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G-protein-coupled receptor 41 (GPR41) is a Gα i -coupled receptor activated by short-chain fatty acids (SCFAs). Here, we tested that GPR41 is also expressed in cardiomyocytes and exerts a direct negative inotropic effect when activated by SCFA butyrate. Primary cardiomyocytes were isolated from wild-type (WT) and GPR41 knockout (GPR41 -/- ) adult mice and intracellular Ca 2+ concentration and cell shortening were measured using the IonOptix system. RNA localization (RNAScope), quantitative real-time polymerase chain reaction (qRT-PCR), immunofluorescence staining, and western blot were used to examine the expression of GPR41 in adult primary cardiomyocytes of WT and GPR41 -/- mice. The effect of butyrate on shortening and intracellular Ca 2+ transient via GPR41 was also tested in cardiomyocytes. We demonstrated for the first time the presence of GPR41s on cardiomyocytes. Butyrate dose-dependently decreased cell shortening and the amplitude of intracellular Ca 2+ transients in cardiomyocytes from WT but not GPR41 -/- mice. In WT cardiomyocytes, butyrate decreased caffeine-mediated amplitudes of intracellular Ca 2+ transients from the sarcoplasmic reticulum (SR). Moreover, the inhibitory effects of butyrate on cell shortening and intracellular Ca 2+ were pertussis toxin (PTX)-sensitive. Finally, butyrate decreased the activity of sarcoendoplasmic reticulum Ca 2+ -ATPase (SERCA) and cellular 3'-5'-cyclic adenosine monophosphate (cAMP) content. In conclusion, GPR41 is expressed on cardiomyocytes. Butyrate, a known GPR41 agonist, reduces cardiomyocyte shortening and intracellular Ca 2+ transient via decreasing Ca 2+ content in the SR by inhibiting SERCA activity in a PTX-dependent manner. These findings establish that GPR41 is directly activated by SCFAs to decrease contraction and intracellular Ca 2+ transient, highlighting the potential inhibitory role of GPR41 in cardiomyocytes., (© 2024 Federation of American Societies for Experimental Biology.)

The gut microbiome regulates many important host physiological processes associated with cardiovascular health and disease; however, the impact of the gut microbiome on aldosterone is unclear. Investigating whether gut microbiota regulate aldosterone can offer novel insights into how the microbiome affects blood pressure. In this study, we aimed to determine whether gut microbiota regulate host aldosterone. We used enzyme-linked immunosorbent assays (ELISAs) to assess plasma aldosterone and plasma renin activity (PRA) in female and male mice in which gut microbiota are intact, suppressed, or absent. In addition, we examined urinary aldosterone. Our findings demonstrated that when the gut microbiota is suppressed following antibiotic treatment, there is an increase in plasma and urinary aldosterone in both female and male mice. In contrast, an increase in PRA is seen only in males. We also found that when gut microbiota are absent (germ-free mice), plasma aldosterone is significantly increased compared with conventional animals (in both females and males), but PRA is not. Understanding how gut microbiota influence aldosterone levels could provide valuable insights into the development and treatment of hypertension and/or primary aldosteronism. This knowledge may open new avenues for therapeutic interventions, such as probiotics or dietary modifications to help regulate blood pressure via microbiota-based changes to aldosterone. NEW & NOTEWORTHY We explore the role of the gut microbiome in regulating aldosterone, a hormone closely linked to blood pressure and cardiovascular disease. Despite the recognized importance of the gut microbiome in host physiology, the relationship with circulating aldosterone remains largely unexplored. We demonstrate that suppression of gut microbiota leads to increased levels of plasma and urinary aldosterone. These findings underscore the potential of the gut microbiota to influence aldosterone regulation, suggesting new possibilities for treating hypertension.

Competing Interests: Disclosures None.

Sex differences in blood pressure are well-established, with premenopausal women having lower blood pressure than men by ~10 millimeters of mercury; however, the underlying mechanisms are not fully understood. We report here that sex differences in blood pressure are absent in olfactory receptor 558 knockout (KO) mice. Olfr558 localizes to renin-positive cells in the kidney and to vascular smooth muscle cells. Female KOs exhibit increased blood pressure and increased pulse wave velocity. In contrast, male KO mice have decreased renin expression and activity, altered vascular reactivity, and decreased diastolic pressure. A rare OR51E1 (human ortholog) missense variant has a statistically significant sex interaction effect with diastolic blood pressure, increasing diastolic blood pressure in women but decreasing it in men. In summary, our findings demonstrate an evolutionarily conserved role for OLFR558/OR51E1 to mediate sex differences in blood pressure.

The gut microbiome impacts host gene expression not only in the colon but also at distal sites including the liver, white adipose tissue, and spleen. The gut microbiome also influences the kidney and is associated with renal diseases and pathologies; however, a role for the gut microbiome to modulate renal gene expression has not been examined. To determine if microbes modulate renal gene expression, we used whole organ RNA sequencing to compare gene expression in C57Bl/6 mice that were germ free (lacking gut microbiota) versus conventionalized (gut microbiota reintroduced using an oral gavage of a fecal slurry composed of mixed stool). 16S sequencing showed that male and female mice were similarly conventionalized, although Verrucomicrobia was higher in male mice. We found that renal gene expression was differentially regulated in the presence vs. absence of microbiota and that these changes were largely sex specific. Although microbes also influenced gene expression in the liver and large intestine, most differentially expressed genes (DEGs) in the kidney were not similarly regulated in the liver or large intestine. This demonstrates that the influence of the gut microbiota on gene expression is tissue specific. However, a minority of genes ( n = 4 in males and n = 6 in females) were similarly regulated in all three tissues examined, including genes associated with circadian rhythm ( period 1 in males and period 2 in females) and metal binding ( metallothionein 1 and metallothionein 2 in both males and females). Finally, using a previously published single-cell RNA-sequencing dataset, we assigned a subset of DEGs to specific kidney cell types, revealing clustering of DEGs by cell type and/or sex. NEW & NOTEWORTHY It is unknown whether the microbiome influences host gene expression in the kidney. Here, we utilized an unbiased, bulk RNA-sequencing approach to compare gene expression in the kidneys of male and female mice with or without gut microbiota. This report demonstrates that renal gene expression is modulated by the microbiome in a sex- and tissue-specific manner.

The diverse functions of each nephron segment rely on the coordinated action of specialized cell populations that are uniquely defined by their transcriptional profile. In the collecting duct, there are two critical and distinct cell populations: principal cells and intercalated cells. Principal cells play key roles in the regulation of water, Na + , and K + , whereas intercalated cells are best known for their role in acid-base homeostasis. Currently, there are no in vitro systems that recapitulate the heterogeneity of the collecting ducts, which limits high-throughput and replicate investigations of genetic and physiological phenomena. Here, we demonstrated that the transcription factor Foxi1 is sufficient to alter the transcriptional identity of M-1 cells, a murine cortical collecting duct cell line. Specifically, overexpression of Foxi1 induces the expression of intercalated cell transcripts including Gpr116 , Atp6v1b1 , Atp6v1g3 , Atp6v0d2 , Slc4a9 , and Slc26a4 . These data indicate that overexpression of Foxi1 differentiates M-1 cells toward a non-A, non-B type intercalated cell phenotype and may provide a novel in vitro tool to study transcriptional regulation and physiological function of the renal collecting duct. NEW & NOTEWORTHY Transfection of M-1 cells with the transcription factor Foxi1 generates cells that express V-ATPase and Gpr116 as well as other genes associated with renal intercalated cells. This straightforward and novel in vitro system could be used to study processes including transcriptional regulation and cell specification and differentiation in renal intercalated cells.

The gut microbiome influences host physiology and pathophysiology through several pathways, one of which is microbial production of chemical metabolites which interact with host signaling pathways. Short-chain fatty acids (SCFAs) are a class of gut microbial metabolites known to activate multiple signaling pathways in the host. Growing evidence indicates that the gut microbiome is linked to blood pressure, that SCFAs modulate blood pressure regulation, and that delivery of exogenous SCFAs lowers blood pressure. Given that hypertension is a key risk factor for cardiovascular disease, the examination of novel contributors to blood pressure regulation has the potential to lead to novel approaches or treatments. Thus, this review will discuss SCFAs with a focus on their host G protein-coupled receptors including GPR41 (G protein-coupled receptor 41), GPR43, and GPR109A, as well as OLFR78 (olfactory receptor 78) and OLFR558. This includes a discussion of the ligand profiles, G protein coupling, and tissue distribution of each receptor. We will also review phenotypes relevant to blood pressure regulation which have been reported to date for Gpr41, Gpr43, Gpr109a , and Olfr78 knockout mice. In addition, we will consider how SCFA signaling influences physiology at baseline, and, how SCFA signaling may contribute to blood pressure regulation in settings of hypertension. In sum, this review will integrate current knowledge regarding how SCFAs and their receptors regulate blood pressure.

In this study, we elucidate factors that regulate the trafficking and activity of a well-conserved olfactory receptor (OR), olfactory receptor 558 (Olfr558), and its human ortholog olfactory receptor 51E1 (OR51E1). Results indicate that butyrate activates Olfr558/OR51E1 leading to the production of cAMP, and evokes Ca 2+ influx. We also find olfactory G protein (Golf) increases cAMP production induced by Olfr558/OR51E1 activation but does not affect trafficking. Given the 93% sequence identity between OR51E1 and Olfr558, it is surprising to note that OR51E1 has significantly more surface expression yet similar total protein expression. We find that replacing the Olfr558 N-terminus with that of OR51E1 significantly increases trafficking; in contrast, there is no change in surface expression conferred by the OR51E1 TM2, TM3, or TM4 domains. A previous analysis of human OR51E1 single nucleotide polymorphisms (SNPs) identified an A156T mutant primarily found in South Asia as the most abundant (albeit still rare). We find that the OR51E1 A156T mutant has reduced surface expression and cAMP production without a change in total protein expression. In sum, this study of a well-conserved olfactory receptor identifies both protein regions and specific amino acid residues that play key roles in protein trafficking and also elucidates common effects of Golf on the regulation of both the human and murine OR.

Next-generation sequencing (NGS) technology has advanced our understanding of the human microbiome by allowing for the discovery and characterization of unculturable microbes with prediction of their function. Key NGS methods include 16S rRNA gene sequencing, shotgun metagenomic sequencing, and RNA sequencing. The choice of which NGS methodology to pursue for a given purpose is often unclear for clinicians and researchers. In this Review, we describe the fundamentals of NGS, with a focus on 16S rRNA and shotgun metagenomic sequencing. We also discuss pros and cons of each methodology as well as important concepts in data variability, study design, and clinical metadata collection. We further present examples of how NGS studies of the human microbiome have advanced our understanding of human disease pathophysiology across diverse clinical contexts, including the development of diagnostics and therapeutics. Finally, we share insights as to how NGS might further be integrated into and advance microbiome research and clinical care in the coming years.

G protein-coupled receptors (GPCRs) are cell surface proteins which play a key role in allowing cells, tissues, and organs to respond to changes in the external environment in order to maintain homeostasis. Despite the fact that GPCRs are known to play key roles in a variety of tissues, there are a large subset of GPCRs that remain poorly studied. In this minireview, we will summarize what is known regarding the "understudied" GPCRs with respect to renal function, and in so doing will highlight the promise represented by studying this gene family., (© 2021 S. Karger AG, Basel.)

Olfactory receptor 78 (Olfr78) is a G protein-coupled receptor (GPCR) that is expressed in the juxtaglomerular apparatus (JGA) of the kidney as well as the peripheral vasculature, and is activated by gut microbial metabolites. We previously reported that Olfr78 plays a role in renin secretion in isolated glomeruli, and that Olfr78 knockout (KO) mice have lower plasma renin activity. We also noted that in anesthetized mice, Olfr78KO appeared to be hypotensive. In this study, we used radiotelemetry to determine the role of Olfr78 in chronic blood pressure regulation. We found that the blood pressure of Olfr78KO mice is not significantly different than that of their WT counterparts at baseline, or on high- or low-salt diets. However, Olfr78KO mice have depressed heart rates on high-salt diets. We also report that Olfr78KO mice have lower renin protein levels associated with glomeruli. Finally, we developed a mouse where Olfr78 was selectively knocked out in the JGA, which phenocopied the lower renin association findings. In sum, these experiments suggest that Olfr78 modulates renin, but does not play an active role in blood pressure regulation at baseline, and is more likely activated by high levels of short chain fatty acids or hypotensive events. This study provides important context to our knowledge of Olfr78 in BP regulation., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Short-chain fatty acids (SCFAs) are metabolites produced almost exclusively by the gut microbiota and are an essential mechanism by which gut microbes influence host physiology. Given that SCFAs induce vasodilation, we hypothesized that they might have additional cardiovascular effects. In this study, novel mechanisms of SCFA action were uncovered by examining the acute effects of SCFAs on cardiovascular physiology in vivo and ex vivo. Acute delivery of SCFAs in conscious radiotelemetry-implanted mice results in a simultaneous decrease in both mean arterial pressure and heart rate (HR). Inhibition of sympathetic tone by the selective β -1 adrenergic receptor antagonist atenolol blocks the acute drop in HR seen with acetate administration, yet the decrease in mean arterial pressure persists. Treatment with tyramine, an indirect sympathomimetic, also blocks the acetate-induced acute drop in HR. Langendorff preparations show that acetate lowers HR only after long-term exposure and at a smaller magnitude than seen in vivo . Pressure-volume loops after acetate injection show a decrease in load-independent measures of cardiac contractility. Isolated trabecular muscle preparations also show a reduction in force generation upon SCFA treatment, though only at supraphysiological concentrations. These experiments demonstrate a direct cardiac component of the SCFA cardiovascular response. These data show that acetate affects blood pressure and cardiac function through parallel mechanisms and establish a role for SCFAs in modulating sympathetic tone and cardiac contractility, further advancing our understanding of the role of SCFAs in blood pressure regulation. SIGNIFICANCE STATEMENT: Acetate, a short-chain fatty acid, acutely lowers heart rate (HR) as well as mean arterial pressure in vivo in radiotelemetry-implanted mice. Acetate is acting in a sympatholytic manner on HR and exerts negative inotropic effects in vivo. This work has implications for potential short-chain fatty acid therapeutics as well as gut dysbiosis-related disease states., (Copyright © 2021 by The American Society for Pharmacology and Experimental Therapeutics.)

Sensory GPCRs such as olfactory receptors (ORs), taste receptors (TRs), and opsins (OPNs) are now known to play important physiological roles beyond their traditional sensory organs. Here, we systematically investigate the expression of sensory GPCRs in the urinary bladder for the first time. We find that the murine bladder expresses 16 ORs, 7 TRs, and 3 OPNs. We additionally explore the ectopic expression of these GPCRs in tissues beyond the bladder, as well as the localization within the bladder. In future work, understanding the functional roles of these bladder sensory GPCRs may shed light on novel mechanisms which modulate bladder function in health and disease., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

In November 2019, the NIH held the "Sensory Nutrition and Disease" workshop to challenge multidisciplinary researchers working at the interface of sensory science, food science, psychology, neuroscience, nutrition, and health sciences to explore how chemosensation influences dietary choice and health. This report summarizes deliberations of the workshop, as well as follow-up discussion in the wake of the current pandemic. Three topics were addressed: A) the need to optimize human chemosensory testing and assessment, B) the plasticity of chemosensory systems, and C) the interplay of chemosensory signals, cognitive signals, dietary intake, and metabolism. Several ways to advance sensory nutrition research emerged from the workshop: 1) refining methods to measure chemosensation in large cohort studies and validating measures that reflect perception of complex chemosensations relevant to dietary choice; 2) characterizing interindividual differences in chemosensory function and how they affect ingestive behaviors, health, and disease risk; 3) defining circuit-level organization and function that link and interact with gustatory, olfactory, homeostatic, visceral, and cognitive systems; and 4) discovering new ligands for chemosensory receptors (e.g., those produced by the microbiome) and cataloging cell types expressing these receptors. Several of these priorities were made more urgent by the current pandemic because infection with sudden acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the ensuing coronavirus disease of 2019 has direct short- and perhaps long-term effects on flavor perception. There is increasing evidence of functional interactions between the chemosensory and nutritional sciences. Better characterization of this interface is expected to yield insights to promote health, mitigate disease risk, and guide nutrition policy., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society for Nutrition.)

C1q/TNF-related protein 1 (CTRP1) is an endocrine factor with metabolic, cardiovascular, and renal functions. We previously showed that aged Ctrp1- knockout (KO) mice fed a control low-fat diet develop renal hypertrophy and dysfunction. Since aging and obesity adversely affect various organ systems, we hypothesized that aging, in combination with obesity induced by chronic high-fat feeding, would further exacerbate renal dysfunction in CTRP1-deficient animals. To test this, we fed wild-type and Ctrp1- KO mice a high-fat diet for 8 mo or longer. Contrary to our expectation, no differences were observed in blood pressure, heart function, or vascular stiffness between genotypes. Loss of CTRP1, however, resulted in an approximately twofold renal enlargement (relative to body weight), ∼60% increase in urinary total protein content, and elevated pH, and changes in renal gene expression affecting metabolism, signaling, transcription, cell adhesion, solute and metabolite transport, and inflammation. Assessment of glomerular integrity, the extent of podocyte foot process effacement, as well as renal response to water restriction and salt loading did not reveal significant differences between genotypes. Interestingly, blood platelet, white blood cell, neutrophil, lymphocyte, and eosinophil counts were significantly elevated, whereas mean corpuscular volume and hemoglobin were reduced in Ctrp1 -KO mice. Cytokine profiling revealed increased circulating levels of CCL17 and TIMP-1 in KO mice. Compared with our previous study, current data suggest that chronic high-fat feeding affects renal phenotypes differently than similarly aged mice fed a control low-fat diet, highlighting a diet-dependent contribution of CTRP1 deficiency to age-related changes in renal structure and function.

The recent discovery of sensory (tastant and odorant) G protein-coupled receptors on the smooth muscle of human bronchi suggests unappreciated therapeutic targets in the management of obstructive lung diseases. Here we have characterized the effects of a wide range of volatile odorants on the contractile state of airway smooth muscle (ASM) and uncovered a complex mechanism of odorant-evoked signaling properties that regulate excitation-contraction (E-C) coupling in human ASM cells. Initial studies established multiple odorous molecules capable of increasing intracellular calcium ([Ca 2+ ] i ) in ASM cells, some of which were (paradoxically) associated with ASM relaxation. Subsequent studies showed a terpenoid molecule (nerol)-stimulated OR2W3 caused increases in [Ca 2+ ] i and relaxation of ASM cells. Of note, OR2W3-evoked [Ca 2+ ] i mobilization and ASM relaxation required Ca 2+ flux through the store-operated calcium entry (SOCE) pathway and accompanied plasma membrane depolarization. This chemosensory odorant receptor response was not mediated by adenylyl cyclase (AC)/cyclic nucleotide-gated (CNG) channels or by protein kinase A (PKA) activity. Instead, ASM olfactory responses to the monoterpene nerol were predominated by the activity of Ca 2+ -activated chloride channels (TMEM16A), including the cystic fibrosis transmembrane conductance regulator (CFTR) expressed on endo(sarco)plasmic reticulum. These findings demonstrate compartmentalization of Ca 2+ signals dictates the odorant receptor OR2W3-induced ASM relaxation and identify a previously unrecognized E-C coupling mechanism that could be exploited in the development of therapeutics to treat obstructive lung diseases., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

Sodium/potassium-transporting adenosine triphosphatase (Na + /K + -ATPase) is one of the most abundant cell membrane proteins and is essential for eukaryotes. Endogenous negative regulators have long been postulated to play an important role in regulating the activity and stability of Na + /K + -ATPase, but characterization of these regulators has been elusive. Mechanisms of regulating Na + /K + -ATPase homeostatic turnover are unknown. Here, we report that 5-diphosphoinositol 1,2,3,4,6-pentakisphosphate (5-InsP 7 ), generated by inositol hexakisphosphate kinase 1 (IP6K1), promotes physiological endocytosis and downstream degradation of Na + /K + -ATPase-α1. Deletion of IP6K1 elicits a twofold enrichment of Na + /K + -ATPase-α1 in plasma membranes of multiple tissues and cell types. Using a suite of synthetic chemical biology tools, we found that 5-InsP 7 binds the RhoGAP domain of phosphatidylinositol 3-kinase (PI3K) p85α to disinhibit its interaction with Na + /K + -ATPase-α1. This recruits adaptor protein 2 (AP2) and triggers the clathrin-mediated endocytosis of Na + /K + -ATPase-α1. Our study identifies 5-InsP 7 as an endogenous negative regulator of Na + /K + -ATPase-α1., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

The diversity and near universal expression of G protein-coupled receptors (GPCR) reflects their involvement in most physiological processes. The GPCR superfamily is the largest in the human genome, and GPCRs are common pharmaceutical targets. Therefore, uncovering the function of understudied GPCRs provides a wealth of untapped therapeutic potential. We previously identified an adhesion-class GPCR, Gpr116, as one of the most abundant GPCRs in the kidney. Here, we show that Gpr116 is highly expressed in specialized acid-secreting A-intercalated cells (A-ICs) in the kidney using both imaging and functional studies, and we demonstrate in situ receptor activation using a synthetic agonist peptide unique to Gpr116. Kidney-specific knockout (KO) of Gpr116 caused a significant reduction in urine pH (i.e., acidification) accompanied by an increase in blood pH and a decrease in pCO 2 compared to WT littermates. Additionally, immunogold electron microscopy shows a greater accumulation of V-ATPase proton pumps at the apical surface of A-ICs in KO mice compared to controls. Furthermore, pretreatment of split-open collecting ducts with the synthetic agonist peptide significantly inhibits proton flux in ICs. These data suggest a tonic inhibitory role for Gpr116 in the regulation of V-ATPase trafficking and urinary acidification. Thus, the absence of Gpr116 results in a primary excretion of acid in KO mouse urine, leading to mild metabolic alkalosis ("renal tubular alkalosis"). In conclusion, we have uncovered a significant role for Gpr116 in kidney physiology, which may further inform studies in other organ systems that express this GPCR, such as the lung, testes, and small intestine., Competing Interests: The authors declare no competing interest.

Shifts in the gut microbiome play a key role in blood pressure regulation, and changes in the production of gut microbial metabolites are likely to be a key mechanism. Known gut microbial metabolites include short-chain fatty acids, which can signal via G-protein-coupled receptors, and trimethylamine-N oxide. In this review, we provide an overview of gut microbial metabolites documented thus far to play a role in blood pressure regulation.

Local and systemic factors that influence renal structure and function in aging are not well understood. The secretory protein C1q/TNF-related protein 1 (CTRP1) regulates systemic metabolism and cardiovascular function. We provide evidence here that CTRP1 also modulates renal physiology in an age- and sex-dependent manner. In mice lacking CTRP1, we observed significantly increased kidney weight and glomerular hypertrophy in aged male but not female or young mice. Although glomerular filtration rate, plasma renin and aldosterone levels, and renal response to water restriction did not differ between genotypes, CTRP1-deficient male mice had elevated blood pressure. Echocardiogram and pulse wave velocity measurements indicated normal heart function and vascular stiffness in CTRP1-deficient animals, and increased blood pressure was not due to greater salt retention. Paradoxically, CTRP1-deficient mice had elevated urinary sodium and potassium excretion, partially resulting from reduced expression of genes involved in renal sodium and potassium reabsorption. Despite renal hypertrophy, markers of inflammation, fibrosis, and oxidative stress were reduced in CTRP1-deficient mice. RNA sequencing revealed alterations and enrichments of genes in metabolic processes in CTRP1-deficient animals. These results highlight novel contributions of CTRP1 to aging-associated changes in renal physiology., (© 2019 Federation of American Societies for Experimental Biology.)

Gut microbial metabolites have been implicated in contributing to blood pressure regulation; however, only a few microbial metabolites have been examined to date. In this study, we hypothesized that an unbiased screen for changes in gut microbial metabolites in a chronic Ang II (angiotensin II) infusion model would identify novel microbial metabolites associated with blood pressure regulation. To accomplish this, we used both conventional and germ-free mice, which had been implanted with minipumps to infuse either saline or Ang II. Our aim was to identify metabolites that were altered with Ang II treatment in conventional mice, but not in germ-free mice, indicating that they are dependent on the gut microbiota. Both plasma and feces samples were processed and analyzed using liquid chromatography-tandem mass spectroscopy. In plasma, we identified 4 metabolites that were significantly upregulated and 8 metabolites that were significantly downregulated with Ang II treatment in conventional mice; none of these metabolites changed in germ-free mice. Similarly, in feces, we identified 25 metabolites that were significantly upregulated and 71 metabolites that were significantly downregulated with Ang II treatment in conventional mice; none of these metabolites changed in germ-free mice. Finally, fecal 16S sequencing revealed significant shifts in the microbiome of conventional mice with Ang II treatment, including sex-specific changes. These data demonstrate that the metabolites that are differentially regulated with Ang II are dependent on the gut microbiome.

The kidney uses specialized G protein-coupled receptors, including olfactory receptors (ORs), to act as sensors of molecules and metabolites. In the present study, we cloned and studied seven renal ORs, which we previously found to be expressed in the murine renal cortex. As most ORs are orphan receptors, our goal was to identify ligands for these ORs in the hope that this will guide future research into their functional roles. We identified novel ligands for two ORs: Olfr558 and Olfr90. For Olfr558, we confirmed activation by previously reported ligands and identified 16 additional carboxylic acids that activated this OR. The strongest activation of Olfr558 was produced by butyric, cyclobutanecarboxylic, isovaleric, 2-methylvaleric, 3-methylvaleric, 4-methylvaleric, and valeric acids. The primary in vivo source of both butyric and isovaleric acids is gut microbial metabolism. We also identified 14 novel ligands that activated Olfr90, the strongest of which were 2-methyl-4-propyl-1,3-oxathiane, 1-octen-3-ol, 2-octanol, and 3-octanol. Interestingly, 8 of these 14 ligands are of fungal origin. We also investigated the tissue distribution of these receptors and found that they are each found in a subset of "nonsensory" tissues. Finally, we examined the putative human orthologs of Olfr558 and Olfr90 and found that the human ortholog of Olfr558 (OR51E1) has a similar ligand profile, indicating that the role of this OR is likely evolutionarily conserved. In summary, we examined seven novel renal ORs and identified new ligands for Olfr558 and Olfr90, which imply that both of these receptors serve to detect metabolites produced by microorganisms.

Olfactory receptors are G protein-coupled receptors that serve to detect odorants in the nose. Additionally, these receptors are expressed in other tissues, where they have functions outside the canonical smell response. Olfactory receptor 1393 (Olfr1393) was recently identified as a novel regulator of Na + -glucose cotransporter 1 (Sglt1) localization in the renal proximal tubule. Glucose reabsorption in the proximal tubule (via Sglt1 and Sglt2) has emerged as an important contributor to the development of diabetes. Inhibition of Sglt2 is accepted as a viable therapeutic treatment option for patients with type 2 diabetes and has been shown to delay development of diabetic kidney disease. We hypothesized that Olfr1393 may contribute to the progression of type 2 diabetes, particularly the development of hyperfiltration, which has been linked to increased Na + reabsorption in the proximal tubule via the Sglts. To test this hypothesis, Olfr1393 wild-type (WT) and knockout (KO) mice were challenged with a high-fat diet to induce early-stage type 2 diabetes. After 16 wk on the high-fat diet, fasting blood glucose values were increased and glucose tolerance was impaired in the male WT mice. Both of these effects were significantly blunted in the male KO mice. In addition, male and female WT mice developed diabetes-induced hyperfiltration, which was attenuated in the Olfr1393 KO mice and corresponded with a reduction in luminal expression of Sglt2. Collectively, these data indicate that renal Olfr1393 can contribute to the progression of type 2 diabetes, likely as a regulator of Na + -glucose cotransport in the proximal tubule.

Short-chain fatty acids (SCFAs) are fermentation by-products of gut microbes which have been linked to positive effects on host physiology; the most abundant SCFA is acetate. Exogenous administration of acetate alters host metabolism, immune function, and blood pressure, making it a biologic of interest. The effects of acetate have been attributed to activation of G-protein-coupled receptors and other proteins (i.e., HDACs), often occurring at locations distant from the gut such as the pancreas or the kidney. However, due to technical difficulties and costs, studies have often delivered exogenous acetate without determining if systemic plasma acetate levels are altered. Thus, it is unclear to what extent each method of acetate delivery may alter systemic plasma acetate levels. In this study, we aimed to determine if acetate is elevated after exogenous administration by drinking water (DW), oral gavage (OG), or intraperitoneal (IP) injection, and if so, over what timecourse, to best inform future studies. Using a commercially available kit, we demonstrated that sodium acetate delivered over 21 days in DW does not elicit a measurable change in systemic acetate over baseline. However, when acetate is delivered by OG or IP injection, there are rapid, reproducible, and dose-dependent changes in plasma acetate. These studies report, for the first time, the timecourse of changes in plasma acetate following acetate administration by three common methods, and thus inform the best practices for exogenous acetate delivery., (© 2019 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.)

The recent surge in research on the intestinal microbiota has greatly changed our understanding of human biology. Significant technical advances in DNA sequencing analysis and its application to metagenomics and metatranscriptomics has profoundly enhanced our ability to quantify and track complex microbial communities and to begin understanding their impact on human health and disease. This has led to a better understanding of the relationships between the intestinal microbiome and renal physiology/pathophysiology. In this review, we discuss the interactions between intestinal microbiota and kidney. We focus on select aspects including the intestinal barrier, immunologic and soluble mediators of microbiome effects, and effects of dysbiosis on acute kidney injury. Relevant studies on microbiome changes in other renal diseases are highlighted. We also introduce potential mechanisms of intervention with regard to gut microbiota in renal diseases., (Copyright © 2018. Published by Elsevier Inc.)

Sensory receptors that detect and respond to light, taste, and smell primarily belong to the G-protein-coupled receptor (GPCR) superfamily. In addition to their established roles in the nose, tongue, and eyes, these sensory GPCRs have been found in many 'non-sensory' organs where they respond to different physicochemical stimuli, initiating signaling cascades in these extrasensory systems. For example, taste receptors in the airway, and photoreceptors in vascular smooth muscle cells, both cause smooth muscle relaxation when activated. In addition, olfactory receptors are present within the vascular system, where they play roles in angiogenesis as well as in modulating vascular tone. By better understanding the physiological and pathophysiological roles of sensory receptors in non-sensory organs, novel therapeutic agents can be developed targeting these receptors, ultimately leading to treatments for pathological conditions and potential cures for various disease states.

The kidneys play a critical role in precisely regulating the composition of the plasma to maintain homeostasis. To achieve this, the kidneys must be able to accurately determine or "sense" the concentration of a wide variety of substances and to make adjustments accordingly. Kidneys face a key challenge in the arena of pH balance, as there is a particularly narrow range over which plasma pH varies in a healthy subject (7.35-7.45) and this pH must constantly be protected against a variety of onslaughts (changes in diet, activity, and even elevation). The proximal tubule, the first segment to come into contact with the forming urine, plays an important role in helping the kidneys to maintain pH homeostasis. Recent studies have identified a number of novel proximal tubule proteins and signaling pathways that work to sense changes in pH and subsequently modulate renal pH regulation. In this review, we will highlight the role of novel players in acid-base homeostasis in the proximal tubule.

The investigation of orphan GPCRs (GPRs) has the potential to uncover novel insights into whole animal physiology. In this study, our goal was to determine the renal localization of Gprc5c, a receptor that we previously reported to be highly expressed in murine whole kidney, and to examine physiologic parameters in Gprc5c knockout (KO) mice to gain insight into function. Gprc5c localized to the apical membrane of renal proximal tubules (PTs) in mice, rats, and humans. With the comparison of Gprc5c wild-type (WT) and KO mice, we found that Gprc5c KO mice have altered acid-base homeostasis. Specifically, Gprc5c KO mice have lower blood pH and higher urine pH compared with WT mice, with a reduced level of titratable acids in their urine. In an in vitro GPCR internalization assay, we observed that Gprc5c internalization (an index of activation) was triggered by alkaline extracellular pH. Furthermore, with the use of an in vitro BCECF assay, we observed that Gprc5c increases Na + /H + exchanger 3 (NHE3) activity at alkaline pH. We also find that the NHE3 activity is reduced in Gprc5c KO mice by 2 photon imaging in seminaphthorhodafluors (SNARF)-4F-loaded kidney sections. NHE3 is a primary contributor to apical transport of H + in the renal PT. Together, these data imply that Gprc5c modulates the renal contribution to systemic pH homeostasis, at least in part, by taking part in the regulation of NHE3.-Rajkumar, P., Cha, B., Yin, J., Arend, L. J., Păunescu, T. G., Hirabayashi, Y., Donowitz, M., Pluznick, J. L. Identifying the localization and exploring a functional role for Gprc5c in the kidney.

Obesity is a significant risk factor for both chronic kidney disease and end-stage renal disease. To better understand disease development, we sought to identify novel genes differentially expressed early in disease progression. We first confirmed that mice fed a high-fat (HF) diet exhibit early signs of renal injury including hyperfiltration. We then performed RNA-Seq using renal cortex RNA from C57BL6/J male mice fed either HF or control (Ctrl) diet. We identified 1,134 genes differentially expressed in the cortex on HF vs. Ctrl, of which 31 genes were selected for follow-up analysis. This included the 9 most upregulated, the 11 most downregulated, and 11 genes of interest (primarily sensory receptors and G proteins). Quantitative (q)RT-PCR for these 31 genes was performed on additional male renal cortex and medulla samples, and 11 genes (including all 9 upregulated genes) were selected for further study based on qRT-PCR. We then examined expression of these 11 genes in Ctrl and HF male heart and liver samples, which demonstrated that these changes are relatively specific to the renal cortex. These 11 genes were also examined in female renal cortex, where we found that the expression changes seen in males on a HF diet are not replicated in females, even when the females are started on the diet sooner to match weight gain of the males. In sum, these data demonstrate that in a HF-diet model of early disease, novel transcriptional changes occur that are both sex specific and specific to the renal cortex.

Glucose homeostasis is highly controlled, and the function of the kidney plays an integral role in this process. The exquisite control of blood glucose relies, in part, on renal glucose filtration, renal glucose reabsorption, and renal gluconeogenesis. Particularly critical to maintaining glucose homeostasis is the renal reabsorption of glucose; with ~162 g of glucose filtered by the kidney per day, it is imperative that the kidney have the ability to efficiently reabsorb nearly 100% of this glucose back in the bloodstream. In this review, we focus on this central process, highlighting the renal transporters and regulators involved in both the physiology and pathophysiology of glucose reabsorption., (Copyright © 2017 the American Physiological Society.)

Increased renal reabsorption of sodium is a significant risk factor in hypertension. An established clinical marker for essential hypertension is elevated sodium lithium countertransport (SLC) activity. NHA2 is a newly identified Na + (Li + )/H + antiporter with potential genetic links to hypertension, which has been shown to mediate SLC activity and H + -coupled Na + (Li + ) efflux in kidney-derived MDCK cells. To evaluate a putative role in sodium homeostasis, we determined the effect of dietary salt on NHA2. In murine kidney sections, NHA2 localized apically to distal convoluted (both DCT1 and 2) and connecting tubules, partially overlapping in distribution with V-ATPase, AQP2, and NCC1 transporters. Mice fed a diet high in sodium chloride showed elevated transcripts and expression of NHA2 protein. We propose a model in which NHA2 plays a dual role in salt reabsorption or secretion, depending on the coupling ion (sodium or protons). The identified novel regulation of Na + /H + antiporter in the kidney suggests new roles in salt homeostasis and disease.

Purpose of Review: Microbial short-chain fatty acids (SCFAs) are byproducts of microbial metabolism which can be absorbed into the bloodstream of the host, where they exert effects on host physiology. SCFAs have been known to influence several aspects of host physiology, including the regulation of blood pressure. In this review, we will consider recent studies linking SCFAs to blood pressure regulation., Recent Findings: Several recent studies have found that changes in blood pressure often coordinate with expected changes in SCFAS. Efforts are now well underway to dissect and better understand this potential connection. One way that SCFAs can influence host cells is by interacting with host GPCRs, including Gpr41 and Olfr78, among others. Intriguingly, mice null for Olfr78 are hypotensive, whereas mice null for Gpr41 are hypertensive, implying that these pathways may be physiologically important links between SCFAs and host blood pressure control. In sum, these studies demonstrate that there does indeed appear to be a link between SCFAs and blood pressure, which likely involves host GPCRs, at least in part; however, the details and intricacies of these interactions are not yet fully understood and will greatly benefit from further studies.

A number of recent studies have begun to explore a new and exciting area: the interaction between the gut microbiome and renal physiology. In particular, multiple studies have focused on the role of microbially produced short chain fatty acids, which are generally thought to promote health. This review will focus on what is known to date regarding the influence of the microbiome on renal function, with emphasis on the cell biology, physiology, and clinical implications of short chain fatty acids and short chain fatty acid receptors. It is clear that microbe-host interactions are an exciting and ever-expanding field, which has implications for how we view diseases such as hypertension, acute kidney injury, and chronic kidney disease. However, it is important to recognize that although the potential promise of this area is extremely enticing, we are only the very edge of this new field., (Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Pathways that control, or can be exploited to alter, the increase in airway smooth muscle (ASM) mass and cellular remodeling that occur in asthma are not well defined. Here we report the expression of odorant receptors (ORs) belonging to the superfamily of G-protein coupled receptors (GPCRs), as well as the canonical olfaction machinery (G olf and AC3) in the smooth muscle of human bronchi. In primary cultures of isolated human ASM, we identified mRNA expression for multiple ORs. Strikingly, OR51E2 was the most highly enriched OR transcript mapped to the human olfactome in lung-resident cells. In a heterologous expression system, OR51E2 trafficked readily to the cell surface and showed ligand selectivity and sensitivity to the short chain fatty acids (SCFAs) acetate and propionate. These endogenous metabolic byproducts of the gut microbiota slowed the rate of cytoskeletal remodeling, as well as the proliferation of human ASM cells. These cellular responses in vitro were found in ASM from non-asthmatics and asthmatics, and were absent in OR51E2-deleted primary human ASM. These results demonstrate a novel chemo-mechanical signaling network in the ASM and serve as a proof-of-concept that a specific receptor of the gut-lung axis can be targeted to treat airflow obstruction in asthma.

Short chain fatty acid (SCFA) metabolites are byproducts of gut microbial metabolism that are known to affect host physiology via host G protein-coupled receptor (GPCRs). We previously showed that an acute SCFA bolus decreases blood pressure (BP) in anesthetized mice, an effect mediated primarily via Gpr41. In this study, our aims were to identify the cellular localization of Gpr41 and to determine its role in BP regulation. We localized Gpr41 to the vascular endothelium using RT-PCR: Gpr41 is detected in intact vessels (with endothelium) but is absent from denuded vessels (without endothelium). Furthermore, using pressure myography we confirmed that SCFAs dilate resistance vessels in an endothelium-dependent manner. Since we previously found that Gpr41 mediates a hypotensive response to acute SCFA administration, we hypothesized that Gpr41 knockout (KO) mice would be hypertensive. Here, we report that Gpr41 KO mice have isolated systolic hypertension compared with wild-type (WT) mice; diastolic BP was not different between WT and KO. Older Gpr41 KO mice also exhibited elevated pulse wave velocity, consistent with a phenotype of systolic hypertension; however, there was no increase in ex vivo aorta stiffness (measured by mechanical tensile testing). Plasma renin concentrations were also similar in KO and WT mice. The systolic hypertension in Gpr41 KO is not salt sensitive, as it is not significantly altered on either a high- or low-salt diet. In sum, these studies suggest that endothelial Gpr41 lowers baseline BP, likely by decreasing active vascular tone without altering passive characteristics of the blood vessels, and that Gpr41 KO mice have hypertension of a vascular origin., (Copyright © 2016 the American Physiological Society.)

Olfactory receptors (ORs) are G protein-coupled receptors which serve important sensory functions beyond their role as odorant detectors in the olfactory epithelium. Here we describe a novel role for one of these ORs, Olfr1393, as a regulator of renal glucose handling. Olfr1393 is specifically expressed in the kidney proximal tubule, which is the site of renal glucose reabsorption. Olfr1393 knockout mice exhibit urinary glucose wasting and improved glucose tolerance, despite euglycemia and normal insulin levels. Consistent with this phenotype, Olfr1393 knockout mice have a significant decrease in luminal expression of Sglt1, a key renal glucose transporter, uncovering a novel regulatory pathway involving Olfr1393 and Sglt1. In addition, by utilizing a large scale screen of over 1400 chemicals we reveal the ligand profile of Olfr1393 for the first time, offering new insight into potential pathways of physiological regulation for this novel signaling pathway.

Olfactory receptors (ORs) are chemosensors that are responsible for one's sense of smell. In addition to this specialized role in the nose, recent evidence suggests that ORs are also found in a variety of additional tissues including the kidney. As this list of renal ORs continues to expand, it is becoming clear that they play important roles in renal and whole-body physiology, including a novel role in blood pressure regulation. In this review, we highlight important considerations that are crucial when studying ORs and present the current literature on renal ORs and their emerging relevance in maintaining renal function.

New Findings: What is the topic of this review? This review covers recent findings highlighting roles for renal and vascular sensory receptors that modify blood pressure control in response to changes in gut microbial metabolites. What advances does it highlight? This review highlights the novel roles that G-protein-coupled receptor 41 and olfactory receptor 78 play in blood pressure regulation. The gut microbiota have recently been recognized as an important component of host physiology and pathophysiology. Our recent studies have shown that a subset of gut microbial metabolites, known as short-chain fatty acids, act as ligands for host G-protein-coupled receptors (G-protein-coupled receptor 41 and olfactory receptor 78). Short-chain fatty acid-mediated activation of G-protein-coupled receptor 41 and olfactory receptor 78 modulates blood pressure control, both by modulating renin secretion and by modulating vascular tone directly. Further studies are needed in order to gain a better understanding of the underlying mechanism by which microbiota and microbial metabolites modulate host physiology and their potential implications in health and disease., (© 2015 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

To maintain metabolic homeostasis, the body must be able to monitor the concentration of a large number of substances, including metabolites, in real time and to use that information to regulate the activities of different metabolic pathways. Such regulation is achieved by the presence of sensors, termed metabolite receptors, in various tissues and cells of the body, which in turn convey the information to appropriate regulatory or positive or negative feedback systems. In this review, we cover the unique roles of metabolite receptors in renal and vascular function. These receptors play a wide variety of important roles in maintaining various aspects of homeostasis-from salt and water balance to metabolism-by sensing metabolites from a wide variety of sources. We discuss the role of metabolite sensors in sensing metabolites generated locally, metabolites generated at distant tissues or organs, or even metabolites generated by resident microbes. Metabolite receptors are also involved in various pathophysiological conditions and are being recognized as potential targets for new drugs. By highlighting three receptor families-(a) citric acid cycle intermediate receptors, (b) purinergic receptors, and, ((c) short-chain fatty acid receptors-we emphasize the unique and important roles that these receptors play in renal and vascular physiology and pathophysiology.)

In many sensory organs, specialized receptors are strategically arranged to enhance detection sensitivity and acuity. It is unclear whether the olfactory system utilizes a similar organizational scheme to facilitate odor detection. Curiously, olfactory sensory neurons (OSNs) in the mouse nose are differentially stimulated depending on the cell location. We therefore asked whether OSNs in different locations evolve unique structural and/or functional features to optimize odor detection and discrimination. Using immunohistochemistry, computational fluid dynamics modeling, and patch clamp recording, we discovered that OSNs situated in highly stimulated regions have much longer cilia and are more sensitive to odorants than those in weakly stimulated regions. Surprisingly, reduction in neuronal excitability or ablation of the olfactory G protein in OSNs does not alter the cilia length pattern, indicating that neither spontaneous nor odor-evoked activity is required for its establishment. Furthermore, the pattern is evident at birth, maintained into adulthood, and restored following pharmacologically induced degeneration of the olfactory epithelium, suggesting that it is intrinsically programmed. Intriguingly, type III adenylyl cyclase (ACIII), a key protein in olfactory signal transduction and ubiquitous marker for primary cilia, exhibits location-dependent gene expression levels, and genetic ablation of ACIII dramatically alters the cilia pattern. These findings reveal an intrinsically programmed configuration in the nose to ensure high sensitivity to odors., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

In order to assess the status of the volume and composition of the body fluid compartment, the kidney monitors a wide variety of chemical and physical parameters. It has recently become clear that the kidney's sensory capacity extends well beyond its ability to sense ion concentrations in the forming urine. The kidney also keeps track of organic metabolites derived from a surprising variety of sources and uses a complex interplay of physical and chemical sensing mechanisms to measure the rate of fluid flow in the nephron. Recent research has provided new insights into the nature of these sensory mechanisms and their relevance to renal function., (Copyright © 2015 by the American Society of Nephrology.)

Recent studies have highlighted a myriad of ways in which the activity and composition of the gut microbiota can affect the host organism. A primary way in which the gut microbiota affect host physiology is by the production of metabolites, such as short-chain fatty acids (SCFAs), which are subsequently absorbed into the bloodstream of the host. Although recent studies have begun to unravel the ways in which gut microbial SCFAs affect host physiology, less is understood regarding the underlying cell biological mechanisms. In this review, we will outline the known receptors and transporters for SCFAs, and review what is known about the cell biological effects of microbial SCFAs., (Copyright © 2014 the American Physiological Society.)

Recent studies have highlighted the important roles that "sensory" receptors (olfactory receptors, taste receptors, and orphan "GPR" receptors) play in a variety of tissues, including the kidney. Although several studies have identified important roles that individual sensory receptors play in the kidney, there has not been a systematic analysis of the renal repertoire of sensory receptors. In this study, we identify novel renal sensory receptors belonging to the GPR (n = 76), olfactory receptor (n = 6), and taste receptor (n = 11) gene families. A variety of reverse transcriptase (RT)-PCR screening strategies were used to identify novel renal sensory receptors, which were subsequently confirmed using gene-specific primers. The tissue-specific distribution of these receptors was determined, and the novel renal ORs were cloned from whole mouse kidney. Renal ORs that trafficked properly in vitro were screened for potential ligands using a dual-luciferase ligand screen, and novel ligands were identified for Olfr691. These studies demonstrate that multiple sensory receptors are expressed in the kidney beyond those previously identified. These results greatly expand the known repertoire of renal sensory receptors. Importantly, the mRNA of many of the receptors identified in this study are expressed highly in the kidney (comparable to well-known and extensively studied renal GPCRs), and in future studies it will be important to elucidate the roles that these novel renal receptors play in renal physiology.

Purpose of Review: This review will summarize recent literature highlighting the roles of sensory Gpr receptors and their roles in renal function., Recent Findings: Chemoreceptors play important roles in renal physiology wherein they modulate renal function in response to ligands from a variety of sources., Summary: As specialized chemical detectors, chemoreceptors in the kidney monitor the level of a variety of chemical ligands in the body and adjust renal function accordingly. In addition to olfactory receptors and taste receptors, G-protein coupled receptors of the orphan Gpr family are now being found to play a 'sensory' role in renal physiology. Identifying the physiological roles of these receptors and elucidating the cell biology underlying these signaling pathways can give us novel insights into renal function.