Deuterium stimulated-echo-type PGSE NMR experiments for measuring diffusion: application to a liquid crystal.

The accessibility of molecular self-diffusion coefficients in anisotropic materials, such as liquid crystals or solids, by stimulated-echo-type (2)H PGSE NMR is examined. The amplitude and phase modulation of the signal in the stimulated-echo-type sequence by the static quadrupole coupling during the encoding/decoding delays is suppressed by adjusting the pulse flip angles and the phase cycle. For nuclei that experience both nonnegligible quadrupole and dipole couplings, the application of magic echoes during the evolution periods of stimulated echo is demonstrated as a helpful technique in the case of slow diffusion. These findings are demonstrated by experimental results in the thermotropic liquid crystal of partially deuterated 8CB. The obtained diffusion coefficients are also compared to data obtained by a (1)H homonuclear-decoupling-type PGSE NMR method in the same material.

Fluorescence in situ hybridization (FISH) for rapid detection of aneuploidy: experience in 911 prenatal cases.

Fluorescence in situ hybridization (FISH) was performed with probes specific for chromosomes 13, 18, 21, X and Y on 911 of 11123 (8.2%) amniotic fluid samples submitted to the present authors' laboratory for cytogenetic analysis over an 8-year period. Altogether 3516 hybridizations were performed with an interpretable FISH result on all chromosomes requested in 884/911 (97%) of cases. An uninformative FISH result occurred in 44 hybridizations among 27 cases (3%). Of a total of 89 karyotypically proven cases with aneuploidy that might have been detected by FISH, the overall detection rate was 84%. An inconclusive or incomplete FISH result occurred in 9/89 (10%) of these proven aneuploid cases. In the remaining 80 informative proven aneuploid cases, correct detection of aneuploidy was accomplished in 75/80 (94%) of samples. A false-negative result occurred in the remaining 5/80 (6%) of such informative cases. Eighteen cases had karyotypically proven abnormalities that could not have been detected by the targeted FISH. Aside from these 18 cases, FISH allowed correct detection of normal disomy in 785/804 (98%) of such cases. An incomplete FISH result occurred in 18 normal disomic cases. There was a single possible 'false-positive' FISH result for chromosome 21. Interphase FISH analysis of uncultured amniotic fluid cells has been shown to be a useful laboratory tool for rapid fetal aneuploidy screening during pregnancy. As with all clinical laboratory diagnostic tests, incomplete or inconclusive results (or even interpretive errors) occur in a small percentage of cases. Nevertheless, FISH results accompanied by other data and by appropriate counseling provide clinicians and patients with valuable information for clinical decision-making surrounding family planning and pregnancy management.

nalyot, a mutation of the Drosophila myb-related Adf1 transcription factor, disrupts synapse formation and olfactory memory.

nalyot (nal) is a novel olfactory memory mutant of Drosophila, encoding Adf1, a myb-related transcription factor. Following extended training sessions, Adf1 mutants show normal early memory but defective longterm memory. Adf1 shows widespread spatiotemporal expression, yet mutant alleles reveal no discernible disruptions in gross morphology of the nervous system. Studies at the larval neuromuscular junction, however, reveal a role for Adf1 in the modulation of synaptic growth-in contrast to the role established for dCREB2 in the control of synaptic function (Davis et al., 1996). These findings suggest that Adf1 and dCREB2 regulate distinct transcriptional cascades involved in terminal stages of synapse maturation. More generally, Adf1 provides a novel link between molecular mechanisms of developmental and behavioral plasticity.

Determination of a molecular torsional angle in the metarhodopsin-I photointermediate of rhodopsin by double-quantum solid-state NMR.

We present a solid-state NMR study of metarhodopsin-1, the pre-discharge intermediate of the photochemical signal transduction cascade of rhodopsin, which is the 41 kDa integral membrane protein that triggers phototransduction in vertebrate rod cells. The H-C10-C11-H torsional angles of the retinylidene chromophore in bovine rhodopsin and metarhodopsin-I were determined simultaneously in the photo-activated membrane-bound state, using double-quantum heteronuclear local field spectroscopy. The torsional angles were estimated to be [phi] = 160+/-10 degrees for rhodopsin and phi = 180+/-25 degrees for metarhodopsin-I. The result is consistent with current models of the photo-induced conformational transitions in the chromophore, in which the 11-Z retinal ground state is twisted, while the later photointermediates have a planar all-E conformation.

Epidermal tendon cells require Broad Complex function for correct attachment of the indirect flight muscles in Drosophila melanogaster.

Drosophila Broad Complex, a primary response gene in the ecdysone cascade, encodes a family of zinc-finger transcription factors essential for metamorphosis. Broad Complex mutations of the rbp complementation group disrupt attachment of the dorsoventral indirect flight muscles during pupal development. We previously demonstrated that isoform BRC-Z1 mediates the muscle attachment function of rbp(+) and is expressed in both developing muscle fibers and their epidermal attachment sites. We now report two complementary studies to determine the cellular site and mode of action of rbp(+) during maturation of the myotendinous junctions of dorsoventral indirect flight muscles. First, genetic mosaics, produced using the paternal loss method, revealed that the muscle attachment phenotype is determined primarily by the genotype of the dorsal epidermis, with the muscle fiber and the ventral epidermis exerting little or no influence. When the dorsal epidermis was mutant, the vast majority of muscles detached or chose ectopic attachment sites, regardless of the muscle genotype. Conversely, wild-type dorsal epidermis could support attachment of mutant muscles. Second, ultrastructural analysis corroborated and extended these results, revealing defective and delayed differentiation of rbp mutant epidermal tendon cells in the dorsal attachment sites. Tendon cell processes, the stress-bearing links between the epidermis and muscle, were reduced in number and showed delayed appearance of microtubule bundles. In contrast, mutant muscle and ventral epidermis resembled the wild type. In conclusion, BRC-Z1 acts in the dorsal epidermis to ensure differentiation of the myotendinous junction. By analogy with the cell-cell interaction essential for embryonic muscle attachment, we propose that BRC-Z1 regulates one or more components of the epidermal response to a signal from the developing muscle.

Segment-specific retention of a larval neuromuscular system and its role in a new, rhythmic, pupal motor pattern in Manduca sexta.

At pupation in Manduca sexta, accessory planta retractor muscles and their motoneurons degenerate in segment-specific patterns. Accessory planta retractor muscles in abdominal segments 2 and 3 survive in reduced form through the pupal stage and degenerate after adult emergence. Electromyographic and electrophysiological recordings show that these accessory planta retractor muscles participate in a new, rhythmic 'pupal motor pattern' in which all four muscles contract synchronously at approximately 4 s intervals for extended bouts. Accessory planta retractor muscle contractions are driven by synaptic activation of accessory planta retractor motoneurons and are often accompanied by rhythmic activity in intersegmental muscles and spiracular closer muscles. The pupal motor pattern is influenced by descending neural input although isolated abdominal ganglia can produce a pupal motor pattern-like rhythm. The robust pupal motor pattern first seen after pupal ecdysis weakens during the second half of pupal life. Anemometric recordings indicate that the intersegmental muscle and spiracular closer muscle component of the pupal motor pattern produces ventilation. Accessory planta retractor muscle contractions lift the flexible abdominal floor, to which the developing wings and legs adhere tightly. We hypothesize that, by a bellows-like action, the accessory planta retractor muscle contractions circulate hemolymph in the appendages. Morphometric analysis shows that dendritic regression is similar in accessory planta retractor motoneurons with different pupal fates, and that accessory planta retractor motoneurons begin to participate in the pupal motor pattern while their dendrites are regressed.

Neural mechanisms of behavioral plasticity: metamorphosis and learning in Manduca sexta.

This review summarizes our current understanding of the neural circuit underlying the larval proleg withdrawal reflex (PWR) of Manduca sexta and describes how PWR function changes in two contexts: metamorphosis and learning. The first form of PWR plasticity occurs during the larval-pupal transformation, when the reflex is lost. One mechanism that contributes to this loss is the weakening of monosynaptic excitatory connection from proleg sensory neurons to proleg retractor motor neurons. This change is associated with the hormonally-mediated regression of proleg motor neuron dendrites, which may break synaptic contacts between the sensory and motor neurons. After pupation, some of the proleg motor neurons die in a segment-specific pattern that persists even after individual motor neurons are isolated from the nervous system and exposed to hormones in vitro. The second form of PWR plasticity involves short-term, activity-dependent changes in neural function during the larval stage. The nicotinic cholinergic connections from proleg sensory neurons to motor neurons exhibit several forms of plasticity including facilitation, depression, post-tetanic potentiation and two types of muscarinic modulation. Larval PWR behavior exhibits two simple forms of learning-habituation and dishabituation-which involve alterations in the central PWR circuit. These studies of a simple circuit illustrate neural mechanisms by which behaviors undergo both short- and long-term modifications.

Broad-complex transcription factors regulate thoracic muscle attachment in Drosophila.

The Broad-Complex, a 20-hydroxyecdysone-regulated gene, is essential for the development of many tissues during metamorphosis. In Broad-Complex mutants of the rbp complementation group, dorsoventral indirect flight muscles (DVM) are largely absent, and the dorsal longitudinal indirect flight muscles, tergotrochanteral muscles, and remaining DVM often select incorrect attachment sites. The Broad-Complex encodes a family of zinc-finger-containing transcription factors, and it is hypothesized that Broad Complex proteins containing the Z1 zinc-finger pair (BRC-Z1) mediate rbp+ function. We provide additional strong support for this hypothesis by showing that heat-shock-induced BRC-Z1 expression rescues the thoracic muscle defects of rbp mutants completely. BRC-Z4 induction can also rescue the thoracic musculature, but BRC-Z2 and -Z3 can not. Thus, the effect is specific to BRC-Z1 and its closest relative, BRC-Z4. Formation of muscle primordia from imaginal myoblasts appears normal in rbp mutants. However, the myotendinous junctions linking the DVM to the dorsal epidermis are weak, and the muscles detach during pupal life and subsequently degenerate. The data indicate that rbp mutations disrupt the cell-cell interactions between developing muscles and epidermal tendon cells as they recognize and attach to one another. Using a BRC-Z1-specific monoclonal antibody, we show that both the developing muscles and epidermal tendon cells express BRC-Z1 at the time of pupation, before mutant muscles begin to detach. We conclude that 20-hydroxyecdysone acts through the Broad-Complex to control the development of thoracic myotendinous junctions.

Novel dual innervation of a larval proleg muscle by two similar motoneurons in the tobacco hornworm Manduca sexta.

In Manduca sexta, the accessory planta retractor muscle (APRM), which retracts the larval proleg, is innervated by two excitatory motoneurons, the accessory planta retractor motoneurons (APRs). These muscles and motoneurons have been the focus of a number of developmental and behavioral studies. The present study investigated properties of the pair of APRs that innervate each APRM and determined their pattern of innervation of APRM fibers. Members of APR pairs could not be distinguished by their anatomical or electrical properties (resting membrane potential, input resistance and spike threshold). Spontaneous synaptic inputs to members of APR pairs were highly correlated, whereas spontaneous synaptic inputs to APRs and functionally dissimilar motoneurons were not well correlated. Synaptic inputs from identified mechanosensory neurons and interneurons to the two APRs were qualitatively similar, but the magnitude of the response to sensory stimulation sometimes differed within a pair. Both APRs produced large, rapidly rising excitatory junction potentials in APRM fibers. Within the APRM, some fibers were singly innervated by one or the other APR while the remaining fibers were dually innervated by both APRs. In dually innervated fibers, the motor terminals of the two APRs were spatially segregated. This innervation pattern appears to be unique among insects and shares some properties with the innervation of vertebrate muscle.

Fish oil ameliorates renal injury and hyperlipidemia in the Milan normotensive rat model of focal glomerulosclerosis.

Rats of the Milan normotensive rat strain (MNS) spontaneously develop severe proteinuria and excessive glomerular thromboxane (Tx)A2 production at a young age. These abnormalities are accompanied by podocyte alterations, progressive focal glomerulosclerosis (FGS), and interstitial fibrosis, resembling human FGS. Since it has been shown that pharmacologic Tx-synthase inhibition protects MNS rats from these changes, it was hypothesized that a fish oil (FO) enriched diet, by enhancing TxA3 production instead of TxA2, might afford similar protection, compared with diets enriched in safflower oil (SO) or lard (LD). Rats were pair-fed 11% fat diets from age of 1 to 11 months. Glomerular TxA2 at 11 months was significantly lower in PO-fed rats than in SO- and LD-fed rats (11 +/- 3.0, 69 +/- 3.0, 59 +/- 19.0 nanograms per min/mg, respectively; P < 0.001). At 3 months, urinary albumin excretion was similar among the groups. Over the course of the study, rats fed FO developed significantly less albuminuria than the SO and LD groups (P < 0.001 by analysis of variance for repeated measures), such that the values at 11 months were 25 +/- 5.8, 49 +/- 8.7, and 68 +/- 13.0 mg/24h, respectively. Serum cholesterol and triglycerides were also significantly lower in FO-fed rats than in SO- and LD-fed rats. The extent of FGS was similar in the three groups, but FO-fed rats had less interstitial injury than the other groups. It was observed that a fish-oil diet substantially alleviated albuminuria, normalized nephrotic hyperlipidemia, and reduced interstitial injury, but did not prevent the development of FGS in the MNS model.

Glomerular epithelial and mesangial cells differentially modulate the binding specificities of VLA-1 and VLA-2.

BACKGROUND: Maintenance of glomerular cell interaction with the complex basement membrane is crucial for the normal functioning of the kidney. Because little is known about the receptors utilized by glomerular cells, we examined the attachment of cultured glomerular cells to extracellular matrix proteins. EXPERIMENTAL DESIGN: We produced monoclonal antibodies that inhibited the function of rat VLA-1 and VLA-2 and used these antibodies alone and in combination to explore the attachment of glomerular epithelial cells (GEC) and mesangial cells to extracellular matrix proteins in vitro. RESULTS: Cultured GEC utilize only VLA-2 for attachment to collagen but use it together with VLA-1 for adhesion to laminin. In contrast, mesangial cells use both receptors for their attachment to collagen but utilize only VLA-1 in their interaction with laminin. The use of VLA-1 by GEC and of VLA-2 by mesangial cells was unexpected because the expression of these receptors was barely detectable in an enzyme-linked immunosorbent assay and by immunocytochemistry. CONCLUSIONS: VLA-1, VLA-2, and VLA-3 are integrin receptors with overlapping specificities in that all have the potential to interact with collagen and laminin. Our studies demonstrate that cultured GEC use VLA-1 and VLA-2 almost exclusively for their adhesion to these ligands, and thus VLA-3 appears to play a negligible role in such attachment. Interestingly, GEC and mesangial cells differentially modulate the ligand binding specificities of VLA-1 and VLA-2. In situ, VLA-1 has been localized within the mesangium, whereas VLA-2 has not been detected within the glomerulus leading to the conclusion that GEC do not use VLA-1 or VLA-2 and that mesangial cells fail to utilize VLA-2. However, our studies have shown that, even when such receptors are barely detectable on the surface of cultured cells by sensitive techniques, they can play a functional role. These results suggest either that the levels of expression in situ are too low for the relatively insensitive immunohistochemical techniques employed, and thus the importance of these receptors to glomerular cell attachment in vivo is under appreciated or that such receptors are the result of de novo expression by glomerular cells when they are subjected to in vitro culture conditions. Because it is known that such conditions may mimic pathologic stress, we are presently examining the expression of these receptors by glomerular cells in various disease models.

Cell-mediated immune injury in the kidney: acute nephritis induced in the rat by azobenzenearsonate.

Cell-mediated immune mechanisms have long been suspected of playing an important role in the pathogenesis of various renal diseases. An animal model of active nephritis secondary to an exogenous antigen that requires antigen presentation to immune-competent T cells has not been developed. Consequently, the potential of kidney cells to serve as effective antigen presenting cells after an exposure to a therapeutic, biological, or environmental agent in the intact animal has not been documented. The present experiments were designed to demonstrate the capacity of the kidney to become the target for cell-mediated immune injury. A model system has been developed whereby a chemically reactive form of the hapten azobenzenearsonate is introduced directly into the left kidney of pre-immunized Brown Norway rats. Previous studies have shown that this form of the hapten requires active antigen presentation but no intracellular processing, since the reactive form of the hapten modifies directly surface expressed proteins. Delayed hypersensitivity was demonstrated in the actively immunized animals by standard lymphocyte stimulation index and by in vivo skin testing. Peak foot pad swelling of 220 +/- 13 x 10(-2) mm in response to the hapten was observed between days 11 and 14 as compared to < 10 x 10(-2) mm in the contralateral foot injected with vehicle alone and < 20 x 10(-2) mm in response to azobenzenearsonate injection in animals immunized with adjuvant alone. The exposure of the kidney to the hapten in the primed animal results in an active unilateral granulomatous nephritis with marked destruction of tubules and glomeruli. On average, 71.5 +/- 5.2% of the renal cortex is affected by the inflammatory process in the actively immunized animals, compared to only 8.1 +/- 3.8% in controls. The disease can be reproduced qualitatively by adoptive transfer of T cells but not by passive antibody administration to naive recipients. These studies demonstrate that intrinsic kidney cells can act as effective antigen presenting cells in the intact animal and that the kidney can become the target of a cell-mediated immune injury.

Pathogenesis of glomerular injury in the fawn-hooded rat: early glomerular capillary hypertension predicts glomerular sclerosis.

Fawn-hooded rats spontaneously develop focal and segmental glomerular sclerosis, systemic hypertension, and proteinuria at a young age. Micropuncture and morphological studies were performed in two inbred strains of fawn-hooded rats, FHH and FHL, with different susceptibilities to develop chronic renal failure. FHH rats have higher values for systolic blood pressure and proteinuria and more rapid development of focal and segmental glomerular sclerosis and subsequent chronic renal failure as compared with genetically closely related FHL rats. FHH and FHL strains and a Wistar control strain, WAG, were matched for age and were studied at 16 wk. FHH, FHL, and WAG-old (WAG-O) strains were matched for weight, and the last group was studied at 22 wk. WAG were also matched for weight to a young group of FHH rats (FHH-Y), and these were studied at 8 wk. In comparison with WAG and WAG-O rats, FHH and FHH-Y rats exhibited an increased in mean glomerular capillary hydraulic pressure (WAG, 52 +/- 1 mm Hg; WAG-O, 47 +/- 2 mm Hg; FHH, 60 +/- 2 mm Hg; FHH-Y, 65 +/- 1 mm Hg), whereas values in FHL animals were intermediate (56 +/- 2 mm Hg). No significant differences in glomerular volume were found among groups. Moderate focal and segmental glomerular sclerosis developed in FHH and FHH-Y rats, with values for older FHH rats being significantly greater than those for WAG, WAG-O, and FHL animals. Thus, the genetically determined sensitivity to develop proteinuria, focal and segmental glomerular sclerosis, and chronic renal failure in fawn-hooded rats correlated with early evidence of glomerular capillary hypertension.(ABSTRACT TRUNCATED AT 250 WORDS)

The sensitivity of single-strand conformation polymorphism analysis for the detection of single base substitutions.

Single-strand conformation polymorphism (SSCP) analysis has proven to be a simple and effective technique for the detection of single base substitutions. We have used SSCP to analyze 29 mouse globin mutations, 27 p53 mutations, and 8 rhodopsin mutations contained within different size PCR products. Our results indicate that the type of mutation (transition versus transversion) did not play a major role in determining whether a mutation was detected by SSCP analysis. The position of the base substitution was more important than the precise base substitution in determining whether a mutation was detected. We report that SSCP sensitivity varies dramatically with the size of the DNA fragment being analyzed. The optimal size fragment for sensitive base substitution detection by SSCP is approximately 150 bp. Our results illustrate the need to keep the size of the PCR fragment small when performing SSCP to detect mutations. Larger fragments can be analyzed when screening for polymorphisms when the need to detect every sequence variation is not as critical.

Reidentification of larval interneurons in the pupal stage of the tobacco hornworm, Manduca sexta.

The abdominal prolegs are the primary locomotory appendages of Manduca sexta larvae. After the prolegs are lost at pupation, some of the proleg motoneurons die while the survivors are respecified to carry out different functions in the adult moth. As a first step toward investigating the process of functional respecification at the synaptic level, we searched for larval interneurons that affected the activity of proleg motoneurons, and followed these interneurons into the pupal stage. Interneurons were judged to be individually identifiable based on their effects on proleg motoneuron activity and their anatomical features. Seven larval interneurons were identified and placed in five physiological classes based on their effects on proleg motoneurons: ipsilateral excitors, contralateral excitors, ipsilateral inhibitors, contralateral inhibitors, and bilateral inhibitor-excitors. Four of the larval interneurons produced apparently monosynaptic postsynaptic potentials in proleg motoneuron. Of the five larval interneurons that were reidentified in the early pupal stage, two showed minor but consistent structural modifications from the larval stage. Interneurons that produced unitary postsynaptic potentials in larval motoneurons continued to do so in pupal motoneurons. These studies demonstrate that individually identified interneurons can be followed through the larval-pupal transformation, during the initial stages of motoneuron respecification.

Glomerular hypertrophy and epithelial cell injury modulate progressive glomerulosclerosis in the rat.

The effects of glomerular size and visceral epithelial cell integrity upon the development of progressive glomerulosclerosis was studied by superimposing renal ablation on adriamycin-induced nephropathy in rats. Adriamycin alone caused focal epithelial cell injury and proteinuria but minimal segmental glomerulosclerosis. In normal rats, renal ablation was accompanied by mild progressive proteinuria and glomerulosclerosis. However, renal ablation in rats with adriamycin nephropathy caused a dramatic increase in proteinuria and a disproportionately high frequency of segmental glomerulosclerosis. Accelerated glomerular injury after renal ablation in adriamycin-treated rats was associated with substantial glomerular hypertrophy with near doubling of the tuft volume. Morphometric and autoradiographic studies showed that compensatory glomerular hypertrophy occurs without a proportional increase in the number of visceral epithelial cells, leading to a substantial reduction in the density of these cells within the capillary tuft. The severity of segmental glomerulosclerosis showed a significant correlation with the glomerular volume and the reciprocal of the visceral epithelial cell density. Ultrastructural observations indicate that epithelial defects with detachment of the cell processes from the underlying basement membrane are almost invariably seen in areas of segmental glomerulosclerosis with hyalinosis. These findings suggest that the process of progressive glomerulosclerosis is, to a great extent, contingent upon the development of epithelial cell defects, that result from direct injury or from a reduction in the cell density after inordinate compensatory glomerular hypertrophy.

Binocular visual acuity of neonates: the acuity card procedure.

The acuity card procedure is a variant of the forced-choice preferential looking technique. In this study, four observers each tested 10 infants (all healthy fullterm neonates) with acuity cards containing either a one-aperture or a two-aperture stimulus configuration. For both card types the mean test time was eight minutes. No effect on acuity results was found between the two different configurations. The mean acuity values were within one octave of those reported in most previous behavioral studies of the acuity of neonates. The authors consider that the acuity card procedure provides a rapid, accurate technique for assessment of healthy newborn infants in a hospital setting.

Kinetic perimetry assessment of binocular visual field shape and size in young infants.

Kinetic perimetry was used to measure the extent of the binocular visual field in 8 directions in 77 full-term infants tested as neonates, 4-week-olds, or 8-week-olds. The apparatus consisted of a black, 4-arm arc perimeter, a centrally-located 6-deg stationary white sphere, and an identical sphere that served as the peripheral target. Neonates showed larger visual fields than did 4- and 8-week-olds, perhaps due to the strength of the older infants' fixation of the central target. Infants at all ages showed significantly smaller fields than did adults. However, visual field shape was similar in infants and adults.

Mechanism of proximal tubule brush border loss and regeneration following mild renal ischemia.

Left kidneys of rats were made ischemic for 25 minutes and proximal tubule brush border alterations studied in the S1 and S2 segments. Scanning electron microscopy revealed that brush border microvilli became unstable, fused with one another, and were interiorized into proximal tubule cytoplasm soon after reflow of blood following ischemia. Rapid regeneration followed; scanning electron microscopy showed that regeneration occurred in a fashion whereby clusters of microvilli in flower-like configurations were extruded from the cell interior toward the surface. Such unique patterns of microvillus formation have not been reported before. Activity of the brush border enzymes, alkaline phosphatase and maltase, were not significantly depressed throughout the cycle of brush border loss and regeneration. Likewise, there were no alterations in the activity of beta-glucuronidase, a lysosomal enzyme. Alkaline phosphatase cytochemistry showed that microvillus membranes that were interiorized into the cell cytoplasm retained enzyme activity on their surfaces during the early period of brush border loss as well as during regeneration. These results strongly suggest that in reversibly injured proximal tubule cells regeneration of the brush border occurs primarily by a process of recycling of damaged, previously incorporated membrane. The nature of the initial membrane damage and the mechanism of recycling remain unknown.

The vascular basis for acute renal failure in the rat. Preglomerular and postglomerular vasoconstriction.

Myohemoglobinuric acute renal failure (ARF) was induced in dehydrated, salt-deficient, salt-loaded, and untreated rats by intramuscular injection of glycerol, and the renal vasculature was studied after 24 hours. Kidneys were prepared for examination by rapid freezing in vivo to -160 degrees C and freeze substitution in -80 degrees C alcohol, and by perfusion fixation with 1% glutaraldehyde in Ringer's solution at 120 mm Hg. Frozen kidneys were examined by light microscopy after paraffin and epoxy resin embedding. Techniques used in examining the perfusion-fixed kidneys were: (1) vascular injection with silicone rubber and clearing in glycerol, (2) electron microscopy, and (3) morphometric evaluation of lumen to wall area ratios of glomerular arterioles. Kidneys of all rats with ARF showed renal cortical arterial and glomerular arteriolar (afferent and efferent) vasoconstriction. The degree of constriction, estimated by lumen to wall ratios, correlated with the degree of azotemia (r = -0.71; P less than 0.001). Differences between all ARF groups and respective controls were highly significant (P less than 0.001). Vasoconstriction was maximal in the dehydrated group, intermediate in the untreated and Na-deficient rats, and lowest in the salt-loaded animals. Glomerular and peritubular capillaries were patent and free of endothelial swelling or thrombi. Glomerular basement membranes and epithelial foot processes showed no morphological alterations. The observations suggest that marked pre- and postglomerular vasoconstriction occurs in established myohemoglobinuric ARF, that it is related to azotemia, and that mechanical vascular obstruction does not play a major role in this experimental model.