Renal effects of angiotensin II inhibition during increases in renal venous pressure.

Increases in renal venous pressure have been shown to consistently increase renal interstitial pressure; however, not until renal interstitial pressure is increased threefold is a natriuresis noted in normal animals. Since the intrarenal angiotensin II (Ang II) concentration has been postulated to increase with increasing renal venous pressure, the antinatriuretic action of Ang II could override the natriuretic effect of increased renal interstitial pressure. Therefore, the role of Ang II in the natriuretic response to increased renal venous pressure was examined in 10 pentobarbital-anesthetized dogs. Mean arterial pressure, renal blood flow, renal interstitial pressure, glomerular filtration rate, urinary sodium excretion, plasma renin activity, and prostaglandin E2 excretion were measured at renal venous pressures of 3, 15, and 30 mm Hg. The measurements were repeated after the administration of captopril (1 mg/kg i.v. bolus, n = 5) or [Sar1,Ile8]Ang II (50 micrograms/kg i.v. bolus + 50 micrograms/kg/hr infusion, n = 5). Under control conditions, mean arterial pressure, renal blood flow, plasma renin activity, and prostaglandin E2 excretion remained unchanged when renal venous pressure was increased. The elevations in renal venous pressure increased renal interstitial pressure from 7 +/- 2 to 12 +/- 2 and 22 +/- 4 mm Hg, while sodium excretion remained unchanged until renal venous pressure was 30 mm Hg. In the captopril-treated group, increasing renal venous pressure increased renal interstitial pressure as under control conditions; however, sodium excretion (23 +/- 4, 19 +/- 4, and 27 +/- 6 mueq/min) was not significantly increased even at the highest renal venous pressure.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of morphine on urine output: possible role of atrial natriuretic factor.

We examined the possible role of atrial natriuretic factor (ANF) in morphine-induced diuresis. Morphine (20 micrograms/20 microliters) administered intracerebroventricularly (i.c.v.) in conscious, normally hydrated Sprague-Dawley rats significantly increased urine output from 0.22 +/- 0.16 ml/h/rat to 2.07 +/- 0.46 ml/h/rat, 3 h after injection. Morphine injection also significantly increased the plasma ANF level to a maximum of 364 +/- 46 pg/ml compared to control animals (33 +/- 7 pg/ml) in which 20 microliters of saline was injected i.c.v. (P < 0.001). This peak increase occurred 1 h after morphine administration, however, it continued to remain significantly elevated (171 +/- 36 pg/ml vs. 39 +/- 8 pg/ml in control animals; P < 0.001) 2 h later. The maximal diuresis observed 3 h after administration of morphine i.c.v. was abolished by pretreatment of the animals with 0.8 mg of naloxone given intravenously (i.v.) and also by anti-rat ANF serum (0.4 ml i.v.). A short, transient increase in blood pressure, occurred 2-5 min after morphine administration, but it is unlikely that this increase accounted for the increase in plasma ANF. We conclude that the diuresis induced by i.c.v. administration of morphine is due to enhanced release of ANF.

Role of nitric oxide and prostaglandin in the maintenance of cortical and renal medullary blood flow.

This study was undertaken in anesthetized dogs to evaluate the relative participation of prostaglandins (PGs) and nitric oxide (NO) in the maintenance of total renal blood flow (TRBF), and renal medullary blood flow (RMBF). It was hypothesized that the inhibition of NO should impair cortical and medullary circulation because of the synthesis of this compound in the endothelial cells of these two territories. In contrast, under normal conditions of perfusion pressure PG synthesis is confined to the renal medulla. Hence PG inhibition should predominantly impair the medullary circulation. The initial administration of 25 microM kg-1 min-1 NG-nitro-L-arginine methyl ester produced a significant 26% decrease in TRBF and a concomitant 34% fall in RMBF, while the subsequent inhibition of PGs with 5 mg/kg meclofenamate further reduced TRBF by 33% and RMBF by 89%. In contrast, the initial administration of meclofenamate failed to change TRBF, while decreasing RMBF by 49%. The subsequent blockade of NO decreased TRBF by 35% without further altering RMBF. These results indicate that initial PG synthesis inhibition predominantly alters the medullary circulation, whereas NO inhibition decreases both cortical and medullary flow. This latter change induced by NO renders cortical and RMBF susceptible to a further decrease by PG inhibition. However, the decrease in medullary circulation produced by NO inhibition is not further enhanced by subsequent PG inhibition.

Nitric oxide and renal function.

A wealth of evidence shows that nitric oxide can modulate the autoregulation of renal blood flow, the glomerular surface area available for filtration, the glomerulotubular feedback response, and the release of renin. From an integrative point of view, inhibition of nitric oxide synthesis will alter the function of all of these homeostatic mechanisms and impair the pressure-induced natriuresis secondary to increases in intrarenal vascular resistance and tubular sodium reabsorption. These effects, along with an elevation of both total peripheral resistance and vascular tone of the capacitance vessels, are the most likely determinants of the volume-dependent elevation of blood pressure (ie, salt-sensitive hypertension) that occurs during partial inhibition of nitric oxide synthesis. This observation has important physiological and pathologic implications because it shows for the first time that the blockade of a single endogenous vasodilator substance can produce a sustained increase in blood pressure that can be influenced by changes in blood volume. Because of these characteristics, this review emphasizes in particular the characteristics of the nitric oxide synthesis pathway and briefly describes several known methods of increasing the biologic activity of nitric oxide; these methods eventually may be modified and used as therapeutic interventions in humans with deficient nitric oxide synthesis.

Vascular development in chick embryos: a possible role for adenosine.

We studied the possible role of adenosine in the development of the vasculature using 217 chick embryos. Adenosine (2-32 mumol/day), inosine (16 mumol/day), dipyridamole (0.04-0.4 mumol/day), or aminophylline (400 and 800 micrograms/day) were administered twice each day into the air space on days 11-14. Control embryos received Ringer solution. Whole body vascularity was estimated on day 15 as the whole body structural vascular resistance (SVR), i.e., the hydraulic resistance of the maximally dilated vasculature. Adenosine decreased the SVR in a dose-related manner at the lower dosage amounts but caused a maximum decrease in SVR at the higher dosage amounts averaging 30% below the Ringer control values. Equimolar amounts of adenosine and inosine decreased the SVR by the same extent. Dipyridamole, which potentiates the biological effects of endogenous adenosine, also decreased the SVR in a dose-related manner to values averaging approximately 30% below control. When the effects of endogenous adenosine were blocked by aminophylline, the SVR increased in a dose-related manner to approximately 100% above control at the highest dosage amount. These results suggest that adenosine could have a physiological role in growth regulation of the vascular system in the chick embryo.

Renal kinin antagonism does not impair pressure-induced natriuresis.

We studied the contribution of the renal kallikrein-kinin system to short-term electrolyte and water balance during baseline and during acutely elevated renal perfusion pressure (RPP) in the anesthetized dog. Renal blood flow, glomerular filtration rate, urine flow rate, renin secretion rate, and urinary excretion of sodium, potassium, prostaglandin E2 (PGE2), and kinins were measured at baseline RPP during intrarenal infusion of 0.9% saline or the competitive bradykinin analogue [D-Arg0,Hyp3,Thi5,D-Phe7,Thi8]bradykinin (50 micrograms/min), which blocks vascular and interstitial kinin receptors. RPP was then raised above baseline (control group 25%; kinin analogue group 22%) by ligating the celiac artery, the superior mesenteric artery, and the aorta distal to the renal arteries. Renal parameters were again measured during infusion of saline or the kinin analogue. The analogue had no effect on renal hemodynamic or excretory parameters at baseline perfusion pressures. Increasing RPP significantly increased urine flow rates and urinary sodium excretion rates (control group, 43 mumol/min; kinin analogue group, 55 mumol/min) in both groups of animals. Increasing pressure also tended to decrease renin secretion rate in both groups of animals; however, neither increased pressure nor infusion of the analogue affected urinary excretion of PGE2 or kinins. The results suggest that intrarenal kinins are not powerful short-term regulators of electrolyte and water balance and that an intact kallikrein-kinin system is not necessary to induce pressure diuresis and natriuresis.

Reflex versus autoregulatory control of hindlimb blood flow during treadmill exercise in dogs.

To evaluate the competition between local autoregulation and reflex neurohumoral control of hindlimb blood flow (HLBF), the hindlimb vascular pressure-flow relationship was determined in nine dogs in response to a 10% decrease in mean arterial pressure (AP) imposed during both low (3.0 km/h, 0% grade) and high (5.5 km/h, 14% grade) intensities of treadmill exercise. HLBF was measured with a Doppler flow probe on the left external iliac artery, and AP was controlled with a gravity reservoir connected to the left carotid artery. A 10 +/- 2% reduction in AP for 25 min caused HLBF to decrease 25 +/- 2% during low-exercise intensity but only 10 +/- 2% during high-exercise intensity. The corresponding closed-loop gains (Gc) of HLBF regulation [Gc = 1 - (% delta hindlimb blood flow/% delta hindlimb perfusion pressure) were -1.6 +/- 0.4 and -0.06 +/- 0.2 during low- and high-exercise intensity, respectively. Autonomic ganglionic blockade (hexamethonium) increased the Gc during low-intensity exercise to 0.07 +/- 0.2. Antagonism of adenosine receptors (aminophylline) decreased the Gc of HLBF regulation during high-intensity exercise to -0.57 +/- 0.3. These data demonstrate that in response to an imposed decrease in AP, autonomic vasoconstriction overrides autoregulatory vasodilatory mechanisms during low-intensity exercise. HLBF regulation increases at a higher exercise intensity, in part due to adenosine, but autoregulation does not predominate over arterial pressure regulating mechanisms.

Direct measurement of renal medullary blood flow in the dog.

We studied the responses of total renal blood flow (RBF) and renal medullary blood flow (RMBF) to changes in renal perfusion pressure (RPP) within and below the range of renal autoregulation in the anesthetized dog (n = 7). To measure RMBF, we developed a technique in which the medulla is exposed by excising a section of infarcted cortex and a multiple optical fiber flow probe, connected to a laser-Doppler flowmeter, is placed on the medulla. At the baseline RPP of 120 +/- 1 mmHg, RBF was 2.58 +/- 0.33 ml.min-1.g perfused kidney wt-1, and RMBF was 222 +/- 45 perfusion units. RPP was then decreased in consecutive 20-mmHg steps to 39 +/- 1 mmHg. At 80 +/- 1 mmHg, RBF remained at 89 +/- 4% of the baseline value; however, RMBF had decreased significantly (P < 0.05) to 73 +/- 4% of its baseline value. The efficiency of autoregulation of RBF and of RMBF within the RPP range of 120 to 80 mmHg was determined by calculating an autoregulatory index (AI) for each parameter using the formula AI = (%delta blood flow)/(%delta RPP). An AI of 0 indicates perfect autoregulation, and an index of 1 indicates a system with a fixed resistance. The AI for RBF averaged 0.33 +/- 0.12 over this pressure range and showed a significantly greater (P < 0.05) autoregulatory ability than did the RMBF (0.82 +/- 0.13). Decreasing perfusion pressure < 80 mmHg produced significant decreases in both RBF and RMBF.(ABSTRACT TRUNCATED AT 250 WORDS)

Morphometric measurements of chorioallantoic membrane vascularity: effects of hypoxia and hyperoxia.

We studied the effects of hypoxia and hyperoxia on the angiogenesis process in the chick embryo chorioallantoic membrane (CAM) using four different morphometric measurements of vascularity. Chick eggs were incubated in various oxygen atmospheres (12, 16, 21, 45, or 70% oxygen) beginning on the 7th day of development, and vascularity was measured on the 14th day. Measurements of vascularity included vessel endpoint density (VED), length density, fractional image area, and a vascular density index. All measurements were made on blood vessels in randomly selected areas of CAM using a computerized image analysis system. An opaque colloidal carbon-albumin perfusate was used as a vascular marker. All four measurements showed that vascularity of CAM was inversely related to the oxygen tension to which the embryos were subjected. The VED, an estimate of total number of pre- and postcapillary vessels, exhibited the greatest degree of change, but overall changes in vascularity were modest. Prolonged exposure to a 12% oxygen atmosphere increased VED by approximately 16%, whereas 70% oxygen decreased VED by approximately 19% compared with room air control groups. We also studied the normal growth of CAM vasculature from days 8 to 18 of development. In these studies, the values of VED increased progressively throughout the entire period of development, whereas the other measurements of vascularity reached maximum values by the 14th day. We conclude that hypoxia stimulates angiogenesis in the CAM in a dose-related manner, hyperoxia inhibits CAM angiogenesis in a dose-related manner, and VED provides a sensitive estimate of vascularity in chick CAM throughout its development.

Molecular cloning and nucleotide sequence of the pestivirus bovine viral diarrhea virus.

The RNA genome of the cytopathic NADL isolate of bovine viral diarrhea virus (BVDV) has been molecularly cloned and the nucleotide sequence determined. The cloned sequence was 12,573 nucleotides in length, corresponding to a molecular weight of 4.3 X 10(6), having a base composition of 32.2% A, 25.7% G, 22.1% U, and 20.0% C. However, the sequences at the 5' and 3' termini of the RNA have not been unequivocally established. A single major open reading frame extending the length of the molecule was found in the viral-sense (positive polarity) sequence. This open reading frame was capable of encoding 3988 amino acids, representing 449 kDa of protein.

Pressure dependency of canine intrarenal blood flow within the range of autoregulation.

The mechanism of pressure-induced natriuresis remains controversial. To assess whether intracortical or medullary renal blood flows (RBF) change with changes in renal perfusion pressure (RPP), global and regional RBFs were measured using the dynamic spatial reconstructor, a fast computed tomography scanner, in eight anesthetized dogs (group B) within the range of RBF autoregulation (RPP of 153.5 and 114.4 mmHg). Similar measurements were obtained in seven control dogs (group A) in which RPP was not manipulated. In group B, only inner medullary perfusion decreased (from 0.84 to 0.51 ml/min per cm3 of tissue, P = 0.03) with reduction of RPP, whereas global renal, intracortical, and outer medullary perfusions remained unaltered. In group A there was no change in global or regional renal perfusion. The change in inner medullary perfusion in group B (-34.7%) was significantly different (P = 0.021) from that in group A (+27.4%). Global, cortical, and total medullary RBFs (ml/min) and volumes did not change in either group. These results suggest that with changes in RPP, the only detectable change in intrarenal perfusion occurs in the inner medulla.

Vpr is preferentially targeted by CTL during HIV-1 infection.

The HIV-1 accessory proteins Vpr, Vpu, and Vif are essential for viral replication, and their cytoplasmic production suggests that they should be processed for recognition by CTLs. However, the extent to which these proteins are targeted in natural infection, as well as precise CTL epitopes within them, remains to be defined. In this study, CTL responses against HIV-1 Vpr, Vpu, and Vif were analyzed in 60 HIV-1-infected individuals and 10 HIV-1-negative controls using overlapping peptides spanning the entire proteins. Peptide-specific IFN-gamma production was measured by ELISPOT assay and flow-based intracellular cytokine quantification. HLA class I restriction and cytotoxic activity were confirmed after isolation of peptide-specific CD8(+) T cell lines. CD8(+) T cell responses against Vpr, Vpu, and Vif were found in 45%, 2%, and 33% of HIV-1-infected individuals, respectively. Multiple CTL epitopes were identified in functionally important regions of HIV-1 Vpr and Vif. Moreover, in infected individuals in whom the breadth of HIV-1-specific responses was assessed comprehensively, Vpr and p17 were the most preferentially targeted proteins per unit length by CD8(+) T cells. These data indicate that despite the small size of these proteins Vif and Vpr are frequently targeted by CTL in natural HIV-1 infection and contribute importantly to the total HIV-1-specific CD8(+) T cell responses. These findings will be important in evaluating the specificity and breadth of immune responses during acute and chronic infection, and in the design and testing of candidate HIV vaccines.

Comprehensive epitope analysis of human immunodeficiency virus type 1 (HIV-1)-specific T-cell responses directed against the entire expressed HIV-1 genome demonstrate broadly directed responses, but no correlation to viral load.

Cellular immune responses play a critical role in the control of human immunodeficiency virus type 1 (HIV-1); however, the breadth of these responses at the single-epitope level has not been comprehensively assessed. We therefore screened peripheral blood mononuclear cells (PBMC) from 57 individuals at different stages of HIV-1 infection for virus-specific T-cell responses using a matrix of 504 overlapping peptides spanning all expressed HIV-1 proteins in a gamma interferon-enzyme-linked immunospot (Elispot) assay. HIV-1-specific T-cell responses were detectable in all study subjects, with a median of 14 individual epitopic regions targeted per person (range, 2 to 42), and all 14 HIV-1 protein subunits were recognized. HIV-1 p24-Gag and Nef contained the highest epitope density and were also the most frequently recognized HIV-1 proteins. The total magnitude of the HIV-1-specific response ranged from 280 to 25,860 spot-forming cells (SFC)/10(6) PBMC (median, 4,245) among all study participants. However, the number of epitopic regions targeted, the protein subunits recognized, and the total magnitude of HIV-1-specific responses varied significantly among the tested individuals, with the strongest and broadest responses detectable in individuals with untreated chronic HIV-1 infection. Neither the breadth nor the magnitude of the total HIV-1-specific CD8+-T-cell responses correlated with plasma viral load. We conclude that a peptide matrix-based Elispot assay allows for rapid, sensitive, specific, and efficient assessment of cellular immune responses directed against the entire expressed HIV-1 genome. These data also suggest that the impact of T-cell responses on control of viral replication cannot be explained by the mere quantification of the magnitude and breadth of the CD8+-T-cell response, even if a comprehensive pan-genome screening approach is applied.

Role of nitric oxide and prostaglandin in the maintenance of cortical and renal medullary blood flow

This study was undertaken in anesthetized dogs to evaluate the relative participation of prostaglandins (PGs) and nitric oxide (NO) in the maintenance of total renal blood flow (TRBF), and renal medullary blood flow (RMBF). It was hypothesized that the inhibition of NO should impair cortical and medullary circulation because of the synthesis of this compound in the endothelial cells of these two territories. In contrast, under normal conditions of perfusion pressure PG synthesis is confined to the renal medulla. Hence PG inhibition should predominantly impair the medullary circulation. The initial administration of 25 µM kg-1 min-1 NG-nitro-L-arginine methyl ester produced a significant 26 percent decrease in TRBF and a concomitant 34 percent fall in RMBF, while the subsequent inhibition of PGs with 5 mg/kg meclofenamate further reduced TRBF by 33 percent and RMBF by 89 percent. In contrast, the initial administration of meclofenamate failed to change TRBF, while decreasing RMBF by 49 percent. The subsequent blockade of NO decreased TRBF by 35 percent without further altering RMBF. These results indicate that initial PG synthesis inhibition predominantly alters the medullary circulation, whereas NO inhibition decreases both cortical and medullary flow. This latter change induced by NO renders cortical and RMBF susceptible to a further decrease by PG inhibition. However, the decrease in medullary circulation produced by NO inhibition is not further enhanced by subsequent PG inhibition.