tubuloglomerular feedback in vivo and in vitro were nearly inactive in the aging kidney, which is associated with the significantly reduced expression of adenosine A1 receptor (A1AR) and suppressed vasoconstrictor response to adenosine in Af-Art. In conclusion, this study demonstrates that aging impairs renal autoregulation with blunted myogenic response and inhibited tubuloglomerular feedback response. The underlying mechanisms involve the downregulations of integrin α5 and A1AR in the Af-Art.

Abstract 144: Macula Densa NOS1β-Mediated TGF Response Plays an Important Role in the Sex Differences in Salt-Sensitive Hypertension

Females are resistant to salt-sensitive hypertension than males, but the mechanisms are not completely elucidated. We recently demonstrated that a decisive role of macula densa neuronal nitric oxide synthase β (NOS1β) in the tubuloglomerular feedback (TGF) response, long-term control of glomerular filtration rate (GFR), sodium excretion and blood pressure. Thus, in this study, we tested a hypothesis that high salt diet-induced upregulation of NOS1β in the macula densa is exaggerated in females than males, which induces more inhibition in TGF response, thereby facilitating sodium excretion and contributing to the resistance to salt-sensitive hypertension. We used microperfusion, micropuncture, clearance of FITC–insulin and radio telemetry to examine the sex differences in macula densa NOS1β expression, nitric oxide (NO) generation, TGF response, kidney clearance function and mean arterial pressure (MAP) in response to a high salt diet in C57BL/6 and macula densa–specific NOS1 knockout (KO) mice. In C57BL/6 mice, males and females had comparable basal levels in macula densa NOS1β expression, NO generation, TGF response and MAP. An 2-week high salt diet resulted in more increases of NOS1β expression in female mice than male mice (3.4±0.3-fold vs. 1.7±0.1-fold), greater increases of NO generation by the macula densa in female mice than male mice (99.1±19.8 % vs. 55.6±11.6 %), and more attenuation in TGF responses in vivo (48.7±7.2 % vs. 25.5±2.9 %) as well as TGF response in vitro (58.5±5.6 % vs. 33.7±9.3 % )in female mice than male mice. Additionally, in response to a 2-week high salt diet along with low dose Ang II, the MAP increased by 13.7±4.2 mmHg in females and 35.4±5.6 mmHg in males with a sex difference of 21.7±4.7 mmHg. In the KO mice, the high salt diet had no significant effects on the macula densa NOS1β expression, NO generation, TGF response in vivo or in vitro in both females and males. Moreover, in response to the high salt diet along with low dose Ang II, the MAP increased by 41.7±4.1 mmHg in KO females and 52.6±3.8 mmHg in KO males with a sex difference of 10.9±3.1 mmHg, which was significantly reduced compared with C57BL/6 mice. In conclusion, macula densa NOS1β-mediated TGF response plays an important role in the sex difference in salt-sensitive hypertension.