Leon C. Moore

Carbon monoxide induces vasodilation and nitric oxide release but suppresses endothelial NOS.

The vascular effects of carbon monoxide (CO) resemble those of nitric oxide (NO), but it is unknown whether the two messengers converge or exhibit reciprocal feedback regulation. These questions were examined in microdissected perfused renal resistance arteries (RRA) studied using NO-sensitive microelectrodes. Perfusion of RRA with buffers containing increasing concentrations of CO resulted in a biphasic release of NO. The NO response peaked at 100 nM CO and then declined to virtually zero at 10 μM. When a series of 50-s pulses of 100 nM CO were applied repeatedly (150-s interval), the amplitude of consecutive NO responses was diminished. NO release from RRA showed dependence on l-arginine but notd-arginine, and the responses to CO were inhibited by pretreatment with N G-nitro-l-arginine methyl ester (l-NAME), an inhibitor of NO synthases (NOS). CO (100 nM) also suppressed NO release induced by 100 μM carbachol, a potent agonist for endothelial NOS (eNOS). RRA from rats in which endogenous CO production from inducible HO was elevated (cobalt chloride 12 h prior to study) also showed suppressed responses to carbachol. Furthermore, responses consistent with these findings were obtained in juxtamedullary afferent arterioles perfused in vitro, where the vasodilatory response to CO was biphasic and the response to acetylcholine was blunted. Collectively, these data suggest that the CO-induced NO release could be attributed to either stimulation of eNOS or to NO displacement from a cellular storage pool. To address this, direct in vitro measurements with an NO-selective electrode of NO production by recombinant eNOS revealed that CO dose-dependently inhibits NO synthesis. Together, the above data demonstrate that, whereas high levels of CO inhibit NOS activity and NO generation, lower concentrations of CO induce release of NO from a large intracellular pool and, therefore, may mimic the vascular effects of NO.The vascular effects of carbon monoxide (CO) resemble those of nitric oxide (NO), but it is unknown whether the two messengers converge or exhibit reciprocal feedback regulation. These questions were examined in microdissected perfused renal resistance arteries (RRA) studied using NO-sensitive microelectrodes. Perfusion of RRA with buffers containing increasing concentrations of CO resulted in a biphasic release of NO. The NO response peaked at 100 nM CO and then declined to virtually zero at 10 microM. When a series of 50-s pulses of 100 nM CO were applied repeatedly (150-s interval), the amplitude of consecutive NO responses was diminished. NO release from RRA showed dependence on L-arginine but not D-arginine, and the responses to CO were inhibited by pretreatment with NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthases (NOS). CO (100 nM) also suppressed NO release induced by 100 microM carbachol, a potent agonist for endothelial NOS (eNOS). RRA from rats in which endogenous CO production from inducible HO was elevated (cobalt chloride 12 h prior to study) also showed suppressed responses to carbachol. Furthermore, responses consistent with these findings were obtained in juxtamedullary afferent arterioles perfused in vitro, where the vasodilatory response to CO was biphasic and the response to acetylcholine was blunted. Collectively, these data suggest that the CO-induced NO release could be attributed to either stimulation of eNOS or to NO displacement from a cellular storage pool. To address this, direct in vitro measurements with an NO-selective electrode of NO production by recombinant eNOS revealed that CO dose-dependently inhibits NO synthesis. Together, the above data demonstrate that, whereas high levels of CO inhibit NOS activity and NO generation, lower concentrations of CO induce release of NO from a large intracellular pool and, therefore, may mimic the vascular effects of NO.

Decreased hepatic insulin-like growth factor (IGF)-I and increased IGF binding protein-1 and -2 gene expression in experimental uremia

The imbalance between normal insulin-like growth factor-I (IGF-I) and markedly increased IGF binding protein (IGFBP) plasma levels plays a pathogenic role for growth retardation and catabolism in children with chronic renal failure. To investigate the mechanism of these alterations, experiments were performed in an experimental model of uremia in rats (5/6 nephrectomy) and in pair-fed and ad libitum-fed sham-operated controls. Using a specific solution hybridization/RNase protection assay, we observed a marked reduction of hepatic IGF-I messenger RNA (mRNA) abundance at steady state in uremic animals (37 ± 5% of control) compared both with pair-fed (65 ± 10%) and ad libitum-fed controls (100± 11%) (P < 0.001). Reduced IGF-I gene expression was clearly organ-specific; it was most pronounced in liver (significant vs.. pair-fed controls) and lung and muscle tissue (significant vs.. ad libitum-fed controls); no change was observed in kidney and heart tissue. To determine a potential mechanism of reduced hepat...

Lens intracellular hydrostatic pressure is generated by the circulation of sodium and modulated by gap junction coupling

We recently modeled fluid flow through gap junction channels coupling the pigmented and nonpigmented layers of the ciliary body. The model suggested the channels could transport the secretion of aqueous humor, but flow would be driven by hydrostatic pressure rather than osmosis. The pressure required to drive fluid through a single layer of gap junctions might be just a few mmHg and difficult to measure. In the lens, however, there is a circulation of Na+ that may be coupled to intracellular fluid flow. Based on this hypothesis, the fluid would cross hundreds of layers of gap junctions, and this might require a large hydrostatic gradient. Therefore, we measured hydrostatic pressure as a function of distance from the center of the lens using an intracellular microelectrode-based pressure-sensing system. In wild-type mouse lenses, intracellular pressure varied from ∼330 mmHg at the center to zero at the surface. We have several knockout/knock-in mouse models with differing levels of expression of gap junction channels coupling lens fiber cells. Intracellular hydrostatic pressure in lenses from these mouse models varied inversely with the number of channels. When the lens’ circulation of Na+ was either blocked or reduced, intracellular hydrostatic pressure in central fiber cells was either eliminated or reduced proportionally. These data are consistent with our hypotheses: fluid circulates through the lens; the intracellular leg of fluid circulation is through gap junction channels and is driven by hydrostatic pressure; and the fluid flow is generated by membrane transport of sodium.

Spectral properties of the tubuloglomerular feedback system

A simple mathematical model was used to investigate the spectral properties of the tubuloglomerular feedback (TGF) system. A perturbation, consisting of small-amplitude broad-band forcing, was applied to simulated thick ascending limb (TAL) flow, and the resulting spectral response of the TGF pathway was assessed by computing a power spectrum from resulting TGF-regulated TAL flow. Power spectra were computed for both open- and closed-feedback-loop cases. Open-feedback-loop power spectra are consistent with a mathematical analysis that predicts a nodal pattern in TAL frequency response, with nodes corresponding to frequencies where oscillatory flow has a TAL transit time that equals the steady-state fluid transit time. Closed-feedback-loop spectra are dominated by the open-loop spectral response, provided that γ, the magnitude of feedback gain, is less than the critical value γc required for emergence of a sustained TGF-mediated oscillation. For γ exceeding γc, closed-loop spectra have peaks corresponding to the fundamental frequency of the TGF-mediated oscillation and its harmonics. The harmonics, expressed in a nonsinusoidal waveform for tubular flow, are introduced by nonlinear elements of the TGF pathway, notably TAL transit time and the TGF response curve. The effect of transit time on the flow waveform leads to crests that are broader than troughs and to an asymmetry in the magnitudes of increasing and decreasing slopes. For feedback gain magnitude that is sufficiently large, the TGF response curve tends to give a square waveshape to the waveform. Published waveforms and power spectra of in vivo TGF oscillations have features consistent with the predictions of this analysis.

Ascending myogenic autoregulation: Interactions between tubuloglomerular feedback and myogenic mechanisms

A mathematical model of the renal vascular and tubular systems was used to examine the possibility that synergistic interactions might occur between the tubuloglomerular feedback (TGF) and myogenic autoregulatory mechanisms in the kidney. To simulate the myogenic mechanism, the renal vasculature was modelled with a resistance network where the total preglomerular resistance varies with intravascular pressure. In addition, a steady-state model of glomerular filtration, proximal and Henles loop reabsorption, and TGF-modulation of afferent arteriolar resistance was derived. The results show that, if TGF acts on the distal portion of the preglomerular vasculature, then any TGF-induced vasoconstriction should raise upstream intravascular pressure and, thereby, trigger a myogenic (AMYO) response. The model further predicts that the magnitude of the AMYO response can be similar in magnitude to the TGF-induced increment in afferent resistance. Hence, the effects of TGF excitation on whole kidney hemodynamics may be much greater than predicted from measurements in single nephrons. Moreover, a significant fraction of the intrinsic myogenic autoregulatory response to increased renal perfusion pressure may result from a synergistic interaction between the TGF and myogenic mechanisms.

Attenuation of isoflurane-induced preconditioning and reactive oxygen species production in the senescent rat heart.

BACKGROUND:Although attenuation of anesthetic preconditioning in aged ex vivo heart models has been studied extensively, there are no comparable in vivo studies. To extend previous work and to address a possible mechanism underlying age-related differences, we investigated isoflurane-induced preconditioning and reactive oxygen species (ROS) production in the aged rat heart in vivo. METHODS:Male Fisher 344 rats were assigned from their respective age groups (young, 3–5 mo; old, 20–24 mo) to either receive 30 min of 1.0 minimum alveolar concentration isoflurane or to a control group. Rats were subjected to coronary artery occlusion for 30 min followed by 2 h of reperfusion. A fluorescent probe for superoxide anion production (dihydroethidium, 1 mg) was administered in the absence of the isoflurane or just before isoflurane exposure in four additional groups. Myocardial infarct size and superoxide anion production were assessed using triphenyltetrazolium staining and epifluorescence microscopy, respectively. RESULTS:Isoflurane decreased myocardial infarct size of young rats (26.7% ± 3.0%) compared with young controls (50.9% ± 1.9%; P < 0.001), whereas isoflurane did not significantly affect myocardial infarct size of old rats (39.1% ± 0.9%) compared with old controls (46.5% ± 2.4%; P > 0.05). Isoflurane increased ROS levels in young rats (430.5 ± 95.9 arbitrary units [AU]) compared with young controls (162.7 ± 25.5 AU; P < 0.01). In contrast, no significant changes in ROS levels were observed in old animals (316.4 ± 56.3 AU isoflurane versus 233.8 ± 59.2 AU control). CONCLUSIONS:Reduction in the cardioprotective effects of isoflurane and attenuation of isoflurane-stimulated ROS production were observed in the senescent myocardium in vivo.

A mathematical model of the myogenic response to systolic pressure in the afferent arteriole

Elevations in systolic blood pressure are believed to be closely linked to the pathogenesis and progression of renal diseases. It has been hypothesized that the afferent arteriole (AA) protects the glomerulus from the damaging effects of hypertension by sensing increases in systolic blood pressure and responding with a compensatory vasoconstriction (Loutzenhiser R, Bidani A, Chilton L. Circ Res 90: 1316-1324, 2002). To investigate this hypothesis, we developed a mathematical model of the myogenic response of an AA wall, based on an arteriole model (Gonzalez-Fernandez JM, Ermentrout B. Math Biosci 119: 127-167, 1994). The model incorporates ionic transport, cell membrane potential, contraction of the AA smooth muscle cell, and the mechanics of a thick-walled cylinder. The model represents a myogenic response based on a pressure-induced shift in the voltage dependence of calcium channel openings: with increasing transmural pressure, model vessel diameter decreases; and with decreasing pressure, vessel diameter increases. Furthermore, the model myogenic mechanism includes a rate-sensitive component that yields constriction and dilation kinetics similar to behaviors observed in vitro. A parameter set is identified based on physical dimensions of an AA in a rat kidney. Model results suggest that the interaction of Ca(2+) and K(+) fluxes mediated by voltage-gated and voltage-calcium-gated channels, respectively, gives rise to periodicity in the transport of the two ions. This results in a time-periodic cytoplasmic calcium concentration, myosin light chain phosphorylation, and cross-bridge formation with the attending muscle stress. Furthermore, the model predicts myogenic responses that agree with experimental observations, most notably those which demonstrate that the renal AA constricts in response to increases in both steady and systolic blood pressures. The myogenic model captures these essential functions of the renal AA, and it may prove useful as a fundamental component in a multiscale model of the renal microvasculature suitable for investigations of the pathogenesis of hypertensive renal diseases.

Dynamics of tubuloglomerular feedback adaptation to acute and chronic changes in body fluid volume

Experiments were performed in anesthetized rats to investigate the time course of the adaptive changes in tubuloglomerular feedback reactivity and juxtaglomerular renin activity induced by DOCA and NaCl loading (isotonic saline as drinking fluid plus daily injections of 25 mg/kg DOCA). In these DOCA escaped rats we studied further the effect of acute volume depletion and repletion on the magnitude of feedback responses. A significant reduction of feedback reactivity was found on the fifth day after starting the DOCA-salt treatment with maximum depression being established after two weeks. In contrast, significant reduction of juxtaglomerular renin content required 10 days and maximum depression three weeks of DOCA-salt administration. In DOCA-salt treated animals with severely curtailed feedback responses acute volume depletion was induced by a single injection of furosemide (10 mg/kg) leading to an excretion of 0.49 ±0.17 mEq sodium and a mean decrease of body weight of 2.17±0.22% over a period of 60–70 min. Mean feedback response (expressed as percent change of early proximal flow rate caused by elevating loop of Henle flow rate from zero to 40 nl/min) was −43.9 ±4.0%, significantly greater than in the pre-depletion phase and indistinguishable from responses seen in normal control rats. Repletion of extracellular volume by infusing HCO3-Ringer solution over 20 min induced a reduction in feedback responsivity to −12.0±2.9%. Plasma and juxtaglomerular renin concentration increased with volume depletion and decreased with volume repletion, but remained considerably lower than observed in normal control rats. Our results suggest that the tubuloglomerular feedback system responds to acute alterations in extracellular volume with rapid adaptations in its reactivity which is consistent with its role in the control of sodium excretion.

Kidney Modeling: Status and Perspectives

Mathematical models have played an essential role in elucidating various functions of the kidney, including the mechanism by which the avian and mammalian kidney can produce a urine that is more concentrated than blood plasma, quasi-isosmotic reabsorption along the proximal tubule, and the control and regulation of glomerular filtration by the myogenic and tubuloglomerular feedback mechanisms. This review includes a brief description of relevant renal physiology, a summary of the contributions of mathematical models at various levels and describes our recent work toward the Renal Physiome.

Nonlinear filter properties of the thick ascending limb

A mathematical model was used to investigate the filter properties of the thick ascending limb (TAL), that is, the response of TAL luminal NaCl concentration to oscillations in tubular fluid flow. For the special case of no transtubular NaCl backleak and for spatially homogeneous transport parameters, the model predicts that NaCl concentration in intratubular fluid at each location along the TAL depends only on the fluid transit time up the TAL to that location. This exact mathematical result has four important consequences: 1) when a sinusoidal component is added to steady-state TAL flow, the NaCl concentration at the macula densa (MD) undergoes oscillations that are bounded by a range interval envelope with magnitude that decreases as a function of oscillatory frequency; 2) the frequency response within the range envelope exhibits nodes at those frequencies where the oscillatory flow has a transit time to the MD that equals the steady-state fluid transit time (this nodal structure arises from the establishment of standing waves in luminal concentration, relative to the steady-state concentration profile, along the length of the TAL); 3) for any dynamically changing but positive TAL flow rate, the luminal TAL NaCl concentration profile along the TAL decreases monotonically as a function of TAL length; and 4) sinusoidal oscillations in TAL flow, except at nodal frequencies, result in nonsinusoidal oscillations in NaCl concentration at the MD. Numerical calculations that include NaCl backleak exhibit solutions with these same four properties. For parameters in the physiological range, the first few nodes in the frequency response curve are separated by antinodes of significant amplitude, and the nodes arise at frequencies well below the frequency of respiration in rat. Therefore, the nodal structure and nonsinusoidal oscillations should be detectable in experiments, and they may influence the dynamic behavior of the tubuloglomerular feedback system.