Tubuloglomerular Feedback Signal Transduction in a Short Loop of Henle

In previous studies, we used a mathematical model of the thick ascending limb (TAL) to investigate nonlinearities in the tubuloglomerular feedback (TGF) loop. That model does not represent other segments of the nephron, the water, and NaCl transport along which may impact fluid flow rate and NaCl transport along the TAL. To investigate the extent to which those transport processes affect TGF mediation, we have developed a mathematical model for TGF signal transduction in a short loop nephron. The model combines a simple representation of the renal cortex with a highly-detailed representation of the outer medulla (OM). The OM portion of the model is based on an OM urine concentrating mechanism model previously developed by Layton and Layton (Am. J. Renal 289:F1346–F1366, 2005a). When perturbations are applied to intratubular fluid flow at the proximal straight tubule entrance, the present model predicts oscillations in fluid flow and solute concentrations in the cortical TAL and interstitium, and in all tubules, vessels, and interstitium in the OM. Model results suggest that TGF signal transduction by the TAL is a generator of nonlinearities: if a sinusoidal oscillation is added to constant intratubular fluid flow, the time required for an element of tubular fluid to traverse the TAL is oscillatory, but nonsinusoidal; those results are consistent with our previous studies. As a consequence, oscillations in NaCl concentration in tubular fluid alongside the macula densa (MD) will be nonsinusoidal and contain harmonics of the original sinusoidal frequency. Also, the model predicts that the oscillations in NaCl concentration at the loop-bend fluid are smaller in amplitude than those at the MD, a result that further highlights the crucial role of TAL in the nonlinear transduction of TGF signal from SNGFR to MD NaCl concentration.     

Signal transduction in a compliant thick ascending limb

In several previous studies, we used a mathematical model of the thick ascending limb (TAL) to investigate nonlinearities in the tubuloglomerular feedback (TGF) loop. That model, which represents the TAL as a rigid tube, predicts that TGF signal transduction by the TAL is a generator of nonlinearities: if a sinusoidal oscillation is added to constant intratubular fluid flow, the time interval required for an element of tubular fluid to traverse the TAL, as a function of time, is oscillatory and periodic but not sinusoidal. As a consequence, NaCl concentration in tubular fluid alongside the macula densa will be nonsinusoidal and thus contain harmonics of the original sinusoidal frequency. We hypothesized that the complexity found in power spectra based on in vivo time series of key TGF variables arises in part from those harmonics and that nonlinearities in TGF-mediated oscillations may result in increased NaCl delivery to the distal nephron. To investigate the possibility that a more realistic model of the TAL would damp the harmonics, we have conducted new studies in a model TAL that has compliant walls and thus a tubular radius that depends on transmural pressure. These studies predict that compliant TAL walls do not damp, but instead intensify, the harmonics. In addition, our results predict that mean TAL flow strongly influences the shape of the NaCl concentration waveform at the macula densa. This is a consequence of the inverse relationship between flow speed and transit time, which produces asymmetry between up- and downslopes of the oscillation, and the nonlinearity of TAL NaCl absorption at low flow rates, which broadens the trough of the oscillation relative to the peak. The dependence of waveform shape on mean TAL flow may be the source of the variable degree of distortion, relative to a sine wave, seen in experimental recordings of TGF-mediated oscillations.     

Feedback-mediated dynamics in a model of a compliant thick ascending limb

The tubuloglomerular feedback (TGF) system in the kidney, which is a key regulator of filtration rate, has been shown in physiologic experiments in rats to mediate oscillations in tubular fluid pressure and flow, and in NaCl concentration in the tubular fluid of the thick ascending limb (TAL). In this study, we developed a mathematical model of the TGF system that represents NaCl transport along a TAL with compliant walls. The model was used to investigate the dynamic behaviors of the TGF system. A bifurcation analysis of the TGF model equations was performed by deriving and finding roots of the characteristic equation, which arises from a linearization of the model equations. Numerical simulations of the full model equations were conducted to assist in the interpretation of the bifurcation analysis. These techniques revealed a complex parameter region that allows a variety of qualitatively different model solutions: a regime having one stable, time-independent steady-state solution; regimes having one stable oscillatory solution only; and regimes having multiple possible stable oscillatory solutions. Model results suggest that the compliance of the TAL walls increases the tendency of the model TGF system to oscillate.     

Effect of tubular inhomogeneities on filter properties of thick ascending limb of Henle's loop

We used a simple mathematical model of rat thick ascending limb (TAL) of the loop of Henle to predict the impact of spatially inhomogeneous NaCl permeability, spatially inhomogeneous NaCl active transport, and spatially inhomogeneous tubular radius on luminal NaCl concentration when sustained, sinusoidal perturbations were superimposed on steady-state TAL flow. A mathematical model previously devised by us that used homogeneous TAL transport and fixed TAL radius predicted that such perturbations result in TAL luminal fluid NaCl concentration profiles that are standing waves. That study also predicted that nodes in NaCl concentration occur at the end of the TAL when the tubular fluid transit time equals the period of a periodic perturbation, and that, for non-nodal periods, sinusoidal perturbations generate non-sinusoidal oscillations (and thus a series of harmonics) in NaCl concentration at the TAL end. In the present study we find that the inhomogeneities transform the standing waves and their associated nodes into approximate standing waves and approximate nodes. The impact of inhomogeneous NaCl permeability is small. However, for inhomogeneous active transport or inhomogeneous radius, the oscillations for non-nodal periods tend to be less sinusoidal and more distorted than in the homogeneous case and to thus have stronger harmonics. Both the homogeneous and non-homogeneous cases predict that the TAL, in its transduction of flow oscillations into concentration oscillations, acts as a low-pass filter, but the inhomogeneities result in a less effective filter that has accentuated non-linearities.     

Feedback-mediated dynamics in a model of coupled nephrons with compliant thick ascending limbs

The tubuloglomerular feedback (TGF) system in the kidney, a key regulator of glomerular filtration rate, has been shown in physiologic experiments in rats to mediate oscillations in thick ascending limb (TAL) tubular fluid pressure, flow, and NaCl concentration. In spontaneously hypertensive rats, TGF-mediated flow oscillations may be highly irregular. We conducted a bifurcation analysis of a mathematical model of nephrons that are coupled through their TGF systems; the TALs of these nephrons are assumed to have compliant tubular walls. A characteristic equation was derived for a model of two coupled nephrons. Analysis of that characteristic equation has revealed a number of parameter regions having the potential for differing stable dynamic states. Numerical solutions of the full equations for two model nephrons exhibit a variety of behaviors in these regions. Also, model results suggest that the stability of the TGF system is reduced by the compliance of TAL walls and by internephron coupling; as a result, the likelihood of the emergence of sustained oscillations in tubular fluid pressure and flow is increased. Based on information provided by the characteristic equation, we identified parameters with which the model predicts irregular tubular flow oscillations that exhibit a degree of complexity that may help explain the emergence of irregular oscillations in spontaneously hypertensive rats.     

Immunoexpression of aquaporins 1, 2, 3 and 4 in kidney of yak (Bos grunniens) on the Qinghai-Tibetan Plateau

Aquaporins (AQP) 1, 2, 3 and 4 belong to the aquaporin water channel family and play an important role in urine concentration by reabsorption of water from renal tubule fluid. Renal AQPs have not been reported in the yak (Bos grunniens), which resides in the Qinghai Tibetan Plateau. We investigated AQPs 1?4 expressions in the kidneys of Yak using immunohistochemical staining. AQP1 was expressed mainly in the basolateral and apical membranes of the proximal tubules and descending thin limb of the loop of Henle. AQP2 was detected in the apical plasma membranes of collecting ducts and distal convoluted tubules. AQP3 was located in the proximal tubule, distal tubule and collecting ducts. AQP4 was located in the collecting ducts, distal straight tubule, glomerular capillaries and peritubular capillaries. The expression pattern of AQPs 1?4 in kidney of yak was different from other species, which possibly is related to kidney function in a high altitude environment.     

Urine concentrating mechanism: impact of vascular and tubular architecture and a proposed descending limb urea-Na+ cotransporter

We extended a region-based mathematical model of the renal medulla of the rat kidney, previously developed by us, to represent new anatomic findings on the vascular architecture in the rat inner medulla (IM). In the outer medulla (OM), tubules and vessels are organized around tightly packed vascular bundles; in the IM, the organization is centered around collecting duct clusters. In particular, the model represents the separation of descending vasa recta from the descending limbs of loops of Henle, and the model represents a papillary segment of the descending thin limb that is water impermeable and highly urea permeable. Model results suggest that, despite the compartmentalization of IM blood flow, IM interstitial fluid composition is substantially more homogeneous compared with OM. We used the model to study medullary blood flow in antidiuresis and the effects of vascular countercurrent exchange. We also hypothesize that the terminal aquaporin-1 null segment of the long descending thin limbs may express a urea-Na(+) or urea-Cl(-) cotransporter. As urea diffuses from the urea-rich papillary interstitium into the descending thin limb luminal fluid, NaCl is secreted via the cotransporter against its concentration gradient. That NaCl is then reabsorbed near the loop bend, raising the interstitial fluid osmolality and promoting water reabsorption from the IM collecting ducts. Indeed, the model predicts that the presence of the urea-Na(+) or urea- Cl(-) cotransporter facilitates the cycling of NaCl within the IM and yields a loop-bend fluid composition consistent with experimental data.     

Glycocalyx heterogeneity of rat kidney urinary tubule: demonstration with a lectin-gold technique specific for sialic acid

The distribution of sialic acid residues in rat kidney urinary tubule was investigated by light and electron microscopy with a lectin-gold technique. The application of the sialic acid specific Limax flavus lectin resulted in intense plasma membrane labeling of the epithelium of the entire proximal tubule and thin limbs of loop of Henle. In contrast, the plasma membrane of the epithelium lining the medullary portion of the thick ascending limb of Henle was not labeled. In the cortical portion, however, microvilli-bearing positive and smooth-surfaced negative cells were present. Moreover, all cells of the convoluted distal tubules were labeled along their plasma membrane. These data demonstrate the existence of a gross difference in glycocalyx composition between proximal tubules and thin limbs of loop of Henle on one hand and thick ascending limbs on the other. In addition, fine heterogeneity in glycocalyx composition between medullary and cortical portion of thick ascending limb exists. It is concluded that the differences in sialic acid content of the glycocalyx may be related to the functional diversity exhibited by these tubular regions.     

A SPAK isoform switch modulates renal salt transport and blood pressure

The renal thick ascending limb (TAL) and distal convoluted tubule (DCT) play central roles in salt homeostasis and blood pressure regulation. An emerging model suggests that bumetanide- and thiazide-sensitive NaCl transporters (NKCC2 and NCC) along these segments are phosphorylated and activated by WNK kinases, via SPAK and OSR1. Here, we show that a kidney-specific SPAK isoform, which lacks the kinase domain, inhibits phosphorylation of NCC and NKCC2 by full-length SPAK in?vitro. Kidney-specific SPAK is highly expressed along the TAL, whereas full-length SPAK is more highly expressed along the DCT. As predicted from the differential expression, SPAK knockout in animals has divergent effects along TAL and DCT, with increased phosphorylated NKCC2 along TAL and decreased phosphorylated NCC along DCT. In mice, extracellular fluid volume depletion shifts SPAK isoform abundance to favor NaCl retention along both segments, indicating that a SPAK isoform switch modulates sodium avidity along the distal nephron.     

Fluid waves in renal tubules.

Autoregulation of renal blood flow is ineffective when arterial pressure perturbations occur at frequencies above 0.05 Hz. To determine whether wave propagation velocity to the macula densa is rate limiting, we estimated compliances of the proximal tubule and the loop of Henle, and used these values in a model of pressure and flow as functions of time and distance in the nephron. Compliances were estimated from measurements of pressures and flows in early proximal, late proximal, and early distal tubules in rats under normal and Ringer-loaded conditions. A model of steady pressure and flow in a compliant, reabsorbing tubule was fitted to these results. The transient model was a set of nonlinear, hyperbolic partial differential equations with split, nonlinear boundary conditions, and was solved with finite difference methods. The loop of Henle compliance was larger than the proximal tubule compliance, and impulses in glomerular filtration rate were attenuated in magnitude and delayed in time in the loop of Henle. Simulated step forcings revealed a similar pattern. Periodic variations of GFR were attenuated at frequencies greater than 0.05 Hz, and there was a delay of 5 s between variations in GFR and macula densa flow rate. The high compliance of the loop slows wave propagation to the macular densa and reduces the amplitude of high frequency waves originating in the glomerulus, but other parts of the signal chain also contribute to the slow response of macula densa feedback.     

Acute saline expansion increases nephron filtration and distal flow rate but maintains tubuloglomerular feedback responsiveness: role of adenosine A(1) receptors

Temporal adaptation of tubuloglomerular feedback (TGF) permits readjustment of the relationship of nephron filtration rate [single nephron glomerular filtration rate (SNGFR)] and early distal tubular flow rate (V(ED)) while maintaining TGF responsiveness. We used closed-loop assessment of TGF in hydropenia and after acute saline volume expansion (SE; 10% body wt over 1 h) to determine whether 1) temporal adaptation of TGF occurs, 2) adenosine A(1) receptors (A(1)R) mediate TGF responsiveness, and 3) inhibition of TGF affects SNGFR, V(ED), or urinary excretion under these conditions. SNGFR was evaluated in Fromter-Wistar rats by micropuncture in 1) early distal tubules (ambient flow at macula densa), 2) recollected from early distal tubules while 12 nl/min isotonic fluid was added to late proximal tubule (increased flow to macula densa), and 3) from proximal tubules of same nephrons (zero flow to macula densa). SE increased both ambient SNGFR and V(ED) compared with hydropenia, whereas TGF responsiveness (proximal-distal difference in SNGFR, distal SNGFR response to adding fluid to proximal tubule) was maintained, demonstrating TGF adaptation. A(1)R blockade completely inhibited TGF responsiveness during SE and made V(ED) more susceptible to perturbation in proximal tubular flow, but did not alter ambient SNGFR or V(ED). Greater urinary excretion of fluid and Na(+) with A(1)R blockade may reflect additional effects on the distal nephron in hydropenia and SE. In conclusion, A(1)R-independent mechanisms adjust SNGFR and V(ED) to higher values after SE, which facilitates fluid and Na(+) excretion. Concurrently, TGF adapts and stabilizes early distal delivery at the new setpoint in an A(1)R-dependent mechanism.     

Expression of epidermal growth factor in the rat kidney. An immunocytochemical and in situ hybridization study

The renal localization and the site of synthesis of epidermal growth factor (EGF) were investigated in the rat kidney by immunohistochemistry and in situ hybridization techniques. EGF was localized in the cells of the thick ascending limb of Henle (TAL) and distal convoluted tubule (DCT). At the ultrastructural level, EGF immunoreactivity was distributed on the apical membrane and trans-Golgi complex of the TAL and DCT cells. These segments of the rat nephron also hybridized to prepro-EGF cRNA probes in a specific manner, indicating that TAL and DCT are the sites of EGF synthesis in the rat kidney.     

Expression of epidermal growth factor in the rat kidney

Summary The renal localization and the site of synthesis of epidermal growth factor (EGF) were investigated in the rat kidney by immunohistochemistry and in situ hybridization techniques. EGF was localized in the cells of the thick ascending limb of Henle (TAL) and distal convoluted tubule (DCT). At the ultrastructural level, EGF immunoreactivity was distributed on the apical membrane and trans-Golgi complex of the TAL and DCT cells. These segments of the rat nephron also hybridized to prepro-EGF cRNA probes in a specific manner, indicating that TAL and DCT are the sites of EGF synthesis in the rat kidney.     

Distribution of saccharide residues in glycoconjugates of the kidney in Gallus domesticus using peroxidase-conjugated lectins

A battery of seven different horseradish-peroxidase labelled lectins (DBA, PNA, SBA, UEA I, WGA, ConA, LTA) was used to study the distribution of sugar residues in the glycoconjugates along the nephron and the collecting duct of the kidney of Gallus domesticus. As far as the glomerular components are concerned, we have demonstrated that the podocytes and, with a lesser extent, the mesangial cells are characterised by the presence of D-mannose, D-galactose-(beta 1- greater than 3)-N-acetyl-D-galactosamine and sialic acid. The glomerular capillary wall shows the presence of the disaccharide D-galactose-(beta 1- greater than 3)-N-acetyl-D-galactosamine and sialic acid. With regards to the tubules, the proximal tubule, the descending limb of the loop of Henle, the connecting tubule and the collecting one, are characterised by N-acetyl-D-galactosamine, (1- greater than 6)-alpha-L-fucose, D-mannose, N-acetyl-D-galactosamine and D-galactose-(beta 1- greater than 3)-N-acetyl-D-glucosamine. The cells of the connecting and collecting ducts show the presence of intracellular sialic acid, found also as component of the mucous secretion. The ascending limb of the loop of Henle and the distal tubule contain only three saccharidic residues, i.e. (1- greater than 6)-alpha-L-fucose, D-mannose and N-acetyl-D-glucosamine. Lectin histochemistry was also useful to define the saccharidic components of the mucus, which is normally present within the connecting and collecting ducts of the kidney of the birds. The cellular variability of the connecting and the collecting ducts is similar to that found in the kidney of some mammals. Such a variability seems to suggest a possible cell specialization along a single kidney tubule.     

The distribution of leptospires in the kidney tubules of some British wild mammals

Histological examination of 69 pairs of infected kidneys from 12 species of Rodentia, Lagomorpha, Carnivora and Insectivora revealed that leptospires were confined mainly to the proximal and distal convoluted tubule, were less often found in the thick loop of Henle and only rarely in the collecting duct. On no occasion were the organisms present in the thin loop of Henle. Preliminary observations on the relationship of leptospires to tubule epithelium indicate some degree of physical attachment. It is suggested that the avoidance of the thin loop of Henle might be a reflection of its structural properties.     

Acute and chronic effects of SGLT2 blockade on glomerular and tubular function in the early diabetic rat

Tubuloglomerular feedback (TGF) stabilizes nephron function from minute to minute and adapts to different steady-state inputs to maintain this capability. Such adaptation inherently renders TGF less efficient at buffering long-term disturbances, but the magnitude of loss is unknown. We undertook the present study to measure the compromise between TGF and TGF adaptation in transition from acute to chronic decline in proximal reabsorption (Jprox). As a tool, we blocked proximal tubule sodium-glucose cotransport with the SGLT2 blocker dapagliflozin in hyperglycemic rats with early streptozotocin diabetes, a condition in which a large fraction of proximal fluid reabsorption owes to SGLT2. Dapagliflozin acutely reduced proximal reabsorption leading to a 70% increase in early distal chloride, a saturated TGF response, and a major reduction in single nephron glomerular filtration rate (SNGFR). Acute and chronic effects on Jprox were indistinguishable. Adaptations to 10-12 days of dapagiflozin included increased reabsorption by Henle's loop, which caused a partial relaxation in the increased tone exerted by TGF that could be explained without desensitization of TGF. In summary, TGF contributes to long-term fluid and salt balance by mediating a persistent decline in SNGFR as the kidney adapts to a sustained decrease in Jprox.     

Molecular regulation of NKCC2 in the thick ascending limb

The kidney plays an essential role in blood pressure regulation by controlling short-term and long-term NaCl and water balance. The thick ascending limb of the loop of Henle (TAL) reabsorbs 25-30% of the NaCl filtered by the glomeruli in a process mediated by the apical Na(+)-K(+)-2Cl(-) cotransporter NKCC2, which allows Na(+) and Cl(-) entry from the tubule lumen into TAL cells. In humans, mutations in the gene coding for NKCC2 result in decreased or absent activity characterized by severe salt and volume loss and decreased blood pressure (Bartter syndrome type 1). Opposite to Bartter's syndrome, enhanced NaCl absorption by the TAL is associated with human hypertension and animal models of salt-sensitive hypertension. TAL NaCl reabsorption is subject to exquisite control by hormones like vasopressin, parathyroid, glucagon, and adrenergic agonists (epinephrine and norepinephrine) that stimulate NaCl reabsorption. Atrial natriuretic peptides or autacoids like nitric oxide and prostaglandins inhibit NaCl reabsorption, promoting salt excretion. In general, the mechanism by which hormones control NaCl reabsorption is mediated directly or indirectly by altering the activity of NKCC2 in the TAL. Despite the importance of NKCC2 in renal physiology, the molecular mechanisms by which hormones, autacoids, physical factors, and intracellular ions regulate NKCC2 activity are largely unknown. During the last 5 years, it has become apparent that at least three molecular mechanisms determine NKCC2 activity. As such, membrane trafficking, phosphorylation, and protein-protein interactions have recently been described in TALs and heterologous expression systems as mechanisms that modulate NKCC2 activity. The focus of this review is to summarize recent data regarding NKCC2 regulation and discuss their potential implications in physiological control of TAL function, renal physiology, and blood pressure regulation.     

Renal tubular actions of ANF.

Many of the earliest investigations of the renal effects of atrial natriuretic factor (ANF) pointed to the glomerulus as a major site of the peptide's action. More recently, there have been many reports showing various effects of ANF on renal tubular epithelia, including collecting ducts, thick ascending limbs of Henle's loop, thin limbs of Henle's loops, and proximal tubules. The purpose of this review is to summarize the evidence for renal tubular actions of ANF and analyze it from the perspective of the specialized functions of the individual nephron segments, addressing the question: can renal tubule effects of ANF play a significant role in the precise day-to-day regulation of renal NaCl and water excretion? Based on these considerations, we propose that long-term renal tubular action of ANF may be distinct from its short-term natriuretic effect. The short-term action of ANF to accelerate salt and water excretion may play a role in the overall response to acute volume overload. This action of ANF appears to be largely due to an ANF-mediated increase in glomerular filtration rate accompanied by a blunting of the tubuloglomerular feedback mechanism, perhaps with some contribution from ANF-mediated inhibition of fluid absorption in the proximal tubule. In contrast, contributions of ANF to the precise day-to-day regulation of salt and water excretion are likely to be chiefly due to ANF-mediated inhibition of NaCl and water absorption in collecting ducts, but may also involve actions of ANF on the loop of Henle.     

Feedback-mediated dynamics in two coupled nephrons

Previously, we developed a dynamic model for the tubuloglomerular feedback (TGF) system in a single, short-looped nephron of the mammalian kidney. In that model, a semi-linear hyperbolic partial differential equation was used to represent two fundamental processes of solute transport in the nephron’s thick ascending limb (TAL): chloride advection by fluid flow along the TAL lumen and transepithelial chloride transport from the lumen to the interstitium. An empirical function and a time delay were used to relate glomerular filtration rate to the chloride concentration at the macula densa of the TAL. Analysis of the model equations indicated that stable limit-cycle oscillations (LCO) in nephron fluid flow and chloride concentration can emerge for sufficiently large feedback gain magnitude and time delay. In this study, the single-nephron model was extended to two nephrons, which were coupled through their filtration rates. Explicit analytical conditions were obtained for bifurcation loci corresponding to two special cases: (1) identical time delays but differing feedback gains, and (2) identical gains but differing delays. Similar to the case of a single nephron, our analysis indicates that stable LCO can emerge in coupled nephrons for sufficiently large gains and delays. However, these LCO may emerge at lower values of the feedback gain, relative to a single (i.e., uncoupled) nephron, or at shorter delays, provided the delays are sufficiently close. These results suggest that, in vivo, if two nephrons are sufficiently similar, then coupling will tend to increase the likelihood of LCO.