Shear Stress Blunts Tubuloglomerular Feedback Partially Mediated by Primary Cilia and Nitric Oxide at the Macula Densa

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macula densa, primary cilia, nitric oxide, tubuloglomerular feedback

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The present study tested whether primary cilia on macula densa serve as a flow sensor to enhance nitric oxide synthase 1 (NOS1) activity and inhibit tubuloglomerular feedback (TGF). Isolated perfused macula densa was loaded with calcein red and 4,5-diaminofluorescein diacetate to monitor cell volume and nitric oxide (NO) generation. An increase in tubular flow rate from 0 to 40 nl/min enhanced NO production by 40.0 ± 1.2%. The flow-induced NO generation was blocked by an inhibitor of NOS1 but not by inhibition of the Na/K/2Cl cotransporter or the removal of electrolytes from the perfusate. NO generation increased from 174.8 ± 21 to 276.1 ± 24 units/min in cultured MMDD1 cells when shear stress was increased from 0.5 to 5.0 dynes/cm2. The shear stress-induced NO generation was abolished in MMDD1 cells in which the cilia were disrupted using a siRNA to ift88. Increasing the NaCl concentration of the tubular perfusate from 10 to 80 mM NaCl in the isolated perfused juxtaglomerular preparation reduced the diameter of the afferent arteriole by 3.8 ± 0.1 μm. This response was significantly blunted to 2.5 ± 0.2 μm when dextran was added to the perfusate to increase the viscosity and shear stress. Inhibition of NOS1 blocked the effect of dextran on TGF response. In vitro, the effects of raising perfusate viscosity with dextran on tubular hydraulic pressure were minimized by reducing the outflow resistance to avoid stretching of tubular cells. These results suggest that shear stress stimulates primary cilia on the macula densa to enhance NO generation and inhibit TGF responsiveness.

the macula densa is a group of specialized epithelial cells located at the distal segment of the thick ascending limb (TAL) that serves as a sensor of luminal NaCl concentration. Increased NaCl delivery to the macula densa promotes the release of adenosine or ATP, which constricts the afferent arteriole (Af-Art) and decreases single nephron glomerular filtration rate (SNGFR) via a process known as tubuloglomerular feedback (TGF) (2, 37, 57). TGF is a negative feedback mechanism to prevent fluctuations in GFR and flow in the proximal tubule to deliver excess NaCl to the distal nephron and overwhelm its transport capacity.

Following acute volume expansion or the ingestion of a sodium load, increases in GFR facilitate the rapid elimination of salt load (29, 40). Baroreflex suppression of renal sympathetic tone and the renin-angiotensin system also contribute to the increase in sodium excretion by inhibiting proximal tubular reabsorption (6, 22). All of these factors result in a large increase in flow to the macula densa, which should trigger a TGF-mediated decrease in GFR that opposes the elimination of the sodium load. However, this decrease in GFR does not occur, partially because of inhibition of TGF responsiveness. Modulation of TGF response following volume expansion should be important in maintaining extracellular sodium and volume balance (42, 52, 53). Indeed, GFR typically increases by 20–30% in humans (4) and in dogs (10) during the postprandial period. Daily salt excretion peaks during the postprandial increase in GFR (21). In these situations, especially following ingestion of a salt load, reduced TGF responsiveness permits high distal NaCl delivery, which could facilitate the excretion of NaCl. The mechanisms for this TGF modulation have not been clarified.

Normal luminal flow rate in the distal tubule is about 2–7 nl/min, but it can be temporally reduced to 0 nl/min in salt and volume depletion, and it increases to 25–37 nl/min following acute volume expansion (11, 20, 43). In physiological conditions, increase of tubular flow to the macula densa activates TGF by increasing NaCl delivery (44). However, it also increases shear stress, stretch, and transmural pressure (54, 59); therefore, it possibly activates mechanosensors and other pathways that might modulate TGF responsiveness. The modulation of TGF responsiveness may depend, in part, on increased nitric oxide (NO) production (41, 48, 58), but the source and mechanism for the increase in NO are not known. Primary cilia extend from the surface of many eukaryotic cells (8, 36, 61). While the function of the primary cilia on most cells has remained elusive, they are known to serve as mechanosensors in the mammalian kidney and vascular endothelial cells (8, 36, 61). Primary cilia have been found on the macula densa cells of humans, rats, rabbits, and dogs (17, 33, 47, 50, 56). Little is known about their function (47). The present study examined the hypothesis that primary cilia on macula densa cells serve as a flow sensor to activate NOS1 activity and enhance NO generation, which inhibits TGF responsiveness.

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Citation / Publisher Attribution

American Journal of Physiology-Regulatory, Integrative and Comparative Physiology, v. 309, issue 7, p. R757-R766