Effect of volume expansion on sodium excretion in the presence and absence of increased delivery from superficial proximal tubules.

The role of the proximal tubule in the natriuresis after volume expansion was investigated by evaluating sodium excretion both in the presence and absence of increased delivery from the proximal tubule. Proximal delivery was calculated from fractional reabsorption in superficial proximal tubules determined by micropuncture and glomerular filtration rate of the micropunctured kidney. Infusion of Ringer's solution in six dogs increased delivery from the proximal tubule 4.7+/-1 ml/min (P < 0.01) and increased fractional sodium excretion 3.6+/-1.1% (P < 0.025). Infusion of hyperoncotic albumin into the renal artery during sustained volume expansion decreased delivery from the proximal tubule 6.5+/-0.9 ml/min (P < 0.01). Although proximal delivery was restored to below control levels, fractional sodium excretion was significantly increased 2.5+/-0.5% (P < 0.01) as compared with the hydropenic control period. Fractional phosphate excretion was increased 15.5+/-3.7% (P < 0.01) after Ringer's infusion and was decreased 10.5+/-1.6% (P < 0.005) after intrarenal albumin infusion, suggesting that changes in superficial nephron reabsorption were paralleled by changes in reabsorption in deeper nephrons. Similar results were found in six additional dogs in which other factors known to affect phosphate reabsorption were controlled; however, these studies cannot completely eliminate a role for deep nephrons in the natriuresis after intrarenal albumin infusion. Since 70% of the natriuresis after volume expansion was present without increased delivery from superficial proximal tubules, it is likely that increased delivery from the proximal tubule contributes a relatively minor fraction to the natriuresis of volume expansion.

Proximal tubule reabsorption after hyperoncotic albumin infusion. Role of parathyroid hormone and dissociation from plasma volume.

Preferential expansion of the plasma volume by infusion of salt-poor hyperoncotic albumin solution decreases sodium reabsorption by the proximal tubule. The present micropuncture studies test the thesis that albumin infusion depresses proximal reabsorption by an effect unrelated to expansion of the plasma volume, perhaps due to an effect of parathyroid hormone (PTH) on proximal sodium reabsorption. Infusion of salt-poor hyperoncotic albumin significantly decreased plasma ionized calcium, increased immunoreactive PTH (iPTH) in plasma, decreased sodium reabsorption by the proximal tubule, and increased phosphate clearance. In contrast, infusions of albumin, in which the ionized calcium was restored to normal plasma levels, had no significant effect on ionized calcium, iPTH, proximal reabsorption, or phosphate clearance in intact dogs. Similarly, in parathyroidectomized animals given a constant replacement infusion of PTH, albumin infusion had no significant effect on proximal reabsorption or phosphate clearance. Plasma volume was markedly expanded following albumin infusion in all groups of dogs. These findings (a) indicate that PTH plays a significant role in the decrease in sodium reabsorption by the renal proximal tubule after salt-poor hyperoncotic albumin infusion, and (b) dissociate preferential expansion of the plasma volume from decreases in sodium reabsorption by the proximal tubule.

Renal afferent arteriole in the spontaneously hypertensive rat.

We conducted morphometric studies on the afferent arteriole of spontaneous hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats to gain a better understanding of its changes with the development of hypertension. Differences may be related to the SHRs' increased renal vascular resistance. Methacrylate vascular casts were made of the renal vasculature after perfusion fixation with glutaraldehyde. These vascular casts were then examined and measurements made with the scanning electron microscope. Results from this examination of the scanning electron microscope demonstrated a smaller afferent arteriolar diameter in the SHR, compared to the WKY, for both the inner and outer cortical glomeruli. This difference was seen in the 6-week-old SHR, prior to a statistically different blood pressure from the WKY controls, as well as in the 12-week-old hypertensive SHR. However, this afferent diameter difference between rat strains was more pronounced in rats at 12 weeks of age. The tapering of the afferent arteriole (difference between proximal and distal afferent diameters) was greater in the 12-week-old SHR than in the age-matched WKY or 6-week-old SHR. We conclude that the smaller caliber afferent arterioles of the SHR may predispose and play a role in the pathogenesis of the subsequent hypertension. The increased afferent arteriolar tapering seen in the hypertensive SHR relates to the already present increased blood pressure. Wall thickness/radius ratios are not different between rat strains (SHR and WKY) at either 6 or 12 weeks of age. These results suggest increased vascular constriction or hypoplastic vessels as the cause of the smaller caliber vessels in the SHR rather than increased wall thickness.

The glomerular filtration barrier in the spontaneously hypertensive rat.

To determine whether the differences in the physiological characteristics of SHR and WKY kidneys might be related to differences in renal structure, we studied the kidneys of SHR and WKY rats utilizing scanning and transmission electron microscopy and latex perfusion of the glomerular vasculature. Scanning and transmission electron microscopy revealed a smaller diameter in the glomerular endothelial fenestrae of SHR compared to that in WKY rats even prior to the development of differences in blood pressure. With age, the density and diameter of SHR endothelial fenestrae progressively decreased, which was not true in WKY rats. Latex casts of glomerular vasculature showed the frequent presence of afferent arteriolar constriction in 12-week-old SHR. Thus, the filtration barrier of the SHR is abnormal. THe pathogenesis of hypertension in the SHR is complex and probably multifactorial; however, renal structural changes may contribute to the development and maintenance of hypertension. The afferent arteriolar constrictions may be the structural basis for the increased vascular resistance described in the SHR.

Effect of bradykinin on the renal medullary osmotic gradient in water diuresis.

The effect of bradykinin on the renal medullary osmotic gradient was evaluated in anesthetized dogs which were undergoing water diuresis and which received a unilateral renal arterial infusion of bradykinin. The effect of the peptide on the medullary osmotic gradient was determined by analysis of medullary tissue electrolyte and urea concentrations and by analysis of changes in urine osmolality induced by vasopressin. Bradykinin decreased the total osmolality per kg H2O in tissue from inner medulla and papilla (-18.7 +/- 6% and -19.3 +/- 8%) and increased fractional water excretion (3.8 +/- 1.3%). Furthermore, a direct relationship between changes in free water clearance and changes in papillary tissue, osmolality was found. Finally, the increase in urine osmolality after ADH was significantly less in vasodilated than in control kidneys. These results indicate that bradykinin can diminish the medullary osmotic gradient during water diuresis in the dog. Thus, a bradykinin-induced increase in free water clearance may be accounted for by other than an inhibition of proximal tubular sodium reabsorption.

Effects of histamine-receptor antagonists on histamine-stimulated renin secretion.

The effects of histamine H1- and H2-receptor antagonists on histamine-stimulated renin secretion were examined in anesthetized dogs. Tripelennamine (H1 blocker) further enhanced renin secretion in the presence of exogenous histamine. Moreover, tripelennamine alone increased renin secretion. These effects are probably due to non-specific properties of the drug and not to interaction of tripelennamine with H1 receptors. Conversely, cimetidine (H2 blocker) significantly inhibited histamine-induced increases in renin secretion, renal blood flow, and sodium excretion without any changes in mean arterial blood pressure or glomerular filtration rate. Cimetidine alone had no effect. We conclude that H2 receptors mediate the effect of histamine on renin secretion in dogs with innervated, intact kidneys.

Effect of experience on medical students' attitudes toward animal laboratories in pharmacology education.

BACKGROUND: Medical students' attitudes toward the use of animal laboratories in pharmacology courses may form a useful source of evaluative information about the laboratories' educational effectiveness. METHOD: In 1992-93, 144 second-year students at the Indiana University School of Medicine were surveyed--before and after completing four hands-on laboratories using dogs--for their assessments of educational and moral aspects of animal laboratories. Statistical analysis involved chi-square and Student's t test. RESULTS: Of the 144 students in the course, 143 responded to the first survey and 86 responded to the second. From before to after the lab experiences, the percentage of students who agreed that the labs would reinforce/had reinforced the lecture material increased from 38% to 69%. In both surveys, 10% of the students objected to the use of any animals in labs, and 24% (before) and 21% (after) objected to the use of dogs. Whereas the percentage agreeing that the labs involved a morally wrong use of animals rose from 11% to 22%, the percentage disagreeing with that notion rose from 53% to 61%. Between 50% and 60% of the students in both surveys opposed doing the labs by computer simulation or videotaped demonstration. CONCLUSION: Most students indicated that the laboratory experiences enhanced their understanding of the actions of drugs, were preferable to alternatives that did not use animals, and did not involve an immoral use of animals. On the other hand, the results suggest that the number of students who have negative feelings about the use of animals in laboratories, though small, tends to be larger than the number who express these feelings to faculty.

Characterization of norepinephrine binding to plasma proteins of the dog and the rabbit.

The binding of 3H-norepinephrine (L-3H-NE, 1.0 X 10(-9) M) to plasma proteins of the dog and the rabbit was studied under controlled conditions. Destruction of NE occurred less rapidly at 22 degrees than at 37 degrees. Binding measured at 22 degrees was equivalent to that at 37 degrees, while binding measured at 0 degree was greater than that at 37 degrees. Therefore, losses of plasma NE were minimized by incubation of samples at 22 degrees for no longer than 30 minutes. L-3H-NE binding was examined in the absence and presence of 10(-9) to 10(-2) M non-labeled L-NE, DL-NE, DL-normetanephrine (NM), DL-epinephrine (E), dopamine (DA), and catechol (C). Specific binding of L-3H-NE varied in the range of NE concentrations (L-3H-NE + non-labeled NE) from 10(-9) M (18.7 +/- 3.1%, rabbit; 25.6 +/- 4.8%, dog) to 10(-6) M (10.8 +/- 3.1%, rabbit; 15.2 +/- 3.6%, dog). Calculated binding constants (KD) were consistent with binding to circulating proteins such as globulins or albumin (4.2 +/- 1.2 X 10(-5) M, rabbit; 5.4 +/- 1.7 X 10(-5) M, dog). In plasma from both species, non-labeled DL-NE, L-NE, E, DA, and C, but not NM (from 10(-9) to 10(-2) M) each significantly displaced L-3H-NE from its binding site in a manner similar to displacement produced by non-labeled NE. The results demonstrate that 1) NE is bound to plasma proteins, although to a lesser extent than had been reported by other investigators; and 2) the binding of catecholamines to plasma proteins may be mediated by the catechol ring.

In vivo pressure measurements of lithotripsy shock waves in pigs.

Stone comminution and tissue damage in lithotripsy are sensitive to the acoustic field within the kidney, yet knowledge of shock waves in vivo is limited. We have made measurements of lithotripsy shock waves inside pigs with small hydrophones constructed of a 25-microm PVDF membrane stretched over a 21-mm diameter ring. A thin layer of silicone rubber was used to isolate the membrane electrically from pig fluid. A hydrophone was positioned around the pig kidney following a flank incision. Hydrophones were placed on either the anterior (shock wave entrance) or the posterior (shock wave exit) surface of the left kidney. Fluoroscopic imaging was used to orient the hydrophone perpendicular to the shock wave. For each pig, the voltage settings (12-24 kV) and the position of the shock wave focus within the kidney were varied. Waveforms measured within the pig had a shape very similar to those measured in water, but the peak pressure was about 70% of that in water. The focal region in vivo was 82 mm x 20 mm, larger than that measured in vitro (57 mm x 12 mm). It appeared that a combination of nonlinear effects and inhomogeneities in the tissue broadened the focus of the lithotripter. The shock rise time was on the order of 100 ns, substantially more than the rise time measured in water, and was attributed to higher absorption in tissue.

Suprarenal intraarterial infusion of alloxan and streptozotocin during balloon occlusion of the juxtarenal abdominal aorta: a simple technique for inducing diabetes mellitus in canines with reduced mortality.

RATIONALE AND OBJECTIVES: The authors performed this study to evaluate the mortality and morbidity associated with a simple technique for inducing diabetes in dogs--suprarenal intraarterial infusion of alloxan and streptozotocin during balloon occlusion of the juxtarenal abdominal aorta. MATERIALS AND METHODS: The authors attempted to induce diabetes in six purpose-bred dogs. After the dogs were fasted for 12 hours, the abdominal aorta at the level of the origin of the renal arteries was occluded with an angioplasty balloon introduced by means of a femoral approach. A 3-F microcatheter (n = 1) or infusion wire (n = 5) was introduced via the percutaneous transluminal angioplasty catheter and positioned at the level of the celiac axis, and a mixture of streptozotocin (20-25 mg/kg) and alloxan (20-25 mg/kg) was infused. Diabetes was considered to have been induced if the dogs experienced sustained hyperglycemia. RESULTS: There were no deaths during the follow-up period (range, 7 months to 2 1/2 years). A diabetes-like state was induced in five of the six dogs, and no nephrotoxicity was seen. Diabetes was not induced in one dog owing to caudal migration of an undersized balloon during the infusion; this also resulted in reversible renal damage. CONCLUSION: This simple technique is effective for inducing diabetes in dogs, and morbidity and mortality rates are lower than those reported in the literature with other described techniques.

Effects of SWL on glomerular filtration rate and renal plasma flow in uninephrectomized minipigs.

This study tested the hypothesis that the effects of SWL on hemodynamics in solitary kidneys differ from those in kidneys of binephric animals. Five female miniature pigs (Pitman-Moore, 6 months of age, 30-35 kg) were anesthetized for unilateral nephrectomy. Seven pigs served as binephric controls. Two weeks later, each pig was anesthetized, prepared for unilateral or bilateral urine collections, and subjected to SWL (Dornier HM3, 2000 shocks, 24 kV). Clearances of inulin (glomerular filtration rate; GFR) and para-aminohippurate (renal plasma flow; RPF) were measured 1 hour prior to and 1, 4, and 24 hours after SWL. The GFR and RPF were higher in uninephrectomized than in intact pigs at all time points. In both groups, SWL reduced GFR and RPF. In the binephric pigs, RPF was reduced at all times post-SWL, but in the uninephrectomized pigs, RPF was returning toward baseline by 4 hours post-SWL and was not different from baseline at 24 hours. A comparison of whole-animal GFR and RPF (righ plus left clearances in binephric pigs v solitary renal clearances in uninephrectomized pigs) showed that whole-animal GFR and RPF did not differ between the groups before or after SWL. Compensatory renal hypertrophy and improved hemodynamics in solitary kidneys may acutely attenuate the renal vasoconstrictive effect of SWL. The long-term consequences of the compensatory changes are unknown.

Renal disease potentiates the injury caused by SWL.

PURPOSE: The present study tested the hypothesis that renal disease potentiates the structural/functional changes induced by a clinical dose of shockwaves. MATERIALS AND METHODS: Experimental pyelonephritis was induced in 6- to 8-week-old pigs before treatment with 2,000 shocks at 24 kV. These pigs were divided into two groups according to whether they were infected with a highly virulent (Group 1) or less virulent (Group 2) inoculation of E. coli. All animals were imaged by MR prior to SWL as a means of documenting the extent of pyelonephritis and immediately after SWL to examine the lesion produced by the shockwaves. The glomerular filtration rate (GFR), renal plasma flow (RPF) and para-aminohippurate (PAH) extraction were determined bilaterally on day 30 (Group 1) or day 80 (Group 2). RESULTS: In group 2, urine flow and sodium excretion were reduced by 50% from baseline in the shocked kidneys at both 1 and 4 hours post-SWL. A sustained reduction in RPF through 4 hours post-SWL was noted in the shocked kidneys in Group 1, but RPF was significantly reduced only at the 1-hour determination in Group 2. Large, consistent reductions in GFR were evident at 1 and 4 hours post-SWL in shocked and unshocked kidneys of Group 2 and in the shocked kidneys of Group 1. No significant changes were noted in PAH extraction. CONCLUSION: Acute pyelonephritis exaggerated the effect of a clinical dose of shockwaves on renal hemodynamics. This effect suggests that renal disease may be risk factor for SWL-induced injury.

Effect of an atrial natriuretic peptide, anaritide, on renal function in dogs on high- and low-sodium diets.

These studies examined the effects in dogs of dietary Na intake on plasma "ANF-like" immunoreactivity (PAI), and on the renal response to anaritide, a synthetic natriuretic peptide. There were no significant differences between high-Na (120 mEq/day) and low-Na (3 mEq/day) dogs in endogenous PAI, nor was PAI altered by oral or i.v. volume expansion with 0.9% saline. Renal arterial infusion of anaritide did not alter arterial pressure, renal blood flow or the frational excretion of lithium, but increased the fractional excretion of Na to similar degrees in both groups. The peptide increased GFR only in the low-Na group. Extremes of sodium balance do not seem to alter the renal response to anaritide, which apparently does not inhibit sodium reabsorption in the proximal tubule.

Renal tubular effects of sodium fluoride.

Administration of sodium fluoride results in vasopressin-resistant polyuric "renal failure" resembling nephrogenic diabetes insipidus. However, the renal tubular site of action of fluoride is not clear. Fischer 344 rats received acute i.v. infusions of sodium fluoride (0.3, 1.47 and 2.20 mumol/min/kg b.wt.) for 2.5 hr which resulted in dissipation of the renal medullary tissue osmotic gradient and a sustained, dose-related increase in fractional sodium excretion and urine flow. In additional experiments, free water reabsorption and excretion were decreased by fluoride, but the decrease in free water excretion occurred only when the fluoride-induced polyuria preceded the onset of the water diuresis. Slices of renal medulla from fluoride-treated rats had lower cyclic AMP concentrations than did slices from control rats and the responsiveness of the medullary tissue to vasopressin was markedly reduced. These data indicate that the fluoride ion dissipates the concentration gradient in the renal medulla largely by inhibiting NaCl reabsorption in the ascending limb of Henle's loop and inhibits antidiuretic hormone-mediated water reabsorption across the collecting duct.

Pentobarbital sleeping time and waking blood levels in spontaneously hypertensive rats.

Pentobarbital sleeping times and blood levels on arousal were determined in spontaneously hypertensive rats (SHR) and normotensive controls (WKR). Sleeping time for SHR was significantly less than for WKR, but blood levels of [14C]pentobarbital at awakening were not significantly different. The shorter sleeping time in SHR appears not to result from decreased brain sensitivity to pentobarbital. Instead, SHR appear to differ from WKR in the rate of metabolic clearance of the drug or in the distribution of the drug between blood and brain.

Aldosterone in the exaggerated natriuresis of spontaneously hypertensive rats.

The effect of mineralocorticoid hormones on the urinary responses of spontaneously hypertensive and normotensive rats to oral salt loading was determined. In response to a control salt load, the increase was determined. In response to a control salt load, the increase in urinary sodium excretion by the spontaneously hypertensive rats was significantly greater than that of the normotensive rats [48 +/- 6 (SE) mueq/h vs. 26 +/- 4 mueq/h]. Treatment with spironolactone did not significantly alter the natriuretic response of the spontaneously hypertensive rats (43 +/- 8 mueq/h) to another salt load, but increased the natriuretic response of the normotensive rats (55 +/- 7 mueq/h) to that of the hypertensive rats. D-Aldosterone suppressed the natriuretic response to salt loading of the hypertensive rats to a level which was not significantly different from that of the normotensive rats. Plasma aldosterone concentration was significantly lower in the spontaneously hypertensive rats than in the normotensive rats (18.0 +/- 3.3 and 52.1 +/- 5.2 ng/100 ml, respectively). Neither extracellular fluid volume nor total body water in spontaneously hypertensive and normotensive rats were significantly different. The data support the hypothesis that the exaggerated natriuresis in the spontaneously hypertensive rats is mediated by a relative lack by these rats of aldosterone-mediated distal tubular sodium reabsorption.

Mechanism of renal tubular secretion of norepinephrine in the rabbit.

The present studies examined renal tubular secretory mechanisms for norepinephrine (NE) in the anesthetized rabbit. The application of a saline droplet containing [3H]NE and [14C]inulin to the decapsulated surface of the left kidney was associated with a greater urinary recovery of 3H from that kidney than from the right kidney. The urinary ratio of 3H to 14C was greater than that in the droplet [ratio of urinary 3H/14C to droplet 3H/14C (U/D) = 8.6 +/- 1.2, P < .005], indicating tubular influx of [3H]NE. The urinary recoveries of [14C]inulin from both kidneys were not significantly different from each other. At peak excretion, nonmetabolized [3H]NE represented 74 +/- 8% of the 3H excreted from the left kidney. Probenecid significantly suppressed, but did not abolish, the tubular influx of 3H, but did not significantly alter the fraction of total 3H excreted as nonmetabolized [3H]NE. Cyanine 863 virtually abolished the tubular influx of 3H and significantly reduced the fraction of total 3H excreted as nonmetabolized [3H]NE. Surface application of [3H]-p-aminohippurate or [14C]tetraethylammonium produced U/D ratios which were indicative of tubular influx of the ions. Probenecid abolished the tubular influx of [3H]-p-aminohippurate (P < .001), whereas cyanine 863 significantly reduced the tubular influx of [14C]tetraethylammonium (P < .001). Surface application of [3H]mannitol resulted in no evidence of tubular influx (U/D = 1.2 +/- 0.3, N.S.). The results demonstrate that 1) [3H]NE was secreted by the rabbit renal tubule; 2) passive diffusion probably accounted for little of the tubular influx of [3H]NE; and 3) three-quarters of the secreted [3H]NE was secreted as nonmetabolized NE, most likely via cationic transport.

Estrogen attenuation of the development of hypertension in spontaneously hypertensive rats.

To examine the effects of estrogen on the development of high blood pressure in rats with a genetic predisposition toward hypertension, we administered to rats of the spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) strains 0.05 mg mestranol daily from ages 4 to 13 wks. Control animals of each strain received corn oil placebos. Systolic blood pressure was measured by a microphonic tail-cuff technique twice a week after the rats were 6 wks of age. Estrogen treatment in SHR was associated with a significant reduction in the level of hypertension attained, but estrogen treatment had no effect on blood pressure in WKY. Estrogen prevented normal growth in SHR and WKY, but this effect (reproduced in another group of SHR and WKY by restriction of food intake) was not related to the lower blood pressures seen in estrogen-treated SHR. Thus, it appeared that estrogen administration attenuated the rise in blood pressure normally seen in SHR and that this attenuation was independent of the estrogenic effect on body weight.