Nitric oxide synthesis influences the renal vascular responseto heme oxygenase - 《American Journal of Physiology》美国生理学杂志干细胞之家



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发表于 2009-4-21 13:37 |只看该作者 |倒序浏览 |打印

作者：FranciscaRodriguez, FanZhang, SandraDinocca,  AlbertoNasjletti作者单位：Department of Pharmacology, New York Medical College, Valhalla, NewYork 10595
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      【摘要】
   Westudied the effects of the heme oxygenase (HO) inhibitor stannousmesoporphyrin (SnMP; 40 µmol/kg iv) on renal hemodynamics inanesthetized rats with and without 48-h pretreatment with N G -nitro- L -arginine methyl ester( L -NAME), an inhibitor of nitric oxide (NO) synthesis. SnMPdecreased renal blood flow (RBF) and increased renal vascularresistance (RVR) in both groups. The SnMP-induced reduction of RBF in L -NAME-pretreated rats was more prominent than in ratswithout pretreatment (43 ± 7 vs. 13 ± 3%) as was theSnMP-induced elevation of RVR (87 ± 31 vs. 14 ± 5%). Therenal vasoconstrictor effect of SnMP is linked, in part, to amplification of prevailing neurohormonal constrictor mechanisms, sincein L -NAME-pretreated rats it was prevented by concurrent administration of prazosin or losartan. However, SnMP (15 µmol/l) also elicits vasoconstriction in isolated, pressurized renalinterlobular arteries and the response is more intense in vesselsobtained from L -NAME-pretreated rats than from rats withoutpretreatment. These data indicate that the status of NO synthesisconditions the vascular response to HO inhibition in the rat kidney.
      【关键词】 carbon monoxide renal hemodynamics angiotensin II norepinephrine renal vascular reactivity
   INTRODUCTION" @2 `$ W/ N9 w
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HEME OXYGENASE (HO)-1 and -2 metabolize heme to biliverdin, ferrous iron, and carbonmonoxide (CO) ( 1, 18 ). The heme-HO system is ubiquitousand relevant to many biological processes that are influenced by hemeor by the products of heme metabolism by HO ( 1, 10, 11, 18, 26 ). In the kidney, HO-1 and HO-2 are expressed in both vascularand tubular structures, and HO-1 is upregulated in experimental modelsof renal injury ( 2, 5, 11, 16, 28 ). The renal heme-HOsystem was reported to promote cytoprotective mechanisms( 11 ) and to participate in the regulation of renalfunction ( 2, 19, 28 ) and eicosanoid production( 4 ).

Metalloporphyrins which inhibit HO decrease renal medullary blood flowin normal rats ( 28 ) and total renal blood flow in chronically hypoxic rats ( 19 ), suggesting the contributionof one or more HO products to renal vasodilatory mechanisms. HO-derived CO is a logical candidate for subserving a renal vasodilatory function,because exogenous CO dilates isolated, perfused rat afferent arterioles( 23 ) and decreases the reactivity of renal interlobararteries to constrictor agonists ( 16 ). CO-induced vasorelaxation and inhibition of agonist-induced vasoconstriction havebeen linked to activation of soluble guanylyl cyclase ( 24 ) and/or stimulation of calcium-activated potassium (K Ca )channels ( 16, 24 ) in vascular smooth muscle.

Recent studies have identified areas of interaction between the heme-HOand the L -arginine-nitric oxide (NO) synthase systems thatmay have great functional relevance. On one hand, NO interferes withthe ability of CO to stimulate K Ca channels in vascularsmooth muscle ( 25 ), induces HO-1 expression ( 8, 9 ), and decreases HO activity ( 6, 15 ) as well asferrochelatase activity ( 22 ), the terminal enzyme of theheme synthetic pathway. On the other hand, CO attenuates NO-inducedactivation of soluble guanylyl cyclase ( 12 ) and inhibitsNO synthase ( 23 ). That CO and NO affect the formation andaction(s) of each other raises the possibility that the vasoregulatoryfunctions of the heme-HO and the L -arginine-NO synthasesystems are interdependent.
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To the extent that NO influences the formation and actions ofHO-derived CO, the status of NO synthesis may determine the natureand/or intensity of the regulatory influence of the heme-HO system onthe renal vasculature. To test this hypothesis, we contrasted untreatedrats and rats undergoing treatment with an inhibitor of NO synthase interms of the renal hemodynamic response to the administration ofstannous mesoporphyrin (SnMP), a nonselective inhibitor of HO isoforms( 7 ). In addition, we compared the effect of SnMP and HOproducts, CO and biliverdin, on the internal diameter of isolated,pressurized renal interlobular arteries taken from normal untreatedrats and rats treated with an inhibitor of NO synthesis.

MATERIALS AND METHODS

Chemicals and Solutions/ ]6 A( e& L1 y9 T
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SnMP, an inhibitor of HO, was purchased from Frontier Scientific(Logan, UT). SnMP was dissolved in 50 mmol/lNa 2 CO 3, sonicated, and filtered immediatelybefore use. Because of the photosensitivity of porphyrins, SnMPsolutions were prepared and experiments were performed under reducedlight conditions. CO was purchased from Tech Air (White Plains, NY),and a CO-saturated solution (1 mmol/l) was prepared shortly before use( 17 ). Other chemicals were obtained from Sigma (St. Louis,MO) and diluted in NaCl (0.15 mol/l). The composition of Krebs bufferused in the studies was (in mmol/l) 118.5 NaCl, 4.7 KCl, 2.5 CaCl 2, 1.2 KH 2 PO 4, 1.2 MgSO 4, 25.0 NaHCO 3, and 11.1 dextrose.
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Animals9 _, m" p0 k+ X: K3 h% a& M

All protocols employed in this study were reviewed and approvedby the Institutional Animal Care and Use Committee of New York MedicalCollege. We used male Sprague-Dawley rats (300-325 g body wt;Charles River, Wilmington, DE) with access to a standard chow and tapwater ad libitum. Studies were conducted in animals treated and nottreated with N G -nitro- L -argininemethyl ester ( L -NAME), an inhibitor of NO synthase ( 21 ). L -NAME was provided in the drinkingwater at a daily dose of 70 mg/kg body wt commencing 48 h before experimentation.4 s1 V9 J! c9 l, k6 L# W
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Experimental Design: O# z/ w4 e( F
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Protocol to contrast the effect of SnMP on renal hemodynamics inrats treated and not treated with L -NAME. Animals were anesthetized with thiobutabarbital (100 mg/kg ip) andplaced on a thermostatically controlled board to maintain bodytemperature at 36-37°C. Polyethylene cannulas were placed in thetrachea (PE-205) to aid ventilation, the bladder (PE-60) for urinecollection, the left femoral vein (PE-50) for administration of fluidand drugs, and the left femoral artery (PE-50) for blood sampling andthe measurement of mean arterial pressure by means of a pressuretransducer (model p23 ID, Oxnard, CA) coupled to a polygraph (model 7D,Grass Instruments, Quincy, MA). Next, the left kidney was exposedthrough a midline abdominal incision and a flow probe (model EP102,2.0-mm circumference, Carolina Medical Instruments, King, NC) wasplaced around the left renal artery for measurement of renal blood flowusing a square-wave electromagnetic flowmeter (model FM 501, CarolinaMedical Instruments). After completion of the cannulation andinstrumentation procedures, an infusion of 0.15 mol/l NaCl (2.5 ml/hiv) was initiated and maintained throughout the study. In someexperiments, [ 3 H]inulin (American Radiolabeled Chemicals,St. Louis, MO) was included into the infusion (1 µCi/ml) formeasurement of glomerular filtration rate. To this end, timed urinecollections were made before and during experimental interventions, andarterial blood samples (200 µl) were collected at the midpoint ofurine collections. The concentration of [ 3 H]inulin inurine and plasma was determined by liquid scintillation counting andthe clearance of inulin, calculated using the standard formula, wastaken to reflect the glomerular filtration rate. Renal vascularresistance was calculated by dividing the value of mean arterialpressure by the value of renal blood flow. The filtration fraction wascalculated by the formula: glomerular filtration rate/renal blood flow(1 hematocrit). Data collection was initiated after a 60-minequilibration period.
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Control data on mean arterial pressure and renal hemodynamics werecollected over a 30-min control period immediately before theadministration of SnMP (40 µmol/kg iv) to rats pretreated ( n = 8) and not pretreated with L -NAME( n = 14); data were collected again over a 30-minexperimental period commencing 45 min after the onset of SnMPtreatment. An identical protocol was used to collect data on renalfunction before and during administration of drug vehicle only (50 mmol/l Na 2 CO 3 ) in rats pretreated( n = 8) and not pretreated ( n = 12)with L -NAME.

Inhibition of NO synthesis fosters the activity of constrictormechanisms involving the sympathetic nervous and the renin-angiotensin systems ( 3 ). This may condition the effect of SnMP onrenal hemodynamics, because the response of small arterial vessels to constrictor stimuli is amplified by HO inhibition ( 16 ). Toexamine this possibility, the effect of SnMP (40 µmol/kg iv) on meanarterial pressure, renal blood flow, and renal vascular resistance was also investigated in rats pretreated with L -NAME along witheither prazosin (1 mg/kg sc; n = 6) or losartan (10 mg/kg iv; n = 7), respectively, to block 1 -adrenergic and angiotensin AT 1 receptors ( 20 ). In these experiments, rats undergoing L -NAME pretreatment for 48 h were injected withprazosin or losartan followed, 45 min later, by examination of theeffects of SnMP using a protocol similar to that described above.9 B5 i) W7 R# P" e- E6 T
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A complementary study was conducted in rats without L -NAMEpretreatment to determine whether SnMP (40 µmol/kg iv) affects therenal circulatory response to norepinephrine and ANG II. Rats wereinstrumented with a cannula in the left carotid artery to measure meanarterial pressure and a flow probe on the left renal artery to measurerenal blood flow. Norepinephrine (2 nmol · kg 1 · min 1; n = 5) or ANG II (5 pmol · kg 1 · min 1; n = 8) was administered by infusion over a 15- to20-min period via a polyethylene cannula made of stretched PE-10 tubingthat was inserted into the left femoral artery and advanced through theabdominal aorta into the left renal artery. After discontinuation ofconstrictor agonist infusion, a 30-min recovery period was allowed,SnMP (40 µmol/kg iv) was injected, and 45 min later the infusion ofconstrictor agonist was repeated.# s  ^% d+ K% U. A
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Renal HO activity was measured in a limited number of control untreatedrats, rats undergoing treatment with L -NAME for 48 h,and rats 1.0-1.5 h after administration of SnMP (40 µmol/kg iv).Kidneys were homogenized, the homogenate was centrifuged, and the10,000- g supernatant was analyzed for HO activity using [ 14 C]heme (Leeds Radioporphyrins, Leeds, UK) as substrate( 16 ). HO activity is expressed as picomoles of bilirubingenerated per milligram of protein per hour.
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Protocols to investigate the effects of CO and the HO inhibitorSnMP in isolated renal interlobular arteries taken from rats treatedand not treated with L -NAME. Animals were anesthetized (60 mg/kg ip pentobarbital sodium), thekidneys were removed and placed on a dish containing ice-cold Krebsbuffer, and interlobular arteries were dissected free of surroundingtissue. Vascular segments 1- to 2-mm in length, with an internaldiameter of ~40 µm in the absence of flow and transmural pressure,were mounted between two micropipettes in the chamber (1 ml) of apressure-myograph (Living System Instrumentation, Burlington, VT)filled with Krebs buffer gassed with 95% O 2 -5% CO 2, which was exchanged at a rate of 1 ml/min( 27 ). To measure vascular diameter, the vessel chamber wasplaced on the stage of a microscope fitted with a video camera(Javelin, Newburgh, NY) linked to a video caliper (Texas A & M, CollegeStation, TX) and a recorder ( 27 ). Intraluminal pressurewas increased slowly to 100 mmHg using a pressure servo-controller, andthis level of pressure was maintained throughout the study unlessindicated otherwise. Experiments were conducted after a 60-minequilibration period in vessels that developed a spontaneous tone whilepressurized to 100 mmHg. The internal diameter of vessels was measuredcontinuously before and after inclusion of SnMP (15 µmol/l), CO (0.1 and 1.0 µmol/l), or biliverdin (1 µmol/l) into the Krebs bufferflowing into the myograph chamber.

To gain information on whether NO synthesis remains inhibited during exvivo superfusion of vessels taken from L -NAME-treated rats,a limited number of experiments were conducted to contrast the effectof L -arginine (10 µmol/l) on the diameter of pressurized renal interlobular arteries taken from rats with and without L -NAME pretreatment. The vessels were superfused with Krebsbuffer for 90-120 min before the experimental intervention. Theinclusion of L -arginine in the superfusion buffer increased( P internal diameter of arteries takenfrom rats without L -NAME pretreatment from 69.7 ± 3.1 to 77.5 ± 5.0 µm ( n = 4). In contrast, L -arginine did not change the diameter of arteries takenfrom rats pretreated with L -NAME (68.0 ± 3.9 vs.68.0 ± 3.9 µm; n = 4). That the dilatory effectof L -arginine is blunted in vessels from rats pretreated with L -NAME implies that NO synthesis in such vesselsremains inhibited even after ex vivo superfusion for up to 90-120 min.0 p8 |( o  i! e
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Additional experiments were conducted to contrast the effect of SnMP(15 µmol/l) on the intraluminal pressure-internal diameter relationship in renal interlobular arteries taken from rats treated andnot treated with L -NAME. After equilibration at 100 mmHgfor 60 min, intraluminal pressure was decreased to ~0 mmHg and, after 10 min, it was increased in 20-mmHg steps until it reached 100 mmHg.The pressure was maintained for ~5-10 min at each pressure stepso that the vessels could reach a steady-state diameter. Finally, thevascular preparation was superfused with calcium-free Krebs buffercontaining 1 mmol/l EGTA and the pressure-diameter relationship wasexamined again to obtain the passive diameter of the vessels at eachlevel of intraluminal pressure. The internal diameter duringsuperfusion of the vessel with calcium-containing buffer (absolutediameter) and with calcium-free buffer (passive diameter) is expressedin micrometers. The normalized diameter refers to the absolute diameterexpressed as a percentage of the passive diameter.  p0 z( \! T# ^# N; m

Data Analysis5 B1 z: m) l5 W5 j, M
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Results are expressed as means ± SE. Data on renalhemodynamics in vivo are the average of two consecutive 15-minobservation periods. Data were analyzed by unpaired Student's t -test and by one- or two-way ANOVA followed by theNewman-Keuls post hoc test or the Fisher test. The null hypothesis wasrejected at P
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Effects of SnMP on Renal Hemodynamics
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Table 1 shows data on renalhemodynamics before and after the administration of SnMP or drugvehicle only to rats pretreated and not pretreated with the NOsynthesis inhibitor L -NAME. Relative to data in ratswithout L -NAME pretreatment, rats pretreated with L -NAME displayed elevated mean arterial pressure, decreasedrenal blood flow, and increased renal vascular resistance, filtration fraction, and hematocrit, without significant changes in glomerular filtration rate. Estimates of renal HO activity did not differ significantly in rats with (96 ± 16 pmol · mg 1 · h 1; n = 4) and without (125 ± 14 pmol · mg 1 · h 1; n = 4) L -NAME pretreatment; renal HOactivity was decreased (15 ± 1 pmol · mg 1 · h 1; n = 8; P inhibitor SnMP (40 µmol/kg iv). In ratswithout L -NAME pretreatment, the administration ofSnMP did not change mean arterial pressure but decreased renalblood flow and increased renal vascular resistance without affectingthe glomerular filtration rate or filtration fraction. In ratspretreated with L -NAME, the administration of SnMP loweredarterial pressure slightly, decreased renal blood flow, and increasedrenal vascular resistance; the treatment did not affect glomerularfiltration but greatly increased the filtration fraction. TheSnMP-induced reduction of renal blood flow was greater ( P L -NAME(43 ± 7%) than in rats without L -NAME pretreatment(13 ± 3%). The SnMP-induced increase of renal vascularresistance was also greater ( P L -NAME-pretreated rats (87 ± 31%) than in ratswithout pretreatment (14 ± 5%). Thus the administration of theHO inhibitor SnMP produces renal vasoconstriction and this effect isintensified in rats pretreated with an inhibitor of NO synthesis. Atvariance with the pronounced effects of SnMP on renal hemodynamics, theadministration of the drug vehicle alone affected renal hemodynamics inneither rats with or without L -NAME pretreatment.8 p5 w* F6 ?. ?0 S5 h
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Table 1. Effect of SnMP on renal hemodynamics

The possibility that SnMP elicits renal vasoconstriction by amplifyingthe renal vascular actions of endogenous vasoconstrictors was examinedby contrasting the effects of SnMP on renal hemodynamics in ratspretreated with L -NAME alone, L -NAME plusprazosin to block adrenergically mediated vasoconstriction, and L -NAME plus losartan to block ANG II-inducedvasoconstriction. Relative to corresponding data in rats pretreatedwith L -NAME alone, before the administration of SnMP, ratspretreated with L -NAME and prazosin displayed decreasedmean arterial pressure (112 ± 8 vs. 130 ± 3 mmHg, P 26.2 ± 1.4 mmHg · ml 1 · min · g 1, P to increase (6.2 ± 0.6 vs. 5.1 ± 0.4 ml · min 1 · g 1, P = 0.14). Rats pretreated with L -NAME andlosartan exhibited unchanged mean arterial pressure (129 ± 6 mmHg), diminished renal vascular resistance (15.8 ± 2.1 mmHg · ml 1 · min · g 1, P 1 · g 1, P L -NAMEalone, SnMP decreased mean arterial pressure, reduced renal blood flow,and increased renal vascular resistance but had no effect on any ofthese parameters in rats pretreated with L -NAME andprazosin (Fig. 1 ). In rats pretreatedwith L -NAME and losartan, SnMP slightly reduced mean arterial pressure and renal blood flow but was without effect on renalvascular resistance (Fig. 1 ). Thus the renal vasoconstriction inducedby the HO inhibitor, in rats pretreated with L -NAME, is blunted in animals undergoing pharmacological blockade of 1 -adrenergic or angiotensin AT 1 receptors." d$ h# c; W5 j1 y  ]
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Fig. 1. Mean arterial pressure (MAP), renal blood flow (RBF), andrenal vascular resistance (RVR) before (open bars) and after (filledbars) treatment with stannous mesoporphyrin (SnMP; 40 µmol/kg iv) inrats pretreated with N G -nitro- L -arginine methyl ester( L -NAME) alone ( n = 8) and concurrentlywith prazosin ( n = 6) or losartan ( n = 7). Results are means ± SE. * P P L -NAME alone.. v- Y: R. v& Y+ ^! _$ ?7 }# u9 s6 V
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Figure 2 illustrates the result ofexperiments comparing the effect of renal arterial infusion ofnorepinephrine (2 nmol · kg 1 · min 1 ), or ANGII (5 pmol · kg 1 · min 1 ),on renal hemodynamics in rats before and after treatment with SnMP.Both constrictor agonists decreased renal blood flow and increasedrenal vascular resistance without altering mean arterial pressure. Therenal vasoconstrictor effect of norepinephrine and ANG II wassignificantly magnified after the administration of SnMP. Thusinhibition of HO amplifies the renal vasoconstrictor action ofnorepinephrine and ANG II.

Fig. 2. MAP and RBF before (open bars) and during (filled bars) renalarterial infusion of norepinephrine (NE; n = 5) or ANGII ( n = 8), both before and after administration ofSnMP (40 µmol/kg iv). Results are means ± SE.* P P% s3 M" A1 n3 A  P4 E& L

Effect of SnMP on the Internal Diameter of Isolated, PressurizedRenal Interlobular Arteries" w, b  v9 s& R; B0 h1 E% z

Inclusion of SnMP into the Krebs buffer superfusing pressurizedrenal interlobular arteries elicited a sustained reduction of internaldiameter in vessels taken from rats treated and not treated with L -NAME (Fig. 3 ). In vesselsobtained from rats not treated with L -NAME, SnMP decreased( P by 6.0 ± 0.8, 8.5 ± 1.1, 9.5 ± 1.1, and 9.8 ± 1.1 µm after 5, 10, 20, and 30 min of exposure to the HO inhibitor, respectively. Invessels obtained from rats treated with L -NAME, SnMPdecreased ( P ± 1.3, and 23.3 ± 1.4 µm after 5, 10, 20, and 30 min of exposure to the HO inhibitor,respectively. Importantly, at all time points, the SnMP-inducedreduction of internal diameter was greater ( P L -NAME-treated rats than invessels obtained from untreated rats. Thus exposure to SnMP bringsabout constriction of renal interlobular arteries and this effect is intensified in vessels taken from rats treated with an inhibitor of NOsynthesis.: C' W, e6 @# W& i! ?) x) t' `$ u

Fig. 3. Internal diameter (ID) of pressurized interlobulararteries obtained from rats with and without 48-h pretreatment with L -NAME, before and during superfusion with buffercontaining SnMP (15 µmol/l). Results are means ± SE.* P) r8 p& L( i$ D6 f9 Y5 r
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The vasoconstrictor effect of HO inhibitors in isolated, pressurizedresistance vessels has been linked to magnification of the prevailingmyogenic tone ( 17 ). To investigate whether SnMP promotesmyogenic behavior and, if so, to determine whether this effect isconditioned by the status of NO synthesis, we contrasted the effect ofSnMP on the pressure-diameter relationship in isolated renalinterlobular arteries obtained from rats treated and not treated with L -NAME. As shown in Fig. 4,stepwise elevation of intraluminal pressure elicited apressure-dependent reduction ( P and not treated with L -NAME. Before exposure to SnMP, vessels from rats treatedand not treated with L -NAME did not differ from each otherin terms of internal diameter over the pressure range 20 to 100 mmHg.The inclusion of SnMP into the superfusion buffer enhanced( P diameter in both groups of vessels. In preparations exposed to SnMP,the internal diameter reduction of vessels obtained from L -NAME-treated rats surpassed ( P from rats not treated with L -NAMEover the pressure range of 40 to 100 mmHg. Thus SnMP enhancespressure-induced constriction of renal interlobular arteries andthis effect is further intensified in vessels obtained from ratstreated with a NO synthesis inhibitor.
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Fig. 4. Effect of stepwise increments in intraluminal pressure onthe normalized ID of renal interlobular arteries superfused with buffercontaining and not containing SnMP (15 µmol/l). The studies wereconducted in vessels obtained from rats with and without 48-hpretreatment with L -NAME. Results are means ± SE.* P
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To the extent that the vascular response to an inhibitor of HO resultsfrom decreased vascular formation of an HO product capable ofinfluencing vasomotor function, the increased constrictor effect ofSnMP in renal interlobular arteries of L -NAME-treated ratsmay be linked to alterations in the responsiveness of such vessels tothe products of HO activity. This possibility was investigated bycomparing the effects of CO and biliverdin on the internal diameter ofisolated, pressurized renal interlobular arteries taken from rats withand without L -NAME treatment. As shown in Fig. 5, the inclusion of 1 µmol/l CO intothe superfusion buffer elicited a sustained reduction( P in vessels taken fromrats not treated with L -NAME. In contrast, 1 µmol/l CObrought about augmentation of internal diameter in vessels taken fromrats treated with L -NAME. At 0.1 µmol/l, CO was withouteffect on the diameter of vessels taken from either rats with andwithout L -NAME treatment. Biliverdin at 1 µmol/l didnot affect the internal diameter of renal interlobular arteries obtained from either untreated rats ( n = 4; 68 ± 2 and 69 ± 2 µm before and 30 min after, respectively) or L -NAME-treated rats ( n = 4; 67 ± 4 and 68 ± 4 µm before and 30 min after, respectively).
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Fig. 5. ID of pressurized renal interlobular arteries from ratswith and without 48-h pretreatment with L -NAME, before andduring superfusion with buffer containing exogenous carbon monoxide(CO). Results are means ± SE. * P

DISCUSSION

The present studies provide information on whether the status ofNO synthesis influences the renal vascular response to treatment withan inhibitor of HO. In anesthetized rats, we found that the administration of SnMP increases renal vascular resistance and lowersrenal blood flow and that both responses are more intense in ratspretreated with L -NAME than in control rats withoutpretreatment. Intensification after L -NAMEpretreatment of the renal vasoconstrictor and blood flow-loweringeffects of SnMP was accompanied by a major elevation in filtrationfraction, an observation consistent with SnMP-induced vasoconstrictionat postglomerular sites and/or increase of the ultrafiltrationcoefficient. We also found that the exposure of isolated, pressurizedrenal interlobular arteries to SnMP elicits sustained reduction ofinternal diameter and that this response is more intense in vesselstaken from L -NAME-treated rats than in vessels fromuntreated controls. Hence, the results of both the in vivo and ex vivostudies indicate that the expression of renal vasoconstriction aftertreatment with SnMP is magnified in settings in which NO synthesis isinhibited with L -NAME. Inhibition of NO synthesis alsofacilitates development of vasoconstriction after treatment with aninhibitor of HO in rat hindlimbs in vivo and in isolated gracilismuscle arterioles ( 13 ).' @, j" x& j* e9 y1 t) Y, t6 ?! p
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According to the present study, in vitro estimates of renal HO activityin L -NAME-treated rats do not differ significantly fromestimates in untreated control rats. Yet, there are reports thatexogenous NO decreases HO activity ( 6, 15 ), which prompt expectation of enhanced HO product generation during NO synthesis inhibition. In line with this notion, a preliminary assessment of CO inthe headspace of urine samples, using gas chromatography-mass spectroscopy ( 16 ), indicates that the urinary excretion ofCO in anesthetized rats undergoing pretreatment with L -NAMEfor 48 h (15.3 ± 4.5 pmol/min; n = 4)surpasses ( P rate inuntreated controls (2.2 ± 0.5 pmol/min; n = 4)(Rodriguez F, Kemp R, and Nasjletti A; unpublished observations). Inrats pretreated with L -NAME, augmentation of HO productgeneration in vivo may condition intensification of SnMP-induced renal vasoconstriction.
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Treatment with SnMP magnifies the reduction of renal blood flowproduced by renal arterial infusion of norepinephrine or ANG II. Thisfinding fits well with reports that interventions that attenuate theexpression or activity of HO enhance the sensitivity of renal arterialvessels to constrictor agonists ( 16 ). It is conceivable,then, that the renal vasoconstrictor effect of SnMP is linked, at leastin part, to amplification of prevailing neurohormonal constrictormechanisms. This seems to be the case in anesthetized rats treated with L -NAME, because concurrent treatment with prazosin orlosartan prevents SnMP from increasing renal vascular resistance.# Z( ~) ?  |7 Z7 o( m& j# U

Previous studies indicate that the increase in renal vascularresistance elicited by the systemic administration of an inhibitor ofNO synthesis relies, in part, on a constrictor mechanism involving thesympathetic nervous and renin-angiotensin systems ( 3 ). Inline with this notion, we found that the values of renal vascular resistance in rats treated with L -NAME alone exceed thecorresponding values in rats concurrently treated with L -NAME and prazosin or L -NAME and losartan. Inview of these findings, consideration should be given to thepossibility that intensification of the renal vasoconstrictor effect ofSnMP in L -NAME-treated rats is conditioned by therecruitment, after inhibition of NO synthesis, of a constrictormechanism dependent on the sympathetic nervous and renin-angiotensin systems.) I8 L. M% A  ^. w1 Q5 g. z! H

According to the present study, the conditioning influence of L -NAME pretreatment on SnMP-induced vasoconstriction isalso demonstrable in renal interlobular arteries, isolated andpressurized, under experimental conditions that preclude participationof the sympathetic nervous and renin-angiotensin systems in vasomotor regulation. The constrictor effect of SnMP in these vessels is attributable to amplification of the prevailing myogenic tone, asdocumented previously in gracilis muscle arterioles ( 17, 27 ), because inclusion of SnMP into the superfusion buffermagnifies the reduction of vessel diameter produced by stepwiseelevation of intraluminal pressure. This effect of SnMP enhancingmyogenic constrictor responsiveness is prominently expressed in renalarteries taken from rats pretreated with L -NAME, whichcould explain the greater constrictor effect of the HO inhibitor insuch vessels.

A priori, the renal vasoconstrictor effect of SnMP may be expected toresult from diminished synthesis of one or more HO products capable ofeffecting vasodilation. Biliverdin does not dilate pressurized renalinterlobular arteries and, therefore, it is unlikely that a decreasedproduction of biliverdin contributes to the vasoconstrictor effect ofthe HO inhibitor. Because renal arteries taken from rats without L -NAME pretreatment respond to exogenous CO withconstriction, like gracilis muscle arterioles not treated with aninhibitor of NO synthesis ( 14 ), the constrictor effect ofSnMP cannot be explained on the basis of diminished CO production insuch vessels, unless it is postulated that exogenous CO and COmanufactured by vascular structures have divergent vasomotor activity.On the other hand, exogenous CO was found to cause prompt dilation ofrenal arteries taken from rats pretreated with L -NAME, which is in agreement with the notion that the constrictor effect ofSnMP in such vessels is a consequence of reduced CO production. On thebasis of these observations, intensification of the renal vasoconstrictor effect of SnMP in L -NAME-treated rats maybe conditioned by the preeminence of renal vasodilatory mechanismsmediated by endogenous CO in settings in which NO synthesis is impaired.

The vasodilatory effect of exogenous CO is ascribed to activation ofsoluble guanylate cyclase ( 24 ) and/or stimulation of K Ca channels ( 16, 24 ) in vascular smoothmuscle, whereas the vasoconstrictor effect is attributed to inhibitionof NO production by the endothelium ( 14 ). That renalarteries from rats without L -NAME pretreatment respond toexogenous CO with constriction rather than dilation implies thatCO-induced inhibition of NO synthesis subserves a vasoconstrictoraction that prevails over the direct vasodilatory actions of the gasmediator. On the other hand, when the ability of CO to inhibit NOsynthesis is rendered inconsequential, as in renal arteries taken fromrats pretreated with L -NAME, the vasodilatory actions of COare unmitigated. The preferential expression of CO-induced dilation ofvessels from rats pretreated with L -NAME may also be due tofacilitation of CO-induced stimulation of K Ca channels invascular smooth muscle, an action that is inhibited by NO( 25 ).

From the preceding discussion, it would appear that the net effect ofCO, both exogenous CO and CO manufactured by vascular tissues, isdetermined by the balance between actions on the smooth muscle, toeffect vasodilation, and actions on the endothelium, to inhibit NOsynthesis and bring about vasoconstriction. If so, the nature andintensity of the vasomotor response elicited by exogenous CO and CO ofvascular origin may be determined not only by the status of NOsynthesis but also by the relative access of the gas mediator to sitesof actions in the endothelium and smooth muscle. For example, invessels from rats without L -NAME pretreatment, COmanufactured by the smooth muscle may be expected to preferentiallypromote vasodilation, reflecting unfettered access to sites of actionin smooth muscle coupled to suboptimal access to sites of action in theendothelium, whereas exogenous CO and CO produced by the endotheliumare expected to preferentially promote vasoconstriction.

In summary, this study demonstrates that the HO inhibitor SnMP producesrenal vasoconstriction and that this effect is greatly magnified inrats pretreated with L -NAME. The study also shows that SnMPamplifies the renal vasoconstriction induced by norepinephrine or ANGII, that pretreatment with either prazosin or losartan prevents the HOinhibitor from increasing renal vascular resistance in L -NAME-treated rats, and that CO elicits dilation andconstriction, respectively, of renal interlobular arteries taken fromrats with and without L -NAME pretreatment. Thus the statusof NO synthesis impacts importantly on the regulatory influence of theheme-HO system on the renal vasculature.
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ACKNOWLEDGEMENTS

We thank J. Brown for secretarial assistance.
      【参考文献】; q7 q# j8 Q3 E: j1 d, g* h
1. Abraham, NG,Drummond GS,Lutton JD,andKappas A. The biological significance and physiological role of heme oxygenase. Cell Physiol Biochem 247:725-732,1997.

7 ~0 `2 M# ~, Y/ K$ |  @; r

2. Aizawa, T,Ishizaka N,Taguchi J,Nagai R,Mori I,Tang SS,Ingelfinger JR,andOhno M. Heme oxygenase-1 is upregulated in the kidney of angiotensin II-induced hypertensive rats: possible role in renoprotection. Hypertension 35:800-806,2000 .
8 K; S; K9 d0 N' i" V

3. Baylis, C,andQiu C. Importance of nitric oxide in the control of renal hemodynamics. Kidney Int 49:1727-1731,1996  .# d% F3 X6 [7 D$ |9 F

# A. T* X* A* t% ?* L( I

4. Botros, FT,Laniado-Schwartzman M,andAbraham NG. Regulation of cyclooxygenase- and cytochrome p450-derived eicosanoids by heme oxygenase in the rat kidney. Hypertension 39:639-644,2002 .

5. Da Silva, JL,Zand BA,Yang LM,Sabaawy HE,Lianos E,andAbraham NG. Heme oxygenase isoform-specific expression and distribution in the rat kidney. Kidney Int 59:1448-1457,2001  .

  q0 F3 l; |; `0 d& c5 q
7 ~! c: ~  W3 O. _" {) Y
6. Ding, Y,McCoubrey WK,andMaines MD. Interaction of heme oxygenase-2 with nitric oxide donors. Is the oxygenase an intracellular "sink" for NO? Eur J Biochem 264:854-861,1996.% N9 y/ k- E- Z5 k
  U8 O  `+ Z1 b3 t( l5 \
4 j& \; Z. F. i3 F) l) H# F0 H

7. Drummond, GS,Galbraith RA,Sardana MK,andKappas A. Reduction of the C2 and C4 vinyl groups of Sn-protoporphyrin to form Sn-mesoporphyrin markedly enhances the ability of the metalloporphyrin to inhibit in vivo heme catabolism. Arch Biochem Biophys 255:64-74,1987  .

5 u% b2 W- x/ g+ H
8. Durante, W,Kroll MH,Christodoulides N,Peyton KJ,andSchafer AI. Nitric oxide induces heme oxygenase-1 gene expression and carbon monoxide production in vascular smooth cells. Circ Res 80:557-564,1997 .

( w; p2 \, n- ~4 x0 M7 L* V

9. Foresti, R,andMotterlini R. The heme oxygenase pathway and its interaction with nitric oxide in the control of cellular homeostasis. Free Radic Res 31:459-475,1999  .: h5 b: r0 S% l! y0 M

4 g3 a; `& p- x
10. Haider, A,Olszanecki R,Gryglewski R,Schwartzman ML,Lianos E,Nasjletti A,Kappas A,andAbraham NG. Regulation of cyclooxygenase by the heme-heme oxygenase system in microvessel endothelial cells. J Pharmacol Exp Ther 300:188-194,2002 .

! Q) U; d3 e" r' r
- I! `: `: L, S$ k
11. Hill-Kapturczak, N,Chang SH,andAgarwal A. Heme oxygenase and the kidney. DNA Cell Biol 21:307-321,2002  .7 u% l6 n) b  x* a/ p2 a
( m" C1 `. O% _: b& V. o: e: L
# ~# r( z8 b  t0 V1 Z9 ~; d
1 W* @9 B8 }! _7 g* E" E: D8 H
12. Ingi, T,Cheng J,andRonnett GV. Carbon monoxide: an endogenous modulator of the nitric oxide-cyclic GMP signaling system. Neuron 16:835-846,1996  .
/ l# x% k4 C, A9 k8 e  o/ R. g

13. Johnson, FK,Teran FJ,Prieto-Carrasquero M,andJohnson RA. Vascular effects of a heme oxygenase inhibitor are enhanced in the absence of nitric oxide. Am J Hypertens 15:1074-1080,2002  .

( Z* Z" T! p) l) r* M9 ~

14. Johnson, RA,andJohnson FK. The heme-heme oxygenase-carbon monoxide system and hypertension.In: Carbon Monoxide and Cardiovascular Functions, edited by Wang R.. Boca Raton, FL: CRC, 2002, p. 149-163.

  q2 W0 A$ {0 B( m' }+ D
15. Juckett, M,Zheng Y,Yuan H,Pastor T,Antholine W,Weber M,andVercellotti G. Heme and the endothelium. Effects of nitric oxide on catalytic iron and heme degradation by heme oxygenase. J Biol Chem 273:23388-23397,1998 .3 h! J, j6 U( o6 q
- g* y* I8 h# F5 j2 s
1 V8 ?! M$ J8 e; y6 O

16. Kaide, JI,Zhang F,Wei Y,Jiang H,Yu C,Wang WH,Balazy M,Abraham NG,andNasjletti A. Carbon monoxide of vascular origin attenuates the sensitivity of renal arterial vessels to vasoconstrictors. J Clin Invest 107:1163-1171,2001  .

17. Kozma, F,Johnson RA,Zhang F,Yu C,Tong X,andNasjletti A. Contribution of endogenous carbon monoxide to regulation of diameter in resistance vessels. Am J Physiol Regul Integr Comp Physiol 276:R1087-R1094,1999 .

- `  |( A9 q5 l9 q0 T
18. Maines, MD. The heme oxygenase system: a regulator of second messenger gases. Annu Rev Pharmacol Toxicol 37:517-554,1997  .

19. O'Donaughy, TL,andWalker BR. Renal vasodilatory influence of endogenous carbon monoxide in chronically hypoxic rats. Am J Physiol Heart Circ Physiol 281:H298-H307,2001 .5 W" w( R# ^# Z% e

, c; v& @# K% t) P& v" m
20. Pucci, ML,Lin L,andNasjletti A. Pressor and renal vasoconstrictor effects of N G -nitro- L -arginine as affected by blockade of pressor mechanisms mediated by the sympathetic nervous system, angiotensin, prostanoids and vasopressin. J Pharmacol Exp Ther 261:240-245,1992 .

, v6 D0 S7 z6 X8 a
9 A; d3 V8 o1 D' R9 z8 V
21. Rees, DD,Palmer RM,Schulz R,Hodson HF,andMoncada S. Characterization of three inhibitors of endothelial nitric oxide synthase in vitro and in vivo. Br J Pharmacol 101:746-752,1990  .

22. Sellers, VM,Johnson MK,andDailey HA. Function of the [2FE-2S] cluster in mammalian ferrochelatase: a possible role as a nitric oxide sensor. Biochemistry 35:2699-2704,1996 ., i7 I- {, r" w: J% n
5 L- t' I' \9 P9 P: t$ l

) q0 g0 V& I) g
23. Thorup, C,Jones CL,Gross SS,Moore LC,andGoligorsky MS. Carbon monoxide induces vasodilation and nitric oxide release but suppresses endothelial NOS. Am J Physiol Renal Physiol 277:F882-F889,1999 .; v0 r; z0 f. \$ J2 b3 l, l

24. Wang, R,Wang Z,andWu L. Carbon monoxide-induced vasorelaxation and the underlying mechanisms. Br J Pharmacol 121:927-934,1997  .; C, R4 v7 m, G+ M$ M' Y: j

3 y+ o5 b  |3 @- h
2 `/ v7 K& X) K, V2 N
25. Wu, L,Cao K,Lu Y,andWang R. Different mechanisms underlying the stimulation of K(Ca)  2 channels by nitric oxide and carbon monoxide. J Clin Invest 110:691-700,2002  .

26. Zhang, F,Kaide JI,Rodriguez-Mulero F,Abraham NG,andNasjletti A. Vasoregulatory function of the heme-heme oxygenase-carbon monoxide system. Am J Hypertens 14:62S-67S,2001  .  y1 O4 ~+ c- c+ B/ D/ _+ P

) ]7 P% B5 c6 f& `' W6 B
27. Zhang, F,Kaide JI,Wei Y,Jiang H,Yu C,Balazy M,Abraham NG,Wang W,andNasjletti A. Carbon monoxide produced by isolated arterioles attenuates pressure- induced vasoconstriction. Am J Physiol Heart Circ Physiol 281:H350-H358,2001 .
4 W" q+ v' D& z- G: y
8 ]% H  W) H2 n& k* q1 C$ h

28. Zou, AP,Billington H,Su N,andCowley AW, Jr. Expression and actions of heme oxygenase in the renal medulla of rats. Hypertension 35:342-347,2000 .

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