Surface projections and internal structure of Chlamydia psittaci. The outermost surface of the small infectious forms of Chlamydia psittaci contain geometrically arranged spikes distributed over approximately 50% of the bacterial surface. The spikes are located opposite the concave side of an electron-dense crescent-shaped chlamydial core.


Effect of cell length on gliding motility of Flexibacter. Short Flexibacter FS-1 cells, generated during logarithmic growth in glucose-salts medium or by filament fragmentation during the transition from log to stationary phase in rich medium, are unable to glide. Motility returns when cells elongate. This strain also dissociates stable, short, nongliding variants.


Identification and synthesis of acyl-phosphatidylglycerol in Acinetobacter sp. HO1-N. A minor phospholipid from Acinetobacter sp. HO1-N was identified as acyl-phosphatidylglycerol. Acyl-phosphatidylglycerol synthesis by outer-membrane preparations appeared to be a result of phospholipase A activity.


Human aggression and the lunar synodic cycle. Data on five aggressive and/or violent human behaviors were examined by computer to determine whether a relationship exists between the lunar syndoic cycle and human aggression. Homicides, suicides, fatal traffic accidents, aggravated assaults and psychiatric emergency room visits occurring in Dade County, Florida all show lunar periodicities. Homicides and aggravated assaults demonstrate statistically significant clustering of cases around full moon. Psychiatric emergency room visits cluster around first quarter and shows a significantly decreased frequency around new and full moon. The suicide curve shows correlations with both aggravated assaults and fatal traffic accidents, suggesting a self-destructive component for each of these behaviors. The existence of a biological rhythm of human aggression which resonates with the lunar synodic cycle is postulated.


The alcohol provocation test.

The alcohol provocation test. An alcohol provocation test is described wherein a 20% solution of alcohol was injected intravenously in 57 subjects with three major goals: 1) To determine criminal culpability. 2) To determine alcohol activation of the electroencephalogram. 3) To aid in treatment of the chronic alcoholic through personal feedback. The test was found safe and reliable. Results are interpreted on the basis of behavioral observations and blood alcohol levels. Advantages of such a test are reviewed.


A review of the relationship between dysphoria, pleasure, and human bonding. A model is presented which links up recent findings regarding the biochemical substrates of neurological networks mediating pleasure states with the concept of animal behavior relating to bonding. The forms of human bonding (parent-child, kinship, friendship) have analogies in the animal world and appear to be based on a combination of fixed action patterns, learning behaviors supported by endogenous secretion of "pleasure-producing" endorphines, and social reinforcement. The implications of this model for psychiatric treatment and rehabilitation are discussed.


Evaluation of substance abuse problems in a psychiatric hospital. Hospital-wide rumors and anecdotal reports had suggested that there was a potentially serious problem of alcohol and drug abuse among the psychiatric patients of this hospital. The authors describe a systematic, evaluative approach which gave direct and detailed information on the nature and extent of the problem thus permitting effective administrative solutions. Results of the study indicated that there were two qualitatively different populations of substance abusing patients. A considerable population of patients were routinely using alcohol and "soft" drugs on the hospital grounds, while another disturbing percentage of patients gave evidence of psychological or physical dependence on alcohol or hard drugs. Data from the evaluation was instrumental in implementing additional security measures to deal with casual substance abuse and additional, confidential treatment resources to deal with the more serious substance abuse problems in patients and staff.


Circadian temperature variation and depressive illness. Circadian temperature rhythms were studied at different clinical phases in inpatients admitted with a diagnosis of depression. It was found that patients meeting diagnostic criteria for either bipolar or unipolar II classification had a significantly greater likelihood of having an altered temperature rhythm while clinically depressed. The implications of this finding are discussed and questions for future research are specified.


Apathetic hyperthroidism.

Apathetic hyperthroidism. Among the psychosyndromes least well known to be associated with an endocrinopathy is apathetic hyperthyroidism. This condition is characterized by apathy, lethargy, severe "senile depression" and often a high output cardiac failure occurring as the presenting problem.1,2,9 The depressive component of the clinical picture may occur without cardiac complications. Such depressions in the elderly, while refractory to treatment with antidepressant drugs, often respond dramatically to the correction of the underlying endocrine dysfunction.4 It is therefore important to consider "thyroid melancholia" in the differential diagnosis of such cases. Given this high index of suspicion, the diagnosis can be fairly easily made.


Clinical importance of doxepin antidepressant plasma levels. Tricyclic antidepressant plasma levels have greatly expanded the capabilities of the physician in treating depression. Although still not widely available to the practicing psychiatrist, plasma levels can make an important difference in treating the "drug resistant" depressed patient in particular. The following case report with the use of doxepin underscores the clinical significance of trychclic plasma levels, plus lends support to the previous reports that there is a therapeutic plasma range for the tricyclics which if at too high a level can be as ineffective as too low a level.


Agranulocytosis associated with tricyclics. A case of agranulocytosis associated with Imipramine use is reported. A review of the literature suggests that this is a rare but hazardous complication of tricyclic use. The authors present characteristic findings, treatment suggestions and a possible pathogenesis for this syndrome.


Fractionation of unfixed chromatin by buoyant-density centrifugation in gradients containing 3-iodo-1,2-propanediol and metrizamide. Buoyant-density centrifugation of unfixed chromatin has been performed in a newly devised medium containing 3-iodo-1,2-propanediol and metrizamide. Chromatins were obtained from isotopically labeled mouse hepatoma cells in suspension culture, either grown normally or density labeled in a medium containing bromodeoxyuridine, by mild digestion of isolated nuclei with micrococcal nuclease. When a mixture of normal and density labeled chromatin, marked with [14C]thymidine and [3H]bromodeoxyuridine, respectively, was centrifuged in the medium, chromatin peaks represented by labeled DNA were resolved to the extent expected from their separate banding profiles. Centrifugation of an equivalent chromatin mixture labeled with [14C] and [3H]lysine, respectively, also yielded resolution of chromatin peaks represented by labeled proteins. Only small amounts of labeled proteins were dissociated from chromatin in the gradient medium. Labeled proteins recovered from the gradient fractions were analyzed by electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. The results suggested that most of the histones remained associated with the original stretches of DNA during the centrifual fractionation period. Essentially all of the dissociated proteins were found to be non-histone proteins.


Physicochemical properties of chondroitin sulfate. I. Ion binding and secondary structure. 1. The degree of calcium ion binding to chondroitin sulfate was larger than that of sodium ions. The association of calcium ions with the mucopolysaccharide cannot be ascribed solely to electrostatic forces. 2. No differences in the degree of ion binding of chondroitin sulfate isomers A and C were detectable. 3. The spacing between charged groups of chondroitin sulfate was found to be 0.48 nm, which is consistent with the results of X-ray analysis. 4. The characteristic ratio was large. Therefore, rotation about the backbone bonds is highly restricted.


Physicochemical properties of chondroitin sulfate. II. Molecular weight dependence of intrinsic viscosity and sedimentation coefficient, and analysis as a semi-flexible coil. 1. The relation between intrinsic viscosity in 0.2 M NaCl and molecular weight for samples of chondroitin sulfate isolated from shark cartilage could be expressed by [eeta] = 6.5 X 10(-6) Mr114. 2. Their sedimentation coefficients were linear with respect to the square root of molecular weight, and the intercept was not zero but definitely positive, indicating that chondroitin sulfate in solution can be represented by a semi-flexible coil with a molecular diameter of about 1.2 nm. This value was consistent with the results of X-ray analysis. 3. The molecular weight dependence of the sedimentation coefficient for chondroitin sulfate from ox nasal septa cartilage was similar to that for chondroitin sulfate isolated from shark cartilage.


Enzymatic determination of phospholipase D activity with choline oxidase. A new enzymatic method was developed for the assay of phospholipase D [phosphatidylcholine phosphatidohydrolase EC 3.1.4.4] from cabbage leaves using choline oxidase from Arthrobacter globiformis cells. The method was based on the estimation of choline by the following series of enzymatic reactions after ending the phospholipase D reaction: Choline + 202 + h2o Choline oxidase Betaine + 2H2O2 2H2O2 + Phenol + 4-Aminoantipyrine Peroxidase Quinoneimine dye + 4H2O The amount of choline was proportional to the amount of resulting quinoneimine dye with an absorbance maximum at 500 nm. The phospholipase D reaction (choline liberation) was carried out at pH 5.5 in the presence of Ca2+ ions and ended by adding EDTA in conc. Tris-HCl buffer, pH 8, to give a final pH of around 8. The initial rate of the phospholipase D reaction was proportional to the enzyme concentration over the absorbance change range of 0 to 0.25 (equivalent to 0-21 micron of choline) under the optimal reaction conditions.


Precursors of globin mRNA in erythroid-enriched bone marrow cells of mouse. RNA from erythroid-enriched bone marrow cells of mouse was analyzed under two kinds of denaturing conditions (in 99% formamide and formaldehyde treatment), and globin mRNA sequence-containing RNAs larger than 18S in addition to 16S and 10S were detected by hybridization at a comparatively high Rot with complementary DNA (cDNA) to globin mRNA (10S). The existence of precursor RNAs larger than 18S was confirmed by two different methods; (i) recentrifugation in 99% formamide of RNA collected from the corresponding regions of RNA fractions and following detection of globin mRNA sequences, (ii) the specific isolation of [3H]labeled precursor RNA with a cDNA-cellulose column and fractionation in a 99% formamide-sucrose gradient. Furthermore, pulse-chase experiments indicated that some large RNAs with globin mRNA sequences could be chased to 10S RNA during a 30 min chase in the presence of a high concentration of actinomycin D (10 microgram/ml). From the results it was assumed that 35-37S RNAs with globin mRNA sequences were possible primary transcripts of globin genes and other RNAs with globin mRNA sequences in 16S, 20S, and 26S regions were intermediate precursors of 10S globin mRNA.


Studies on soybean trypsin inhibitors, XII. Linear sequences of two soybean double-headed trypsin inhibitors, D-II and E-I. Soybean inhibitor D-II is an inhibitor of bovine trypsin. Sequence analysis was carried out on the reduced and S-carboxymethylated protein by conventional methods to establish the complete amino acid sequence. The sequence of D-II indicated high homology with other legume inhibitors, but it was unique because of the occurrence of identical residues (arginine) at both of the reactive sites. This structure is thought to reflect that of a prototype double-headed inhibitor. The possible evolutionary process of the legume double-headed inhibitors is discussed on this basis. Comparison with another soybean inhibitor C-II suggested that a single methionine (C-II)-glutamine (D-II) replacement at the P2'position resulted in the loss of alpha-chymotrypsin inhibitory activity of D-II. The results of a hydrogen peroxide oxidation experiment on C-II supported this suggestion. The sequence of the amino-terminal 21 residues of inhibitor E-I was determined using a sequentor. It was shown that this inhibitor lacks the amino-terminal nine residues of D-II.


Isolation of 3beta,7alpha-dihydroxychol-5-enoic acid, an intermediate of chenodeoxycholic acid biogenesis, and 3alpha,7alpha-dihydroxychol-4-enoic acid from bladder bile of hens. Two Lifschütz-positive C24-bile acids were isolated from bladder bile of hens. One of these was identified by isotope dilution experiments after conversion to a 3H-labeled compound, and also by GLC after methoxylation, as 3beta,7alpha-dihydroxychol-5-enoic acid, a key intermediate of chenodeoxycholic acid biogenesis. The other, to which the structure 3beta,7alpha-dihydroxychol-4-enoic acid had been assigned previously, was proved to be its 3alpha-epimer by several experiments. These findings favor the alternative pathway of chenodeoxycholic acid biogenesis proposed by Yamasaki and his associates.


Pyridinoline, a non-reducible crosslink of collagen. Quantitative determination, distribution, and isolation of a crosslinked peptide. Pyridinoline is a crosslink compound isolated from bovine Achilles tendon collagen. It is a 3-hydroxypyridinium derivative with three amino and three carboxyl groups (Fujimoto, D., Akiba, K., & Nakamura, N. (1977) Biochem. Biophys. Res. Commun. 76, 1124-1129). The contents of pyridinoline in collagens from various sources were determined. The pyridinoline content of bovine Achilles tendon was 0.16 residue per 1,000 residues and that of rat Achilles tendon collagen was 0.017 residue per 1,000 residues. Besides Achilles tendon collagens, pyridinoline was found in collagens from costal cartilage, rib and femoral bone of rat. It was not found in collagens from the tail tendon and skin of rat. A crosslinked, triple-chained peptide containing pyridinoline was isolated from bovine Achilles tendon collagen after digestion with pronase. Its amino acid composition suggests that the peptide may be involved in an intermolecular crosslink among a carboxyterminal sequence, a sequence near the aminoterminus and a sequence in the helical region.


Topography of the myosin molecule as visualized by an improved negative staining method. An improved negative staining method has been used to visualize monomeric myosin molecules. Each lobe (subfragment 1) of the molecule looked like an elongated pear, and the widths of the thick and thin portions were about 95 and 55 A, respectively. The length of each lobe was about 210 A. It appeared capable of moving azimuthally and altitudinally, utilizing its juncture with the rod portion as the base. The rod portion of the molecule was about 1400 A long and 30 A wide. It also appeared to possess a considerably flexible region at a point about 680 A from the tail-end.


The use of transparent media in medical illustration. Transparent media, such as watercolor, acrylic, and dyes, have been used by scientific illustrators for centuries. This article gives one artist's view of the highly complex medium. Materials, techniques, short cuts, and potential problem areas are discussed. Illustrations of specific techniques and a step-by-step development of a medical illustration is provided.


Concentration-dependent conformational transition of alpha-elastin in aqueous solution. Circular dichroism measurements on aqueous solutions of alpha-elastin have given evidence of beta-bend structure at elevated concentrations. When Ca2+ was added, the concentration-dependent conformational transition was somewhat inhibited and the binding of the metal ion was shown, by means of equilibrium dialysis, to be essentially independent from alpha-elastin concentration in the range of 1 to 10 mg/ml. The results obtained are discussed in connection with the elasticity and calcification of elastin.


Biosynthesis of S-N6-methyladenosylhomocysteine, an inhibitor of RNA methyltransferases. This paper demonstrates that N6-methyladenosine (6-methylaminopurine ribonucleoside) will condense in vitro with homocysteine to form S-N6-methyladenosylhomocysteine in a reaction catalyzed by mouse liver S-adenosylhomocysteine hydrolase. Injection of mice with N6-methyladenosine is followed by accumulation of S-N6-methyladenosylhomocysteine in the liver. Studies from other laboratories have shown that S-N6-methyladenosylhomocysteine is nearly as potent an RNA methyltransferase inhibitor as S-adenosylhomocysteine. This indicates that administration of N6-methyladenosine may be a general method for blocking in vivo RNA methylation in studies to determine the role of methylation in RNA processing and translational function.


Kinetics of oxygen-18 exchange between inorganic phosphate and water catalyzed by myosin subfragment 1, using the 18O shift in 31P NMR. The time course of oxygen-18 exchange between [18O]Pi and normal water, catalyzed by myosin subfragment 1 in the presence of MgADP, was followed using the shift in 31P NMR caused by the presence of oxygen-18 bound to the phosphorus. Essentially all molecules of [18O]Pi that bind to the enzyme undergo complete exchange and are released as [16O4]Pi. Exchange probably occurs by formation of myosin.ATP from a myosin.ADP.Pi complex and is rapid relative to release of Pi from this complex. The kinetics of exchange give a value for the rate constant for binding Pi to myosin.ADP of 0.23 M-1 S-1 (pH 8.0, 22 degrees C). This value is consistent with exchange occurring by reversal of the ATP-ase reaction back to the myosin.ATP complex.


A comparison of molecular properties of hepatic triglyceride lipase and lipoprotein lipase from human post-heparin plasma. Hepatic triglyceride lipase was isolated from human post-heparin plasma by the method of Ehnholm et al. using modifications which increased the specific activity 12-fold to approximately 3,000 mumol of free fatty acid/h/mg of protein. Lipoprotein lipase with similar specific activity was prepared from the same plasma samples using heparin and concanavalin A affinity chromatography. The molecular weight of hepatic triglyceride lipase (69,000) was slightly greater than that of lipoprotein lipase (67,000) as determined by polyacrylamide electrophoresis in sodium dodecyl sulfate-containing buffers. These proteins had identical amino acid compositions, terminal amino acid residues, and tryptic peptide maps. However, the differences previously described regarding optima of pH and ionic strength and the requirement for apolipoprotein CII (only for lipoprotein lipase) were maintained in the highly purified state. It was found that both proteins contain approximately 8% carbohydrate. Antisera prepared in goats selectively precipitated each activity. Other antisera prepared in chickens reacted with both enzymes, suggesting a common antigenic determinant.


Hepatic microsomal epoxide hydrase. Involvement of a histidine at the active site suggests a nucleophilic mechanism. The effects of a wide variety of chemical modification reagents on the activity of purified rat liver microsomal epoxide hydrase have been investigated. Alkylating agents, such as the phenacyl bromides and benzyl bromide are potent inhibitors of epoxide hydrase. 2-Bromo-4'-nitroacetophenone (p-nitrophenacyl bromide) specifically and irreversibly inactivates epoxide hydrase. Pseudo-first order kinetics of inhibition is observed at higher inhibitor/enzyme ratios. The rate of inactivation is controlled by a group on the enzyme with an apparent pKa of 7.6. Inactivation of the enzyme with 14C-labeled 2-bromo-4'-nitroacetophenone leads to the incorporation of approximately 1 mol of radioactive inhibitor/mol of protein. Epoxide hydrase can be protected against this inactivation by the substrate phenanthrene-9,10-oxide. These results are consistent with the interpretation that 2-bromo-4'-nitroacetophenone acts as an active site-directed inhibitor. The site of alkylation by 2-bromo-4'-nitroacetophenone is a histidine residue of epoxide hydrase. The N-alkylated histidine derivative has been identified as 1-(p-nitrophenacyl)-4-histidine. A possible mechanism for the enzymatic hydration catalyzed by epoxide hydrase is discussed which involves a histidine residue of the enzyme serving as a general base catalyst for the nucleophilic addition of water.


The green hemoproteins of bovine erythrocytes. II. Spectral, ligand-binding, and electrochemical properties. The two green hemoproteins isolated from bovine erythrocytes (form I and form II) have been characterized as to spectral, electrochemical, and chemical properties. The absorption spectra of the isolated hemoproteins are typical of high spin ferric states. Reduction of the hemoproteins yields high spin ferrohemoproteins. Complexation of the ferrohemoproteins with CO and the ferrihemoproteins with cyanide yields low spin complexes, demonstrating the presence of an exchangeable weak field ligand in both the ferrous and ferric states of the hemoproteins. The differences in position and intensity of the absorption peaks of the visible spectra allow the two forms to be distinguished from one another. The midpoint potential of forms I and II were found to be +0.075 and +0.019 V, respectively, at pH 6.4 and +0.038 and -0.005 V, respectively, at pH 7.0. This is consistent with the gaining of 1 proton/electron during the reduction. The Nernst plot reveals an unusual 0.5-electron transfer, whereas a quantitative titration demonstrates a 1-electron transfer. Form I binds cyanide more tightly than form II (KD of 84 and 252 micrometer, respectively). The observed spectral, electrochemical, and ligand-binding differences between forms I and II can be explained in terms of a greater electron-withdrawing ability of the side chains of the heme of form I relative to the heme of form II.


Binding and functional properties of concanavalin A and its derivatives. III. Interactions with indoleacetic acid and other hydrophobic ligands. The binding of concanavalin A to various structures via hydrophobic interactions has been studied using a variety of physicochemical assays. It was found that concanavalin A binds to nonpolar compounds such as the plant auxin beta-indoleacetic acid and its structural analogue tryptophan and that this binding is independent of the saccharide-binding activity normally associated with the lectin. The results of equilibrium dialysis experiments on the binding of beta-indoleacetic acid were consistent with the presence of a single weak binding site per subunit of protein, having an association constant of about 7 X 10(2) M-1. Competition experiments using various nonpolar compounds such as o-iodobenzoic acid suggested that this hydrophobic binding site is located in the same cavity which binds the iodine-containing ligand as shown by x-ray crystallography. Concanavalin A also binds to lipid vesicles composed of dipalmitoylphosphatidylcholine or 12-O-tetradecanoyl phorbol-13-O-acetate. This binding to lipid membranes raises the possibility that the synergistic effects of concanavalin A and tetradecanoyl phorbol acetate on lymphocyte mitogenesis may be due in part to an interaction between lectin and the phorbol ester.


A comparison of the substrate specificities of cathepsin D and pseudorenin. Cathepsin D, purified from hog spleen, releases angiotensin I from tetradecapeptide renin substrate and from protein renin substrates purified from hog and human plasma. However, the enzyme does not act on the naturally occurring renin substrate as it exists in plasma nor on purified substrate in the presence of plasma. Cathepsin D releases angiotensin I quantitatively from tetradecapeptide renin substrate and does not further degrade the angiotensin I on prolonged incubation. The pH optimum for cathepsin D prolonged incubation. The pH optimum for cathepsin D acting on tetradecapeptide renin substrate is 4.5, and there is very low activity above pH 7. These properties are very similar to those of pseudorenin, an angiotensin-forming enzyme originally isolated from human kidney, indicating that cathepsin D and pseudorenin may be identical.


Alteration of 86Rb+ influx and efflux following depletion of membrane sterol in L-cells. Ouabain-sensitive uptake of 86Rb+ (an analogue of K+) was enhanced in L-cells that had been treated with 25-hydroxycholesterol or 7-ketocholesterol in order to deplete their sterol concentration. Ouabain-insensitive Rb+ efflux also increased in the sterol-depleted cells and the intracellular concentration of K+ diminished while the concentration of Na+ increased. All of these effects of 25-hydroxycholesterol were counteracted by the addition of mevalonate to the culture medium. Despite the evidence for increased active Rb+ transport in the 25-hydroxycholesterol-treated cells, the level of sodium and potassium ion-activated adenosine triphosphatase ((Na+ + K+)-activated ATPase) activity measured in homogenates and plasma membrane preparations from the treated cells was not significantly different from the control values. Rb+ uptake was more sensitive to ouabain inhibition in sterol-depleted cells than in control cells, although ATPase activity in plasma membrane fractions isolated from treated cells was not more sensitive to ouabain inhibition than was that from control cells. It is possible that the ability of the oxygenated sterols to inhibit DNA synthesis and cell division (Kandutsch, A. A., and Chen, H. W. (1977) J. Biol. Chem. 252, 409-415) is related to their effects upon cellular ion transport.


Purification of mammalian filamin. Similarity to high molecular weight actin-binding protein in macrophages, platelets, fibroblasts, and other tissues. We have purified the high molecular weight actin-binding protein, filamin from guinea pig vas deferens. We find this mammalian filamin is very similar to chicken gizzard filamin in subunit molecular weight, amnio acid composition, actin-binding properties, immunological cross-reactivity, and the ability to be phosphorylated by cyclic AMP-dependent protein kinase. Anti-filamin antibodies cross-react with a high molecular weight macrophage actin-binding protein, and with a high molecular weight protein in platelets and fibroblasts. Furthermore like filamin, these proteins are also phosphorylated and cyclic AMP stimulates their phosphorylation. Anti-filamin antibodies do not cross-react with the erythrocyte membrane protein spectrin or with high molecular weight proteins in brain extracts. We conclude that filamin from avian and mammalian smooth muscle are very similar proteins and furthermore that many, but not all, non-muscle cells contain a protein closely related to filamin.


Neurohypophyseal hormone-responsive renal adenylate cyclase. IV. A random-hit matrix model for coupline in a hormone-sensitive adenylate cyclase system. A "random-hit" matrix model is proposed to account for the dynamic and steady state relationship between occupation of bovine renal medullary membrane receptors by [Lys8]vasopressin (LVP) and neurohypophyseal hormones (NHH) and the associated activation of membrane-bound adenylate cyclase. The model was developed by systematic introduction of specific rules concerning receptor coupling into a general structural model which consists of two square matrices of identical size, one composed of homogeneous R ("receptor") units, the second of homogeneous C ("cyclase") units. R units are either occupied (RO) or unoccupied (RU); C units are either active (CA) or inactive (CI). Hormone molecules are envisioned to "collide" with R units randomly; collision with RU leads to "binding", and occupation is maintained for a characteristic mean occupancy time, TO. In this structure, each R unit has an "interaction field" which consists of the "twin" unit in the "C" matrix, and the 4 nearest neighbor C units surrounding the twin. Occupation of an R unit leads to activation of all CI units in the interaction field of that R; CA units in the interaction field are refractory. Thus binding at a given R may "recruit" a variable number of inactive neighboring C units (5, 4, 3, 2, 1, or 0). The model requires that there be individual coupling delays between the moment of binding at a given R and subsequent activation of CI units (mean coupling delay (Td) approximately 10% To). Activation of C units persists as long as the "parent" R is occupied and is maintained for an additional short time interval (Tp) after RO reverts to RU, corresponding to hormone dissociation from receptor. The model accounts for the following previously demonstrated relations between LVP occupation of receptors and adenylate cyclase activation in bovine renal medullary membranes: 1) the shape of the nonlinear steady state relation between normalized (percentage maximal) receptor occupation (O) and cyclase activation (A), uniformly observed in different membrane preparations: 2) variable hormone concentration-dependent trajectories of approach to the final steady state A:O value (A:Oss) which may be either monophasic or biphasic; 3) the loss of intrinsic adenylate cyclase activity observed in bovine membranes for a series of NHH analogs with progressively diminishing affinity for receptors. The model represents an explicit theory of coupling where a successive series of temporal events are quantitatively related to each other and privide major constraints to any interpretation of the molecular organization of receptors and adenylate cyclase units in membranes. The model excludes a number of mechanistic proposals and suggests a new hypothesis for membrane coupling with features which may be generally applicable to other hormone-sensitive adenylate cyclase systems.


Studies on macromomycin, an antitumor protein. Macromomycin is a protein isolated from the culture filtrate of Streptomyces macromomyceticus. It is an antibiotic and also cytotoxic to a broad spectrum of carcinoma cells, the ID50 for P388 leukemia cells being 1 X 10(-9) M. Macromomycin binds rapidly and tightly to the P388 cell membrane and the eventual death of the cell cannot be reversed by either washing the toxin away or treating the cell with trypsin. The cytotoxicity does not appear to be specific for any phase of the P388 cell cycle. Macromomycin is a single polypeptide, pI 5.38, devoid of methionine and arginine residues and contains 4 cysteine residues joined by two intramolecular disulfide bonds. The cytotoxicity results in inhibition of DNA, RNA, and protein synthesis in P388, the latter inhibition occurring a few hours after the inhibition of nucleic acid synthesis. The antibiotic and antitumor activities are destroyed rapidly by ultraviolet light, which gives a product that differs little in amino acid composition, molecular weight, and antigenic property, but can be separated from the native macromomycin by ion exchange chromatography. It is proposed that macromomycin has an ultraviolet-sensitive prosthetic group upon which much of the biological activity is based.


Complications after cuneiform osteotomy for moderately or severely slipped capital femoral epiphysis. Wedge osteotomy of the femoral neck for slipped capital femoral epiphysis was performed in seventy-seven hips of seventy-one children at Gillette Children's Hospital during the period 1938 to 1973. The major complications were avascular necrosis (28.5 per cent) and cartilage necrosis (37.6 per cent). This high incidence of severe complications led to the use of an osteotomy through the base of the neck for the treatment of severely slipped capital femoral epiphysis which has given satisfactory results in six hips to date.


Legg-Perthes disease in children less than four years old. In children with Legg-Perthes disease who were less than four years old, better results were seen after treatment by recumbency and abduction bracing than with no treatment. All results were better in children having mild epiphyseal involvement (Catterall Groups I and II) than in those with more extensive disease.


The effect of hip reduction on function in patients with myelomeningocele. Potential gains and hazards of surgical treatment. We studied seventy-six patients with myelomeningocele who were more than five years old and ahd had no hip surgery during the two previous years. Of these seventy-six patients, forty-one had had no operative treatment intended to reduce the dislocation of their hips and thirty-five had been operated on one or more times to reduce or maintain reduction of one or both hips. The presence of the femoral head in the acetabulum did not improve range of hip motion or ability to walk, nor did it reduce the amount of bracing required or decrease pain. The complications of surgical treatment to gain reduction were numerous and included failure to obtain stability in 40 per cent of the hips, loss of hip motion, and fractures. A level pelvis and a good range of hip motion were found to be more important for function than reduction of the hips. The goal of treatment should be maximum function, not roentgenographic reduction of the dislocated hip.


Scoliosis: incidence and natural history. A prospective epidemiological study. A prospective study was carried out of the incidence and natural history of adolescent idiopathic scoliosis in 26,947 students. Data were obtained on 1,122 students with idiopathic scoliosis. The incidence of idiopathic scoliosis was 4.5 per cent. The female-to-male ratio was 1.25:1.0 over-all, but the ratio varied directly with the severity of the curve--that is, 1:1 for curves of 6 to 10 degrees, and 5.4:1 for curves of more than 20 degrees. Progression of the curve was determined by a two-year follow-up of 603 patients. Progression was observed in 6.8 per cent of the students and in 15.4 per cent of the skeletally immature girls with scoliosis of more than 10 degrees at the initial examination. In 20 per cent of the skeletally immature children with curves of 20 degrees at the initial examination, there was no progression. Spontaneous improvement of the curve occurred in 3 per cent and was seen more frequently in curves milder than 11 degrees. Treatment was required in 2.75 students per 1,000 screened.


Unicompartmental tibiofemoral resurfacing arthroplasty. Thirty-seven patients with unicompartmental osteoarthritis were treated by replacement arthroplasty using the Marmor modular prosthesis and each patient was followed for at least two years. Despite rigid preoperative criteria for the selection of cases, the over-all results were definitely inferior to those seen after either bicompartmental or tricompartmental replacement arthroplasty. Complications included recurrent pain and degeneration of the other compartment, often associated with abrasion of the remaining cartilaginous surfaces caused by polyethylene particles. There was also more settling of the tibial component in these patients than in patients treated by bicompartmental replacement arthroplasty. Because of these findings, we no longer use unicompartmental replacement of the medial side of the knee. The results after the three lateral compartment replacements, on the other hand, were good, and use of this procedure for post-traumatic arthritis is still under investigation.


Factors influencing manual muscle tests in physical therapy. To determine whether it is the amount or the duration of the force applied manually by the tester, or both, that determines the tester's perception of the strength of the hip flexor or abductor muscles, an electromechanical device was designed which was placed between the tester's hand and the subject's limb. With the device we measured the force applied to the limb, the time interval during which it was applied, and the angular position of the limb during the entire test. In 240 such tests, the testers' ratings of the differences in strength between the right and left sides were correlated with seven variables involving force and time. It was found statistically that the impluse--that is, the duration of the tester's effort multiplied by the average applied force during each test--was the factor that most influenced the tester in the ratings.


Osteonecrosis of the knee. A clinicopathological study in twenty-eight patients. Specimens from thirty-four knees in twenty-eight patients with histologically proved osteonecrosis were reviewed. Twelve knees had the clinical diagnosis of so-called idiopathic osteonecrosis; eight, osteoarthrosis; and fourteen, rheumatoid arthritis. Pathologically, the necrosis was localized in the subchondral region of the medial femoral condyle in 67 per cent of the knees. There were pathological fracture and collapse and fragmentation of the necrotic segment leading to marked deformities in most cases. The response to necrosis consisted of histiocytic resorption of necrotic material and formation of granulation tissue and reactive new bone surrounding it. The response was relatively less prominent in the knees with degenerative joint disease and rheumatoid arthritis.


Neuroma scar formation in rats following peripheral nerve transection. Utilizing a sciatic nerve-transection model in rats, we studied the collagen content of normal control nerves, the long-term collagen accummulation in the distal stump of the transected nerve, the content of collagen of the posterior tibial nerve distal to the transection, and the myofibrosis of the gastrocnemius muscle. The distal stump of the sciatic nerve accumulated significant amounts of collagen with time but the posterior tibial nerve at a distance from the site of transection showed no progressive collagen accumulation. The histologically evident myofibrosis of the denervated gastrocnemius muscle was shown to be the result of a selective loss of muscle elements and not of an accumulation of collagen. cis-Hydroxyproline, a proline analogue, limited neuroma formation following neurorrhaphy. The findings suggest that the results of delayed repair of nerves should not be impaired by intraneural scarring if enough of the transected nerve beyond the point of intraneural scar formation is resected.


Total hip replacement with and without osteotomy of the greater trochanter. Clinical and biomechanical comparisons in the same patients. Twelve patients with bilateral hip replacement, one side performed with a trochanteric osteotomy and the other without osteotomy, were analyzed by review of the clinical records, the findings on physical examination, preoperative and postoperative Harris hip scores, each patient's evaluation, biomechanical analysis of the preoperative and postoperative roentgenograms of the hips and pelvis, and force-plate gait analysis. All operations yielded good results clinically but the majority of patients preferred the replacement done without osteotomy. Blood loss and trochanteric bursitis were greater problems in patients who had an osteotomy. There were no differences between the two groups in the postoperative Harris scores or in the biomechanical measurements made on the preoperative and postoperative roentgenograms except for the change in the abductor-muscle length. Changes in this parameter correlated well with the patients' symptoms and clinical findings.


Reattachment of the greater trochanter in total hip-replacement arthroplasty. A new technique. For certian total hip-replacement operations osteotomy of the greater trochanter is essential, and for others it is advantageous. A system of management of trochanter reattachment is described that takes into account multiple factors, such as the need to remove large or small amounts of the trochanter depending on the patient's anatomy and on the style of the femoral component used, the need in some cases to advance the trochanter to lie against the cortex of the femur, the need in other instances to deal with markedly porotic trochanters, and the fact that a small number of the wires will fatigue, regardless of the technique used. The basic technique has one horizontal and two separate vertical wires, all passing medial to the femoral stem and each tied to itself in a square knot; All wires pass through the trochanter to prevent rotation. Wire mesh is used prophylactically if the trochanter is porotic. All patients use two crutches for two months postoperatively; In a consecutive series of 136 total hip replacements in which the trochanter was reattached using this technique, none of the trochanters migrated and all united.


Complications of trochanteric osteotomy in total hip replacement. Nearly 5 per cent of 728 hips showed separation and migration of the trochanteric fragment after total hip replacement using the transtrochanteric approach and a cruciate two-wire technique of reattachment. In the 728 consecutive hips, thirty-five trochanters showed separation and eighteen patients had a positive Trendelenburg test. The abductor weakness was correlated with the amount of separation especially if it exceeded two centimeters. Trochanteric separation was attributed to osteoporosis or poor technique in all but two hips, and trauma was implicated in one.


Dislocations after total hip-replacement arthroplasties. In a series of 300 total hip replacements, nine (3 per cent) dislocated. Precise measurements of the orientation of the acetabular cup were made and it was found that anterior dislocations were associated with increased acetabular-component anteversion. There was no significant correlation between cup-orientation angle and posterior dislocation. The dislocation rate for cup orientation with anteversion of 15 +/- 10 degrees and lateral opening of 40 +/- 10 degrees was 1.5 per cent, while outside this "safe" range the dislocation rate was 6.1 per cent. Other factors that were documented include time after surgery (with the greatest risk in the first thirty days) and surgical history (with a greater risk in hips that have had prior surgery).


Influence of age and sex on the strength of bone-ligament junctions in knee joints of rats. The bone-ligament junction strength of femur-medial collateral ligament-tibia complexes in rats was measured in situ at various ages during a two-year period. Male rats had a higher junction strength than female rats, a difference that became apparent when the animals were sixty days old and in male but not female animals subsequently paralleled the changes in body weight. However, on a bodyweight basis, the junctions were stronger in female than in male rats and this sex difference was evident at fifteen days old and persisted thereafter. Regression analysis between body weight and junction strength indicated that female rats had a significantly higher slope than males, which suggested that the sex differences were due to a hormonal factor or factors. Other measurements showed that elastic stiffness, failure energy, and collagen concentration in the ligament increased, whereas the water content of the ligament decreased with age. Most of these changes could be attriubted to the aging process and not the sex of the animal. It was concluded, however, that the strength of the insertion sites of ligaments on bones are responsive to the hormonal fluctuations that occur with aging.


Osteochondritis dissecans of the knee with loose fragments. Treatment by replacement and fixation with readily removed pins. We reviewed six patients with a five to fifteen-year follow-up and a seventh with shorter follow-up who underwent replacement and fixation with easily removable pins of large osteocartilaginous fragments resulting from osteochondritis dissecans. Excellent or good results were achieved in all but one patient. Preparation of the fragments to fit into their vascularized beds prior to internal fixation is advisable, and in some cases the bed should be partially filled with cancellous bone before the fragments are replaced. Immobilization is prescribed until the pins are removed three to six weeks after surgery. Weightbearing is delayed until there is roentgenographic evidence of beginning union of the fragment to the underlying bone.


Coccidioidal spondylitis.

Coccidioidal spondylitis. A review of twelve cases in which disseminated coccidioidomycosis caused localized infection of the spine showed that eight of the twelve patients were alive and well with no evidence of active infection an average of eleven years after onset (range, two to thirty-five years). One patient who was followed for more than twenty-three years had a slowly developing neurological impairment in the lower extremities as a result of lumbosarcral destruction instability. One patient died early in the course of the disease from fulminating cervical spondylitis and quadriplegia. A second patient had a paraplegia from thoracic spondylitis. On patient had no evidence of active spondylitis five years after the onset of the disease, but then died of coccidioidal meningitis. All patients were treated with intravenous amphotericin at some time in the course of their illness, although its effect was not always dramatic. The three patients with neurological impairment did not undergo spine fusion, but most of the others had that operation. Surgical evacuation of abscesses and debridement of infected bone was also performed in many cases.


Simultaneous and staged bilateral total knee arthroplasty. Bilateral total knee arthroplasties were performed in twenty-six patients with severe rheumatoid arthritis and osteoarthritis. Twelve had simultaneous operations bilaterally and fourteen had separate, staged procedures. The improvement in the two groups of patients was statistically comparable to the relief of pain and the over-all restoration of function. There were no intraoperative complications. The postoperative morbidity was minimum and was not predominant in either group. Costs were greatly increased with the staged procedure.


Deyerle treatment for femoral neck fractures. Thirty-eight displaced intracapsular fractures of the femoral neck treated by the Deyerle method of internal fixation have been added to the previously reported series of sixty-three cases from the Walter Reed Army Medical Center Orthopaedic Service. A significant change in the incidence of non-union was found. In the over-all series of 101 patients, the incidence of non-union was 13.8 per cent and the incidence of segmental collapse due to avascular necrosis was 13.8 per cent. The roentgenographic and functional over-all end results were rated excellent in 67.7 per cent and poor in 32.3 per cent of the fractures. Some difficulties arising from the multiple-pin and plate technique and the associated morbidity are discussed. We now think that Deyerle method produces no better results than other methods of internal fixation of displaced intracapsular fractures.


Radiologic findings in acute childhood asthma. One hundred and seventy-eight admissions of 142 children with acute asthma were reviewed and graded for clinical severity. Abnormalities including consolidation (21%), atelectasis (3%) and pneumomediastinum (2%) were found on chest radiographs, most commonly in patients with clinically moderate or severe acute asthma. Hyperinflation proved difficult to assess, but was present in only 67% of admissions using the criteria described. The frequency of complications detected justifies the use of chest radiographs in assessing children admitted to hospital for acute asthma.


Discogenic vertebral sclerosis.

Discogenic vertebral sclerosis. Narrowing of a disc space, sclerosis of the adjacent vertebral bodies, and irregularity of the vertebral end plates constitute a roentgen complex classically associated with infectious spondylitis. However, narrowing of a disc space may be the result of a herniated nucleus pulposus, and in some individuals the reactive sclerosis is sufficiently intense to resemble infection. Roentgenographic recognition of discogenic vertebral sclerosis may obviate the need for further evaluation and biopsy. Forty-four cases of discogenic vertebral sclerosis, nine with surgical proof and four developing after disc surgery, are presented. Roentgen signs helping to distinguish it from infection include: (i) rounded sclerosis most frequently in the anterior vertebral body (100%), (ii) central lucency in the sclerosis (77%), (iii) associated degenerative changes at the intervertebral disc, e.g. vacuum phenomena (44%), (iv) absence of a para-spinal mass and (v) maintenance of vertebral body height.


Metoclopramide in gastrointestinal radiology. Since the late 1960's, metoclopramide has been used in Europe for the treatment of gastrointestinal motility disorders. It is less well known in North America and its application in diagnostic radiology is unfamiliar to most radiologists. By comparing a control group of 103 patients with a study group of 86 patients, we found that intramuscular injection of metoclopramide is effective in reducing both gastric emptying time and small bowel transit time. Side effects were negligible in 125 patients. We also found the drug useful in gastric stasis, to differentiate between mechanical obstruction and motility disorders, in fluoroscopy of the terminal ileum, and in duodenal intubation.


Diagnostic ultrasound in a community hospital the first two years. The implications of setting up a diagnostic ultrasound service in a community hospital are described and present no insuperable difficulties. The escalation in the utilization indicates growing awareness of the value and safety of the technique among community physicians, and from the present workload (2.4 examinations per 100 persons per year) it appears that the service is no less valid and valuable than in major medical centres.


Ultrasonic evaluation of extremity soft-tissue lesions with arthrographic correlation. Diagnostic ultrasound is a fast and accurate method to detect extremity soft-tissue lesions, including popliteal (Baker) cysts, abscess, rupture of the suprapatellar bursa, peripheral artery aneurysms, and soft-tissue tumors.


How many views to a liver scan?

How many views to a liver scan? In a prospective study of 500 consecutive patients, the addition of oblique scans increased the accuracy of the colloid liver scans used to detect metastases. The oblique views changed the interpretation in 12 (2.4%) patients; objective evidence to support this change was obtained in 11 of the 12. The addition of at least a right anterior oblique scan should, on this evidence, form part of the radiocolloid evaluation of the liver for the presence of metastatic disease.


Visualization of hepatic veins on excretory urography. The hepatic veins were visualized on the early nephrogram film of an excretory urogram in a patient with tetralogy of Fallot. A nuclear angiocardiogram showed pulsatile reflux of blood from the right atrium into the hepatic veins.


Pulmonary hemorrhage in systemic lupus erythematosus. Pulmonary hemorrhage is a rare and often fatal manifestation of systemic lupus erythematosus (SLE), and enters the differential diagnosis of diffuse lung disease in patients with SLE. Pulmonary hemorrhage results from deposit of immune complexes in the alveolar basement membrane, which in turn damages capillary endothelium. Similar immune-complex deposition is present in the glomerular basement membrane of patients with systemic lupus erythematosus and pulmonary hemorrhage.


Annual oration. Diagnostic clues in the growing ends of bone. Abnormalities of the growth plate account for many of the radiologic manifestations in non-neoplastic pediatric bone disease. A good understanding of the physiology of this area is helpful in radiologic diagnosis.


In vitro and in vivo actions of zinc ion affecting cellular substances which influence host metabolic responses to inflammation. Glycogen-stimulated rabbit peritoneal exudate cells (polymorphonuclear leukocytes, PMN) produce prostaglandins (PG) and substances which induce alterations (mediators) in experimental animals characteristic of host metabolic responses to infection and other acute inflammatory stresses. The effect of Zn2+ on mediator production and PG synthesis was examined because: Zn homeostasis is perturbed during infection, Zn is known to regulate some cellular functions, and there appears to be an interrelationship between PG synthesis and mediator production. Using exudate cells, 2 mM Zn2+ caused complete inhibition of in vitro PG synthesis as assessed by conversion of [1-14C] arachidonic acid into PG. This concentration of Zn2+ also inhibited production of substances mediating plasma Zn depression, hepatic amino acid "uptake", fever, and neutrophil release from bone marrow. Conversely, Zn2+ did not inhibit in vivo metabolic responses to these mediators. Zn-pretreatment of rabbits or simultaneous injection of Zn2+ and crude PMN-derived pyrogenic activity resulted in prolongation of fever. It is suggested that this action of Zn2+ may be attributed to either stabilization of cyclic AMP through inhibition of phosphodiesterase or a Zn-mediator interaction which stabilizes crude endogenous pyrogen. The potential physiological significance of these results includes: possible potentiation of the host's defense mechanisms by Zn2+ and its utilization for prolongation of fever to determine its effect on potentially temperature-dependent host defense mechanisms.


Reciprocal regulation of glucose and glutamine utilization by cultured human diploid fibroblasts. Human diploid fibroblasts utilize both glucose and glutamine as energy sources. The utilization of glutamine by fibroblasts is regulated by glucose, and vice versa. This conclusion is supported by the following observations: (1) essentially identical growth rates were observed in Eagle's minimum essential medium (MEM)3 in which the glucose concentration was either 5.5 mM or was maintained between 25 and 40 micrometer, (2) the total glutamine utilization by fibroblasts increase at least 30% in medium with 25 micrometer to 70 micrometer glucose compared to medium with 5.5 mM glucose, while the rate of glutamine-1 or 5-14C oxidation to CO2 increased 5-fold as the glucose concentration was decreased to zero, (3) 2 mM glutamine inhibited glucose-6-14C oxidation by 88% and stimulated glucose-1-14C by 77% in log phase cells and (4) glutamine oxidation in normal medium contributed approximately 30% of the energy requirement of human diploid fibroblasts.


Affinity chromatography of lactate dehydrogenase and wheat germ lectin on new gels bearing carboxylic functions. The suitability of two new functionalized copolymer gels for use in affinity chromatography has been examined. Both gels were substituted by two ligands, one being specific for lactate dehydrogenase and the other for wheat germ lectin. The derivatives thus obtained were used successfully for the purification of two proteins with different biological activities.


Determination of 1,2-benzisoxazole-3-acetamidoxime hydrochloride (PE-257) in plasma using electron-capture gas chromatography. A gas chromatographic method has been developed that permits the accurate and specific determination of a new psychotropic agent, PF-257, in plasma. PF-257 is extracted with ethyl acetate from alkaline plasma and, after a clean-up procedure, derivatized with heptafluorobutyric anhydride to form 3-[(5-n-heptafluoropropyl-1,2, 4-oxadiazol-3-yl)methyl]-1,2-benzisoxazole (HOMB). The HOMB is assayed on a gas chromatograph equipped with an electron-capture detector. Accurate determinations of PF-257 are possible in the concentration range from 1-40 ng/ml with a relative standard deviation of 6.8%. The minimum detectable concentration in plasma is 0.1 ng/ml. Plasma levels of PF-257 in rats receiving intravenous or oral dosing (10 mg/kg) were determined.


[Gas chromatographic determination of diethylcarbamazine (Hetrazan) in serum of laboratory rodents (author's transl)]. After chloroform extraction in an alkaline solution, diethylcarbamazine (DEC) is measured in the presence of dipropylacetamid (internal standard), treated in the same manner, by a gas chromatographic procedure using a nitrogen detector in a 10% BDS column. Results can be easily reproduced for analysis of DEC in sera up to levels of 10 mg/1 and sensibility allows determination of approximately 0.5 mg/l levels on 50-microliter samples of serum. This method is suitable for pharmacokinetic studies.


Separation and determination of lipids by one-dimensional micro-thin-layer chromatography followed by densitometry. A method is described for the rapid analysis of a mixture of phospholipids and neutral lipids, which was used for the analysis of extracts obtained from a nuclear fraction isolated from rat liver. The lipids are separated by one-dimensional thin-layer chromatography on microchromatoplates (48 X 24 mm), using three solvents for development. After spraying the plates with phosphoric acid and heating, the amount of carbon from the charred compounds is measured densitometrically. Only 2-10 microgram of lipid mixture are needed for the determination of the relative amounts of the separated compounds.


Analysis of LSD in human body fluids by high-performance liquid chromatography, fluorescence spectroscopy and radioimmunoassay. A scheme of analysis is described in which the particular advantages of high-performance liquid chromatography (HPLC), fluorescence spectroscopy and radioimmunoassay (RIA) are exploited to the greatest effect. RIA affords a rapid and sensitive preliminary screening method, while the subsequent HPLC analysis using fluorimetric detection yields quantitative chromatographic evidence together with characteristic fluorescence spectra. Fractionation of samples by HPLC followed by RIA of the fractions gives further confirmation of the presence of LSD and its metabolites. The combined methodology has been applied to the analysis of LSD in body fluids for forensic and clinical purposes. Levels down to 0.5 ng of LSD per ml can be detected using the minimum of sample.


Gas-liquid chromatographic method for the assay of aminoglycoside antibiotics in serum. A gas-liquid chromatographic (GLC) method is presented for the rapid analysis of gentamicin, tobramycin, netilmicin, and amikacin from human serum. This procedure may have application to all aminoglycoside drugs. The three isomers of gentamicin are resolved as two bands, while tobramycin, netilmicin, and amikacin appear in this system as single bands. Normal serum constituents do not interfere with chromatograms. Thus far, no assay interference has been found in cases where other drugs and antibiotics were administered concurrently with aminoglycoside therapy. Dose-response data demonstrating linear recovery are included for all four aminoglycosides as well as a comparison of the GLC method with the microbiological method for the assay of gentamicin and amikacin. Quantitation is based upon the relative response of the antibiotics to a fixed amount of the internal standards, either kanamycin A or paromomycin B. These standards are clearly resolved as symmetrical peaks from the antibiotics of assay interest. Isothermal chromatographic analysis time is less than 8 min, while total assay time per single serum specimen is approximately 50 min. Preparation of serum includes: precipitation, evaporative drying of the supernatant, a two-stage derivatization (N-trimethylsilylimidazole, N-heptafluorobutyrylimidazole), and a single hexane extraction with a water wash. The methodology described may be applied to the analysis of other compounds (e.g., saccharides, amino-saccharides, amino acids, etc.) which do not rapidly partition into an organic phase.


Detection and determination of N-nitrosamino acids by thin-layer chromatography using fluorescamine. A novel procedure is described for the detection and determination of N-nitrosamino acids (NAAs) on activated silica gel thin-layer chromatographic plates. N-Nitrososarcosine, N-nitrosoproline, and N-nitroso-4-hydroxyproline could be detected as fluorophors at the 200-pmole level (20-30 ng) after being irradiated with ultraviolet light and sprayed with fluorescamine reagent. Spectrophotometric determination of the relative fluorescence of 0.4-40 nmoles of NAAs gave rise to similar calibration curves when plotted on a log-log scale. An application of this method to the detection of NAAs in uncooked bacon is described.


A high performance liquid chromatographic system for the analysis of tetracycline drug standards, analogs, degradation products and other impurities. This paper describes the high performance liquid chromatographic (HPLC) analysis of eight parent tetracycline standards: tetracycline, chlortetracycline, rolitetracycline, oxitetracycline, minocycline, doxycycline, democlocycline, methacycline; and three tetracycline epimers: epitetracycline, epianhydrotetracycline, and anhydrotetracycline. The HPLC system employs an octadecylsilane reverse phase column and an isopropanol-diethanolamine-phosphate-ammonium EDTA-water mobile phase. This system produced at least partial resolution of all eight parent compounds and many of their degradation products.


Long-term preservation of ischemic myocardium after experimental coronary artery occlusion. The results of experiments with indirect methods have suggested that various interventions reduce infarct size after coronary artery occlusion. To determine and quantify directly both the short- and long-term effects of several interventions on myocardial salvage without relying on indirect methods, the left coronary artery was occluded in 880 rats; they were then given either no treatment or one of the following interventions: (a) hyaluronidase, an enzyme that hydrolyzes interstitial glycoproteins, 1,500 National Formulary (NF) U/kg i.v. 5 min and 24 h after occlusion; (b) cobra venom factor, a protein that depletes the third component of complement, 20 U/kg i.v. 5 min after occlusion; (c) a glucocorticoid: hydrocortisone, 50 mg/kg i.v. 5 min after occlusion; or the five-fold more potent methylprednisolone (MP): (i) 50 mg/kg i.v. 5 min after occlusion or (ii) 50 mg/kg i.v. 5 min after occlusion followed by 50 mg/kg i.m. 3, 6, and 24 h after occlusion; or (d) reserpine, an agent that depletes the heart of catecholamines, 0.5 mg/kg i.m. once on each of the 3 days before occlusion. The animals were sacrificed either 2 days after occlusion, i.e., at the time of peak necrosis, or after 3 wk, i.e., after the infarct was completely healed. The amount of preserved myocardium was then assessed by two independent techniques: planimetric measurement of serial histologic sections and creatine kinase activity of the whole left ventricle. The amount of normal myocardium preserved at 21 days postocclusion was significantly increased, by 22.3+/-7.8% (P < 0.025) after the administration of hyaluronidase, by 25.3+/-5.8% (P < 0.005) after cobra venom factor, by 14.5+/-6.9% (P < 0.05) after hydrocortisone, by 20.8+/-8.2% (P < 0.025) after the single dose of MP, by 20.9+/-3.9% (P < 0.001) after the four doses of MP, and by 10.2+/-3.7% (P < 0.05) as a result of pretreatment with reserpine. The four doses of MP significantly thinned the infarct-by 25.6+/-2.9% (P < 0.001)-and although ventricular rupture did not occur, the intervention caused distension of the left ventricle as a result of stretching of the infarcted tissue during scar formation. Thus, myocardium acutely jeopardized by ischemia can be preserved on a long-term basis.


Effect of phosphate depletion on magnesium homeostasis in rats. The effects of phosphate depletion on magnesium (Mg) homeostasis were evaluated in rats fed a diet containing 0.03% phosphorus for periods up to 8 wk. Plasma phosphorus fell significantly (P < 0.01) from 10.1+/-0.27 (SE) to 5.0+/-0.54 mg/100 ml within 1 day and continued to fall gradually to a level of 1.2+/-0.21 mg/100 ml by the end of the 8th wk. A significant (P < 0.01) increment in urinary Mg excretion (UMgV) from 46+/-2.7 to 126+/-24 mueq/24 h occurred during the 1st day of phosphate depletion; UMgV reached a peak of 300+/-24 mueq/24 h by the 3rd day and remained high ranging between 150-300 mueq/24 h, thereafter. The magnitude of the magnesuria was related to the degree of hypophosphatemia and was not affected by lowering the calcium intake and reducing the hypercalciuria. The concentration of plasma Mg fell significantly (P < 0.01) from 1.2+/-0.02 to 0.79+/-0.10 meq/liter by the 1st day of the study and remained low throughout.Mg balance became negative during the 1st day of phosphate depletion and remained so during the entire study. This occurred despite a significant increment in the fraction of ingested Mg absorbed which became evident by the 3rd wk of phosphate depletion. Mg content of muscle, kidney, and liver were not affected but bone Mg was reduced significantly. The change in bone Mg was not due to an overall reduction in bone mineral content because bone calcium content was not affected. Supplementation of large amounts of Mg (800-1,000 mueq/day) in the drinking water produced a normalization of serum Mg but did not bring about restoration of bone Mg despite a positive Mg balance. The disturbances in Mg metabolism were independent of the age or weight of the animals. Our results indicate that phosphate depletion is associated with (a) magnesuria due to a decrease in the net renal tubular reabsorption of Mg with the main source of the urinary losses being bone Mg; (b) hypomagnesemia secondary to the renal leak of Mg; (c) negative Mg balance; and (d) increase in the intestinal fractional absorption of Mg. The latter was not adequate to compensate for the urinary losses of Mg.


Role of hemodynamic changes in the increased cation excretion after acute unilateral nephrectomy in the anesthetized dog. Studies were carried out in anesthetized dogs to characterize the increase in cation excretion which occurs after acute unilateral nephrectomy (AUN). 60 min after AUN, cation excretion had increased from 31.5+/-2.7 to 66.3+/-12.0 mueq/min (P < 0.005) and fractional cation excretion had increased from 0.56+/-0.05 to 1.03+/-0.14% (P < 0.005), as the glomerular filtration rate was unchanged and renal blood flow fell. The increased cation excretion was accompanied by an increase in fractional phosphate excretion, no change in chloride excretion, and a fall in renin secretion. These alterations in renal function were associated with marked changes in systemic hemodynamics: cardiac output fell from 2.52+/-0.24 to 1.85+/-0.16 liters/min (P < 0.001), as diastolic pressure rose without an overall increase in mean arterial pressure, and heart rate fell. To assess the importance of these hemodynamic changes in the renal response, AUN in a separate group of dogs was accompanied by the simultaneous opening of a surgically created femoral artery-to-vein fistula at flow matching the blood flow to the removed kidney. When this was done, no alterations in systemic or renal hemodynamics were observed, and cation excretion did not differ from control. Subsequent closure of the fistula then caused a fall in cardiac output from 2.15+/-0.25 to 1.77+/-0.20 liters/min (P < 0.05), and an increase in cation excretion from 34.6+/-9.5 to 52.3+/-13.7 mueq/min (P < 0.01), thus mimicking the findings with AUN alone.These results demonstrate that AUN causes hemodynamic changes resembling those seen on closure of a chronic arteriovenous fistula. Prevention of these hemodynamic changes after AUN also prevents the functional adjustment of the remaining kidney, suggesting that they may be important in initiating the renal response. The increased electrolyte excretion after AUN may occur through mechanisms similar to that seen on closure of an arteriovenous fistula.


Increased fluidity of human platelet membranes during complement-mediated immune platelet injury. Complement appears to be involved in the destruction of platelets in certain clinical disorders, such as quinidine purpura and post-transfusion purpura. In both disorders, the classical complement sequence is activated by antigen-antibody complexes. It has been suggested that the terminal components of the complement sequence insert into the hydrophobic core of cell surface membranes and that this process leads to cell lysis. Fluidity is a fundamental property of lipids within the membrane's hydrophobic core. To examine the interaction of complement with membranes, we investigated the effect of complement activation on the fluidity of human platelet membranes. Complement was fixed to platelets using a post-transfusion purpura antibody, and membrane lipid fluidity was assessed in terms of fluorescence anisotropy using two fluorescent probes, 1,6-diphenyl-1,3,5-hexatriene and 9-(12-anthroyl) stearic acid. Microviscosity, expressed in poise, was derived from the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene.Post-transfusion purpura antibody plus complement made platelet membranes more fluid as evidenced by a 21% decrease in anisotropy and a 35% decrease in microviscosity of platelets at 37 degrees C, and this was associated with platelet lysis ((51)Cr release). Complement damage to platelets was accompanied by a 10-15% increase in DeltaE, the fusion activation energy for microviscosity, indicating that complement not only decreased membrane microviscosity but also made membrane lipids less ordered. These changes were consistent and rapid, with platelet lysis and the reduction in microviscosity being half-maximal by 6 min. They were prevented by inactivation of complement with heat or with EDTA, and they were not observed when C5-deficient plasma was used as the complement source. Qualitatively similar changes in platelet membrane fluidity were observed when complement was fixed to platelets by a quinidine-dependent anti-platelet antibody rather than by post-transfusion purpura antibody. Post-transfusion purpura antibody plus complement also decreased the microviscosity of isolated platelet membranes. Moreover, the lipids extracted from platelets lysed by complement had a 22% decrease in microviscosity (P < 0.01), with no associated changes in the amount of cholesterol relative to phospholipid or in the amounts of the various phospholipids. These studies demonstrate that lipids within the hydrophobic core of platelet membranes damaged by complement become more fluid, and this is associated with platelet lysis. These findings are consistent with the concept that the insertion of the terminal complement components into the platelet membrane bilayer perturbs lipid-lipid interactions within the membrane's hydrophobic core.


The key role of peptidoglycan in the opsonization of Staphylococcus aureus. In an effort to determine the staphylococcal cell surface component(s) of importance in opsonization, cell walls (peptidoglycan and teichoic acid) and peptidoglycan were isolated from Staphylococcus aureus strain H grown in [3H]glycine-containing broth. After incubation of the cell walls and peptidoglycan with various opsonic sources, uptake by human polymorphonuclear leukocytes was measured. The opsonic requirements for phagocytosis of cell walls and peptidoglycan were found to be similar to those of intact bacteria. Removal of teichoic acid from the cell wall did not affect opsonization. Likewise, a teichoic acid-deficient mutant strain of S. aureus H was opsonized in a manner similar to that of the parent strain. Immunoglobulin G functioned as the major heat-stable opsonic factor and both the classical and alternative pathways participated in opsonization. Kinetic studies revealed that opsonization of peptidoglycan, as well as C3-C9 consumption by peptidoglycan, proceeded at a slower rate via the alternative pathway (C2-deficient serum) than when the classical pathway was present (normal serum). The ability of peptidoglycan to activate C3-C9 was significantly reduced when normal and C2-deficient sera were preabsorbed with peptidoglycan at 2 degrees C suggesting that antibodies to peptidoglycan may be involved in activation of both the classical and alternative complement pathways. Thus, peptidoglycan appears to be the key cell wall component involved in staphylococcal opsonization, and it is suggested that host response to peptidoglycan, a major cell wall component of most gram-positive bacteria, may be related to the development of "natural immunity" to this group of microorganisms.


The effect of altered sodium balance upon renal vascular reactivity to angiotensin II and norepinephrine in the dog. Mechanism of variation in angiotensin responses. The mechanism whereby the vasoconstrictor response to angiotensin II (AII) is influenced by sodium balance or disease is unclear. To explore this question, the renal vascular responses (RVR) to intrarenal injections of subpressor doses of AII and norepinephrine were studied in dogs with an electromagnetic flowmeter. Acute and chronic sodium depletion increased plasma renin activity (PRA) and blunted the RVR to AII, while acute sodium repletion and chronic sodium excess plus desoxycorticosterone acetate decreased PRA and enhanced the RVR to AII. The magnitude of the RVR to AII was inversely related to PRA. The RVR to norepinephrine was unaffected by sodium balance and was not related to PRA. Inhibition of the conversion of angiotensin I to AII by SQ 20,881 during sodium depletion lowered mean arterial blood pressure (MABP), increased renal blood flow (RBF), and enhanced the RVR to AII but not to norepinephrine. Administration of bradykinin to chronically sodium-depleted dogs also lowered the MABP and increased RBF but had no effect on the RVR to AII. SQ 20,881 had no effect on MABP, RBF, or the RVR to AII in the dogs with chronic sodium excess and desoxycorticosterone acetate. Administration of indomethacin to chronically sodium-depleted dogs lowered RBF but did not influence the RVR to AII. The results indicate that the RVR to AII is selectively influenced by sodium balance and that the magnitude of the response is inversely related to the availability of endogenous AII. The data did not suggest that the variations in the RVR to AII were because of direct effects of sodium on vascular contraction, changes in the number of vascular AII receptors, or the renal prostaglandins. The results are consistent with the hypothesis that the vasoconstrictor effect of AII in the renal vasculature is primarily dependent upon the degree to which the AII vascular receptors are occupied by endogenous hormone.


Alcohol-induced ovarian failure in the rat. The effect of ethanol feeding on ovarian function and structure in female rats was studied in alcohol-fed animals, isocalorically fed controls, and two ad libitum-fed control groups. Ovarian weight was reduced by 60% in alcohol-fed animals compared with the control groups. Gross disruption of ovarian architecture was noted, characterized by the absence of any corpus hemorrhagicum and corpus albicans. Moreover, plasma levels of estradiol were significantly reduced in the alcohol-fed animals (P < 0.01) compared with the levels found in isocaloric controls. Plasma levels of estrone and corticosterone were increased in alcoholfed and isocaloric control animals relative to those of ad libitum-fed animals suggesting a primarily adrenal, rather than ovarian, origin for these two steroids. Despite the increase in estrone, the secondary sex organs (uterus and fallopian tubes) reflected marked estrogen deprivation presumably as a result of estradiol insufficiency. Progesterone levels in the alcohol-fed animals were significantly less than levels in the isocaloric and intact ad libitum-fed controls but were not significantly different compared to oophorectomized animals. Plasma follicle-stimulating hormone levels were similar in alcohol-fed, isocaloric controls, and ad libitum-intact controls. They were, however, one-third the level of oophorectomized controls. Both alcohol-fed and isocaloric controls had increased levels of plasma luteinizing hormone, although levels were below those seen in oophorectomized controls (P < 0.01). The results establish that ingestion of a diet containing 5% ethanol for periods as short as 6 wk produces functional and histologic ovarian failure in the female rat.


Studies on the mechanism of reduced urinary osmolality after exposure of renal papilla. Studies were performed in Munich-Wistar rats to determine whether changes in papillary plasma flow might be responsible for the concentrating defect which occurs after exposure of the extrarenal papilla. Papillary plasma flow was measured by (125)I-albumin accumulation. Initial studies in hydropenic animals revealed that papillary plasma flow was 40% higher in the kidney with the exposured papilla, 41 vs. 29 ml/min per 100 g of papilla (P < 0.001). This increase in papillary plasma flow was detectable 15 or 45 min after removing the ureter. Because it was unclear whether the rise in papillary plasma flow was a cause or the result of the fall in urine osmolality, similar studies were performed in animals undergoing a water diuresis. In this setting, papillary plasma flow still increased on the exposed side compared to the control side, 81 vs. 60 ml/min per 100 g, despite similarly low urine osmolalities of 155 and 174 mosmol/kg, respectively. This finding is compatible with the possibility that papillary exposure per se causes an increase in papillary plasma flow and that this hemodynamic alteration may lead to a reduction in urinary osmolality secondary to washout of the medullary interstitium. A final group of hydropenic rats was given either indomethacin or meclofenamate before removing the ureter. In these studies, there was no difference in either the papillary plasma flow or the urine osmolality between control and exposed kidneys. It is therefore suggested that opening the ureter induces an increase in papillary plasma flow by some mechanism which may involve an alteration in prostaglandin synthesis.


Ricinoleic acid effect on the electrical activity of the small intestine in rabbits. Using myoelectric recording techniques, we examined the myoelectric effects of castor oil; ricinoleic acid (cis isomer), the active ingredient of castor oil; and ricinelaidic acid (trans isomer) in the small intestine of New Zealand white rabbits. Ricinoleic acid, 2 microgram/kg per min (6mM), was perfused into a distal 12-cm ileal loop. An abnormal myoelectric pattern developed that was similar to the alteration in the electrical activity that has previously been reported for cholera enterotoxin. Castor oil, 0.85 ml/kg, had a similar effect. Ricinelaidic acid, 2 microgram/kg per min, induced no activity. A second preparation consisted of an intraluminal perfusion of ricinoleic acid, 2 microgram/kg per min, into the first section of the duodenum. The abnormal myoelectric pattern was observed in the jejunum and the ileum but not the duodenum. The mean onset time for the development of this altered myoelectric state for all experiments was 3.5 h. These studies suggest that an active motility component in addition to the secretory state exists throughout the small intestine that is exposed to castor oil or ricinoleic acid.


Homocystinuria. Evidence for three distinct classes of cystathionine beta-synthase mutants in cultured fibroblasts. We have compared in vivo pyridoxine responsiveness with in vitro cystathionine beta-synthase activity in extracts of confluent fibroblasts from 14 synthase-deficient patients. Enzyme activity was measured with and without addition of its cofactor, pyridoxal-5'-phosphate, using a radioisotopic assay which detects as little as 0.25% of control activity. Six of seven lines from responsive patients had measurable activity without the added cofactor (0.6-15% of mean control). Two of these lines showed a five- and sevenfold stimulation of cystathionine beta-synthase activity with added pyridoxal-5'-phosphate; in the other four, the cofactor addition increased activity only modestly, as in controls. Two of seven lines from nonresponsive patients had measurable activity (each 3% of mean control) which increased two- and fivefold with the added cofactor. Cystathionine beta-synthase activity was undetectable in one line from a responsive patient and in five lines from nonresponsive ones. To characterize control and mutant synthase further, dissociation constants for pyridoxal-5'-phosphate were estimated and thermostability (54 degrees C) was studied in two control and five mutant lines. In one mutant, both parameters were normal; in the others, the affinity for the cofactor was reduced 3-to 11-fold and thermostability was much impaired. We conclude that at least three general classes of cystathionine beta-synthase mutants exist: those with no residual activity; those with reduced activity and normal affinity for pyridoxal-5' phosphate; and those with reduced activity and a reduced affinity for the cofactor. Pyridoxine responsiveness in vivo cannot be correlated simply with the presence or absence of residual synthase activity in vitro or with stimulation of in vitro enzyme activity by cofactor.


Reduction by cobra venom factor of myocardial necrosis after coronary artery occlusion. Components of the complement system are known to play an important role in the cytolytic process and in chemotaxis of leukocytes. Cobra venom factor specifically cleaves C3 activity via activation of the alternative (properdin) complement pathway. It does not act directly on C3. If C3 is involved in tissue necrosis after ischemic injury, cobra venom factor might reduce tissue damage after acute coronary occlusion. Accordingly, in 14 control dogs occlusion of the left anterior descending artery was carried out for 24 h. Epicardial electrograms were recorded 15 min after occlusion, and 24 h later transmural specimens for creatine phosphokinase activity (CPK) and for histological analysis were obtained from the same sites. In another 14 experimental dogs, 20 U/kg cobra venom factor was given intravenously 30 min after occlusion. Serum complement levels fell within 2-4 h to <20% of normal. In the control dogs, the relationship between ST-segment elevation and CPK activity 24 h later was: log CPK = -0.06 ST + 1.48 (n = 111 specimens, 14 dogs, r = 0.77). In the experimental dogs, log CPK = -0.024 ST + 1.46 (n = 111 specimens, 14 dogs, r = 0.60), showing significantly different slopes (P < 0.001), i.e., less CPK depression for any level of ST-segment elevation. Histologically, 69 of 71 sites (97%) with ST-segment elevation exceeding 2 mV in the control dogs showed signs of necrosis 24 h later, whereas in the experimental group only 43 of 79 sites (54%) with abnormal ST-segment elevations showed signs of necrosis (P < 0.0005). At the same time, it was shown that the administration of cobra venom factor did not alter cardiac performance, collateral blood flow to the ischemic myocardium or the clotting system, but infiltration of polymorphonuclear leukocytes into the myocardium was decreased. It is concluded that cobra venom factor, by reducing the amount of C3 and C5 substrate available for chemotactic factor generation, or other as yet undefined mechanisms, protects the ischemic myocardium from undergoing necrosis, as judged by histology and local CPK activity. Hence, a new approach to limiting the extent of myocardial infarcts after experimental coronary occlusion, based upon inhibition of complement-dependent inflammatory processes, is demonstrated.


Comparative adherence of granulocytes to endothelial monolayers and nylon fiber. Adherence of granulocytes to tissue culture monolayers of endothelium averaged 26.2 +/- 1.3% SEM, which was similar to their adherence on 50-mg nylon fiber columns (27.7 +/- 3.6%). In contrast, adherence to epithelial cells, fibroblasts, kidney cells, and plastic Petri dishes without monolayers was only 12.4, 9.9, 11.1, and 4.3%, respectively. Cyclic nucleotides and adherence-modifying plasma factors induced changes of adherence to endothelium similar to those in nylon fiber columns. Adherence of granulocytes in whole blood was the same as for purified granulocytes in Hank's balanced salt solution. Exposure of endothelial monolayers to 0.18% trypsin for 10 min reduced subsequent granulocyte adherence to 25.2% of control values. Incubation of trypsin-treated monolayers with nutrient medium for 4 h did not improve adherence, but values returned to normal or above by 24 h, with or without serum proteins present in the nutrient medium. The similarity of granulocyte adherence to nylon fiber and to endothelial monolayers in vitro suggests that results with the nylon fiber assay reflect in vivo granulocyte-endothelium interaction. Furthermore, the endothelial monolayer offers a new model for studying this cell-cell relationship in vitro.


Depression of ventilation by dopamine in man. Evidence for an effect on the chemoreceptor reflex. Dopamine is present in the carotid body and has been postulated to be an inhibitory neurotransmitter. The purpose of this study was to determine the effects of dopamine on ventilation in man and to examine its mechanism of action. Dopamine (0.5-10 mug/kg per min) was infused in eight normal men at different levels of arterial chemoreceptor activity, produced by varying the inspired Po(2). During normoxia dopamine produced a small decrease in minute ventilation (Ve) and an increase in arterial Pco(2). When arterial chemoreceptors were stimulated by hypoxia, infusion of dopamine produced a marked initial depression of Ve followed by a sustained although less pronounced decrease in Ve. An increase in Pa(co) (2) and a decrease in Pao(2) were also observed. When arterial chemoreceptor activity was suppressed by hyperoxia, infusion of dopamine did not affect ventilation. Subjects also breathed a hypercarbic, hyperoxic gas mixture. The hypercarbia produces hyperventilation by stimulating central chemoreceptors, whereas the hyperoxia suppresses peripheral chemoreceptors. Dopamine did not alter ventilation while the subjects were breathing this gas mixture. These studies suggest that dopamine suppresses ventilation in man through an action on the arterial chemoreceptor reflex. These findings support the hypothesis that dopamine is an inhibitory neurotransmitter in the carotid body, and that release of dopamine may modulate the sensitivity of peripheral arterial chemoreceptors.


Ventromedial hypothalamic lesions and the mobilization of fatty acids. We have explored the effects of ventromedial hypothalamic lesions on the mobilization of free fatty acids in rats exposed to several stresses. The rise in free fatty acids and glycerol in response to norepinephrine had the same time-course and dose-response characteristics in the sham-operated and lesioned animals, indicating comparable degrees of peripheral responsiveness to this hormone. Forced swimming significantly lowered insulin and increased glycerol and free fatty acids more in control than in ventromedial hypothalamic-lesioned rats. During fasting, the rise in glycerol and free fatty acids was smaller in the lesioned rats, but the fall in insulin was greater. Exposure to cold raised fatty acids and glycerol more in the control than in the sham-operated animals, but had no significant effect on plasma insulin or glucose concentration. Injection of 2-deoxyglucose was done on lesioned or control rats with intact or removed adrenal medullas. The rise in free fatty acids and glycerol was less in the lesioned rats than in the controls, and was not affected by adrenodemedullation. The rise in glucose, however, was completely blocked in the adrenodemedullated rats. Changes in insulin were small and not statistically significant. The reduced mobilization of fatty acids from adipose tissue depots after ventromedial hypothalamic injury is consistent with the hypothesis that the ventromedial hypothalamic region serves to modulate activation of the sympathetic nervous system.


Selective induction of an immune response in human external secretions by ingestion of bacterial antigen. Ingestion of capsules which contained killed Streptococcus mutans by four healthy human subjects led to the appearance of specific antibodies in external secretions. Salivary and lacrymal antibodies were detected within 1 wk of ingestion and continued to increase throughout a 14-day immunization period, with a gradual decline during the 2 ensuing months. A second period of immunization resulted in a pronounced increase of specific antibody levels which occurred earlier than in the primary immunization period and reached peak levels by day 10. No change was detected in serum antibody levels throughout either immunization period. The antibody activity in all secretions was associated with the immunoglobulin A class, as determined by immunochemical analyses. These data indicate that ingestion of bacterial antigens selectively stimulates the immune response in secretions.


The role of renal nerves and prostaglandins in control of renal hemodynamics and plasma renin activity during hypotensive hemorrhage in the dog. The effects of hypotensive hemorrhage (HH) on renal hemodynamics and plasma renin activity (PRA) during prostaglandin (PG) synthesis inhibition were examined in three groups of dogs. In each group of animals arterial blood pressure was lowered by a 30% decrement. In the first group of eight control animals, HH was not associated with a significant change in glomerular filtration rate (GFR, 42-36 ml/min, NS); renal blood flow (RBF) declined significantly, from 234 to 171 ml/min, P < 0.05. In the second group of eight animals, pretreated with RO 20-5720 (RO, 2 mg/kg), a competitive inhibitor of PG synthesis, HH was associated with a significant fall in GFR (43-17 ml/min, P < 0.001) and RBF (195-89 ml/min, P < 0.001). In the third group of eight animals, pretreatment with indomethacin (IN, 10 mg/kg), a chemically dissimilar PG inhibitor, HH was also associated with a significant fall in GFR (38-8 ml/min, P < 0.001) and RBF (150-30 ml/min, P < 0.001). Renal denervation attenuated this renal ischemic effect of HH in the presence of PG inhibition. In the RO group, GFR (34 vs. 17 ml/min, P < 0.005) and RBF (145 vs. 89 ml/min, P < 0.025) were significantly greater in denervated vs. innervated kidneys during HH. Similarly, in animals treated with IN, a significantly higher GFR (28 vs. 8 ml/min, P < 0.005) and RBF (101 vs. 30 ml/min, P < 0.005) occurred in denervated as compared to innervated kidneys during HH. With HH, the increase in PRA in the control group (3.34-11.68 ng/ml per h, P < 0.005) was no different than that observed in the RO group (4.96-18.9 ng/ml per h, P < 0.001) or IN group (4.71-17.8 ng/ml per h, P < 0.001). In summary, the present results indicate that renal PG significantly attenuate the effect of HH to decrease GFR and RBF. Furthermore, renal denervation exerts a protective effect against the enhanced renal ischemic effects which occur in the presence of PG inhibition during HH. Finally, PG inhibition does not alter the effect of HH to cause an increase in PRA.


Dominant inheritance of large molecular weight immunoreactive glucagon. Plasma from some individuals contains substances which are reactive with glucagon antiserum, are larger than 3,500-dalton glucagon, and have been proposed as possible precursors of glucagon. We have evaluated three generations of a kindred in which 9 of 15 members evaluated had elevated plasma levels of large molecular weight immunoreactive glucagon (L-IRG) with an average concentration of 822 pg/ml. The distribution of individuals with elevated L-IRG levels in this pedigree is consistent with autosomal dominant inheritance. Gel filtration of plasma revealed that all affected family members had excessive amounts of two L-IRG peaks, one with a molecular weight of approximately 9,000 daltons and another in the 10,000 to 20,000-dalton range. Oral glucose tolerance tests were nondiabetic and elicited a fall in L-IRG levels, whereas L-IRG concentrations rose dramatically during the infusion of arginine. These L-IRG species may be precursors of 3,500-DALTON GLUCAGON AND MAY BE ELEVATED in this kindred because of an inherited defect in either their synthesis or degradation.


Production of a low molecular weight eosinophil polymorphonuclear leukocyte chemotactic factor by anaplastic squamous cell carcinomas of human lung. A peptide of approximately 300-400 daltons exhibiting in vitro chemotactic activity for human polymorphonuclear (PMN) leukocytes, with a preference for the eosinophil series, was isolated from extracts of anaplastic lung carcinomas of the large squamous cell type obtained from three patients with marked peripheral blood hypereosinophilia and eosinophilic infiltration of the tumors and surrounding normal pulmonary tissues. This chemotactic factor was termed ECF-LSC (eosinophil chemotactic factor of lung squamous cell carcinoma). ECF-LSC appeared in the urine of two of the patients in increasing quantities late in the course of their disease and was also elaborated by long-term cultures of dispersed tumor cells from the same two patients. Three anaplastic large cell bronchogenic carcinomas which were not associated with tumor tissue or peripheral blood eosinophilia, a bronchogenic adenocarcinoma from a patient with only peripheral eosinophilia, and a renal cell carcinoma metastatic to the lungs and associated with transient pleural tissue and fluid eosinophilia were all devoid of ECF-LSC. ECF-LSC from tumor tissue extracts, urine, and tumor cell culture medium was comparable to the mast cell-associated tetrapeptides of the eosinophil chemotactic factor of anaphylaxis (ECF-A) in size, but eluted from Dowex-1 at pH 5.0-3.5 in contrast to the more acidic ECF-A tetrapeptides which eluted at pH 3.2-2.2 ECF-LSC, like the tetrapeptides of ECF-A, had a secondary chemotactic activity for neutrophil PMN leukocytes, but not mononuclear leukocytes, and deactivated both eosinophil and neutrophil PMN leukocytes so that they would not respond to a subsequent in vitro chemotactic stimulus. Eosinophils from the two patients with urinary excretion of ECF-LSC and the highest concentrations in tumor extracts were hyporesponsive in vitro to homologous and heterologous chemotactic stimuli, suggesting that ECF-LSC had deactivated the eosinophils in vivo.


Vagal, cholinergic regulation of pancreatic polypeptide secretion. THE EFFECT OF EFFERENT, PARASYMPATHETIC STIMULATION UPON PANCREATIC POLYPEPTIDE (PP) SECRETION WAS STUDIED IN THREE WAYS: (a) Plasma PP concentrations increased in response to insulin-induced hypoglycemia in both normal subjects, from 11 pM (9.5-12.5) to 136 pM (118-147), n = 8 (median and interquartile range) and in duodenal ulcer patients, from 33 pM (21-52) to 213 pM (157-233), n = 7. The PP response to hypoglycemia was diminished by atropine in normal subjects (P < 0.005) and completely abolished by vagotomy in the duodenal ulcer patients. (b) Electrical stimulation, 8 Hz, of the vagal nerves in anesthetized pigs induced an increase in portal PP concentrations within 30 s from 32 pM (28-39) to 285 pM (248-294), n = 12. Minimal stimulatory frequency was 0.5 Hz and maximal stimulatory frequency 8-12 Hz. Atropine inhibited the PP response to electrical stimulation. Median inhibition with 0.5 mg of atropine/kg body wt was 74%, range 31-90%, n = 6. The response was eliminated by hexamethonium. Adrenergic alpha and beta blockade did not influence the release of PP in response to vagal stimulation. (c) Acetylcholine stimulated, in a dose-dependent manner, the secretion of PP from the isolated perfused porcine pancreas, half-maximal effective dose being 0.19 muM; maximal PP output in response to 5 min stimulation was 228 pmol, range 140-342 pmol, n = 5. Atropine completely abolished this response.The results of the present study together with the previously demonstrated poor PP response to food in vagotomized patients, indicate that vagal, cholinergic stimulation is a major regulator of PP secretion.


Renal tubular transport of organic acids. Studies with oxalate and para-aminohippurate in the rat. The renal handling of oxalate was examined by free-flow micropuncture, intratubular microinjection, and droplet precession techniques in the rat. After the sustained i.v. infusion of [(14)C]oxalate, the fractional delivery of oxalate from the early portions of the proximal tubule was 120.1+/-4.4%, indicating net secretion. Fractional delivery rates from the late proximal tubule (124.6+/-6.1), distal tubule (120.9+/-2.9), and final urine (126.2+/-2.9%) were not different from that of the early proximal tubule. Direct intratubular microinjections of oxalate into the early proximal tubule and late proximal tubule yielded urinary recovery rates of 85+/-3% and 101+/-2%, respectively, suggesting that oxalate absorption does occur in the mid-portions of the proximal tubule. Droplet precession studies confirmed a secretory flux for oxalate. In contrast to oxalate, para-aminohippurate (PAH), the more traditional marker for organic acid transport, was secreted in the late portions of the proximal tubule and in large measure at a site between the late proximal and distal tubules, presumably the pars recta. Probenecid inhibited PAH secretion but was without effect on net oxalate transport, oxalate absorption, or oxalate secretion. These studies demonstrate that net oxalate secretion occurs in the early portions of the proximal convoluted tubule, undergoes bidirectional transport of approximately equal magnitude in later segments of the proximal tubule, and probably is not transported in more distal nephron sites. The secretory mechanism for oxalate differs from that of PAH in that it is located in a different segment of the nephron and is not inhibited by probenecid. These differences suggest that the early portions of the proximal tubule are important in the renal metabolism of some organic acids.