Oxidative status in the macula densa modulates tubuloglomerular feedback responsiveness in angiotensin II-induced hypertension.

AIM: Tubuloglomerular feedback (TGF) is an important mechanism in control of signal nephron glomerular filtration rate. The oxidative stress in the macula densa, primarily determined by the interactions between nitric oxide (NO) and superoxide (O2-), is essential in maintaining the TGF responsiveness. However, few studies examining the interactions between and amount of NO and O2- generated by the macula densa during normal and hypertensive states. METHODS: In this study, we used isolated perfused juxtaglomerular apparatus to directly measure the amount and also studied the interactions between NO and O2- in macula densa in both physiological and slow pressor Angiotensin II (Ang II)-induced hypertensive mice. RESULTS: We found that slow pressor Ang II at a dose of 600 ng kg(-1) min(-1) for two weeks increased mean arterial pressure by 26.1 ± 5.7 mmHg. TGF response increased from 3.4 ± 0.2 µm in control to 5.2 ± 0.2 µm in hypertensive mice. We first measured O2- generation by the macula densa and found it was undetectable in control mice. However, O2- generation by the macula densa increased to 21.4 ± 2.5 unit min(-1) in Ang II-induced hypertensive mice. We then measured NO generation and found that NO generation by the macula densa was 138.5 ± 9.3 unit min(-1) in control mice. The NO was undetectable in the macula densa in hypertensive mice infused with Ang II. CONCLUSIONS: Under physiological conditions, TGF response is mainly controlled by the NO generated in the macula densa; in Ang II induced hypertension, the TGF response is mainly controlled by the O2- generated by the macula densa.

Antioxidants block angiotensin II-induced increases in blood pressure and endothelin.

Chronically infusing a subpressor dose of angiotensin (Ang) II increases blood pressure via poorly defined mechanisms. We found that this hypertensive response is accompanied by increased oxidant stress and is prevented by blocking endothelin (ET) receptors. Thus, we now tested whether blocking oxidant stress decreases both blood pressure and ET levels. We infused Sprague-Dawley rats (via osmotic pumps) with either vehicle (group 1) or Ang II (5 ng. kg(-1). min(-1); groups 2 to 4) for 15 days. Groups 3 and 4 also received either tempol in the drinking water (1 mmol/L) or vitamin E (5000 IU/kg diet), respectively, for 15 days. We measured systolic blood pressure (SBP) and urinary nitrite excretion every 3 days, and on day 15 we measured systemic and renal venous plasma levels of ET, isoprostanes, and thiobarbituric acid reactive substances (TBARS). SBP in Group 1 did not change throughout the study, whereas Ang II increased SBP (from 132+/-5 to 151+/-7 mm Hg). In addition, Ang II increased the systemic and renal venous levels of isoprostanes, TBARS, and ET and caused a transient decrease in urinary nitrites (that returned to control levels by day 9). Both tempol and vitamin E prevented Ang II-induced hypertension and either prevented or tended to blunt the increase in systemic and renal isoprostanes, TBARS, and ET. Finally, both antioxidants abolished the transient decrease in urinary nitrites. These results together with our previous study suggest that subpressor-dose Ang II increases oxidant stress (and isoprostanes). This in turn increases ET levels, which participate in the hypertensive response to Ang II.

Chronic renal failure secondary to oxalate nephropathy: a preventable complication after jejunoileal bypass.

Enteric hyperoxaluria is a commonly seen adverse event after the jejunoileal bypass procedure. The increased concentration of urinary oxalate predisposes bypass patients to various renal complications such as nephrolithiasis and oxalate nephropathy. If not diagnosed and appropriately treated, these complications can lead to irreversible renal damage. We describe 3 patients in whom severe renal complications developed with irreversible compromise of renal function after a jejunoileal bypass. Patients who undergo a jejunoileal bypass require lifelong follow-up with close monitoring of their renal function. Marked decline in renal function mandates prompt investigation and aggressive intervention, including reversal of the jejunoileal bypass if necessary. Chronic renal failure secondary to oxalate nephropathy is preventable and treatable but may require conversion of a jejunoileal bypass to a more current form of bypass.

Role of endothelin and isoprostanes in slow pressor responses to angiotensin II.

We tested the hypothesis that angiotensin II (Ang II)-induced stimulations of endothelin (ET) and isoprostanes are implicated in the slow pressor responses to Ang II. We infused either vehicle (group 1) or Ang II (groups 2 to 4) intravenously at 5 ng/kg per minute via osmotic pumps for 15 days into Sprague-Dawley rats. Groups 3 and 4 received 30 mg/kg per day of either losartan (Ang II type 1 receptor blocker) or bosentan (ET(A) and ET(B) receptor blocker) in their drinking water. We measured systolic blood pressure (SBP) every 3 days during the infusion. Plasma levels of Ang II, ET, isoprostanes, and urinary nitrites were determined at 15 days. Vehicle infusion did not change SBP (from 138+/-13 to 136+/-2 mm Hg at day 15). Circulating Ang II, ET, and isoprostane levels were 35+/-9, 39+/-3, and 111+/-10 pg/mL, respectively, whereas urinary nitrites were 2.3+/-0.4 microgram/d. Ang II increased SBP (from 133+/-10 to 158+/-8 mm Hg), plasma Ang II (179+/-77 pg/mL), and isoprostanes (156+/-19 pg/mL) without altering ET levels (38+/-5 pg/mL) or urinary nitrites (1.8+/-0.5 microgram/d). Losartan prevented Ang II-induced increases in SBP and isoprostanes (SBP went from 137+/-5 to 120+/-4 mm Hg; isoprostanes were 115+/-15 pg/mL) while increasing urinary nitrite levels (5.2+/-1.1 microgram/d). Losartan did not alter Ang II (141+/-57 pg/mL) or ET (40+/-4 pg/mL) levels. Bosentan also blocked Ang II-induced hypertension (from 135+/-4 to 139+/-3 mm Hg) but did not decrease isoprostanes (146+/-14 pg/mL). Ang II (63+/-11 pg/mL), ET levels (46+/-2 pg/mL), and urinary nitrites (2.8+/-0.4 microgram/d) were not altered. In conclusion, our results suggest that low-dose Ang II increases isoprostanes via its Ang II type 1 receptor and causes an ET-dependent hypertension, without altering circulating ET levels.

Microcomputed tomography of kidneys following chronic bile duct ligation.

BACKGROUND: In hepatic cirrhosis, renal sodium and water retention can occur prior to decreases in renal blood flow (RBF). This may be explained in part by redistribution of the intrarenal microcirculation toward the juxtamedullary nephrons. To appreciate this three-dimensional spatial redistribution better, we examined the intrarenal microcirculatory changes using microcomputed tomography (micro-CT) in rats subjected to chronic bile duct ligation (CBDL). METHODS: Six kidneys from control rats and eight kidneys from rats that had undergone CBDL for 21 days were perfusion fixed in situ at physiological pressure, perfused with silicon-based Microfil containing lead chromate, embedded in plastic, and scanned by micro-CT. The microvasculature in the reconstructed three-dimensional renal images was studied using computerized image-analysis techniques. To determine the physiological condition of the rats, parallel experiments were conducted on six control and six CBDL rats to measure mean arterial pressure (MAP), RBF, glomerular filtration rate (GFR), urine flow (UF) rate, and sodium excretion by conventional methods. RESULTS: The percentage of vasculature in the renal cortex from CBDL rats was significantly decreased (10.8 +/- 0.4% vs. 16.8 +/- 2.7% control values). However, the vascular volume fractions of the medullary tissues were not significantly altered. There were no significant differences in the number of glomeruli between groups (36,430 +/- 1908 CBDLs, 36,609 +/- 3167 controls). The CBDL rats had a similar GFR than the controls but a reduced MAP, RBF, UF, and sodium excretion. CONCLUSIONS: The results indicate that after CBDL, there is a selective decrease in cortical vascular filling, which may contribute to the salt and water retention that accompanies cirrhosis.

Systemic hemodynamics and renal function in hemorrhaged dogs resuscitated with cross-linked hemoglobin.

Cross-linked hemoglobin (XL-Hb) infused into dogs increases mean arterial pressure (MAP) but decreases blood flow to the renal (RBF), mesenteric (MBF), and iliac (IBF) circulations. These actions differ markedly from dextran infusion (which increases RBF, MBF, and IBF without altering MAP) and may be due to scavenging of nitric oxide by XL-Hb. However, because the hormonal milieu regulating regional circulation is altered during hemorrhage (when XL-Hb may be used), we studied whether systemic hemodynamics, RBF, MBF, IBF, and renal excretory function in hemorrhaged dogs was altered when resuscitated with XL-Hb compared with dextran (n = 6 each). Hemorrhage decreased MAP by 25% due to a 75% decline in cardiac output. RBF, MBF, and IBF all fell by 33, 64, and 72%, respectively (P<0.05 each). There was also a fall in glomerular filtration rate (GFR), urinary flow, and sodium excretion (P<0.05 each). After resuscitation, MAP, cardiac output, RBF, MBF, IBF, and GFR all recovered to basal values with either XL-Hb or dextran. Urinary flow and sodium excretion increased to above basal levels with dextran (both by 3.5-fold; P<0.05) or XL-Hb (by 7.5- and 10-fold, respectively; P<0.05). We conclude that resuscitation with XL-Hb after hemorrhage not only increases MAP, but also restores RBF, MBF, IBF, GFR, and urinary sodium and volume excretion analogously to dextran. The results contrast with those in normal dogs and suggest that nitric oxide inhibition does not impair hemodynamic and renal function recovery during hemorrhage.

Low-dose angiotensin II increases free isoprostane levels in plasma.

Chronic intravenous infusion of subpressor doses of angiotensin II causes blood pressure to increase progressively over the course of several days. The mechanisms underlying this response, however, are poorly understood. Because high-dose angiotensin II increases oxidative stress, and some compounds that result from the increased oxidative stress (eg, isoprostanes) produce vasoconstriction and antinatriuresis, we tested the hypothesis that a subpressor dose of angiotensin II also increases oxidative stress, as measured by 8-epi-prostaglandin F(2alpha) (isoprostanes), which may contribute to the slow pressor response to angiotensin II. To test this hypothesis, we infused angiotensin II (10 ng/kg per minute for 28 days via an osmotic pump) into 6 conscious normotensive female pigs (30 to 35 kg). We recorded mean arterial pressure continuously with a telemetry system and measured plasma isoprostanes before starting the angiotensin II infusion (baseline) and again after 28 days with an enzyme immunoassay. Angiotensin II infusion significantly increased mean arterial pressure from 121+/-4 to 153+/-7 mm Hg (P<0. 05) without altering total plasma isoprostane levels (180.0+/-24.3 versus 147.0+/-29.2 pg/mL; P=NS). However, the plasma concentrations of free isoprostanes increased significantly, from 38.3+/-5.8 to 54.7+/-10.4 pg/mL (P<0.05). These results suggest that subpressor doses of angiotensin II increase oxidative stress, as implied by the increased concentration of free isoprostanes, which accompany the elevation in mean arterial pressure elevation. Thus, isoprostane-induced vasoconstriction and antinatriuresis may contribute to the hypertension induced by the slow pressor responses of angiotensin II.

Abnormal renal vasodilation to an amino acid infusion in congestive heart failure: normalization by enalapril.

In congestive heart failure (CHF), the neurohormonal mechanisms that cause renal vasoconstriction, particularly those depending on the renin-angiotensin system, could interfere with renal vasodilating mechanisms. To elucidate this issue, we studied the kidney response to an amino acid infusion (known to cause renal vasodilation in healthy individuals) in eight patients with CHF. We found that the amino acid infusion (0.7 mL/kg/h of a 10% solution) elicited no renal hemodynamic response, in marked contrast to healthy subjects. We next hypothesized that the renin-angiotensin system (known to be activated in heart failure) has a role in the lack of response to the amino acid infusion. To test this hypothesis, we repeated the study after two 5-mg doses of enalapril, an inhibitor of the angiotensin-converting enzyme, administered 12 hours apart. After enalapril treatment, the amino acid infusion caused a 45% increase in mean renal blood flow (RBF) from 383 +/- 55 to 557 +/- 51 mL/min at the fifth hour (P < 0.05). This normalization of the renal response to the amino acid infusion occurred without changes in cardiac output or in systemic vascular resistance. Hence, the renal fraction of the cardiac output increased during the amino acid infusion. The recovery of the renal vascular response was not accompanied by an increase in glomerular filtration rate (GFR; filtration fraction decreased), suggesting a predominant efferent arteriole dilatation. Our study shows that, in heart failure, the kidney loses its ability to increase RBF in response to an amino acid load. This lack of renal vascular response can be restored by inhibiting the renin-angiotensin system and is unrelated to changes in systemic hemodynamics.

Biphasic effect of bradykinin on rabbit afferent arterioles.

Bradykinin plays an important role in the regulation of renal hemodynamics. However, there have been few studies of the effect of bradykinin on isolated afferent arterioles, vascular segments that are important for the regulation of renal blood flow and glomerular filtration rate. Our purpose was to study (1) the effects of bradykinin on isolated perfused rabbit afferent arterioles and (2) the mechanisms of actions. Afferent arterioles dissected from rabbits were perfused in vitro at 60 mm Hg. In afferent arterioles preconstricted with phenylephrine, 10(-12) to 10(-10) mol/L bradykinin increased luminal diameter from 9.0+/-1.0 to 14.3+/-1.2 microm (P<0.003). In contrast, 10(-9) and 10(-8) mol/L bradykinin decreased luminal diameter to 10.8+/-1.4 and 9.7+/-1.2 microm, respectively (P<0.001). Bradykinin added to the bath had no effect on preconstricted afferent arterioles. The addition of [des-Arg9]-bradykinin (10(-9) and 10(-8) mol/L), a B1 receptor agonist, to the lumen decreased diameter from 9.7+/-1.2 to 6.7+/-1.2 microm at 10(-8) mol/L (P<0.002). Icatibant (Hoe 140), a B2 receptor antagonist, blocked both the vasodilation and vasoconstriction induced by bradykinin as well as the vasoconstriction induced by [des-Arg9]-bradykinin. L-NAME had no effect on bradykinin-induced dilation or constriction. Indomethacin blocked both the dilation induced by 10(-12) to 10(-10) mol/L bradykinin and the constriction induced by 10(-9) to 10(-8) mol/L bradykinin. In fact, in the presence of indomethacin, 10(-9) and 10(-8) mol/L bradykinin increased luminal diameter from 6.2+/-0.7 to 10.7+/-0.6 microm at 10(-8) mol/L (P<0.001), which was attenuated by L-NAME. Finally, in the presence of SQ29548, a prostaglandin H2/thromboxane A2 receptor antagonist, bradykinin caused dilation at all concentrations tested. In conclusion, bradykinin has a biphasic effect on afferent arterioles. Both dilation and constriction may be mediated by bradykinin B2 receptors. The mechanisms of vasodilation and vasoconstriction are due to cyclooxygenase products, not nitric oxide.

Abnormal endothelium-dependent responses in early radiation nephropathy.

While arterial hypertension and renal dysfunction are well recognized complications of renal irradiation, the mechanisms that trigger the development of these complications are unknown. Recently, it was reported that the endothelium is a major target in radiation injury. Because dysfunction of the endothelial cells may lead or contribute to the development of hypertension and renal dysfunction in radiation nephropathy, we tested the hypothesis that endothelium-dependent vasodilation is impaired in radiated kidneys prior to the onset of hypertension. To test this hypothesis, we used Long-Evans rats that had undergone left nephrectomy (3 weeks earlier) and irradiation (3000 r's) to the right kidney 8 days earlier (mean blood pressures in the irradiated rats were not different than in the controls). We then measured the changes in renal blood flow (RBF) induced by endothelium-dependent (acetylcholine and bradykinin) and -independent (nitroprusside, norepinephrine, and angiotensin II) vasoactive agents. We found that the increases in RBF induced by the endothelium-dependent but not independent vasodilators were markedly impaired in the irradiated kidneys. Blocking nitric oxide synthesis with nitro L-arginine methyl ester in sham rats mimicked the blunted responsiveness of the irradiated rats, whereas indomethacin (an inhibitor of prostaglandin synthesis) had no effect on either sham or irradiated rats. Finally, the RBF responses to the endothelium-independent vasoconstrictors, norepinephrine and angiotensin II, were not altered in the irradiated kidneys. These results suggest that renal irradiation causes endothelial dysfunction (prior to the onset of hypertension) but spares the vascular smooth muscle cells.

Angiotensin II action in isolated microperfused rabbit afferent arterioles is modulated by flow.

We have recently presented evidence that endogenous nitric oxide (NO) and prostaglandins (PGs) modulate angiotensin II (Ang II) action in microperfused afferent arterioles (Af-Arts). Because flow may be a physiological stimulus of endothelial release of NO and PGs, we tested the hypothesis that flow through the lumen of the Af-Art stimulates the endothelium to produce NO and PGs, which in turn modulate the action of Ang II. We microdissected the terminal segment of an interlobular artery together with two Af-Arts, their glomeruli and efferent arterioles (Ef-Art). The two Af-Arts were perfused simultaneously from the interlobular artery, while one Ef-Art was occluded. Since the arteriolar perfusate contained 5% albumin, oncotic pressure built up in the glomerulus with the occluded Ef-Art and opposed the force of filtration, resulting in little or no flow through the corresponding Af-Art. Thus this preparation allowed us to observe Ang II action in free-flow and non-flow Af-Arts simultaneously. Ang II-induced constriction was weaker in free-flow than non-flow Af-Arts, with the luminal diameter decreasing by 8 +/- 2% and 23 +/- 3% at 10(-9) M, respectively (P < 0.013 free-flow vs. non-flow; N = 9). Disrupting the endothelium augmented Ang II action in free-flow (33 +/- 5.1%; P < 0.01 vs. intact endothelium) but not non-flow Af-Arts (31 +/- 5.3%), thus abolishing the differences between them (N = 8). Pretreatment with an inhibitor of either NO synthase (N-nitro-L-arginine methyl ester) or cyclooxygenase (indomethacin) augmented Ang II action more in free-flow than non-flow Af-Arts, likewise abolishing the differences between them. These results suggest that intraluminal flow modulates the vasoconstrictor action of Ang II in Af-Arts via endothelium-derived NO and PGs. Thus flow may be important in the fine control of glomerular hemodynamics.

Platelet-activating factor dilates efferent arterioles through glomerulus-derived nitric oxide.

UNLABELLED: Despite evidence that platelet-activating factor (PAF) is produced by the glomerulus, its direct action on the glomerular microcirculation is poorly understood. It was recently reported that at picomolar concentrations, PAF dilates isolated microperfused afferent arterioles (Af-Art) via nitric oxide (NO). The present study tested the hypothesis that PAF acts on the glomerulus to release NO, which in turn controls the resistance of the efferent arteriole (Ef-Art). Rabbit Ef-Art were perfused from the distal end (retrograde perfusion [RP]) to eliminate the influence of the glomerulus, or through the glomerulus from the end of the Af-Art (orthograde perfusion [OP]) to maintain the influence of the glomerulus. Ef-Art were preconstricted by approximately 40% with norepinephrine and increasing doses of PAF were added to both the arteriolar perfusate and bath. Only with OP did PAF at picomolar concentrations cause significant dilation: at 400 pmol, the diameter increased by 64 +/- 11% from the preconstricted level (N = 6, P < 0.01). This dilation was completely abolished by pretreatment with an NO-synthesis inhibitor. To study its possible constrictor action, PAF was added to nonpreconstricted Ef-Art. At nanomolar concentrations, PAF constricted Ef-Art similarly in both RP and OP: at 40 nM, the diameter decreased by 24 +/- 4% (N = 6, P < 0.01) and 20 +/- 2% (N = 6, P < 0.01), respectively. This constriction was attenuated by pretreatment with indomethacin (Indo) in both RP (14 +/- 2%, N = 7; P < 0.02 versus without Indo) and OP (10 +/- 2%, N = 6; P < 0.02 versus without Indo). IN CONCLUSION: (1) at picomolar concentrations, PAF stimulates the glomerulus to release NO, which in turn dilates the Ef-Art; and (2) at nanomolar concentrations, PAF constricts the Ef-Art partly through release of cyclooxygenase metabolites. Thus, PAF may play a role in glomerular hemodynamics under various physiological and pathological conditions.

Flow modulates myogenic responses in isolated microperfused rabbit afferent arterioles via endothelium-derived nitric oxide.

Flow may be a physiological stimulus of the endothelial release of nitric oxide (NO) and prostaglandins (PGs). We tested the hypothesis that pressure-induced constriction of the glomerular afferent arteriole (Af-Art) is modulated by luminal flow via endothelial production of NO. We microdissected the terminal segment of an interlobular artery together with two Af-Arts, their glomeruli (GL) and efferent arterioles (Ef-Art). The two Af-Arts were perfused simultaneously from the interlobular artery, while one Ef-Art was occluded. Since the arteriolar perfusate contained 5% albumin, oncotic pressure built up in the glomerulus with the occluded Ef-Art and opposed the force of filtration, resulting in little or no flow through the corresponding Af-Art. Thus this preparation allowed us to observe free-flow and no-flow Af-Arts simultaneously during stepwise 30-mmHg increases in intraluminal pressure (from 30 to 120 mmHg). Pressure-induced constriction was weaker in free-flow than no-flow Af-Arts, with the luminal diameter decreasing by 11.1 +/- 1.7 and 25.6 +/- 2.3% (n = 30), respectively, at 120 mmHg. To examine whether flow modulates myogenic constriction through endothelium-derived NO and/or PGs, we examined pressure-induced constriction before and after (a) disruption of the endothelium, (b) inhibition of NO synthesis with NW-nitro-L-arginine methyl ester (L-NAME), or (c) inhibition of cyclooxygenase with indomethacin. Both endothelial disruption and L-NAME augmented pressure-induced constriction in free-flow but not no-flow Af-Arts, abolishing the differences between the two. However, indomethacin had no effect in either free-flow or no-flow Af-Arts. These results suggest that intraluminal flow attenuates pressure-induced constriction in Af-Arts via endothelium-derived NO. Thus flow-stimulated NO release may be important in the fine control of glomerular hemodynamics.

Glomerular prostaglandins modulate vascular reactivity of the downstream efferent arterioles.

The balance of vascular resistance in afferent (Af-) and efferent arterioles (Ef-Arts) is a crucial factor that determines glomerular hemodynamics. We have recently reported that when Ef-Arts were perfused from the distal end of the Af-Art through the glomerulus (orthograde perfusion; OP), both angiotensin II (Ang II) and norepinephrine (NE) induced much weaker constriction than they did when Ef-Arts were perfused from the distal end (retrograde perfusion; RP). This difference was not affected by inhibiting synthesis of nitric oxide. In the present study, we tested the hypothesis that glomerular prostaglandins (PGs) may modulate vascular reactivity of the downstream Ef-Art. In addition, we examined the possible modulatory role of PGs in the Af-Art responses to Ang II or NE. Both Ang II and NE caused dose-dependent constriction of Ef-Arts with either OP or RP; however, the constriction was stronger in RP. At 10(-8) M, Ang II decreased Ef-Art diameter by 35 +/- 3.5% in OP (N = 9) compared to 73 +/- 3.9% in RP (N = 5), while 10(-6) M NE decreased the diameter by 25 +/- 3.6% in OP (N = 9) compared to 62 +/- 7.2% in RP (N = 5). Pretreatment with 5 x 10(-5) M indomethacin (Indo) did not alter basal diameter with either method of perfusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Removal of endothelium-dependent relaxation by antibody and complement in afferent arterioles.

Studies of endothelial regulation of microvascular function have been hampered by the technical difficulty of removing the endothelium without damaging the vascular smooth muscle cells. This study presents a novel method of endothelial damage and lysis based on the facts that endothelial cells express specific antigens and that complement reacts with antibody/antigen complexes, causing cell lysis. We isolated and perfused rabbit glomerular afferent arterioles in vitro and examined vascular responses before and after treating them with an antibody against factor VIII-related antigen and complement. The treatment consisted of perfusing afferent arterioles with medium containing the antibody and complement for 10 minutes, followed by a 20-minute washout period. Before treatment, acetylcholine and the calcium ionophore A23187 (receptor- and nonreceptor-mediated endothelium-dependent vasodilators, respectively) dilated norepinephrine-preconstricted afferent arterioles, whereas neither dilated the arterioles after treatment, suggesting loss of endothelium-dependent vasodilation. In contrast, responses to nicardipine and norepinephrine (endothelium-independent vasodilator and constrictor, respectively) were not altered by the treatment, indicating intact vascular smooth muscle cell function. Transmission electron microscopy revealed that the antibody- and complement-treated arterioles had various degrees of endothelial damage, including areas of detachment from the basement membrane and marked loss of the number and structure of mitochondria, but no evidence of vascular smooth muscle cell damage. These results indicate that treatment with anti-factor VIII-related antigen antibody and complement is an effective method for eliminating endothelium-dependent vasodilation without altering endothelium-independent responses. Thus, this method may be useful for studying the functional role of the endothelium in microvessels.

Disparate effects of insulin on isolated rabbit afferent and efferent arterioles.

Despite evidence that insulin per se may be an important regulator of glomerular hemodynamics, little is known about its direct action on the glomerular afferent arterioles (Af-Art) and efferent arterioles (Ef-Art), the crucial vascular segments that control glomerular hemodynamics. In the present study, we examined the direct effect of physiological concentrations of insulin on isolated microperfused rabbit Af- and Ef-Arts. After cannulation, vessels were equilibrated in insulin-free medium for 30 min. To determine whether insulin causes vasodilation or constriction, increasing doses (5, 20, and 200 microU/ml) were added to the bath and lumen of arterioles that were either preconstricted to 50% of control diameter with norepinephrine or left nonpreconstricted. Insulin caused no vasoconstriction in either Af- or Ef-Arts, but it reversed norepinephrine-induced constriction in Ef-Arts but not Af-Arts (suggesting a vasodilator action selective to the Ef-Art): at 200 microU/ml, insulin increased Ef-Art luminal diameter by 75.8 +/- 7.0% from the preconstricted level (n = 6; P < 0.008). The vasorelaxant effect of insulin on Ef-Arts was not affected by blockade of either endothelium-derived relaxing factor/nitric oxide or prostaglandin synthesis. Despite the lack of effect of insulin on Af-Art when added after the equilibration period, when Af-Arts were equilibrated in the presence of either 20 or 200 microU/ml insulin, their basal diameter was significantly reduced (11.7 +/- 0.9 microns; P < 0.025, n = 6, and 12.0 +/- 0.9 microns; P < 0.025, n = 7, respectively) compared with nontreated Af-Arts (16.2 +/- 1.3 microns; n = 7). In conclusion, this study demonstrates that at physiological concentrations, insulin dilates NE-constricted Ef-Arts, while insulin pretreatment enhances Af-Art tone. The disparate actions of insulin on the Af- vs the Ef-Art may contribute to its beneficial effect on glomerular hypertension.

Endothelium-derived relaxing factor/nitric oxide modulates angiotensin II action in the isolated microperfused rabbit afferent but not efferent arteriole.

It has been reported that sensitivity to angiotensin II (Ang II) is higher in efferent (Ef) than afferent (Af) arterioles (Arts). We tested the hypothesis that this is due to arteriolar differences in the interaction between Ang II and endothelium-derived relaxing factor/nitric oxide (EDNO). Rabbit Af-Arts with glomerulus intact were microperfused in vitro at a constant pressure. Ef-Arts were perfused from the distal end of either the Af-Art (orthograde perfusion) or the Ef-Art (retrograde perfusion) to eliminate influences of the Af-Art or glomerulus, respectively. Ang II did not alter Af-Art luminal diameter until the concentration reached 10(-9) M, which decreased the diameter by 11 +/- 2.6% (n = 11; P < 0.002). In contrast, Ef-Arts became significantly constricted at concentrations as low as 10(-11) M with either perfusion. Surprisingly, the decrease in Ef-Art diameter at 10(-10), 10(-9), and 10(-8) M was significantly greater with retrograde perfusion (44 +/- 6.9%, 70 +/- 5.6%, and 74 +/- 4.1%, respectively; n = 5) than with orthograde perfusion (16 +/- 4.2%, 25 +/- 2.9%, and 35 +/- 3.5%; n = 9). ENDO synthesis inhibition with 10(-4) M nitro-L-arginine methyl ester (L-NAME) decreased the diameter to a greater extent in Af-Arts (22 +/- 3.0%; n = 11) compared to Ef-Arts with either orthograde (9.5 +/- 2.3%; n = 8) or retrograde perfusion (1.2 +/- 2.1%; n = 6). With L-NAME pretreatment, Af-Art constriction induced by 10(-10) M (14 +/- 4.0%, n = 9) and 10(-9) M Ang II (38 +/- 3.9%) was significantly greater compared to nontreated Af-Arts. In contrast, L-NAME pretreatment had no effect on Ang II-induced constriction in Ef-Arts with either perfusion. In conclusion, this study demonstrates higher sensitivity of Ef-Arts to Ang II, particularly with retrograde perfusion. Our results suggest that EDNO significantly modulates the vasoconstrictor action of Ang II in Af-Arts II but not Ef-Arts, contributing to the differential sensitivity to Ang II.

Vasodilator and constrictor actions of platelet-activating factor in the isolated microperfused afferent arteriole of the rabbit kidney. Role of endothelium-derived relaxing factor/nitric oxide and cyclooxygenase products.

It has been suggested that platelet-activating factor (PAF) plays a prominent role in the control of glomerular hemodynamics in various physiological and pathological conditions. We examined the direct effect of PAF on rabbit glomerular afferent arterioles (Af-Arts) microperfused in vitro and tested whether endothelium-derived relaxing factor/nitric oxide (EDNO) and cyclooxygenase products are involved in its actions. In nanomolar concentrations PAF caused dose-dependent constriction of Af-Arts, with the maximum constriction being 34 +/- 10% at 4 x 10(-8) M (n = 9, P < 0.001). The constriction was blunted by cyclooxygenase inhibition (11 +/- 6%, n = 7, P < 0.05) but augmented by EDNO inhibition (76 +/- 14%, n = 8, P < 0.005). To study a possible vasodilator effect of PAF, Af-Arts were preconstricted with norepinephrine and increasing concentrations of PAF added to the lumen. At picomolar concentrations (lower than those that caused constriction), PAF produced dose-dependent vasodilation that was unaffected by cyclooxygenase inhibition but was abolished by EDNO synthesis inhibition. Both PAF-induced constriction and dilation of Af-Arts were blocked by a PAF receptor antagonist. This study demonstrates that PAF has a receptor-mediated biphasic effect on rabbit Af-Arts, dilating them at low concentrations while constricting them at higher concentrations. Our results suggest that PAF's vasodilator action may be due to production of EDNO, while its constrictor action is mediated at least in part through cyclooxygenase products.

Pressure-induced constriction of the afferent arteriole of spontaneously hypertensive rats.

In uncomplicated essential hypertension, renal blood flow, glomerular filtration rate, and glomerular capillary pressure are within the normal range despite elevated renal perfusion pressure, suggesting abnormally high resistance of the preglomerular vessels. Among various preglomerular vascular segments, the afferent arteriole (Af-Art) is thought to be the site responsible for most resistance. However, little is known about the vascular reactivity of the Af-Art or its alteration in hypertension. In this study, we tested the hypothesis that pressure-induced constriction is exaggerated in Af-Arts from spontaneously hypertensive rats (SHRs). Single Af-Arts were microdissected from kidneys of SHRs and normotensive control Wistar-Kyoto (WKY) rats and were microperfused in vitro. When pressure in the Af-Art was increased stepwise from 20 to 80 mm Hg, luminal diameter increased similarly in both WKY and SHR Af-Arts (from 10.0 +/- 0.8 to 18.6 +/- 1.3 microns and from 10.1 +/- 1.2 to 16.9 +/- 1.5 microns, respectively). However, when pressure was further increased to 140 mm Hg, the diameter remained unchanged in WKY Af-Arts (19.2 +/- 1.9 microns), whereas it decreased significantly to 11.1 +/- 0.9 microns in those from SHRs. We conclude that pressure-induced constriction is exaggerated in SHR Af-Arts, which may contribute to the development and maintenance of hypertension.