Transcription of DNAs of known sequence after injection into eggs and oocytes of Xenopus laevis.

1. When the synthetic polynucleotide, poly[d(A-T) - d(A-T)] is injected into the eggs and oocytes of Xenopus laevis, a stimulation of RNA synthesis results. Analysis of extracted RNA by high-voltage electrophoresis, shows that this stimulation of RNA synthesis is due to the production of poly[r(A-U)] transcripts. The rate of poly[r(A-U)] synthesis is calculated to be at least ten-fold greater in eggs than in oocytes. The amount of poly[r(A-U)] detectable in injected eggs has reached a maximum by 1.5 h after injection; in oocytes, however, poly[r(A-U)] continues to accumulate between the third and 16th hour after injection. The transcripts range in length from less than 80 nucleotides up to over 2000 nucleotides long. The co-injection of poly[d(A-T)] - d(A-T)] and alpha-amanitin into oocytes, has shown that the synthesis of poly[r(A-U)] is approximately 90% inhibited at a concentration of alpha-amanitin which has no effect on the capacity of the oocyte to synthesize ribosomal and 4-S RNA; thus the nucleoplasmic RNA polymerases IIa and/or IIb, are implicated as playing a major role in poly[r(A-U)] synthesis in oocytes. 2. When poly(dG) - poly(d-C), poly(dA), poly(dA) - poly(dT) and poly[d(I-C) - d(I-C)] are individually injected into eggs only poly[d(I-C) - d(I-C)] is transcribed as efficiently as poly[d(A-T) - d(A-T)]. 3. When calf thymus native or denatured DNA, polyoma, T2, T4 and phiX DNAs are individually injected into eggs only the injection of calf thymus native DNA causes a detectable stimulation of RNA synthesis. 4. The activities of crude preparations of egg and oocyte RNA polymerases are tested in vitro with different DNA templates. In contrast to the situation in vivo, it is found that poly[d(A-T1 - d(A-T)] is as efficiently transcribed in vitro by oocyte polymerase as by egg polymerase. Additionally poly[d(A-T) - d(A-T)] is transcribed ten-fold more efficiently in vitro than calf thymus native DNA. When poly(dA) - poly(dT), poly(dA), phiX, T2, and calf thymus denatured DNA are tested in vitro, only calf thymus denatured DNA is transcribed to a significant extent. The above results are discussed in relation to the known synthetic activities of Xenopus eggs and oocytes.


Hydroxylation of testosterone at carbons 1, 2, 6, 7, 15 and 16 by the hepatic microsomal fraction from adult female C57BL/6J mice.

The metabolism of a mixture of [4-14C]- and [7 beta-2H]testosterone by the hepatic microsomal fraction from adult femal C57BL/6J mice has been investigated. The following metabolites were identified by their mass spectra and by their retention times on gas chromatography on one or two phases: 1epsilon-, 2beta-, 6alpha-, 6beta-, 7alpha-, 15alpha-, 15beta-, 16alpha- and 16beta-hydroxytestosterone; 6alpha-, 6beta- and 7alpha-hydroxy-4-androstene-3,17-dione; and 4-androstene-3,17-dione. A compound tentatively identified as 6- or 7-oxotestosterone was also isolated. 17beta-Hydroxy-4,6-androstadien-3-one, 17beta-hydroxy-1,4-androstadien-3-one and 4,6-androstadiene-3,17-dione were identified but are considered to arise non-enzymatically from 7alpha-hydroxytestosterone, 1epsilon-hydroxytestosterone and 7alpha-hydroxy-4-androstene-3,17-dione, respectively.


Studies on the role and mode of operation of the very-lysine-rich histone H1 (F1) in eukaryote chromatin. Histone H1 in chromatin and in H1 - DNA complexes.

The nuclear magnetic resonance (NMR) spectrum of chromatin at ionic strengths below about 0.5 M may be attributed solely to its histone H1 component. The effect of various ions and urea on the complex has been investigated using NMR and confirm that the contraction of the complex on increase of ionic strength is largely due to electrostatic interactions. A detailed study of the H1 - DNA complex has also been undertaken. The behaviour of H1 in the two cases is virtually identical, implying that in chromatin the H1 is complexed with the DNA rather than with the other histones. Microcalorimetric measurements reveal that the binding of H1 to DNA is athermic or involves a heat of reaction which is very small indeed.


Human luteinizing hormone. Isolation and characterization of the native hormone and its alpha and beta subunits.

A new procedure is described for the isolation of the alpha and beta chains of the hormone. In this method, thenative hormone is incubated in acidic urea and the chains are then separated by ion-exchange chromatography. The amino-terminal residue of the alpha subunit is valine. The carboxy-terminal end of the alpha subunit is of variable length. No amino-terminal residue was detected for the beta chain; glycine was found at its carboxy-terminal end by the selective titration method. The amino acid and carbohydrate compositions of the hormone and both subunits are presented. The beta chain contains sialic acid and is devoid of galactosamine in contrast to the beta subunits of other species. Contamination of our human lutenizing hormone preparation by other pituitary glycoprotein hormones such as thyroid-stimulating hormone and follicle-stimulating hormone amounted to 0.5 and 0.25 percent by weight respectively. Cross-contamination of the initial alpha and beta subunit preparations was measured by specific radioimmunoassays and amounted to 4.1 and 2 percent by weight respecitively. Further extensive purification of these subunit preparations was then performed by means of affinity chromatography using immunosorbants. The final preparations exhibited a residual cross-contamination amounting to 0.2 and 0.02 percent by weight for the alpha and beta subunits respectively.


Kinetics of interaction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and beta-lactam antibiotics. A choice of models.

The simplest model for the interaction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and beta-lactam antibiotics involves the three following steps: (a) the formation of a reversible equimolar enzyme - antibiotic complex; (b) the irreversible transformation of this complex into a modified enzyme - antibiotic complex; and (c) the breakdown of this latter complex and the concomitant release of a regenerated enzyme and a modified antibiotic molecule. The dissociation constant for step 1 and the rate constants for steps 2 and 3 were measured with various beta-lactam antibiotics. With antibiotic such as benzylpenicillin, which behaves as a good 'substrate', steps 1 and 2 occur at enzymic velocities, whereas step 3 occurs at a very low velocity and hence is responsible for the low efficiency of the overall process.


DNA replication in methotrexate-treated human lymphoblasts.

The rate of DNA synthesis in cultures of human lymphoblasts decreased more than 80% within 30 min after the cells were exposed to methotrexate, a potent inhibitor of dihydrofolate reductase. Despite this rapid initial inhibition, DNA continued to be synthesized for at least an additional 6 h. The mode of this subsequent replication appeared to be semiconservative, as indicated by the buoyant density of 5-bromodeoxyuridine-substituted DNA in alkaline CsCl gradients. The growth rates of DNA chains in cells exposed to methotrexate were determined by sedimentation rate analysis in alkaline sucrose gradients. DNA synthesized during 2-min or 10-min pulses with labeled deoxycitidine in the presence of methotrexate had about the same sedimentation coefficient, 35 S, as controls. When methotrexate-treated cultures were pulse-labeled for 10 min and then chased for various times, DNA fragments of about 80 S accumulated. DNA synthesized in the presence of methotrexate was stable and elongated to bulk-size DNA after methotrexate inhibition of growth was removed by addition of thymidine and deoxycytidine. The data suggest that methotrexate reduces the rate of DNA replication by inhibiting chain initiation independently of chain elongation.


Immunologically active and structurally similar fragments of protein A from Staphylococcus aureus.

To study the active site(s) in protein A, partial tryptic digestions of the protein and of intact Staphylococcus aureus were performed. Fragments which bind to the Fc-part of human IgG were isolated by affinity chromatography on IgG-Sepharose 4B and purified by ion-exchange chromatography on phosphocellulose. From a partial tryptic digest of pure protein A at 30 degrees C, pH 8.2 for 30 min we have isolated and characterized six active fragments with molecular weights ranging from 6000 to 8000. Two active fragments, obtained in high yields by digestion at pH 7.2 of intact protein-A-containing bacteria, were shown to be similar to two of the six characterized fragments from the digest of pure protein A. All fragments appeared to have similar amino acid sequences, judged by peptide mapping, specific staining and amino acid analysis; some are very possibly overlapping peptides. Each fragment probably contains only one active site region since all are monovalent in the Fc-reaction when studied with a hemagglutination technique. The maximal molar yield of active fragments obtained from the digestion of pure protein A accounts for about 210% of the amount of protein A used. Thus protein A, suggested to consist of repeating units, should exhibit at least three similar if not identical active regions.


Similarities and differences of five peroxidases from turnip and horseradish. Peptide mapping studies on glycoproteins.

Four isoperoxidases of turnip root and isoperoxidase C of horseradish root were digested with trypsin, and their peptide maps, prepared by high-voltage paper electrophoresis, were compared. All five tryptic digests were completely soluble at pH 8. The maps were developed with a variety of general and specific reagents: ninhydrin, histidine, tyrosine, tryptophan and arginine reagents. Cystine peptides and cysteic acid derivatives have also been characterized. All detected half-cystine residues seemed engaged in disulfide bridges. For each individual peroxidase the number of specifically staining peptides agreed very well with the amino acid composition. The two most acidic peroxidases of turnip, P1 and P2, only differ significantly in one peptide. The P2 gene is tentatively proposed to have developed from the P1 gene by a single base mutation, changing an asparagine residue to alysine residue. A less acidic turnip peroxidase, P3, is distinct, although related to peroxidases P1 and P2. Horseradish isoperoxidase C also belongs to this group which appears to be closely related in the amino acid sequences around four disulfide bridges. Peroxidase P7 differs from this group, at least around two of its disulfide bridges, and therefore, may differ from the other four in parts of its three dimensional structure. Sequences of particular importance to peroxidase function must be present in all peroxidases. From the peptide mapping studies we only find two highly homologous sequences present in all five examined peroxidases. Both contain histidine. This finding corroborates previous suggestions of two histidine sequences near the peroxidase heme prosthetic group. The rules applied in relating peptides of different proteins are outlined, and the sources of errors in mapping of glycoproteins of high carbohydrate content (about 20%) are discussed in detail.


Analysis of the pancreatic-ribonuclease-digestion products of alfalfa-mosaic-virus ribonucleic acid. Sequence homologies between the different RNAs.

Alfalfa mosaic virus (AMV) genome consists of three pieces of RNA (24-S, 20-S and 17-s RNA). For infectivity these three RNAs and the coat protein are required. In the absence of coat protein, infectivity is obtained by adding the 12-S RNA also normally present in the virus. This 12-S RNA represents the message for coat protein. Thus a redundancy of the gene for coat protein exists between 12-S RNA and one of the other RNAs. Sequence analysis of the oligonucleotides resulting from pancreatic ribonuclease digestion of the AMV RNAs indicates that the nucleotide sequence of 12-S RNA occurs in 17-S RNA. Analysis of the pancreatic ribonuclease digestion products of the two larger alfalfa mosaic virus RNAs (20-S and 24-S RNA) shows some oligonucleotides containing seven, eight and nine nucleotides with the same structure present in both RNAs. The possibility of a limited nucleotide sequence homology between these two RNAs is discussed. The comparison of the RNase digestion products of 20-S and 24-S RNA with those of 12-S or 17-S RNA revealed no homologous oligonucleotides, thus the origin of 12-S RNA appears to be 17-S RNA.


Identification of NADH-specific and NADPH-specific FMN reductases in Beneckea harveyi.

Distinct FMN reductases specific for NADH and NADPH were identified in extracts of Beneckea harveyi. These enzymes differ in their physical (molecular weight, thermostability) as well as in their chemical properties (binding constants for NADH and NADPH). The NADH-specific enzyme is more efficient than the NADPH-specific one with respect to the bioluminescent reaction.


Multiple molecular forms of purified human erythrocyte acetylcholinesterase.

1. Human erythrocyte acetylcholinesterase was solubilized by Triton X-100 and purified by affinity chromatography to a specific activity of 3800 IU/mg of protein. The yield of the purified enzyme was 25--45%. 2. Gel filtration on Sepharose 4-B in the presence of Triton X-100 revealed one peak of enzyme activity with a Stokes' radius of 8.7 nm. Density gradient centrifugation in 0.1% Triton X-100 showed one peak of enzyme activity with an S4 value of 6.3S. 3. Isoelectric focusing in Triton X-100 resolved the enzyme into five molecular forms with isoelectric points of 4.55, 4.68, 4.81, 4.98 and 5.18. Upon incubation with neuraminidase the enzyme activity in the first four forms was decreased with a concommitant increase in activity in the form with the higher isoelectric point. 4. After removal of excess Triton X-100 on Bio-Gel HTP, polyacrylamide gel electrophoresis showed seven bands of protein and corresponding bands of enzyme activity. Density gradient centrifugation of the detergent-depleted enzyme at high ionic strength revealed five multiple molecular forms with S4 values of 6.3 S, 10.2 S, 12.2 S, 14.2 S and 16.3 S. At low ionic strength, higher aggregates were observed in addition to the other forms. Dodecylsulfate-polyacrylamide gel electrophoresis gave one subunit only with an apparent molecular weight of 80 000. 5. These results suggest that human erythrocyte acetylcholinesterase, solubilized by Triton X-100, exists in various forms differing in net charge but of apparently similar molecular dimensions. After removal of the detergent, forms with different molecular sizes are observed.


Isolation and characterization of active ribosomal subunits from human placenta.

We have developed a method for the large-scale isolation of active ribosomal subunits from human placenta. The technique involves incubating crude ribosomes for 15 min at 37 degrees C with 0.2 mM puromycin in 50 mM Tris-HCl buffer, pH 7.6, 500 mM KCl and 3 mM MgCl2 followed by centrifugation at 5 degrees C in a BXV zonal rotor using an equivolumetric sucrose gradient in the same buffer, upon which 80--90% of all ribosomes are dissociated into subunits. The purified subunits differ in their chemical composition, the 60-S particle containing no more than 36% protein whereas the 40-S subunit consists of 43% protein. In poly(U)-directed protein synthesis, tested in a completely homologous cell-free system, one recombined couple polymerizes at 37 degrees C 12 to 17 phenylalanine residues at an initial rate of 0.7 residues per minute. However, free 80-S ribosomes obtained by puromycin treatment of the crude ribosomes and reassociation of the subunits without prior isolation, have an even higher incorporating activity (20--25 mol phenylalanine/mol of ribosome). At least 55% of the subunits were estimated to actively participate in the polyphenylalanine synthesis.


Hydroperoxide-metabolizing systems in rat liver.

1. Metabolism of added hydroperoxides was studied in hemoglobin-free perfused rat liver and in isolated rat hepatocytes as well as microsomal and mitochondrial fractions. 2. Perfused liver is capable of removing organic hydroperoxides [cumene and tert-butyl hydroperoxide] at rates up to 3--4 mumol X min-1 X gram liver-1. Concomitantly, there is a release of glutathione disulfide (GSSG) into the extracellular space in a relationship approx. linear with hydroperoxide infusion rates. About 30 nmol GSSG are released per mumol hydroperoxide added per min per gram liver. GSSG release is interpreted to indicate GSH peroxidase activity. 3. GSSG release is observed also with added H2O2. At rates of H2O2 infusion of about 1.5 mumol X min-1 X gram liver-1 a maximum of GSSG release is attained which, however, can be increased by inhibition of catalase with 3-amino-1,2,4-aminotriazole. 4. A contribution of the endoplasmic reticulum in addition to glutathione peroxidase in organic hydroperoxide removal is demonstrated (a) by comparison of perfused livers from untreated and phenobarbital-pretreated rats and (b) in isolated microsomal fractions, and a possible involvement of reactive iron species (e.g. cytochrome P-450-linked peroxidase activity) is discussed. 5. Hydroperoxide addition to microsomes leads to rapid and substantial lipid peroxidation as evidenced by formation of thiobarbituric-acid-reactive material (presumably malondialdehyde) and by O2 uptake. Like in other types of induction of lipid peroxidation, malondialdehyde/O2 ratios of 1/20 are observed. Cumene hydroperoxide (0.6 mM) gives rise to 4-fold higher rates of malondialdehyde formation than tert-butyl hydroperoxide (1 mM). Ethylenediamine tetraacetate does not inhibit this type of lipid peroxidation. 6. Lipid peroxidation in isolated hepatocytes upon hydroperoxide addition is much lower than in isolated microsomes or mitochondria, consistent with the presence of effective hydroperoxide-reducing systems. However, when NADPH is oxidized to the maximal extent as evidenced by dual-wavelength spectrophotometry, lipid peroxidation occurs at large amounts. 7. A dependence of hydroperoxide removal rates upon flux through the pentose phosphate pathway is suggested by a stimulatory effect of glucose in hepatocytes from fasted rats and by an increased rate of 14CO2 release from [1-14C]glucose during hydroperoxide metabolism in perfused liver.


Oxidation of cytochrome b5 by hydroperoxides in rat liver.

1. Spectral changes following the addition of hydroperoxides to isolated hepatocytes and to perfused rat liver were observed. Cytochrome b5 is the major, if not the only, hemoprotein exhibiting redox changes under these conditions; cytochrome b5 is oxidized by added hydroperoxides, e.g. tert-butyl or cumene hydroperoxides. No spectral changes attributable to cytochrome b5 were observed with tert-butanol. 2. The effect is present also when the mitochondrial respiratory chain is inhibited by antimycin A, and it is not observable with isolated mitochondria. On the other hand, the oxidation of cytochrome b5 by hydroperoxides is readily demonstrable in microsomal fractions in presence of NADH. 3. Spectral evidence for a participation of the other microsomal hemoprotein, cytochrome P-450, in the hydroperoxide-linked effects was not obtained. Thus, in hepatocytes from phenobarbital-pretreated rats, no formation of cytochrome P-420, no displacement of a type I substrate, hexobarbital, and no major steady state redox change of cytochrome P-450 was detectable. However, when cytochrome P-450 was dithionite-reduced, an oxidation of this cytochrome occurred upon subsequent hydroperoxide addition. 4. Hydrogen peroxide addition to hepatocytes also leads to a lower steady-state degree of reduction of cytochrome b5. Evidence is provided with hepatocytes from rats pretreated with 3-amino-1,2,4-triazole that H2O2 generated intracellularly, e.g. from added glycolate, also causes a detectable oxidation of cytochrome b5. 5. The mechanism of these hydroperoxide effects remains to be established, and it is not clear whether cytochrome b5 reacts directly or indirectly. However, it is suggested that these effects may be of significance for the further study of cytochrome-b5-linked metabolic pathways.


Studies on the role and mode of operation of the very-lysine-rich histone H1 (F1) in eukaryote chromatin. The properties of the N-terminal and C-terminal halves of histone H1.

Restricted chymotrypsin digestion of calf thymus H1 histone gives two fragments, residues 1--106 and 107--C-terminal. These were studied by proton magnetic resonance and circular dichroism. The N-terminal fragment exhibited some salt-induced structure in aqueous solution, but this did not parallel the globular structure of the intact H1 molecule. Comparison of circular dichroism results with helix predictions for this portion of the molecule suggests that the secondary structure may be the same in this fragment as it is in the corresponding region of the whole molecule. The C-terminal fragments show very little salt-induced structure. The N-terminal fragments binds to DNA very weakly, but the C-terminal fragment binds as strongly as the whole molecule. In the C-terminal fragment, about one quarter of the lysine residues are not bound to the DNA in water, but initial increase of salt concentration causes them to become bound. This increasing binding occurs under the same ionic conditions that cause chromatin condensation and condensation of H1 - DNA complexes, and it is suggested that there may be a connection between these phenomena.


GTP degradation to guanine catalyzed by ribosomal subunits and microsomal-wash factors.

Ribosomes from stringent strains of bacteria generate (p)ppGpp if incubated with uncharged tRNA, a ribosomal wash fraction, GTP and ATP. By contrast, an analogous system from rat liver does not transform GTP to (p)ppGpp but degrades it to guanine. The reaction requires the ribosomal subunits, a 40 000-Mr and a 60 000-Mr microsomal wash protein factor and is inhibited if the ribosomal A-site is charged with aminoacyl tRNA. The degradation of GTP to guanine occurs in the following four distinct reaction steps: (a) hydrolysis of GTP to GDP plus Pi, (b) hydrolysis of GDP to GMP plus Pi, (c) hydrolysis of GMP to guanosine plus Pi, (d) hydrolysis of guanosine to guanine plus ribose. The reaction step (a) is inhibited by fusidic acid, cycloheximide, emetine, tetracycline and puromycin. The hydrolysis of GDP is inhibited strongly by fusidic acid, emetine and tetracycline. A putative physiological significance of this ribosome-dependent pathway in the processes of growth control of animal cells under conditions of amino acid deprivation is discussed.


Evidence for essential histidine residues in bovine-liver mitochondrial monoamine oxidase.

Ethoxyformic acid anhydride, amino-1H-tetrazole, and photooxidation in the presence of rose bengal, which are reagents known to react with histidine residues of proteins, were shown to inactivate monoamine oxidase. Ethoxyformic acid anhydride reacted with about 6 histidine residues per 100 000 g of protein under the experimental conditions adopted and completely inactivated the enzyme. However, NH2OH reactivated the ethaxyformic acid derivative t only. Since NH2OH specifically deacylates N-ethoxyformylimidazole, it was shown that at least some of the histidine residues are essential for activity. In addition, photooxidation experiments in the presence of 0.01% rose bengal confirmed that only histidine residues of bovine hepatic monoamine oxidase are destroyed under the designated experimental conditions. About 9 histidine residues per 100 000 g of protein were destroyed during the photooxidation experiments. In the presence of substrate, kynuramine or benzylamine, only 7 histidine residues were destroyed, which indicates that 2 histidine residues per 100 000 g of protein are essential for activity.


Evidence for a halothane-dependent cyclic flux of calcium in rat-liver mitochondria.

The previously reported (Hall et al., Biochem. Soc. Trans. 1973) halothane-dependent, calcium-induced loss of respiratory control in rat liver mitochondria is relatively specific to calcium; the effect of strontium ions is much smaller, and comparable additions of potassium salts have no effect on mitochondrial respiration on succinate in the presence of halothane. The calcium-dependent loss of respiratory control can be prevented, or reversed, respectively, by the prior or subsequent addition of agents that either chelate extramitochondrial Ca2plus or inhibit calcium accumulation, or that inhibit the efflux of accumulatec calcium. These results suggest that the halothane-dependent, calcijm-induced loss of respiratory control is due to a cyclic flux of calcium uptake and release.


Radioimmunoassay of prostaglandins Falpha, E1 and E2 in human plasma.

Antibodies against prostaglandins (PG)F2alpha, E1 and E2 were obtained inrabbits immunized with respectively PG F2alpha, PG E1 and PG E2 conjugated to bovine serum albumin by carbodiimide. A radioimmunoassay capable of measuring 7 pg of PG Falpha, 2 pg of PG E2 and 14 pg of PG E1 in human peripheral plasma is described. Plasma samples (pH 3, citric acid) are extracted with cyclohexane: ethyl acetate, 1:1 and then chromatographed on silicic acid columns to separate the prostaglandins into three fractions: fraction I, PG A, PG B and some unknown immunoreactive compounds; fraction II, PG E and fraction III, PG Falpha. The recovery is 80+ +/- 6.2. Mean plasma levels in adults of PG Falpha and PG E, expressed in pg/ml: -PG Falpha 12 +/- 2.8 (n- 25 men), 8 +/- 2.3 (n= 18 women, follicular phase), 7 +/- 1.4 (n= 18 women, luteal phase).  -PG E1 40.5 +/- 7.6 (n= 13 men), 38 +/- 17.1 (n= 10 women). -PG E2 4.5 +/- 1 (n= 12 adult subjects). The major characteristics of the method described herein are the following: - a large volume of plasma has to be processed (10 ml or more for PG Falpha and PG E1, 5 ml or more ofr PG E2). - a chromatographic step is necessary to separate the different prostaglandins which makes it possible to circumvent probelms of immunological cross reactivity and interference with unknown immunoreactive compounds. - great care has been taken in collection of blood samples, especially to insure complete removal of blood cells namely platelets.


The mechanism of action of mercuric chloride on the isolated perfused rat kidney.

Studies on 80 rat kidneys, perfused at constant flow, showed that mercuric chloride produced a marked increase in perfusion pressure within five minutes of administration. Adrenergic blocking agents (phentolamine and propranolol), angiotensin sensitivity depletion, bradykinin and low concentrations of mannitol had no effect on this increase in resistance. Perfusion of the kidney with 5 percent mannitol solution, however, significantly reduced the increase of vascular resistance induced by mercuric chloride. Evidence is presented that mercuric chloride may evoke an increase in vascular resistance by inducing endothelial cell swelling, an action antagonised by the hypertonic effect of mannitol. Other possible contributory mechanisms are also discussed.


Effects of burn injury on insulin secretion and on sensitivity to insulin in the rat in vivo.

Both insulin resistance and impairment of insulin secretion are know to occur in man after injury. The relative importance of these effects was studied in rats 2 h after a non-lethal 20 percent dorsal scald. No impairment of insulin secretion was found after this injury. Concentrations of both blood glucose and plasma insulin were elevated in scalded rats. Scalded rats responded to intravenous glucose injection (1-0 g/kg) with a further rise in plasma insulin concentration, which remained normal for the prevailing blood glucose concentration. However, marked impairment of glucose tolerance was observed, indicating the presence of insulin resistance. After intravenous insulin injection (1-0 U/kg) the initial rate coefficient for fall of blood glucose concentration was significantly lower (p less than 0-02) in scalded (mean 3-9 percent min.(-1) than in control rats (mean 6-3 percent min.(-1). The minimum in blood glucose concentration after insulin injection was reached at 10 min. in control rats, but not until 60 min. after injection in scalded rats. This difference was due to a delay in compensation for the hypoglycaemia in the scalded rats, since the rate of disappearance of insulin measured by injection of a tracer of 125I-labelled bovine insulin was not decreased after this injury. It was concluded that the impairment of glucose utilization in scalded rats (Heath and Corney, 1973) is due to decreased sensitivity to insulin rather than to suppression of insulin release.


The role of calcium in insulin action. III. Calcium distribution in fat cells; its kinetics and the effects of adrenaline, insulin and procaine-HCl.

The effects of adrenaline, insulin and procaine-HCl on Ca distribution in intact fat cells and on Ca binding to fat cell ghost membranes have been investigated. 1. Fat cells incubated in 45Ca containing media till isotopic equilibrium indicated that the exchangeable Ca in these cells averages 25.7 +/-3.2nmol/mg protein, which represents approximately 9.8% of their ttotal Ca content. 2. Perifusion of 45Ca prelabelled fat cells gave washout curves whose analysis conformed with three kinetically distinct Ca pools (Fig. 1). The fast exchangeable pool (Compartment A) had an efflux rate constant of 0.193 +/-0. 013 min.-. The release of Ca from the second and thrid pools (Compartments B and C) was much slower with efflux rate constants of 0.032 +/-0.0018 min.-1 and 0.0042 +/- 0.0006 min.-1 respectively. Changing the Ca concentration in the perifusing medium modified the initial fast phase and its rate constant, while added dinitrophenol (DNP) inhibited the efflux rate from the later compartments...


The role of calcium in insulin action. IV. Mechanism of insulin resistance in adipose tissue of obese (ob/ob) mice and old Wistar rats.

The in-vitro antilipolytic response to insulin and procaine hydrochloride by adipose tissue from young rats (150 - 180 g) and lean mice has been compared with that from aged Wistar rats (600 g) and obese (ob/ob) hyperglycaemic mice. 1. The adipose tissue from the obese mice showed diminished responsiveness to insulin and to procaine hydrochloride. Response to these agents, however, was restored by prewashing the tissue, suggesting that the apparent resistance in this tissue reflected saturation of the insulin receptors to endogenous insulin. 2. In adipose tissue of old Wistar rats the antilipolytic effect of insulin was also impaired, but this was not restored after extensive washing. Unlike adipose tissue ghosts prepared from young rats, insulin did not decrease the binding of calcium to ghost membrane preparations from old rats. Neither did insulin inhibit the adrenaline stimulated 45 calcium efflux from perifused isolated fat cells prepared from old rats. These results suggest that the insulin response of fat cells from old rats is impaired because of a defect either in their insulin receptors or in their post-receptor responses. 3. Procaine-hydrochloride, however, when added to the medium perifusing fat cells of these old rats inhibited the adrenaline stimulated lipolysis, reduced the Ca efflux and decreased the binding of Ca to fat cell ghosts; as it did with similar preparations of young rats. Thus the cells from old rats still show full post-receptor responsiveness to an insulin-like stimulus, provided the stimulus for such a response is given at a point beyond the insulin receptor itself. The results suggest that the insulin resistance observed in old rat fat cells may be related to some by deficiency in the insulin receptors, possibly due to their lower replacement with age.


Synthesis of immunoglobulins and 15 other proteins by diseased human kidneys.

In an effort to determine the synthetic capacity of human kidneys for serum proteins, pieces of both kidneys removed at operation prior to transplantation were placed in organ culture. Incubation with 14C labelled amino acids and subsequent radioimmunoelectrophoresis with antisera to all five classes of immunoglobulins showed synthesis of IgG by all 5 pairs of kidneys of IgA and IgE by 4, and of IgM by 2. None synthesized IgD. Radioimmunoelectrophoresis with specific antisera to 15 non-immunoglobulin proteins revealed synthesis of lactoferrin and uromucoid while several other serum proteins, such as alpha1 antitrypsin and alpha2 macroglobulin, known for their binding properties, showed apparent synthesis. No evidence for secretory component synthesis was seen despite positive immunofluorescent localization in the tubular cells. Further, Zn alpha2 and beta2GP I also showed positive immunofluorescence but negative synthesis. It seems possible that these latter proteins may not be synthesized by the tubular cells but rather may be actively excreted or reabsorbed by them.


The effect of yohimbine on the turnover of brain catecholamines and serotonin.

Yohimbine moderately increased the depletion of brain dopamine (DA) after alpha-methyl-p-tyrosine (AMT) only when the two drugs were given at the same time; the baseline concentration of brain homovanillic acid (HVA) and its accumulation after probenecid were strongly increased by yohimbine. Yohimbine markedly decreased the concentration of brain noradrenaline (NA), both when given alone and before or at the same time as AMT; when it was given at an increasing interval after AMT, the effect became progressively smaller. The baseline concentration of brain 3-methyoxy-4-hydroxyphenylethyleneglycol sulphate and its accumulation after probenecid were increased by yohimbine; this effect was not as marked as that on HVA and was proportional to the quantity of NA depleted in the Amt method. The accumulation of brain k-hydroxyindoleacetic acid after probenecid was decreased by yohimbine pretreatment.


Centrally mediated hypotension and bradycardia by methysergide in anesthetized dogs.

In anesthetized dogs, methysergide (1 and 3 mg/kg i.v.) caused reductions in systolic and diastolic blood pressure, heart rate, left ventricular pressure and peripheral resistance. Caardiac output was unchanged because of an increase in stroke volume. Methysergide exhibited no alpha-receptor, ganglion, or adrenergic neuron-blocking properties nor did it have marked direct vasocilator action. The BCO, but not the orthostatic, reflex was severely inhibited by the drug, evidence for a central inhibitory action. Atropine, vagotomy or carotid sinus debuffering had little or no effect on the hypotension and bradycardia produced by methysergide, whereas guanethidine pretreatment essentially abolished these effects. Direct intracerebronventricular administration of small doses of methysergide (0.2 mg/kg) caused significant hypotension and bradycardia. It is concluded that methysergide causes centrally mediated hypotension and bradycardia, the mechanism of which is not clearly understood.


An autoradiographic method of mapping the distribution and density of monoamine neurons in mouse brain.

A combined in vitro uptake and autoradiographic procedure as an important complement to the histochemical fluorescence method is described. Slabs of fresh mouse brain were incubated with 14C-NE, 14C-DA or 14C-5-HT, freeze-dried, and placed against X-ray film for autoradiography, Catecholamine nerve terminals were labeled by in vitro incubation with 14C-NE or 14C-DA. Dopaminergic terminals were labeled by 14C-NE incubation preceded by desipramine (to block uptake into NE terminals). With 14C-5-HT incubation, the uptake pattern indicated the possibility that 5-HT nerve terminals were being labeled. Advantages of this method are that it allows the visualization of overall density and distribution of selected monoamine nerve terminals or uptake sites of other putative neurotransmitters in whole coronal or sagittal sections, so that data are obtained from many areas of brain or spinal cord rather than in only those areas preselected for microscopic viewing.


Increased binding of quinidine to serum albumin and lipoproteins in anuric rats.

The purpose of this work was to identify the main quinidine binding molecules of rat serum and those macromolecules responsible for increased quinidine binding in serum from rats with acute anuria. Rat sera were fractionated by gel filtration, ultracentrifugation and anion exchange chromatography. The binding of quinidine to sera and serum fractions was determined by equilibrium dialysis. The experiments demonstrated that albumin and lipoproteins are the main quinidine binding molecules in serum from normal and anuric rats. The increased binding by serum from anuric rats is due to both serum lipoproteins and albumin, even though the concentration of albumin is decreased from 2.8 to 1.7 g/100 ml. The increased binding to albumin may be due to conformation changes induced by endogenous substances. The increased binding to lipoproteins may be caused by an increased concentration of pre-beta-lipoprotein.


The effect of catecholamine depletion on the bradykinin-induced relaxation of isolated smooth muscle.

The effect of depletion of catecholamines by tyramine or reserpine on the bradykinin-induced relaxation of the rat duodenum aan rabbit ileum was the object of this study. The relaxation was not affected by catecholamine depletion due to repeated addition of tyramine to the isolated organs from either normal or reserpine-treated animals. From these findings and on the basis of other data in the literature it can be concluded that the bradykinin relaxation is not due to a release of catecholamines, but rather to a direct effect on the smooth muscle cells.


Antagonism of corticosteroids against the lethal effects of hemicholinium-3 in rats and mice.

The corticosteroids prednisolone or dexamethasone administered simultaneously with hemicholinium-3 (HC-3) by i.p. injection, increased the LD50 of HC-3 in rats and mice about twofold. Daily treatment with prednisolone for 7 days also increased the LD50 of HC-3 in rats. Choline completely protected rats and mice against HC-3. A sublethal dose of physostigmine did not change the LD50 of HC-3 in rats but increased the LD50 in mice to a small extent. The possible implications of these experiments for the explanation of the beneficial effects of corticosteroids in myasthenia gravis are discussed.


Effect of dopamine on renin secretion in the anesthetized dog.

Intrarenal perfusion of dopamine (6 mug/kg/min for 10 min) caused a significant increase of renin secretion, together with a significant increase in renal blood flow. This renin hypersecretion is not accompanied by any significant alteration in renal perfusion pressure, kalemia or natriuresis. The role of intrarenal dopaminergic receptors has been studied: (a) Haloperidol (intrarenal perfusion of 50 mug/kg/min for 20 min) suppresses the renal vasodilation and renin hypersecretion induced by dopamine. (b) Propranolol (intrarenal perfusion of 1 mg/kg in 15 min, then of 4 mg/kg/hr) alters neither the renal vasodilation nor the renin hypersecretion induced by dopamine. These observations support the assumption that the dopaminergic receptors are brought into play in the two renal responses to dopamine studied by us.


Mechanisms of selective depletion of brain regional noradrenaline by systemic 6-hydroxydopamine in newborn rats.

6-Hydroxydopamine injected systemically into newborn rats caused permanent depletions of the noradrenaline content of only certain brain regions. When 6-hydroxydopamine was given i.p. on days 1 and 2 after birth only the cortex, hippocampus and spinal cord were permanently depleted. 6-Hydroxydopamine given on days 9 and 10 after birth caused a permenent depletion predominantly from the cerebellum. Other regions were unaffected or were depleted for 4 days or less. The hypothalamus showed a gradual recovery of noradrenaline levels over a period of more than 20 days after injection, suggesting that only in this region was the apparent lack of effect of 6-hydroxydopamine due to neuronal regrowth. 6-Hydroxydopamine appeared to gain access to all regions except the pons-medulla when injected up to 10 days after birth, since at least temporary effects on noradrenaline storage were observed. 3H-Noradrenaline after i.p. injection was found in similar relative amounts in all brain regions in rats up to 20 days old. Thus regional differences due to age in the ability of 6-hydroxydopamine to reach various brain regions apparently do not explain the selective susceptibility to permenent depletion. It is suggested that the selective depletion must be due to interactions between the maturity of the adrenergic neurones and other factors such as their ability to take up or retain the 6-hydroxydopamine.


The evaluation of anti-parkinson drugs on reserpine-induced rigidity in rats.

The effect of anti-parkinson drugs on reserpine-induced rigidity was examined using a technique which measured rigidity by hind limb palpation. Centrally acting dopaminergic and anti-cholinergic drugs were able to antagonize reserpine-induced rigidity, whereas peripherally acting drugs had no effect. Results are discussed in terms of possible modes of action. Relative potencies of the drugs studied in this model were well correlated with the dose ranges of clinically established anti-parkinson drugs. It is concluded that measurement of reserpine-induced rigidity by hind limb palpation is a rapid and sensitive technique to evaluate potential anti-parkinson drugs.


Blockade by metal complexing agents and by catalase of the effects of arachidonic acid on platelets: relevance to the study of anti-inflammatory mechanisms.

Metal-chelating agents inhibited platelet aggregation and the accompanying generation of rabbit aorta contracting and PG-like activities, when platelets were challenged with arachidonic acid. Inhibition required the presence of the chelating agents in the medium, and was insured by reagents avid for free or protein-bound copper. Catalase also prevented aggregation and generation of pharmacologically active substances; its activity was reversed by aminothiol agents and by Cu2+ and Zn2+, shown previously to potentiate the platelet effects of arachidonic acid. Inhibition by indomethacin was not prevented by amino-thiol drugs nor by Cu2+ or Zn2+. The catalase-induced inhibition was not affected by scavenging of thiol groups; this rules out, as a mechanism of action of catalase, the increased destruction of popoperoxides by glutathione peroxidase, which requires reduced glutathione as hydrogen donor. The results are compatible with the hypothesis that the agent that mediates platelet aggregation by arachidonic acid is a popoperoxide, requiring the presence either of H2O2 or of a similarly catalase-sensitive substance to be generated.


The stimulation of renin secretion by diazoxide in the isolated rat kidney.

The intrarenal effect of diazoxide on renin secretion was examined in the isolated perfused rat kidney. Mean renin secretion measured at 2 min intervals during the continuous infusion of diazoxide for 6 min was consistently higher than the corresponding control values, although this was significantly different at the end of the infusion only. Since renal perfusion pressure during diazoxide infusion and control studies decreased to a similar extent, it is suggested that diazoxide stimulates renin secretion by a direct effect on the juxtaglomerular cell.


Effect of chronic pentobarbital treatment on blood-cerebrospinal fluid kinetics.

Cats were made tolerant and physically dependent by twice daily equieffective anesthetic dosing with sodium pentobarbital intragastrically for 5 weeks. Serial blood and CSF samples were simultaneously collected on day 1 and day 35 and analyzed for pentobarbital by a gas chromatographic method. Pentobarbital penetration into CSF was rapid and unchanged by chronic treatment. The concentration in blood and CSF increased significantly with chronic treatment; however, the relative CSF/blood concentration ratios were unchanged.


The influence of piribedil (ET495) on components of locomotor activity.

Recent evidence suggests piribedil affects both cerebral NA neurones and DA neurones and receptors since it increases brain MOPEG-SO4, an observation confirmed presently. In the reserpinised mouse piribedil and apomorphine caused a reversal of akinesia, which was significantly enhanced by concurrent administration of clonidine. However, piribedil neither reduced nor increased the motor effects of apomorphine or L-DOPA in the reserpinised mouse, but slightly enhanced that of amphetamine. Phenoxybenzamine reduced the motor action of L-DOPA but had no effect on that of apomorphine or amphetamine. These results suggest piribedil has no significant NA receptor blocking or stimulating action in the reserpinised animal. They confirm the view that piribedil increases cerebral NA turnover due to a presynaptic action on NA neurones.


Prevention of 6-hydroxydopamine neurotoxicity.

1-Phenyl-3-(2-thiazolyl)-2-thiourea (PTTU) prevented the neurotoxic actions of 6-hydroxydopamine on adrenergic nerves in the mouse atrium and iris. This is the first reported 6-hydroxydopamine antagonist that does not act by blocking uptake of catecholamines into nerve terminals. PTTU also prevented the diabetogenic action of alloxan.


Changes in acetylcholine and noradrenaline sensitivity of chick smooth muscle wholly innervated by sympathetic nerve during development.

Developmental changes in sensitivity of the isolated expansor sedumdariorum muscle of posthatching chicks to noradrenaline (NA), Acetylcholine (ACh) and some other drugs were investigated. This muscle responded to both NA and ACh in early life. The sensitivity to ACh decreased progressively with increasing age and disappeared on day 40 after hatching, however, a corresponding elevation of cholinesterase activity was not observed. The sensitivity to NA remained at the same level during the period of 2-60 days after hatching. The Contractile action of ACh on this muscle was not affected by d-tubocurarine, hexamethonium or phentolamine, but was completely abolished by atropine. These results suggest that there are at least two kinds of responsive sites on the expansor secumdariorum muscle in the new-born chick and that the sites sensitive to ACh degenerate progressively during the developmental processes. The cholinergic sensitive sites of this muscle in the new-born chick may be muscarinic.


Effects of N-acetylprocainamide as compared with procainamide in isolated rat atria.

The actions of procainamide and its major metabolite N-acetylprocainamide were tested and compared on isolated rat atria. While procainamide exerted a negative chronotropic and iontropic effect, N-acetylprocainamide had the opposite effect. It is suggested that a N-acetylprocainamide-induced increase in myocardial work can counteract the negative inotropic action of procainamide and thus to some extent explain the variable results with the latter compound on myocardial performance reported from in vivo experiments. Procainamide increased the refractory period and reduced the excitability of isolated rat atria. N-acetylprocainamide, on the other hand, caused negligible effects on these parameters.


The effect of dopaminergic stimulation and blockade on the nociceptive and antinociceptive responses of mice.

Agents which stimulate dopaminergic receptors directly or indirectly such as apomorphine and L-dopa, increased the reactivity of mice to a nociceptive stimulus. The increased reactivity was pharmacologically quantitated by estimating the hyperalgesic ED50 to be 4.4 and 115 mg/kg for apomorphine and L-dopa, respectively. This hyperalgesia was blocked by the dopamine receptor blocking agents, haloperidol and pimozide, but not by the narcotic antagonist, naloxone. Apomorphine antagonizes morphine analgesia. However the induced hyperalgesia only accounts for part of the antagonistic activity of apomorphine. The majority of the antagonistic activity of apomorphine appears to be by means other than action on dopaminergic receptors.


Lidoplazine and Na and K distribution in normo-and hypokalaemic rats.

The effects of lidoplazine on K-metabolism were investigated in two types of experiments. (1) K and Na concentrations were measured in plasma, heart, diaphragm and erythrocytes in normo- and hypokalaemic rats with or without chronic (1 month) lidoplazine treatment. Total body weight, heart weight, dry weight/wet weight ratios and extracellular space were also determined. (2) Radioactive K influx in the heart was measured in acute in vitro experiments with or without lidoplazine. It is demonstrated that while lidoflazine has no effect on the Na and K distribution of normokalaemic animals, it has rather a K-saving effect in conditions of severe loss of intra- and extracellular K-ions. In normal in vitro preparations the unidirectional K-influx is not affected by lidoflazine.


Localization of the central sympatho-inhibitory effect of a narcotic analgesic agent, fentanyl, in cats.

Fentanyl (10 and 30 mug/kg), a narcotic analgesic, induced in cats a decrease in blood pressure and heart rate and reduced spontaneous splanchnic nerve activity. Fentanyl reduced the pressor response to medullary stimulation, but did not change the pressor response to hypothalamic or cervical spinal cord stimulation. Fentanyl reduced the potential evoked in the splanchnic nerve by stimulation at low frequency of a pressor area of the medulla oblongata. The potentials evoked in the splanchnic nerve by hypothalamic or cervical spinal cord stimulation were only slightly changed. Nalorphine (0.5 mg/kg) or naloxone (30 mug/kg) induced a recovery in blood pressure, heart rate and spontaneous splanchnic discharges which had been reduced by fentanyl, but nalorphine or naloxone did not restore pressor response to medullary stimulation or potentials evoked in the splanchnic nerve by medullary stimulation, which had been decreased by fentanyl.


[Mercury fixation in different subfractions of human serum albumin separated by polyacrylamide gel electrophoresis].

Electrophoresis of mercurial serumalbumin allows isolation of 5 proteic sub-fractions, whose respective importance is appraised by densitometry. In a parallel direction mercury dosages are performed on each sub-fraction. They disclose that there is a relation between mercury fixation and the sub-fractions migration speeds: the 2 sub-fractions with the lowest migration speeds had fixated more mercury than the others.


[Distribution of organophosphate pesticides in two artificial plasmas: FSB-FSP].

Here is an attempt to establish that a close relation exists between the chemical formulae of organophosphates and their affinities for lipids, proteins and water. We carried out this study on two systems similar to human plasma. First, each component was sorted, then the organophosphates were detected and titrated by gas chromatography with flame photometry detector. Thiophosphates had a greater affinity for lipids than phosphates. Fixation of proteins was only observed on three hydrophilous organophosphates: phosphamidon, dichlorvos, ciodrin.


Fine structure of lymphocytes from an immune deficient child before and after administration of transfer factor.

Lymphocytes from a male infant delivered by Cesarean section and placed into a germ-free environment were examined by electron microscopy (EM). The child had a sex-linked severe combined immunodeficiency. The lymphocytes were atypical, having sparse cytoplasm with little rough endoplasmic reticulum (ER) but abundant smooth ER. The nuclear membrane was pulled away from the nuclear space, and no evidence of nuclear pores or aggregated ribosomes was found. Mitochondria were intact. Repeated injections of the subject during the 9-month period with KLH, typhoid vaccine, and diphtheria toxoid yielded no significant observable change in the fine structure of the lymphocytes. At 11 months, the subject was given transfer factor. Following repeated injections of this material, the original cell type was still present but a new type of lymphocyte was also observed by EM examination. The new cell type resembled a more normal lymphocyte. It had a higher density of cytoplasmic material, in comparison with cells prior to administration of transfer factor. It was smaller in size with some aggregated ribosomes, had detectable amounts of rough ER, and more intact nuclear membranes. This new type of lymphocyte may represent a small population of B lymphocytes perhaps stimulated by T cells made immunocompetent by transfer factor.


[The influence of dietary regimen on the stimulation of microsomal monooxygenases in rat liver induced by 2,3-dimethylquinoxaline].

Activities, on the hepatic microsomal fraction, of the following enzymes: aniline aromatic hydroxylase, p-nitroanisole O-demethylase, aminopyrine N-demethylase, N-methylaniline N-demethylases, as well as the P-450 cytochrome level, have been evaluated on female rats. During a period of 26 days, they are fed on 4 diets including either: 20% or 30% proteins, or: 1% or 30% lipids. A parallel study is carried out: animals fed on the same diets are given, orally on the 22nd day and for 4 days, a bactericide, the dimethyl-2.3 quinoxaline; the dose is 500 mg/kg/day, with an interval of 24 hours. 1) A stimulation of the microsomal monooxygenase activity and an increase in the P-450 cytochrome level are induced by the dimethyl-2.3 quinoxaline on all animals regardless the diet they are on. 2) The hyperlipidic diet is the only one to induce an increased activity of the same enzymes, but the P-450 cytochrome level remains unchanged. 3) In rats fed on the hyperlipidic diet and treated with dimethyl-2.3 quinoxaline, the effects of this substance and of this diet, on the drug-metabolizing enzymes, become additive.


[Reactivation by 1-hydroxyimino-4-methyl-2-pentamone of plasma cholinesterases in the dog inhibited by different orgenophosphates].

The hydroxyimino-1 methyl-4 pentanone-2 was selected among several oximes because of its highly successful restoring action on dog plasmatic cholinesterases inhibited by DFP (diisopropyl fluorophosphate). During the in vitro study, this storing action was tested on 10 organophosphates. Results are compared to those obtained when contrathion is used.


Estimation of the LD1 and extrapolation of the LD0.1 for five organothiophosphate pesticides.

The oral LD50's for five organothiophosphate pesticides have been determined in CD-1 strain male and female mice. The values in mg/kg are: parathion, 15.0 and 14.3; methyl parathion, 14.5 and 19.5; guthion, 7.15 and 6.35; imidan, 25.2 and 23.1; sumithion, 1,045 and 1,220 respectively. Toxicity was greater parathion but there was no difference in the males. The relationship between chemical structure and toxicity has been discussed. The addition of a methyl group in the meta position of the nitrophenyl group (sumithion) decreased toxicity 72 times compared to methyl parathion. The predicted LD1 and extrapolated LD0.1's have been determined for the five organothiophosphates and the conditions required for accurate results have been established. It has been shown that the slopes of the curves (males vs. females) obtained with 50, 100 and 660 animals are parallel for all compounds health implications of exposure to low levels of environmental pesticides have been discussed.


[Ultrastructural study of a renal biopsy in a patient poisoned by paraquat].

It is possible through investigation under the electronic microscope to determine the location and nature of lesions with accuracy. These lesions are mainly found on the epithelium of proximal and distal convoluted tubules. However, the extent of lesions together with the deterioration process of the cell-constituents vary from one structure to the other and even from cell to cell. On the other hand, while blood capillary vessels pertaining to tubules may also show moderate lesions, glomerular and interstitial deterioration is rare. Kidney deterioration which may either result directly from one of the poison effects or from related phenomena, is under discussion.


Suicidal poisoning by nitrite.

The authors relate a suicidal poisoning by nitrites. Post mortem findings are reported as well as a method for the quantitative assessment of NO2- in blood, urine, gastric and intestinal contents, kidney and liver. The concentration values obtained are correlated with the methemoglobin figures which indeed are lethal. The highest concentrations of NO2- are found in the small intestine where the nitrites seem to resist chemical decomposition owing to the reducing properties of the bacterial flora.


Effect of different sedative drugs on the proliferation of bone marrow cells. Preliminary results.

Several drugs were assayed for their effect on bone marrow hematopoïetic cells proliferation. Chlorpromazine, barbiturates and benzopiperidine were found active only at toxic dosage. Propranolol and fluanison seem in the preliminary studies to lessen 3 H-thymidine incorporation in vitro at non toxic concentration.


(S)-3-p-Methoxyphenyl-3-acetylaminopropan-1-ol, a New Metabolite of an Actinomycete.

(S)-3-p-methoxyphenyl-3-acetamidopropan-1-ol was isolated from cultures of an actinomycete (Streptomyces michiganensis). Its structural determination by spectroscopic means and its synthesis are described.


The absolute configuration of the enantiomers of glutethimide and aminoglutethimide.

Aminoglutethimide (Elipten¿ CIBA) was resolved into the optical antipodes I and II. The endocrinological properties and the absolute configuration of both enantiomers I and II were determined. Most of the steroidal synthesis inhibition was found in the (+) enantiomer II. On the basis of circular dichroism, the R-configuration was assigned to the (+) enantiomer II.


The effect of skim milk on plasma cholesterol in rats.

From the 15th day of birth, newborn rats were offered rat chow mixed with water or skim milk. The former also received tap water, the latter skim milk. In the latter group, plasma cholesterol was lowered in 43- and 64-day-old males and in 64-day-old females.


Chlorodimeform and its effect on monoamine oxidase activity in the cattle tick, Boophilus microplus.

The action of the acaricide, chlorodimeform and its metabolite. N-desmethylchlorodimeform, on the activity monoamine oxidase (MAO) from the cattle tick, Boophilus microplus, were studied. Both compounds were found to be potent in vitro and in vivo inhibitors of the enzyme. However the inhibition of MAO does not seem to be related to the toxic action of the acaricide.


Binding of D-alpha-tocopherol to rat liver nuclear components.

When D-alpha-tocopherol is administered i.v. to vitamin E deficient rats, significant amounts of this vitamin are bound to a nucleoprotein complex in hepatic nuclei, and this complex can be solubilized by high concentrations of sodium chloride (0.6 M). The bound vitamin in this complex, extractable by ethanol, was found to be identical with authentic alpha-tocopherol by thin layer chromatography.


Lactate dehydrogenase isozymes of mouse epidermis.

Five isozymes of LDH are demonstrable in the epidermis of the ear pinnae, hind feet, trunk dorsa, and tails of adult C57BL, C57HR, and C3HB mice by polyacrylamide gel electrophoresis. LDH-5 activity predominates in electropherograms. The ratio of LDH-1 to LDH-K is greater in the epidermis of ear pinna and trunk dorsum than in that of tail and hind foot. The region-specific patterns of epidermal LDH isozymes are not correlated with melanin pigmentation or "hairiness' of the skin.


Manganese action potentials in mammalian cardiac muscle.

The membrane potential in guinea-pig's papillary muscles from right ventricle was recorded by glass microelectrodes and stimulation was effected by current pulses applied through a sucrose-gap. Action potentials with overshoot were recorded in the solution lacking Na+ and Ca++ but containing 2-95 mM Mn++. The overshoot was increased with the increase of [Mn++]o by about 30 mV/decade. Similar Mn++ dependent action potentials were also obtained in Na-free solution containing 0.6 mM Ca++. The results indicate that Mn inward current is sufficient to generate action potentials in cardiac muscle.


Tendon shortening in striated muscle.

Tendon shortening in the digastric muscle of the rabbit resulted in a short term increase in gross muscle length and sarcomere length. Subsequently, muscle and sarcomere lengths decreased to less than control values. Long-term measurements suggested that a return to control sarcomere length may have been achieved by a reduction in sarcomere number.


Facilitation by imidazole of the aqueous flare response to alpha-melanocyte stimulating hormone.

MSH, like traumata to the eye, cause a permeability disturbance in rabbits, with protein leakage into the aqueous. The MSH effect was enormously increased by instillation of imidazole or pilocarpine. The MSH effect seems to engage a different mechanism than the prostaglandin-dependent action of other agents.


Inhibition of isoprenaline-induced increase in plasma renin concentration by vasoconstrictors.

The vasoconstrictors angiotensin II, vasopressin and the alpha-sympathomimetic phenylephrine significantly inhibit the renin release caused by the beta-sympathomimetic isoprenaline. The mechanism of the inhibition is discussed.


Scanning electron microscope observations of the canaliculi in the rat pineal gland.

The scanning electron microscope has shown rich ramifications of the parenchymal canaliculi forming a three-dimensional network of anastomosing intercellular spaces in the rat pineal gland. Every pineal cell seems to be in contact with this channel system. An abundance of cellular processes can be found within the canaliculi which may play an important role in the histophysiology of the pineal body.


Effects of glucagon and insulin on the Paneth cells of the mouse duodenum.

The effect of glucagon and insulin on the paneth cells (PC) of the duodenum of the mouse was investigated using light microscopy. Both glucagon and insulin were able to increase significantly the number of the secretory granules of PC. This possibly means that these hormones are capable of inhibiting the secretion of PC.


Role of sialic acid in the maintenance of cell surface rigidity.

Removal of the cell surface sialic acid with neuraminidase brings about cell deformation in amoeba. The membranes of these deformed cells are eventually ruptured leading to the liberation of the cell mass.


Degeneration of adrenergic axons in the longitudinal muscle coat of the rat duodenum following treatment with 6-hydroxydopamine.

Following systemic treatment with 6-OHDA, electron microscopy of adult rat duodenum showed degenerating adrenergic axons in the longitudinal muscle coat making neuromuscular relationships with gaps of varied widths.


[The early differentiation of the perinotochordal connective tissue. A scanning and transmission electron microscopic study on chick embryos (author's transl)].

The early differentiation of the connective tissue was investigated in the perinotochordal zone of 2-3 day-old chick embryos. After characterizing the different tissue components by transmission electron microscopy, their arrangement and distribution were examined by SEM. The results are discussed with regard to the role of the extracellular material in embryonic tissue interactions.


Differentiation of quail Hensen's node in chick coelomic cavity.

Quail Hensen's nodes were cultured in the chick coelomic cavity. They differentiated into various tissues, intimately mixing with host cells, which in many cases formed the supporting mesoderm of graft epithelial or nervous tissue.


Activities of kidney decarboxylases of histidine and ornithine in adrenalectomized mice substituted with cortisone.

In adrenalectomized mice, cortisone inhibited histidine decarboxylase of the kidney in a dose-related manner. The effect of cortisone on ornithine decarboxylase was diphasic: small doses elevated, high doses inhibited.


A simple method for collection and purification of chicken peritoneal macrophages.

The authors describe a simple method for collection and culture of chicken peritoneal macrophages. The macrophages are collected from peritoneum with polystirol disks on the surface of which they are subsequently cultured in vitro.


New prenylated chalcones from Lonchocarpus neuroscapha Benth. (Cordoa piaca).

The isolation of eight prenylated chalcones (cordoin, isocordoin, psi-isocordoin, derricin, lonchocarpin, 4-hydroxyderricin, 4-hydroxylonchocarpin, 4-hydroxyisocordoin) and of the flavanone and dihydrochalcone corresponding to cordoin is described. These substances are biogenetically correlated. The structures of the above mentioned substances were established through the examination and comparison of spectral data (U.V., I.R., N.M.R., M.S.) and of the chemical behaviour. Particular interest is shown by psi-isocordoin and 4-hydroxy derivatives. The latter and 4-hydroxyderricin in particular show also a marked inhibition on the growth of gram-positive bacteria.


The reaction of phthalazino(2,3-b)phthalazine-5,12(7H, 14H)-diones with nitrous acid.

3,4-Dihydrophthalazin-1(2H)-one (I) was oxidized to phthalazin-1(2H)-one (III) with nitrous acid or with ferric chloride . Phthalazino [2,3-b] phthalazine-5,12(7H, 14H)-diones (IV) did not react with ferric chloride but they were oxidized with nitrous acid to 2-[1(2H)-oxo-2-phthalazinyl] methylbenzoic acids (V) and (VI). The formation of (V) or (VI) depends upon the substituents of compounds (IV). Strucutres (V) and (VI) were established by pKa measurements in methylcellosolve and by mass and N.M.R. spectra.


Biological properties of a new non-steroidal anti-inflammatory drug: etoclofene.

Etoclofene, the ethoxy methyl ester of N-(2,6-dichloro-m-tolyl)anthranilic acid, showed potent anti-inflammatory, anti-pyretic and peripheral analgesic activity in several experimental models of inflammation. There was no evidence of adrenal dependence or corticoid-like effects at effective and non-toxic dose levels. Etoclofene showed a low level of toxicity and was well tolerated in animals.


[Isoxazoles substituted with 4-pyridyl and o-chlorophenyl radicals].

By reaction of suitable hydroxamic acids chlorides with beta-ketoesters, we have prepared the ethyl esters of isoxazol-4-carboxylic acids, substituted in the 3- and 5-positions with 4-pyridyl and o-chlorophenyl groups, and some of their esters and amides of pharmaceutical interest. 3-(4-Pyridyl)-5-(o-chlorophenyl)isoxazole was obtained either by decarboxylation of the acid or by oxidation of the syn and anti oximes of 3-(o-chlorophenyl)-1-(4-pyridyl)-2-propen-1-one; from the syn oxime the corresponding isoxazoline was also obtained. Pharmacological screening shows that some of the new compounds have a myolytic activity.


[Esters and amides of salicyloylamino acids].

N-Salicyloylamino acids, esters and amides have been prepared by the usual condensation methods and also by methods which may demonstrate 2-(o-hydroxyphenyl)oxazoline-5-ones as possible reaction intermediates. Some derivatives display pharmacological activities.


[N 1-nicotinoyl and isonicotinoylpyrazoles].

Synthesis of twelve N 1-nicotinoyl and isonicotinoylpyrazoles by reaction of 4,5-tetra-, penta-, hexamethylenepyrazoles, dihydronaphtopyrazole, bornylenepyrazole and cineolylenepyrazole with nicotinoyl and isonicotinoylchloride, is described. The structure of N 1-derivatives of these compounds was ascertained by comparison of N.M.R. spectra with those of 1-nicotinoyl and 1-isonicotinoyl-4,5-pentamethylene-5-hydroxy-2-pyrazoline, obtained by reaction of 2-hydroxymethylenecycloheptanone with nicotinoyl and isonicotinoylhydrazide, respectively. N 1-isonicotinoylbornylenepyrazole shows weak C.N.S. depression, muscle relaxation and anticonvulsant activity.


[Reaction of 1,2-diphenyl-4-bis-(methylthio)methylenepyrazolidin-3,5-dione with serine, cysteine and their derivatives].

The reaction between 1,2-diphenyl-4-bis-(methylthio)methylenepyrazolidin-3,5-dione with serine and cysteine and their derivatives is described. The structure of the resulting compounds is assigned spectrophotometrically.


Determination of structure-activity relationships for salicylaldehydes using CNDO and EHT calculations.

EHT and CNDO calculations have been performed on a series on fungicidal salicylaldehyde derivatives. Correlations are found between the inhibitory activity and several quantum-mechanical indices.


Structure-activity correlations for acaricidal hydrazones.

The acaricidal activity of hydrazones was studied by means of the Free-Wilson method. The importance of both lipophilic and electronic effects is pointed out.


Metabolic and myogenic factors in local regulation of the microcirculation.

Patterns of flow were recorded from individual capillaries of mesentery and muscle during autoregulation and reactive hyperemia. In cat mesentery at normal arterial pressure capillary blood flow was often periodic in nature. When arterial pressure was reduced periodicity was abolished and in certain cases mean flow increased. Elevation of venous pressure at this time caused restoration of flow periodicity and simultaneously a large fall in mean flow. Vasomotion and autoregulation in mesentery appear to be dependent on intravascular pressure per se. In cat sartorius muscle substantial increase in flow was seen in most capillaries during reactive hyperemia. In certain capillaries the pattern resembled the gross flow pattern while others showed a brief hyperemia and then a period of flow arrest that is presumably due to a strong precapillary vasoconstriction. The latter response is suggestive of a myogenic control while the former may be due to accumulation of metabolites. In frog pectoralis muscle reactive hyperemia was very prolonged in comparison to cat sartorius muscle. The general pattern of flow was consistent with the notion of a strong metabolic control mechanism. The three tissues studied provide examples of strong myogenic, strong metabolic, and combined metabolic and myogenic control of the microcirculation.


Metabolic models of microcirculatory regulation.

The functions and integrity of body tissues are critically dependent on an adequate oxygen supply. Because the transport of oxygen to the cells is intimately linked to the microcirculation, the concept of microcirculation-metabolism coupling has received much attention. In essence, the metabolic theory of intrinsic control of the microcirculation states that microvascular tone is locally modulated to maintain adequate oxygen levels in the parenchymal cells. We propose a two-component control system for the regulation of tissue O2 delivery in accordance with metabolic needs. A precapillary sphincter control mechanism maintains tissue PO2 by governing the number of perfused capillaries. Functional capillary density in turn determines surface area available for diffusion and capillary-to-cell diffusion distance. On the other hand, the arteriolar control system modulates local blood flow in accordance with parenchymal O2 utilization and thereby minimizes changes in capillary PO2 when the O2 availability/demand ratio is decreased. We propose that the precapillary sphincters are more sensitive to changes in tissue PO2 than are the flow-regulating arterioles. Consequently, for mild stresses, adequate tissue oxygenation is maintained mainly by precapillary sphincter control of diffusion parameters without the need for changes in blood flow. However, as metabolic stresses become greater, blood flow regulation becomes the dominant factor in the control of tissue O2 delivery. Thus, by working in concert, the local mechanisms regulating microvascular resistance and effective capillary density provide a wide margin of safety against the development of cellular hypoxia.


Biological interactions between selenium and a number of other elements occur that render selenium much less toxic than when it is present alone. These elements are arsenic, mercury, cadmium, and copper. Furthermore, the presence of selenium reduces the toxicity of mercury and cadmium. These are general biological interactions and have been found to occur in a number of animal species under a variety of conditions. It has been shown that the reaction products of selenium with mercury and cadmium are less toxic than an equal amount of selenium fed alone to chicks. The presence of arsenic shifts the excretion of selenium to the bile. There is no conclusive evidence that the presence of other elements reduces the absorption or retention of selenium. It is possible that some of the interactions are caused by the formation of a compound by selenium and other elements which has less affinity for active groups on biologically active compounds.--Hill, C.H. Interrelationships of selenium with other trace elements.


Pressure regulation in the microcirculation.

The results of direct pressure measurements are described which demonstrate that pressures in a certain fraction of mesenteric capillaries remain remarkably constant during large changes in systemic pressure. The results of isogravimetric studies, reported in the literature, are also described which indicate that this phenomenon may also occur in the intestine. The question is raised whether capillary pressures may therefore be regulated. Pressures recorded from mesenteric arterioles and capillaries are shown which indicate that maintenance of a constant capillary pressure is primarily the consequence of the vascular architecture peculiar to this tissue, and is merely a secondary reflection of mechanisms associated with flow regulation. The results of direct pressure measurements recorded in the microcirculation of intestinal muscle are also shown. These data indicate that capillary pressures in innervated, denervated, and xylocaine-treated intestinal muscle change in direct proportion to variations in arterial pressure. It is concluded that capillary pressures in the intestinal muscle layers are therefore not regulated, so that the observation that capillary pressures may be maintained is probably a phenomenon unique to the mesentery. Pressures recorded from capillaries in the mucosal villi are also shown and compared to capillary pressures measured in the microvasculature of mesentery and intestinal muscle. When systemic pressure was normal (107 +/- 10 mm Hg), capillary pressure in the mesentery averaged 30 to 33 mm Hg; capillary pressures in the intestinal muscle averaged 22 to 24 mm Hg; and capillary pressures in the mucosal villi averaged 13 to 15 mm Hg. These data suggest that mesenteric capillaries are primarily a filtering network; intestinal muscle capillaries are normally in fluid balance; whereas at rest mucosal capillaries are primarily absorptive. These pressures, recorded from the three major regions of the rat intestine, were used to calculate a weighted average for the whole organ. The calculated value, based on assumed values for relative capillary densities, was 17 mm Hg. This result compares favorably with data from whole organ, isogravimetric studies, and may clarify some of the apparent discrepancies between previous isogravimetric and servopressure studies.


On the role of haemopoietic tissue in interstrain differences in radiosensitivity of mice.

Mice of three different strains were classified according to the percentage of survival after a whole-body exposure of 600R as radioresistant (strain C57BL/10), radiosensitive (strain BALB/c) and intermediate (strain BFU). Haemopoietic recovery after a whole-body exposure of 200R was followed in the three groups. It was found that the radiosensitivity of bone marrow and spleen haemopoietic stem cells was equal in all the mouse strains tested. The extent of radiation damage to haemopoiesis, assessed by the changes in nucleated cell counts in bone marrow and spleen, as well as by bone marrow and spleen 59Fe uptake, was smaller and the rate of recovery higher in strain C57BL/10 than in strains BALB/c or BFU. An analogical difference among the strains was also observed during realimentation in the recovery of bone marrow and spleen haemopoiesis inhibited by fasting. The role of the capacity of haemopoietic cell population to repair the induqed haemopoietic damage is discussed and its decrease in BALB/c mice is held responsible for their higher radiosensitivity.


Adherent and non-adherent mouse peritoneal exudate cells.

Administration of proteose peptone to donors of peritoneal exudate cells resulted not only in quantitative increase in the mono-histiocytic cells but also in their activation as judged by their rapid adherence to surfaces, strong acid phosphatase reaction and increased numbers of lysosomes and phagocytic vacuoles. The three-hour adherent fraction of peritoneal exudate cells (from normal as well as peptone-stimulated mice) contains a large number of activated lymphocytes and fibroblastoid cells and is not enriched in macrophages. Moreover, the adherent cells, when collected by scraping, are unsuitable for immediate biological use since they lose the enzymatic equipment. For purification of histiocytes and macrophages by adherence, the 2-day cultivation of peritoneal cells was found to be optimal, as judged by the number of acid phosphatase positive cells. Their decrease after 4-day cultivation may indicate over-growth of the fibroblast and epithelial cell lineages. The non-adherent fraction was found to be enriched in lymphocytes as well as in mast cells; numerous macrophages were observed, too.


Biological characteristics of cell lines derived from the respiratory tract of a bovine foetus.

Bovine foetal cell lines were derived from foetal bovine lung, foetal bovine nose and foetal bovine trachea and kept in serial cultivation for 2 years. During their development they showed marked changes in morphological and growth characteristics of the cells accompanied by changes in the karyotype. The FBL line consisting of fibroblasts as well as the epithelial to epitheloid cultures of FBN and FBT have shown a good capacity to grow in vitro.


Determination of protein levels by quantitative immunoelectrophoresis in rats subjected to continuous irradiation.

Changes in the concentration of some serum proteins are determined by quantitative two-dimensional immunoelectrophoresis in rats subjected to continuous irradiation with 400R/day for 11 days. At this dose rate albumin levels were decreased by 36% and IGG levels by 70% in comparison with the control (C2) values. The concentration of haptoglobin, haemopexin and caeruloplasmin was increased by 75-140% on day 10 as compared with the control (C2) values.


The functional wholes of cells. Nucleolar coefficients, mitoses and multinuclearity in neighbouring HEp-2 cells.

Preparations of HEp-2 cell cultures seeded in concentrations of 120,000 per cm.2 and 15,000 per cm.2 and growing for 15 and 25 hours were studied with respect to local differences in the frequencies of monucleolar cells, seven types of which were distinguished by the values of nucleolar coefficient, multinuclear cells and mitotic cells in the neighbourhood of some cells. The latter exhibited the following features such as mononuclearity with NC ranging from 1 to 4, or mitosis. The observed differences are a) higher between the groups than between the levels of cells, b) higher in the distribution of cells than in the average nucleolar coefficient, c) most frequently in the neighbourhoods of mitotic and mononuclear cell with NC 4, and d) as a rule, complementarily dependent on the initial concentration of cells and the time of cultivation. The term "functional wholes" is proposed to describe those unique formations as well as the ones stated in the introductory remarks. The functional whole is a group of cells of the same type which are identical or closely similar in a variable feature.


Analysis of programmed gene action by means of radiation chimaeras.

Agglutinability by H-2 antisera of neonatal erythrocytes appears early in some mouse strains (such as C57BL/10) and late in others (A, CBA, etc). This might be due to interstrain differences of the postulated temporal genes controlling in this case the developmental expression of H-2 gene products A similarr effect might be simulated by interstrain variation in neonatal size of erythrocytes (possibly reflecting their stage of maturation from the generally larger blasts) which could indirectly affect agglutinability through a different density of H-2 sites on the cell surface. Such correlation was, however, not found in this study. A possible involvement of an extracellular control mechanism was also ruled out by demonstrating that in allogeneic radiation chimaeras created reciprocally from late and early strain mice (used as irradiated recipients and donors of curative cell inocula) the appearance of donor-specific H-2 agglutinability conforms to the donor's timing type being unaffected by the contrasting host's environment. The delayed agglutinability in a late strain was further shown to concern not only neonatal erythrocytes, but also those recruited from stem cells in "adult" bone marrow. Finally, two strains (C67BL/10 and CBA) classified as, respectively, early and late by H-2 agglutinability were typed similarly on the basis of the time course of 59Fe-incorporation rate/mg. spleen in newly formed syngeneic radiation chimaeras. The question whether this coincidence is due to chance or to H-2 linkage of a common or rather separate control mechanisms is discussed.


Effect of continuous irradiation upon bone marrow haemopoietic stem cells in mice.

The effect upon the stem cells in mouse bone marrow of continuous irradiation at different daily exposure rates ranging from 1R--1,000R was investigated by means of the exogenous spleen colony assay. At exposure rates between 1 and 50R/day, a stabilized state in the CFU value becomes established, its level decreasing with the increase in the daily exposure. Exposure rates between 80 and 1,000R/day lead to an initial steep, but later on flatter, decrease in CFU number until death of the animals. Mass deaths occur in mice when the CFU number falls below 5%. The results further delimit the range of exposure doses for which a dependence was found of the CFU number upon the total exposure dose of continuous radiation.


Effect of large doses of turkey herpes virus on antibody response in chickens.

During vaccination of chickens, repeated administration of large doses of a cell-associated turkey herpes virus compared to a single large dose did not affect markedly the virus-neutralizing antibody titre but accelerated the formation of agaroprecipitating antibodies. The formation of antibody types proceeded independently of each other. Health condition of chickens was not influenced.


Capacity of contact cytotoxic reaction to distinguish targets presented by F1 and somatic hybrid cells of the same H-2 parentage.

51Cr-release from prelabelled target cells caused by contact with effector lymphocytes was greater when the H-2d/H-2k cells used for sensitization (i.e. production of effector cells) were also phenotypically similar to the target cells. The contact cytotoxic reaction thus turned out to be sensitive enough to distinguish a putative phenotypic difference between F1 and somatic hybrid cells of the same H-2 parentage. A possible involvement of non-H-2 differences was controlled and ruled out.


Identification of chromosomes in intraspecific hybrid cells by the G-banding method.

The karyotype of HY 5/3--4 hybrid cells formed by fusion of diploid and heteroploid murine cells (LS/BLxR-AG/20) consisted of 60--75 chromosomes (modal number 72). The G-banding technique made it possible to identify the origin of the chromosomes; it is concluded that due to chromosomal segregation the hybrid genome lost preferentially the chromosomes derived from the diploid parental line LS/BL. Even after long-term culture in vitro, the chromosomal complement of hybrid cells retained all bi-armed chromosomes and most of the telocentrics, derived from the heteroploid parental cells.


Early changes in the number of haemopoietic stem cells in irradiated young rats.

The frequency of exogenous haemopoietic spleen colonies in whole-body irradiated rats, aged 4--5 weeks, was found to be linearly dependent upon the dose of inoculated bone marrow or spleen nucleated cells. The recovery rate of the number of haemopoietic stem cells in rats exposed to 300R assessed on day 12 postirradiation was found to be faster in bone marrow than in spleen.


The effect of antilymphocyte globulin (ALG) on precipitin formation to human serum albumin in rabbits.

The effects of pig anti-rabbit ALG on precipitin formation to human serum albumin were studied in rabbits with an aim to contribute to an elucidation of the effects of ALG on the humoral antibody response. Antibody levels were assayed quantitatively after the first, second and third local injection of antigen with adjuvant. ALG was found to suppress distinctly, though not completely precipitin formation throughout the immunization cycle in the experimental group compared to the control groups injected with pig IgG or saline at the same time intervals. The immunosuppressive effect was particularly pronounced after the third injection of antigen which did not further increase antibody levels in experimental animals as compared to the controls. The system appears to be effective in partial suppression of the humoral antibody response when a good preparation with appropriate dosage and timing is used.


Induction of rat pituitary thymidine kinase: another physiological response to oestradiol in the male?

Subcutaneous injection of oestradiol-17beta enhanced the thymidine kinase activity in the anterior pituitary of immature male rats, but did not alter the thymidylate synthetase level. The kinase activity reached a sharp maximum 36 hours after injection of the steroid and then decreased to its original value. The estimated minimum active dose was 0.020 mug per rat. The 17alpha isomer was inactive. Cycloheximide and actinomycin D, according to injection schedule, prevented the rise in activity, suggesting a regulation of pituitary thymidine kinase at the level of protein biosynthesis. The induced enzyme exhibited the same Km and the same thermal inactivation profile as the constitutive enzyme. With the doses of oestradiol required for induction of the enzyme, no significant variations of serum and pituitary LH and FSH concentrations were detected. Based on these and previous results it is suggested that, in the male pituitary, the concentration ratio of circulating oestrogens over androgens controls not only the secretory function but also the production of specific enzymes.


Evaluation of the three-hour oral glucose tolerance test in detection of significant hyperglycemia and hypoglycemia in pregnancy.

Analysis of 2,000 consecutive patients who had a three-hour 50-gm. oral glucose tolerance test done in the third trimester of pregnancy has shown that the three-hour reading was not necessary for the diagnosis of gestational diabetes. It was found that hyperglycemia and hypoglycemia (95th and 5th percentiles, respectively, for plasma glucose levels) were significantly associated with an increased risk for perinatal mortality. Furthermore, hyperglycemia was associated with an increased incidence of large-for-dates placentas and hypoglycemia with small-for-dates infants and small-for-dates placentas. These associations with hypoglycemia were seen to be greatest when this occurred at the three-hour level, and it was concluded that the three-hour measurement should be retained until the clinical significance of hypoglycemia in pregnancy is fully determined.


Metabolic studies in the pyelonephritic rat.

Insulin antagonism characterizes infection, but the mechanism is unknown. Previous studies have been performed during the acute catabolic stage of infection, and the resultant metabolic changes reflect this decreased food intake and weight loss. To delineate metabolic alterations due to infection itself, rats with pyelonephritis induced by tail-vein injection of 1 ml. of Streptococcus faecalis (10(9) bacteria per milliliter) were studied two weeks later during a period of near-normal weight gain and food intake. Fasting growth hormone concentrations (nanograms per milliliter) in the pyelonephritic rats were nearly five times normal (45.8 vs. 9.9). Intra-arterial glucose and insulin tolerance tests were impaired. Early glucose-induced insulin release was depressed. Fat pads from infected rats manifested higher basal lipolysis per cell. Glycerol-mediated gluconeogenesis by liver slices was decreased. This pathway was unaffected by insulin in infected rats but readily inhibited in control rats. The following metabolic parameters were similar in control and infected animals: (in vivo) fasting concentrations of plasma glucose, free fatty acids, triglycerides, total corticoids, creatinine, insulin, glucagon, molar ratios of insulin and glucagon, glucose and insulin responses to tolbutamide, and glucagon and free fatty acid suppression after glucose; (in vitro) glucose metabolism by muscle and fat, epinephrine- and theophylline-stimulated lipolysis and re-esterification by epididymal fat pads, fasting hepatic glycogen content, glucose production by liver slices with and without alanine. No plasma insulin antagonist was found in the infected rats. Metabolic alterations in infected rats can be demonstrated independently of the associated catabolism. Increased growth hormone secretion cannot explain all of these changes.


Insulin metabolism in hypothyroidism.

Insulin resistance has been invoked to explain the glucose intolerance observed in hypothyroid patients. This possibility was studied by determining fractional and metabolic clearances of intravenously administered porcine crystalline insulin (0.1 U./kg.) and its effect on plasma glucose concentration in ten hypothyroid patients, ten normal subjects, and six treated euthyroid patients. Following administration of porcine insulin, serum immunoreactive insulin concentrations during the period of observation were similar in hypothyroid patients, in normal control subjects, and in treated euthyroid patients. Similarly, no significant differences in the mean half-life, distribution space, or fractional and metabolic clearances of insulin were observed among any of the three groups. In response to insulin administration, plasma glucose concentrations declined to the nadir of 36 +/- 4, 43 +/- 3, and 38 +/- 4 mg. per 100 ml. in hypothyroid patients, normal control subjects, and treated euthyroid patients, respectively. Thereafter, plasma glucose steadily increased and approached the baseline value at ninety minutes in normal subjects and treated euthyroid patients. In contrast, the plasma glucose values remained significantly lower than the baseline for the rest of the procedure in hypothyroid patients. The present study demonstrates that there is no evidence of resistance to the action of insulin in hypothyroid patients. The observation of prolonged hypoglycemic action of exogenously administered insulin in hypothyroid patients might in fact suggest increased sensitivity to insulin action. These findings indicate that glucose intolerance of the hypothyroid state is not characterized by insulin resistance.


Antiketogenic action of fructose, glyceraldehyde, and sorbitol in the rat in vivo.

The purpose of this study was to compare the metabolism and antiketogenic properties of fructose, glyceraldehyde, and sorbitol. Fructose, glyceraldehyde, and sorbitol were readily metabolized and exhibited an antiketogenic effect in both blood and liver when injected intramuscularly to starved (forty-eight hours) rats. Sorbitol had the most pronounced antiketogenic effect and produced an 80 to 90 per cent decrease in the blood ketone bodies sixty minutes after administration. Fructose and glyceraldehyde were equally effective and produced about a 60 to 70 per cent decrease in ketone bodies. Fructose, glyceraldehyde, and sorbitol caused a significant decrease in the concentration of hepatic ketone bodies. In liver, sorbitol was found to be most effective in its antiketogenic action. The concentration of plasma free fatty acids remained unchanged after injection of all three antiketogenic substrates. Fructose, glyceraldehyde, or sorbitol caused increased blood lactate and pyruvate concentrations, and fructose was the most effective of the three substrates. Fructose administration resulted in a significant decrease in hepatic lactate/pyruvate and beta-OH-butyrate/acetoacetate concentration ratios, whereas sorbitol caused an increase in the concentration ratio of these two substrat pairs. Decreases in blood and liver ketone body levels were associated with lowering of liver acetyl-CoA concentration . However, the decrease in hepatic acetyl-CoA produced upon the administration of antiketogenic substrates was not pronounced. Sorbitol administration resulted in the most pronounced increase in hepatic alpha-glycerophosphate concentration. Fructose or glyceraldehyde also caused an increase in alpha-glycerophosphate content. Administration of each of the three antiketogenic substrates produced an increase in hepatic dihydroxyacetone phosphate concentration. All three antiketogenic compounds increased liver glycogen and blood glucose concentrations. No significant changes were observed in hepatic ATP, ADP, or AMP concentrations sixty minutes after the injections of any of the antiketogenic substrates. Although decreased liver acetyl-CoA levels were associated with the antiketogenic effects of the compounds tested, the increased liver alpha-glycerophosphate content best explains the differences between fructose or glyceraldehyde and sorbitol.


Cardiac changes during 2-deoxy-d-glucose test. A study in patients with selective vagotomy and pyloroplasty.

2-Deoxy-D-glucose (2-DG), a potent vagal stimulator, is used for testing the completeness of surgical vagotomies. 2-DG has however not yet been employed routinely for the postoperative testing of bagotomized patients because of possible cardiotoxic effects. In a previous study, we have shown the effects of 2-DG on the resting ECG in 48 patients. We did not observe any significant changes besides nonspecific T wave flattening and QT prolongation. The present investigation included 37 additional patients. In 27 patients, a continuous ECG tape recording for detection of arrhythmias was performed and in 10 patients, the systolic time intervals for non-invasive assessment of the myocardial contractility was recorded before and after 2-DG. Serious arrhythmias were not provoked by 2-DG. Premature contractions were, as a whole, not incrased by the drug. 2-DG did not have any cardiodepressive action. On the contrary myocardial contractility evaluated by the systolic time intervals was enhanced.


Gastric acid responses to graded vagal stimulation in the anaesthetized cat.

Graded acid secretory responses were obtained in anesthetized cats with intact pyloric region when the frequency of electrical stimulation of the neck vagi was increased from 0.5 to 8 Hz; 4-8 Hz induced maximal acid secretion. Antrectomy reduced the response to vagal stimulation but the reduction reached statistical significance only after additional resection of the duodenum. Administration of per se subthereshold or threshold doses of exogenous gastrin restored the acid response to vagal stimulation in antrectomized cats. Combination of a high dose of histamine and vagal stimulation in antrectomized output in anaesthetized cats which roughly amounted to two thirds of the reported maximal acid secretory rate during histamine stimulation in non-anaesthetized cats. The present results suggest that the physiological vagal discharge rate to the acid secreting glands does not reach higher frequencies than aroung 8 Hz. It is also suggested that an intact antroduodenal region is essential for acid responses to physiological vagal stimulation in the cat. Finally, the reduced sensitivity to gastric secretory stimulants under anaesthesia appears to be due in part to a reduced vagal tonus.


Psychosomatic aspects of defecation and its disturbances.

Constipation and diarrhea are usually defined in terms of frequency of defecation and consistency of stool. However, the subjective component in these symptoms is so great that the objective criteria cannot be satisfactory. An important additional aspect is provided by the analysis of sensations and postures in the three phases of the act of defecation (announcing, emptying, subsequent phase). Taking account of the subjective aspects may contribute to a better differentiation and classification not only of constipation and diarrhea but also of such disturbances as organic rectal disease and irritable colon.


Chronic pancreatitis, alcoholic liver cirrhosis and salivary secretion.

By comparison with control subjects, patients with chronic pancreatitis and alcoholic liver cirrhosis displayed in basal condition a reduction in salivary volume, bicarbonate and an amylase concentration and outputs. These reductions disappeared during stimulating by chewing parafilm. It is concluded that these reductions are not the consequence of a specific process affecting both the salivary glands and the pancreas. It is suggested that they are more likely due to a decreased sensitivity of the salivary nervous control than to an actual alteration in the secretory capacities.


75Se-selenomethionine excretion in bile and pancreatic juice.

The radioactivity in the bile and pancreatic juice of two patients was measured after intravenous injection of 75Se-l-selenomethionine. The concentration of radioactivity in both fluids is almost equal. Thin-layer chromatography shows that the main part of radioactivity in pancreatic juice is due to unbound 75Se-selenomethionine, while free 75Se-selenomethionine is only a minor part of the total radioactivity appearing in the bile.


Effects of secretin and cholecystokinin on motor activity of human jejunum. A radiotelemetering study of jejunal motility during secretin and cholecystokinin intravenous infusion.

The effects of intravenous administration of secretin and cholecystokinin (CCK) on motility of the human jejunum were investigated with pressure-sensitive radiotelemetering capsules. Secretin inhibited and CCK stimulated the spontaneous motor activity of this part of the small intestine. Raising the dosages of the two hormones caused increasing effects. It seems likely, therefore, that both hormones play a role in the regulation of intestinal motility in man.