Article

Luminal flow induces eNOS activation and translocation in the rat thick ascending limb.

Division of Hypertension and Vascular Research, Department of Internal Medicine, Henry Ford Hospital, 2799 W. Grand Blvd., Detroit, MI 48202, USA.
American journal of physiology. Renal physiology (Impact Factor: 3.61). 08/2004; 287(2):F274-80. DOI: 10.1152/ajprenal.00382.2003
Source: PubMed

ABSTRACT Nitric oxide (NO) produced by endothelial NO synthase (eNOS) acts as an autacoid to inhibit NaCl absorption in the thick ascending limb of the loop of Henle (THAL). In the vasculature, shear stress activates eNOS. We hypothesized that increasing luminal flow activates eNOS and enhances NO production in the THAL. We measured NO production by isolated, perfused THALs using a NO-sensitive microelectrode. Increasing luminal flow from 0 to 20 nl/min increased NO production by 43.1 +/- 4.1 pA/mm of tubule (n = 10, P < 0.05), and this response was blunted (92%) by the NOS inhibitor L-(omega)nitro-methylarginine (P < 0.05). We studied the effect of flow on eNOS subcellular localization. In the absence of flow, eNOS was diffusely localized throughout the cell (basolateral = 33 +/- 4%; middle = 27 +/- 3%; apical = 40 +/- 4% of total eNOS). Increasing luminal flow induced eNOS translocation to the apical membrane, as evidenced by a 60% increase in eNOS immunoreactivity in the apical membrane (from 40 +/- 4 to 65 +/- 2%; n = 6; P < 0.05). Disrupting the actin cytoskeleton with cytochalasin D (10 microM) reduced flow-induced NO production by 62% (from 37.1 +/- 3.4 to 14.0 +/- 2.4 pA/mm tubule, n = 7, P < 0.04) and blocked flow-induced eNOS translocation. Flow also increased the amount of phosphorylated eNOS (Ser1179) at the apical membrane (from 25 +/- 2 to 56 +/- 2%; P < 0.05). We conclude that increasing luminal flow induces eNOS activation and translocation to the apical membrane in THALs. These are the first data showing that flow regulates eNOS in epithelial cells. This may be an important mechanism for regulation of NO levels in the renal medulla.

0 Bookmarks
 · 
100 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: The involvement of purinergic signalling in kidney physiology and pathophysiology is rapidly gaining recognition and this is a comprehensive review of early and recent publications in the field. Purinergic signalling involvement is described in several important intrarenal regulatory mechanisms, including tuboglomerular feedback, the autoregulatory response of the glomerular and extraglomerular microcirculation and the control of renin release. Furthermore, purinergic signalling influences water and electrolyte transport in all segments of the renal tubule. Reports about purine- and pyrimidine-mediated actions in diseases of the kidney, including polycystic kidney disease, nephritis, diabetes, hypertension and nephrotoxicant injury are covered and possible purinergic therapeutic strategies discussed.
    Purinergic Signalling 11/2013; · 2.64 Impact Factor
  • Wiener Medizinische Wochenschrift 07/2008; 158:365-369.
  • [Show abstract] [Hide abstract]
    ABSTRACT: Pathways that contribute to TNF production by the kidney are not well defined. Mice given 1% NaCl in the drinking water for 3 days exhibited a 2.5-fold increase in urinary, but not plasma, TNF levels compared with mice given tap water. Since furosemide attenuated the increase in TNF levels, we hypothesized that hypertonic NaCl intake increases renal TNF production by a pathway involving NKCC2. A 2.5-fold increase in NKCC2A mRNA accumulation was observed in medullary thick ascending limb (mTAL) tubules from mice given 1% NaCl; a concomitant 2-fold increase in NFAT5 mRNA and protein expression was observed in outer medulla. Urinary TNF levels were reduced in mice given 1% NaCl after an intrarenal injection of a lentivirus construct designed to specifically knockdown NKCC2A (EGFP-N2A-ex4); plasma levels of TNF did not change after injection of EGFP-N2A-ex4. Intrarenal injection of EGFP-N2A-ex4 also inhibited the increase of NFAT5 mRNA abundance in outer medulla of mice given 1% NaCl. TNF production by primary cultures of mTAL cells increased approximately 6-fold in response to an increase in osmolality to 400 mosmol/kg H2O produced with NaCl, and was inhibited in cells transiently transfected with a dnNFAT5 construct. Transduction of cells with EGFP-N2A-ex4 also prevented increases in TNF mRNA and protein production in response to high NaCl concentration and reduced transcriptional activity of a NFAT5 promoter construct. Since NKCC2A expression is restricted to the TAL, NKCC2A-dependent activation of NFAT5 is part of a pathway by which the TAL produces TNF in response to hypertonic NaCl intake.
    AJP Renal Physiology 12/2012; · 4.42 Impact Factor

Full-text

Download
0 Downloads