Phosphonoacetic acid-resistant mutants of herpes simplex virus: effect of phosphonoacetic acid on virus replication and in vitro deoxyribonucleic acid synthesis in isolated nuclei. Phosphonoacetic acid (PAA) inhibits the replication of herpes simplex virus in BSC-1 cells and the in vitro synthesis of deoxyribonucleic acid (DNA) in isolated nuclei. Phosphonopropionic acid at a concentration of 100 mug/ml had no effect on herpes simplex virus replication. PAA-resistant mutants were obtained at a rate of 1 in 10(4) plaque-forming units after 5-bromodeoxyuridine mutagenization of the virus. These mutants replicate in BSC-1 cells in the presence of 100 mug of PAA per ml and induce a PAA-resistant DNA polymerase that synthesizes DNA in vitro in the presence of PAA.


Inhibition of herpes simplex virus strains isolated from herpetic keratitis by polyinosinic acid-polycytidylic acid. Fifty strains of herpes simplex virus, isolated from patients with herpetic keratitis, were examined in vitro for susceptibility to polyinosinic acid-polycytidylic acid [poly(I:C)] in the presence of a constant concentration of diethylaminoethyl-dextran. The minimal inhibitory concentration of poly(I:C) for 44 of these strains ranged from 0.0001 to 0.1 mug/ml; for the remaining six strains, the minimal inhibitory concentration stood at 1 to 2 mug/ml. Fifteen isolates from primary infections were more susceptible to poly(I:C) than 35 isolates from recurrent infections. Isolates acquired at different points of a given clinical episode showed similar susceptibilities to poly(I:C). In two patients, isolates from consecutive recurrences of infection exhibited reduced susceptibilities. The implications of the above observations for the therapeutic use of poly(I:C) are discussed.


Induction of bacteriocins from Clostridium perfringens by treatment with mitomycin C. Six of ten bacteriocinogenic strains of Clostridium perfringens were inducible by treating the cultures with mitomycin C. Induction did not occur in the continued presence of mitomycin C but only when excess mitomycin C was removed. Cell death was associated with induction of bacteriocin.


Cyclic adenylic acid-dependent and -independent production of chloramphenicol acetyltransferase in Escherichia coli carrying R plasmids. A plasmid-specified, inducible, but cyclic adenylic acid (cAMP)-independent resistance to chloramphenicol (CM) is reported. The resistance level to CM was increased two- to fourfold by treatment with a small amount of CM for a short period in Escherichia coli strains carrying the R plasmid pJY1, which was obtained from a clinical isolate of Vibrio cholerae. Though cAMP was required for production of CM acetyltransferase (CATase) in cAMP-deficient mutants of E. coli carrying the R100 plasmid, the same species harboring pJY1 did not require cAMP for production of the enzyme. The possibility of a mechanism other than CATase activity for the CM resistance conferred by R plasmids is discussed.


Inactivation of poliovirus in wastewater sludge with radiation and thermoradiation. Sludge is highly protective of poliovirus against both heat and ionizing radiation. This protection could be largely overcome when these two treatments were given simultaneously.


Hutchinson's Archives of Surgery revisited. I present a bried review of Hutchinson's Archives of Surgery. I have listed most of his well-known and not-so-well-known early clinical descriptions. To this list can be added descriptions of Peutz-Jegher syndrome, lipomelanic reticulosis, telangiectatic lupus erythematosus, temporal arteritis, and recurrent herpes simplex on the thigh and buttocks. The characteristics of Hutchinson's writing are the use of catchy, descriptive terms, his use of patients' names for diseases, his belief in the effectiveness of illustrations, superb clinical observation but faulty conclusions, and a lack of humor. Seventy-two years after his death, there is still much that can be learned about clinical disease from Hutchinson.


Pulmonary involvement with cytomegalovirus infections in children. Among 40 hospitalized infants and children with cytomegalovirus infection, 14 (35%) had interstitial pneumonitis, 4 (10%) had wheezing or tachypnoea but without x-ray evidence of classical interstitial pneumonia, the remaining 22 (55%) were free of pulmonary involvement. Most patients had tachypnoea and nonproductive cough of varying durations: those with underlying pulmonary pathology tended to have persistent and prolonged respiratory symptoms. Mortality and severity of the lung disease were related to the underlying immunodeficiency or concomitant pulmonary process.


Intestinal damage in rotavirus and adenovirus gastroenteritis assessed by d-xylose malabsorption. The absorption of D-xylose infused into the duodenum was assessed in infants with acute gastroenteritis. 1-hour blood-xylose levels were low in 6 patients found to harbour rotavirus in the small intestinal aspirate. Normal levels (greater than 1-26 mmol/l) were obtained in the absence of virus particles in the small intestine in a further 6 patients: in 3 of these adenovirus or rotavirus was recovered from the stools. Three patients with adenovirus in the small intestinal juice and ill with acute gastroenteritis also had low xylose levels. This finding supports earlier epidemiological studies that adenovirus may be a causative agent of acute infantile gastroenteritis. The association between virus in the small intestine and xylose malabsorption may indicate mucosal damage. Formal proff of this is awaited.


Variations in adrenocortical responsiveness during severe bacterial infections. Unrecognized adrenocortical insufficiency in severe bacterial infections. Plasma cortisol levels and their response to .25 mg synthetic A.C.T.H. (Cortrosyn) were studied in 26 septic patients. Four (15.4%) of these patients appeared to have greatly increased adrenocortical activity with plasma cortisol levels averaging 65.4 +/- 14.8 microgram/dl (normal = 8-18 microgram/dl. All four of these patients were agonal and died within five days. Seventeen (65.4%) of these 26 patients appeared to have an appropriate adrenocortical response to severe infection in that their plasma cortisol levels increased (averaging 19.2 +/- 6.0 microngram/dl) following synthetic A.C.T.H. The remaining five patients, who constituted 19.2% of the 26 patients studied, appeared to have some impairment of adrenocortical function. In spite of severe bacterial infections and no history to support Addison's disease, their plasma cortisol levels (averaging 13.8 +/- 3.3 microgram/dl) were not increased above normal and their response to Cortrosyn was much less than would be expected; the increase in plasma cortisol levels in these patients following the synthetic A.C.T.H. averaged 1.1 +/- 3.6 microgram/dl. It is reemphasized that patients with severe sepsis who are not responding adequately to standard therapy should be suspected of having adrenocortical insufficiency and treated accordingly.


Regional pancreatectomy: en bloc pancreatic, portal vein and lymph node resection. Eighteen patients are reported who have had a regional pancreatectomy. The pancreatic segment of portal vein was excised with en bloc total pancreatectomy and regional lymph node dissection in all 18. Venous repair was by end-to-end anastomosis without a graft. Five of the 18 also had various arterial resections and reconstructions. Sixteen of the 18 had been explored and deemed nonresectable elsewhere. This operation has doubled the resectability rate in this institution. The 30-day operative mortality rate was 16.6%. Acurarial survival is 62% at one year compared with 36% one year survival rate for patients undergoing pancreaticoduodenectomy for less advanced cancer in previous years. A more valid comparison would be between those who had a palliative procedure since most patients in the present series were initially considered unresectable. One year survival for these patients was 22%. The quality of life was good for most patients.


The beta-adrenoceptors responsible for a direct relaxation of the uncontracted nictitating membrane of the anaesthetized cat. Direct relaxation of the nictitating membrane of the anaesthetized cat was produced by close intra-arterial injections of single doses of isoprenaline and other beta-adrenoceptor agonists. This response was relatively resistant to blockade by doses of propranolol that almost abolished the corresponding positive chronotropic and vasodepressor responses. Construction of dose-response curves, however, revealed antagonism of all three parameters by propranolol and different sensitivities of the parameters to isoprenaline. The absence of blockade by practolol, although negative evidence, permitted the nictitating membrane response to be classified as a beta2-adrenoceptor effect, which was supported by the full agonist activity of salbutamol. The blockade of the nictitating membrane response by propranolol was however weaker than of the beta2-adrenoceptor mediated vasodepressor response. Contractile responses were observed with higher doses of isoprenaline and after intravenous injection. These were probably indirect effects induced by local changes in blood pressure at the nictitating membrane due to the concommitant vasodepressor response.


Influence of apomorphine on sympathetic neural transmission in the nictitating membrane of the cat. Apomorphine inhibited the nictitating membrane contractions elicited by pre- or postganglionic stimulation of the cervical sympathetic nerve but did not influence even in higher doses the effect of noradrenaline and adrenaline on this organ. The inhibition evoked by apomorphine could be antagonized by haloperidol but was not influenced by phentolamine and propranolol. Dopamine and noradrenaline inhibited the nictitating membrane contractions elicited by nerve stimulation, but clonidine was ineffective in our experiments. Haloperidol antagonized the inhibition evoked by dopamine but did not influence the similar effect of noradrenaline.


Pharmacology of the hemodynamic and myocardial response to occlusion of the mesenteric artery in the cat. Occlusion of the superior mesenteric artery in the cat produces increases in heart rate, systemic pressure, cardiac output and muscle blood flow. Total peripheral resistance is reduced and regional resistance in skin increased. Blockade of the right and left cardiac nerves with xylocaine lowered basal heart rate and reduced cardiac output. The increase in heart rate produced by occlusion of the artery (MAO) was prevented by blockade of the stellate ganglia and cardiac nerves. Propranolol lowered basal heart rate and cardiac output and subsequently blocked the increase in both parameters during MAO. The increase in muscle blood flow was also blocked resulting in an increase in total resistance. We conclude that, in the cat, MAO reflexly increases myocardial tone and enhances venous return from muscle, the latter by a reflex beta adrenergic vasodilation. Both factors provide for the increase in cardiac output. A reflex beta adrenergic vasodilation in muscle, evoked by vasoconstriction in the mesenteric bed, may contribute to survival in the hemorrhaged animals, since capillary exchange in muscle may be increased by this reflex and fluid shifted from the extra--to the intravascular compartment.


Primary and secondary cytomegalovirus infection. Forty-seven patients who underwent renal transplants were followed clinically and were examined for serologic or virologic evidence of cytomegalovirus (CMV) infection. There were 18 cases of primary infection and ten cases of secondary infection. These findings were based on whether the patient was seronegative or seropositive prior to transplantation. Thirteen patients with primary infection and only one patient with secondary infection had two or more of the following manifestations that are temporally associated with laboratory evidence of infection: fever, leukopenia, atypical lymphocytes, lymphocytosis, hepatosplenomegaly, myalgia, arthralgia, and pneumonitis. Five patients with primary infections, one of whom died with disseminated disease, were recognized by attending physicians as having CMV disease. Since primary infection is though to be largely due to virus transmitted by the kidney of a seropositive donor, it may be possible to prevent symptomatic primary infection by using only seronegative donors for seronegative recipients.


Psychoses precipitated by psychotomimetic drugs. A follow-up study. Fifteen patients who developed prolonged psychotic reactions following psychotomimetic drug use (probably primarily LSD) were followed up 1.9 to 5.8 years later. Two patients had committed suicide. Approximately half of the patients had a relatively good outcome and half did poorly. Aspects of the initial clinical picture that correlated with outcome measures are discussed. The possibility is considered that vulnerability to a prolonged psychotic reaction following psychotomimetic drug use may be related to a genetic vulnerability to illnesses in the manic-depressive/schizo-affective spectrum. In some instances this vulnerability may implicate central serotonergic neuronal systems.


Cytochemical studies of respiratory enzymes in carcinoma of the cervix uteri after irradiation by radium and cobalt 60. Enzymological studies of succinic dehydrogenase and cytochrome oxidase were made on human squamous cell carcinoma of the cervix uteri before, during and after radiation therapy by radium and 60CO. The enzymatic activities of the two enzymes were higher in cells of stage III than in the corresponding cells of stage II. In addition, after treatment, no enzymatic activity could be detected. These changes may be explained on the basis of mitochondrial damage as a result of radiation effect.


Elimination of Proteus mirabilis 51Cr endotoxin from the liver in rats. Using isotope methods, elimination of the endotoxin of Proteus mirabilis labeled with chromium (CrEPm) from the liver of rats was studied. The following studies were carried out: intravital exploration of the liver with a scintillation probe, measurements of radioactivity of organs and excreted urine and stools, scrintigraphy of the liver, binding of CrEPm by subcellular fractions of hepatocytes, and the influence of selected drugs (polymyxin and hydrocortisone) on elimination of CrEPm from the liver and organelles of hepatocytes.


Mallory bodies compared with microfilament hyperplasia. The binding characteristics of Mallory bodies (MBs) induced in livers of mice fed a long-term diet of griseofulvin were compared with the hyperplastic microfilaments induced by chronic phalloidin injections and with the MBs derived from livers of alcoholic patients at autopsy. Using the indirect immunofluorescence technique, smooth muscle antibody was shown to bind the hyperplastic microfilaments but not the human or mouse MBs. Peroxidase-antiperoxidase was shown to bind the human and mouse MBs but not the hyperplastic microfilaments. The evidence supports the idea that MBs are not composed of microfilaments (actin). The affinity of mouse and human MBs for peroxidase-antiperoxidase suggests that the MBs from these two species are chemically similar.


Superficial cystic gastritis with alopecia. A forme fruste of the Cronkhite-Canada syndrome. The Cronkhite-Canada syndrome is a rare and often fatal condition. A case of superficial cystic gastritis was suspected on peroral gastric biopsy and was later confirmed at autopsy. The selective involvement of the stomach together with the presence of alopecia leads us to conclude that this case is unique and represents a forme fruste of the Cronkhite-Canada syndrome. A complicating feature was the presence of an opportunistic cytomegalovirus infection of the gastrointestinal tract and lungs.


C bodies and R bodies in the epithelial cells of normal and diseased human rectum. Small spherical, extracellular bodies (C bodies) measuring 20 to 80 nm in diameter and enclosed by a trilaminar membrane 7 nm wide surrounded the microvilli of human rectal epithelial cells. Similar bodies were observed within membrane-bound vesicles (R bodies) in the apical cytoplasm. The C bodies associated with the mature epithelial cells of the luminal surface were more numerous than those associated with the immature cells of the crypts, where the R bodies predominated. A decreased number of C bodies was observed in ulcerative colitis. We concluded that the membrane-bound C bodies surrounding the microvilli were cellular in origin and arose from R bodies within the cytoplasm of the rectal epithelial cells.


Fine structure of Sertoli and Leydig cells in azoospermic human testis. Testicular biopsy specimens taken from seven patients with azoospermia were studied by electron microscopy. We considered the possibility of a relationship between the annulate lamellae in the Sertoli cells and azoospermia. In the Leydig cells, we observed intranuclear and intracytoplasmic paracrystalline inclusions and considered their relationship with the Reinke crystalloid.


Tendon reflex latencies in the upper extremity. This study was aimed at establishing a baseline for the latency values of the biceps-brachii, triceps-brachii and brachioradialis tendon reflexes from the moment of elicitation by percussion of the tendon, to the appearance of the muscle action potential. The percussion hammer triggered the electronic sweep on the electromyograph oscilloscope, showing latency to the onset of the potential. Standard skin electrodes were used. Certain precautions were taken to avoid artifacts caused by skin movement. Thirty normal volunteer subjects, between the ages of 17 and 55 years, were examined. Biceps reflex latency values varied between 9 and 14 msec; with a mode of 11 msec and an average of 11 msec. Triceps latency values varied between 9 and 13 msec; with a mode of 11 msec and an average of 10 msec, and brachioradialis latency values varied between 13 and 20 msec, with a mode of 16 msec and an average of 16 msec.


Bacteriology of the human biliary tract and the duodenum. Using the modern anaerobic transport media and meticulous culture techniques, 74 patients undergoing biliary tract surgery were studied. The biliary system was found to be sterile in 58 patients (78%). Fifteen patients had 35 isolates of aerobic and facultative bacteria. The most common ones were Klebsiella, Enterococcus, and Escherichia coli. The only anaerobe isolated was Clostridium perfringens. Eight of 17 patients (47%) with acute cholecystitis and five of 49 patients (10%) with chronic cholecystitis, harbored bacteria in the biliary system. This study suggests that anaerobes are rare in the human biliary system; therefore, if antibiotic therapy is considered, aerobic coverage should suffice.


Chromatography by silica-gel chromatobars. Prospects of its application in the field of forensic toxicology. A new type of chromatographic column of silica gel 20 mm in diameter and 180 mm long has been investigated in the field of forensic analytical toxicology. In order to evaluate the usefulness of chromatobars in isolating organic compounds from biological extracts, sodium phenyl-ethyl-barbiturate, atropine, morphine and strychnine were added to the aqueous extract obtained with Stas-Otto's isolation method. Afterwards the residues of the acidic ether extract, alkaline ether extract and alkaline chloroform extract, were chromatographed on chromatobar. This chromatographic device seems able to avoid the drawbacks and limitations of paper and thin-layer chromatography, mainly from the preparative aspect. The method is particularly advantageous in analysing the whole extract obtained with the Stas-Otto process, without bringing about its partition, especially when slight traces of toxic substances are present.


[Acute leukemia and generalized cytomegaly in a 10-year-old boy]. A case of acute myelomonocytic leukemia in a 10-year-old boy who received massive cytostatic therapy is described. At autopsy a picture of therapeutically determined pathomorphism and cytostatic disease with generalized cytomegalia and terminal staphylococcal sepsis was revealed. Multiple thrombosis of major venous trunks, necroses of the cecum and subtotal necrosis of the spleen were observed; this apparently was associated with generalized cytomegalia and massive cytostatic therapy. Peculiar features of the case were dystrophic changes in cytomegalic cells subjected to necrobiosis, which, probably, was due to cytostatic therapy.


Recurrent encephalitis with elevated titers for herpes simplex. An 8-year-old boy was seen on two occasions 14 months apart with signs and symptoms of encephalitis. The clinical course and laboratory studies were highly suggesive of herpes simplex infection during both episodes.


Diagnostic value of crossed vs uncrossed acoustic reflexes: eighth nerve and brain stem disorders. Acoustic reflexes to crossed (sound in one ear; probe in opposite ear) and uncrossed (sound and probe in the same ear) stimulation were obtained from 17 patients with eighth nerve or brain stem disorders and 20 normal control subjects. Comparison of crossed and uncrossed reflex thresholds provided an important diagnostic tool for differentiating the eighth nerve from the brain stem site. Egihth nerve disorders were usually characterized by diagonal reflex patterns. In contrast, brain stem disorders were usually characterized by horizontal reflex patterns. Detailed evaluation of the reflex time course in two selected patients with intra-axial brain stem disorder showed unusual abnormalities in the crossed acoustic reflex.


The fine structure of lipofuscin in the human inner ear. Lipofuscin inclusions in the human membranous labyrinth were studied by electron microscopy. Lipofuscin is morphologically an irregularly shaped, membrane-bound inclusion consisting of an electron-dense structure. The most common component was a fine, granular, osmiophilic substance which was always associated with a homogenous, spherical structure resembling a lipid droplet. The combination of these two components was frequently observed in the human inner ear. Distended inclusions containing lipofuscin components were also observed within the supporting cells, saccular, utricular and ampullar wall, the epithelial cells of the transitional zone and in the dark cells. Lipofuscin is closely associated with lysosome and is known to accumulate in the tissue as a result of aging. The high lysosomal activity possibly may result in lipofuscin formation in the human inner ears. Also some other unknown metabolic conditions may provide the deposits of lipofuscin.


Increased responsiveness of heart rate to beta-adrenergic stimulation in cold-adapted rats. The responsiveness of heart rate of cold-adapted rats (10-12 weeks, 5degrees C.) to a specific beta-adrenergic agonist was studied. Two hours after removal from cold, s.c. administration of  l-isoproterenol (4, 8, or 16 microng/kg body weight) to female rats was accompanied by a greater responsiveness in cold-treated animals. The increase in heart rate of cold-treated rats above that of warm-adapted controls was related linearly to the dose of isoproterenol administered. An increased sensitivity of heart rate to isoproterenol (8 microng/kg body weight, s.c.) was also observed in male, cold-treated rats. The results suggest that adaptation to cold air is accompanied by increased beta-adrenergic responsiveness in rats.


Actions of cholinergic agonist and antagonists on the adrenocortical response of basal, hypoxic, and hypercapnic rats. Cholinergic neuronal influences on the function of male rat's hypothalamo-hypophyseal-adrenocortical (HHA) system both during basal and stressful situations (hypoxia and hypercapnia) were  investigated using a cholinergic agonist (eserine) and  antagonists (atropine, methyl atropine, mecamylamine and 4-(1-naphthylvinyl) pyridine). The results indicate that the transmitter, acetylcholine (ACh), plays a partial role in the regulation of the HHA system. The muscarinic (m) effects of ACh were stimulatory peripherally and inhibitory centrally. The nicotinic (n) effects were stimulatory and possibly affected the HHA system by inhibiting the central m-inputs. The cholinergic regulation of the HHA system for both non-stressed and hypercapnic animals is probably mediated via a common nm-cholinergic pathway.


Adenylate kinase 2, a mitochondrial enzyme. The subcellular compartmentalization of the isoenzymes of ATP:AMP phosphotransferase (adenylate kinase) was analyzed in HeLa cells, RAG cells, and RAG-human hybrids that express human AK-2. In HeLa cells and in the hybrids, human AK-2 was present in a mitochemical fraction prepared from cell extracts and in mitochondria purified by density gradient centrifugation. Human AK-1 was, as expected, distributed in the soluble cytoplasmic fraction of the cells. The rodent isozymes which are homologous to human AK-1 and AK-2 have been determined.


Rabbit tissue peptidases that hydrolyse the peptide hormone bradykinin. Differences in enzymic properties and concentration in rabbit tissues. The distribution and properties of neutral peptidases acting on the peptide hormone bradykinin (Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg) were determined in several rabbit tissues. The supernatant and particulate fractions prepared from tissue homogenates (25000g for 60min) were studied. Bradykinin inactivation (kininase activity) was measured by bioassay with the isolated guinea-pig ileum. The sites of peptide-bond cleavage were determined in the amino acid analyser, which permits detection and measurement of amino acids and peptides derived from bradykinin. The results indicate that kininases are present in a wide range of concentrations in different tissues, kidney and lung having the most activity. Kininases present in different tissues were distinguished on the basis of sensitivity to the effects of EDTA, dithiothreitol and ZnCl2 and by the site of peptide-bond hydrolysis in bradykinin.


Generation of hydrogen peroxide, superoxide and hydroxyl radicals during the oxidation of dihydroxyfumaric acid by peroxidase. 1. Dihydroxyfumarate slowly autoxidizes at pH6. This reaction is inhibited by superoxide dismutase but not by EDTA. Mn2+ catalyses dihydroxyfumarate oxidation by reacting with O2 leads to to form Mn3+, which seems to oxidize dihydrofumarate rapidly. Cu2+ also catalyses dihydroxyfumarate oxidation, but by a mechanism that does not involve O2 leads to. 2. Peroxidase catalyses oxidation of dihydroxyfumarate at pH6; addition of H2O2 does not increase the rate. Experiments with superoxide dismutase and catalase suggest that there are two types of oxidation taking place: an enzymic, H2O2-dependent oxidation of dihydroxyfumarate by peroxidase, and a non-enzymic reaction involving oxidation of dihydroxyfumarate by O2 leads to. The latter accounts for most of the observed oxidation of dihydroxyfumarate. 3. During dihydroxyfumarate oxidation, most peroxidase is present as compound III, and the enzymic oxidation may be limited by the low rate of breakdown of this compound. 4. Addition of p-coumaric acid to the peroxidase/dihydroxyfumarate system increases the rate of dihydroxyfumarate oxidation, which is now stimulated by addition of H2O2, and is more sensitive to inhibition by catalase but less sensitive to superoxide dismutase. Compound III is decomposed in the presence of p-coumaric acid. p-Hydroxybenzoate has similar, but much smaller, effects on dihydroxyfumarate oxidation. However, salicylate affects neither the rate nor the mechanism of dihydroxyfumarate oxidation. 5. p-Hydroxybenzoate, salicylate and p-coumarate are hydroxylated by the peroxidase/dihydroxyfumarate system. Experiments using scavengers of hydroxyl radicals shown that OH is required. Ability to increase dihydroxyfumarate oxidation is not necessary for hydroxylation to occur.


Characterization of serum lipoproteins of the shark Centrophorus squamous. 1. Blood serum from the shark Centrophorus squamosus (Bonnaterre) was shown to contain VLD (very-low-density), LD (low-density) and HD (high-density) lipoproteins. 2. In shape, size and general physical properties, these lipoproteins were very similar to those described for other animals. The VLD lipoproteins were the major components of the mixture, and HD lipoproteins were present at the lowest amount. 3. In addition to the usual lipid components, the shark lipoproteins also contain substantial amounts of hydrocarbon, probably mainly squalene, and monoalkyldiacylglycerols. Only trace amounts of wax ester were detected. 4. The protein moiety of the VLD and LD lipoproteins contained a component which, in its solubility and electrophoretic properties, molecular weight and amino acid composition, resembled the B apolipoprotein of man and other mammals. This accounted for a large part of the total shark apolipoprotein. 5. There were also present smaller amounts of proteins which were soluble in 8M-urea. In their electrophoretic mobility on basic polyacrylamide gel, some of these were like the A and C apoproteins of man. 6. The electrophoretic distribution of the soluble proteins from the VLD and LD lipoproteins resembled that in higher mammals, but in the HD lipoproteins the similarity was less.


A comparison of the association of yeast phosphoglycerate mutase (EC2.7.5.3) with that of haemoglobin. An ultracentrifuge study. 1. Previous work showed that yeast phosphoglycerate mutase (EC 2.7.5.3) has a mol.wt. of between 107000 and 110000. Preliminary examination showed that at dilutions less than 0.1 g/1 the enzyme dissociated into its subunits. 2. This dissociation was quantitatively examined by both equilibrium and velocity centrifugation. 3. The mathematical analysis of the equilibrium records was tested against oxyhaemoglobin in a variety of ionic strengths and at two temperatures. 4. The estimated L2,4 (interaction coefficient) for oxyhaemoglobin generally agreed with published values except at 6 degrees C in 0.9 M-NaCl, when it was 2.5 times larger than the published value. 5. Statistical analysis of ultracentrifugal-equilibrium experiments showed that the predominant reaction for phosphoglycerate mutase was monomer in equilibrium tetramer, to give an L1,4 of 40.3+/-23.4 (S.D.)1(3)-g(-3) at 20 degrees C. Decreasing the temperature decreased the association to given an enthalpy of between 40 and 60kJ/mol. 6. Analysis of velocity experiments carried out with concentrations varying from 0.3 to 17 g/1 gave an L1,4 of 3111(3)-g(-3). Incorporating errors from estimating S20,w into the analysis showed that this estimate could range from 893 to 1421(3)-g(-3). 7. The concentration-dependence of S20,w was 0.95 litre-g-1 and s020,w for the tetramer was 66.9ps. 8. These results are discussed in relation to the activity of the enzyme.


The reduction of Pseudomonas cytochrome c551 oxidase by chromous ions. The reduction  of cytochrome c551 oxidase from Pseudomonas aeruginosa by Cr2+ ions was followed in the stopped-flow apparatus at a number of wavelengths. The c-haem reduction proceeded in a biphasic fashion with second-order rate constants of 2.6 X 10(5)M-1-S-1 and 4.8 X 10(4)M-1-S-1 at 25 degrees C, whereas the biphasic reduction of the d1-haem appeared to be independent of reductant concentration with rate constants of approx. 1.0S-1 and 0.25S-1 respectively. The kinetically determined difference spectra (reduced minus oxidized) for the c- and d1-haems are presented.


Aldehyde dehydrogenase in 2-acetamidofluorene-induced rat hepatomas. Ontogeny and evidence that the new isoenzymes are not due to normal gene de-repression. The pre- and post-natal ontogeny of Sprague-Dawley rat liver aldehyde dehydrogenase [aldehyde-NAD(P)(+) oxidoreductase, EC 1.2.1.5] is described. At no time in its ontogenetic development does normal liver aldehyde dehydrogenase exhibit any of the characteristics of a series of unique aldehyde dehydrogenases that can be isolated from 2-acetamidofluorene-induced rat hepatomas. Enzyme activity is first detectable in 15-day foetal liver and gradually increases throughout pre- and post-natal development until adult activities are attained by day 49 after birth. Electrophoretically, normal aldehyde dehydrogenase, throughout its ontogeny, exists as the same single isoenzyme found in normal adult liver. Isoelectric points for two normal liver isoenzymes demonstrable by isoelectric focusing are pH5.9 and 6.0. The immunochemical properties of aldehyde dehydrogenase during its ontogeny are identical with those of normal adult liver aldehyde dehydrogenase when tested against anti-(hepatoma aldehyde dehydrogenase) serum in Ouchterlony double-diffusion tests. The results indicate that the hepatoma-specific aldehyde dehydrogenases are not the result of the de-repression of genes normally repressed in adult rat liver or in some other adult tissue.


Adenosine as a constituent of the brain and of isolated cerebral tissues, and its relationship to the generation of adenosine 3':5'-cyclic monophosphate. 1. Adenosine was determined in rapidly frozen rat and guinea-pig brain and in guinea-pig cerebral tissues after incubation in vitro. Adenosine concentrations were approx. 2nmol/g wet wt. in frozen tissue, diminished at room temperature, and returned to 2nmol/g on incubation in oxygenated glucose/salines. 2. Superfusion with noradrenaline then increased the tissue's adenosine concentration 2.5-fold, and hypoxia caused an 8-fold increase. 3. Electrical stimulation alone or in the presence of noradrenaline or histamine increased the tissue's adenosine and cyclic AMP, but adenosine concentrations reached their peak later and were maintained for longer than those of cyclic AMP. 4. Superfusion with l-glutamate with and without electrical excitation raised adenosine concentrations to 15-34nmol/g. The increases in cyclic AMP on electrical stimulation, superfusion with glutamate or a combination of these treatments were diminished by addition of adenosine deaminase or theophylline. 5. It is concluded that adenosine can be produced endogenously in cerebral systems, in sufficient concentrations to accelerate an adenosine-activated adenylate cyclase, and by this route can contribute to the cerebral actions of electrical stimulation and of the neurohumoral agents. In certain instances cyclic AMP as substrate contributes to an increase in adenosine.


Endogenous phosphorylation of microsomal proteins in bovine corpus luteum. Tenfold activation by adenosine 3':5'-cyclic monophosphate. Free ribosomes and a smooth-microsomal fraction were prepared from bovine corpus luteum. Both preparations will self-phosphorylate when incubated with Mg(2+) and ATP, but at low concentrations of Mg(2+) and ATP the self-phosphorylation of the smooth-microsomal fraction was much more dependent on cyclic AMP than was that of free ribosomes, stimulation by the nucleotide being up to 10-fold in the former case. The self-phosphorylation of the smooth-microsomal fraction was studied further. The reaction bears similarities to that brought about by soluble cyclic AMP-dependent protein kinase, being inhibited by Ca(2+) and the heat-stable inhibitor protein from skeletal muscle. Cyclic GMP will activate the reaction at concentrations higher than those required for full activation by cyclic AMP. In the presence of cyclic AMP, phosphate bound to protein is found almost exclusively as phosphoserine. Several proteins are phosphorylated, as judged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, and the phosphorylation of all of them is markedly stimulated by cyclic AMP. If the reaction is carried out at high concentrations of Mg(2+) and ATP, a distinct cyclic AMP-independent phosphorylation is observed. This activity is not inhibited by the heat-stable inhibitor protein, and phosphate is found esterified with both threonine and serine residues.


Phosphatidylinositol-degrading enzymes in liver lysosomes. The major pathway by which liver lysosomal enzymes degrade phosphatidylinositol is through an EDTA-insensitive formation of phosphorylinositol. This is in distinct contrast with the Ca2+-dependent production of phosphorylinositol from phosphatidylinositol, which is located in the cytosol. Lysosomal enzymes can also totally deacylate phosphatidylinositol, producing glycerophosphorylinositol.


Net activity of phospholipase A2 in brain and the lack of stimulation of the phospholipase A2-acylation stem. Certain observations reported previously from this laboratory have not proved reproducible. These are (1) the relatively rapid hydrolysis of added phosphatidylcholine by phospholipase A2 of tissue from the cerebral cortex of the guinea pig and (2) the stimulation by 10 micron-noradrenaline and by 1.0nM-cyclic AMP of the phospholipase A2-acylation system of isolated synaptic membranes.


Feedback regulation of vitamin D metabolism by 1,25-dihydroxycholecalciferol. Many factors influence the production of 1,25(OH)2D3 (1,25-dihydroxycholecalciferol) by the kidney. One important factor seems to be feedback regulation by 1,25(OH)2D3 itself. Administration of 1,25(OH)2D3 to vitamin D-deficient chicks abolishes renal 25(OH)D3(25-hydroxycholecalciferol)1-hydroxylase activity and induces the appearance of 25(OH)D3 24-hydroxylase activity. It is likely that these effects are mediated via a nuclear effect, as they are prevented by pretreatment with actinomycin D and alpha-amanitin. Further, 1,25(OH)2D3 has a marked effect on gene transcription in the kidney cell, as assessed by measurement of RNA polymerase activities. RNA polymerase I and II activities are 80-90% inhibited by 12.5nmol of 1,25(OH)2D3 within 30min of subcutaneous administration, indicating an immediate and massive decrease in total gene transcription. By 4h RNA polymerase II activity has returned to control values, but RNA polymerase I activity is markedly enhanced. These results are consistent with the view that regulation of cholecalciferol metabolism in the kidney is associated with an effect of the active metabolite on the kidney nucleus.


The nature of controlled respiration and its relationship to protonmotive force and proton conductance in blowfly flight-muscle mitochondria. 1. To determine whether controlled (State 4) pyruvate oxidation can support a high energy state, measurements of the redox span NAD-cytochrome c, phosphorylation potential and protonmotive force (the gradient in electrochemical activity of protons across the mitochondrial inner membrane) were made as indices of energy status. For comparison, these three measurements were also made with glycerol 3-phosphate, an alternative substrate. The two substrates gave essentially identical values for the redox span NAD-cytochrome c in State 4, and the phosphorylation potential was of sufficient magnitude to be considered in equilibrium with the redox span over the first two phosphorylation sites. The magnitude of the protonmotive force in State 4 was much less and the implications of this finding are discussed. 2. Measurements made during the controlled (State 4) to active (State 3) transition indicated that with glycerol 3-phosphate as substrate, both the redox span NAD-cytochrome c and the protonmotive force were diminished; the State 4 --> State 3 transition with pyruvate as substrate was accompanied by an increase in the redox span but a decrease in protonmotive force. The contrary behaviour of these two energetic parameters in the presence of pyruvate was ascribed to a transient excess in the flux of protons through the adenosine triphosphatase relative to the protonpumping respiratory chain, in spite of the increased dehydrogenase activity. 3. The lower protonmotive force seen in State 3 relative to State 4 with pyruvate as substrate was due to a diminution of both the electrical (DeltaPsi) and the chemical (DeltapH) components; with glycerol 3-phosphate, the magnitude of the decrease in protonmotive force during the State 4 --> State 3 transition was similar to that seen with pyruvate, but was due to a large decrease in the electrical component (DeltaPsi) and a small rise in the chemical component (DeltapH). The reason for the difference seen in the behaviour of the components of the protonmotive force was investigated but not established. 4. In the presence of oligomycin and ADP, oxidation of pyruvate, but not of glycerol 3-phosphate, supported a greater protonmotive force than in State 4, in keeping with the dehydrogenase activation and increased redox span NAD-cytochrome c found under these conditions. 5. Experiments involving the use of uncoupling agent to stimulate respiration are compared with those in which limiting concentrations of ADP were used. Estimates of the proton conductance of the inner membrane indicate a similar non-linear dependence on uncoupler concentration with the two substrates. 6. A model is proposed as an explanation of the high rates of controlled glycerol 3-phosphate oxidation. The model relies on a high permeability of the inner membrane to protons and other ions being induced by glycerol 3-phosphate oxidation in State 4.


Uridine kinase activities and pyrimidine nucleoside phosphorylation in fluoropyrimidine-sensitive and -resistant cell lines of the Novikoff hepatoma. Uridine kinase, the rate-limiting enzyme in the activation (phosphorylation) of uridine and the corresponding chemotherapeutic analogues, is present as two isoenzymes localized exclusively in the cytosol of rapidly growing neoplasms, including the S-37 sarcoma, EL-4 leukaemia, HeLa cells (a human carcinoma) and the Novikoff hepatoma. The activities of the isolated isoenzymes are markedly decreased when the concentrations of ATP, phosphate or Mg2+ that are optimum in vitro are replaced by concentrations of ATP, phosphate or Mg2+ that are optimum in vitro are replaced by concentrations approximating to those found in vivo. Further, comparisons of the Km values of isolated uridine kinases with those for cellular uptake of pyrimidine nucleosides and their rate of intracellular phosphorylation suggest that nucleoside-transport systems play a rate-limiting role in nucleoside analogue activation and consequently that it is impossible to estimate the Km of uridine kinase in the intact cell. During the development of tumour-cell resistance to 5-fluorouracil or 5-fluorouridine in vivo there was an early differential increase in the activity of a low-affinity (high-Km) uridine kinase isoenzyme, as measured in cell extracts, and a 7-fold increase in the Km values for the uptake of both uridine and 5-fluorouridine into the intact resistant cells.


Inhibition of binding of corticotropin-(1-24)-tetracosapeptide (Synacthen) to membrane receptors of adrenal cortex by cortisol. The binding of Synacthen to partially purified bovine adrenocortical plasma membranes was shown to be inhibited by cortisol. The findings suggest that cortisol is involved in a peripheral feedback mechanism for the control of its release.


1,4-Bis-[1-methyl-5-nitroimidazolyl-(2-methyleneimino)]-piperazine, a new drug with marked activity against Entamoeba histolytica. Short communication. 1,4-Bis-[1-methyl-5-nitroimidazolyl-(2-methyleneimino)]-piperazine (HOE 316) administered orally produces an effect which is clearly superior to that of metronidazole against Entamoeba histolytica in the golden hamster and slightly inferior against Trichomonas fetus and T. vaginalis in the mouse. The acute tolerance of the compound is very good (LD 50% greater than 10 g/kg body weight). It is assumed that the possible main metabolite of HOE 316 is a monohydrazone: 1-methyl-2-(4-amino-piperazino-1-imino-methyl)-5-nitroimidazole.


Purification and characterization of high-molecular-weight forms of adrenocorticotropic hormone of ovine pituitary glands. A highly purified preparation of high-molecular-weight adrenocorticotropic hormone (ACTH) was prepared from ovine pituitary glands by dilute acetic acid extraction, oxycellulose fractionation. Sephadex gel filtration, and affinity chromatography on immobilized alphap(1-39)ACTH antibodies. Two ACTH peptides of molecular weights of 24 000 and 34 000 were detected by sodium dodecyl sulfate-acrylamide gel electrophoresis in this preparation. It appeared that the immobilized antibodies adsorbed two forms equally well and could not distinguish between them under the conditions used. These two ACTH peptides were found to be present in crude extracts of ovine pituitary glands, indicating that they were not artifacts produced by the purification procedure. The high-molecular-weight forms of ACTH were found to be susceptible to degradation by tissue enzymes. They could be easily destroyed during the extraction, if precautions were not taken. Moreover, they were poorly adsorbed by oxycellulose which had been used for the adsorption of ACTH activity from crude preparations by most investigators. These properties probably accounted for the fact that high-molecular-weight forms of ACTH remained undetected until very recently.


A comparison of membrane components of normal and transformed BALB/c cells. Membrane glycolipids, glycoproteins, and surface proteins of normal and transformed BALB/c cell lines have been compared. Several virally and spontaneously transformed cell lines showed differences in membrane components compared to normal A31 cells. These differences consisted of increased amounts of simpler gangliosides, absence of the large external transformation sensitive (LETS) protein, and the appearance of a major new glycoprotein band of about 105 000 molecular weight. In contrast, the spontaneously transformed cell line that caused the fastest growing tumors in vivo and the most rapid animal death (3T12T) did not have these changes. A31 and 3T12T glycolipid profiles appear similar as did glycoproteins and cell surface proteins detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. When Pronase-generated glycopeptides were analyzed by Sephadex G-50 chromatography, and enrichment in faster-eluting species was seen in two killing tumor lines (c5T and 3T12T) compared to A31. Regressing tumor lines (MSC, c5) did not show this change. Isolated membrane glycoproteins yield glycopeptides of different sized after Pronase digestion. In addition, several 3T12T glycoproteins yield glycopeptides that are larger than those from the corresponding glycoproteins of A31 cells. It appears that glycopeptide alterations associated with transformation occur in several membrane glycoproteins.


Characterization of collagen precursors found in rat skin and rat bone. Two genetic types of collagenous proteins, type I and type III, were isolated by extraction and differential salt precipitation from rat skin. The yield of collagen precursors was increased by injecting animals with colchicine 30 min before sacrifice to inhibit secretion of collagen. DEAE-cellulose chromatography was used to separate collagen from collagen precursors. Although these preparations contained more type I collagen than type III collagen, there were always more type III than type I precursors. The precursor chains of type I fractions were separated on CM-cellulose chromatography after denaturation. Three precursor forms were found for each collagen alpha chain, a complete chain (proalpha chain), and a precursor chain with only an amino-terminal (pNalpha chain) and carboxy-terminal extension (pCalpha chain). Species differences were demonstrated between rat collagen precursors and other species using rat calvaria (frontal and parietal) bones extracted with either 0.5 N acetic acid or neutral salt buffers containing protease inhibitors. Native rat procollagen elutes earlier than chicken or human procollagen on DEAE-cellulose chromatography and does not separate significantly from the pC collagen form. The collagenase resistant amino terminal peptides of rat pNalpha1 and pNalpha2 were the same size (16 000) but could be separated by DEAE-cellulose chromatography.


Kinetic studies on redox reactions of hemoproteins. I. Reduction of thermoresistant cytochrome c-552 and horse heart cytochrome c by ferrocyanide. The oxidation-reduction reaction of horse heart cytochrome c and cytochrome c (552, Thermus thermophilus), which is highly thermoresistant, was studied by temperature-jump method. Ferrohexacyanide was used as reductant. (Formula: see text.) Thermodynamic and activation parameters of the reaction obtained for both cytochromes were compared with each other. The results of this showed that (1) the redox potential of cytochrome c-552, + 0.19 V, is markedly less than that of horse heart cytochrome c. (2) deltaHox of cytochrome c-552 is considerably lower than that of horse heart cytochrome c. (3) deltaSox and deltaSred of cytochrome c-552 are more negative than those of horse heart cytochrome c. (4) kred of cytochrome c-552 is much lower than that of horse heart cytochrome c at room temperature.


Uptake of ATP analogs by isolated pea chloroplasts and their effect on CO2 fixation and electron transport. 1. The ATP analog, adenylyl-imidodiphosphate rapidly inhibited CO2-dependent oxygen evolution by isolated pea chloroplasts. Both alpha, beta- and beta, gamma-methylene adenosine triphosphate also inhibited oxygen evolution. The inhibition was relieved by ATP but only partially relieved by 3-phosphoglycerate. Oxygen evolution with 3-phosphoglycerate as substrate was inhibited by adenylyl-imidodiphosphate to a lesser extent than CO2-dependent oxygen evolution. The concentration of adenylylimidodiphosphate required for 50% inhibition of CO2-dependent oxygen evolution was 50 micronM. 2. Although non-cyclic photophosphorylation by broken chloroplasts was not significantly affected by adenylyl-imidodiphosphate, electron transport in the absence of ADP was inhibited by adenylyl-imidodiphosphate to the same extent as by ATP, suggesting binding of the ATP analog to the coupling factor of phosphorylation. 3. The endogenous adenine nucleotides of a chloroplast suspension were labelled by incubation with [14C]ATP and subsequent washing. Addition of adenylyl-imidodiphosphate to the labelled chloroplasts resulted in a rapid efflux of adenine nucleotides suggesting that the ATP analog was transported into the chloroplasts via the adenine nucleotide translocator. 4. It was concluded that uptake of ATP analogs in exchange for endogenous adenine nucleotides decreased the internal ATP concentration and thus inhibited CO2 fixation. Oxygen evolution was inhibited to a lesser extent in spinach chloroplasts which apparently have lower rates of adenine nucleotide transport than pea chloroplasts.


Aerobic respiration in mutants of Escherichia coli accumulating quinone analogues of ubiquinone. The ability of three naturally occurring analogues of ubiquinone to function in aerobic respiration in Escherichia coli has been studied. The compounds, which differ from ubiquinone in terms of the substituents on the quinone ring, accumulate in the cytoplasmic membranes of ubiE-, ubiF- and ubiG- mutants. One of the analogues (2-octaprenyl-3-methyl-6-methoxy-1,4-benzoquinone, NMQ), which lacks the 5-methoxyl group of the benzoquinone ring of ubiquinone promoted the oxidation of NADH, D-lactate and alpha-glycerophosphate but not succinate. Electron transport supported by MMQ was found to be coupled to phosphorylation. In contrast, 2-octaprenyl-6-methoxy-1,4-benzoquinone, which lacks both the 3-methyl and 5-methoxyl groups of ubiquinone, and 2-octaprenyl-3-methyl-5-hydroxy-6-methoxy-1,4-benzoquinone, in which the 5-methoxyl group of ubiquinone is replaced by an hydroxyl group, were virtually inactive in the oxidases tested. The ability of MMQ to function in respiration in isolated membranes is consistent with the findings that the growth rate and yield of a ubiF- strain, unlike other ubi- strains, were only slightly lower than those of a ubiF+ strain. The fact that MMQ is active in some but not all oxidases provides further support for the concept that the quinones link the individual dehydrogenases to the respiratory chain and that each dehydrogenase has specific structural requirements for quinone acceptors.


Role of quinones in electron transport to oxygen and nitrate in Escherichia coli. Studies with a ubiA- menA- double quinone mutant. A ubiA- menA- double quinone mutant of Escherichia coli K12 was constructed together with other isogenic strains lacking either ubiquinone or menaquinone. These strains were used to study the role of quinones in electron transport to oxygen and nitrate. Each of the four oxidases examined (NADH, D-lactate, alpha-glycerophosphate and succinate) required a quinone for activity. Ubiquinone was active in each oxidase system while menaquinone gave full activity in alpha-glycerophosphate oxidase, partial activity in D-lactate oxidase but was inactive in NADH and succinate oxidation. The aerobic growth rates, growth yields and products of glucose metabolism of the quinone-deficient strains were also examined. The growth rate and growth yield of the ubi+menA- strain was the same as the wild-type strain, whereas the ubiA-men+ strain grew more slowly on glucose, had a lower growth yield (30% of wild type) and accumulated relatively large quantities of acetate and lactate. The growth of the ubiA-menA- strain was even more severely affected than that of the ubiA-men+ strain. Electron transport from formate, D-lactate, alpha-glycerophosphate and NADH to nitrate was also highly dependent on the presence of a quinone. Either ubiquinone or menaquinone was active in electron transport from formate and the activity of the quinones in electron transport from the other substrates was the same as for the oxidase systems. In contrast, quinones were not obligatory carriers in the anaerobic formate hydrogenlyase system. It is concluded that the quinones serve to link the various dehydrogenases with the terminal electron transport systems to oxygen and nitrate and that the dehydrogenases possess a degree of selectivity with respect to the quinone acceptors.


Modulation of drug permeability in Chinese hamster ovary cells. Possible role for phosphorylation of surface glycoproteins. We have previously reported that the uptake of colchicine and other drugs in Chinese hamster ovary (CHO) cells can be greatly enhanced by the addition of metabolic inhibitors such as cyanide (See, Y.P., Carlsen, S.A., Till, J.E. and Ling, V. (1974) Biochim. Biophys. Acta 373, 242-252). This has led us to postulate the presence of an active drug permeability barrier in these cells. In this paper we provide evidence for the dependence of this permeability barrier on intracellular ATP levels. Colchicine-resistant mutants of CHO cells exhibiting a reduced drug permeability, however, can maintain this drug permeability barrier at much lower ATP levels, suggesting that they possess an altered active drug permeability barrier. We have also observed a membrane-associated protein kinase-phosphoprotein phosphatase system in the isolated membranes of mutant and wild-type cells. Differences in the intrinsic protein phosphorylation patterns between the membranes of these cells have led us to conclude that the control of the drug permeability barrier may be mediated via the phosphorylation of at least two high molecular weight surface glycoproteins.


Cyclic AMP modulation of calcium accumulation by sarcoplasmic reticulum from fast skeletal muscle. The role of cyclic 3',5'-AMP in modulating sarcoplasmic reticulum from fast skeletal muscle was studied. The rate of Ca2+ uptake was stimulated in the presence of protein kinase plus 1 micron cyclic AMP. The stimulation was absent when denatured protein kinase was used. When an adenylate cyclase inhibitor was added, the uptake rates fell to 55% of control. This decrease in rate was partially overcome by 1 micron cyclic AMP. A modulating role for cyclic AMP in fast skeletal muscle is proposed.


Decreased iodination of the red cell surface following phospholipase C treatment. Human red blood cells were treated with phospholipase C from Clostridium welchii. Lipase concentrations which produced less than 1% hemolysis and 10-15% hydrolysis of the membrane phospholipids reduced markedly (greater than 80%) the accessibility of membrane proteins to the external surface as measured by lactoperoxidase-catalyzed iodination.


Genetic regulation of the constitutive D-ribose operon in Escherichia coli B/r. Merodiploid complementation analysis of the constitutive synthesis of the D-ribokinase and the D-ribose permease in Escherichia coli B/r has shown that the constitutive D-ribose operon is genetically controlled by a transdominant regulatory gene closely linked to the D-ribokinase and D-ribose permease structural genes. The regulatory mechanism for this operon shows no requirement for operator-repressor interaction, rather a truly positive control mechanism and thus suggests an extension of the operon model in its application to constitutive enzyme regulation in bacteria.


Involvement of NAD  in induced synthesis of colicin E1. Escherichia coli K-12 nadC13 (ColE1) was starved for nicotinic acid and cellular NAD levels decreased to less than 10% of the normal. Under these conditions, induction of colicin E1 synthesis decreased to about 1% of the normal value, while 30% of the total protein synthesis remained intact. Addition of nicotinic acid reversed both the ability of the colicin induction and cellular NAD level. Induced replication of colicin E1 DNA was greatly reduced in the NAD-deprived cells.


Stimulation of synthesis of the proteins of 30-S nuclear ribonucleoprotein particles in human amnion U cells by viral infection. Early increase in RNA synthesis induced in human amnion U cells by infection with poliovirus is accompanied by an increased incorporation of amino acids into non-histone nuclear proteins with an approximate molecular weight of 40 000. These proteins are the main polypeptides of the 30-S nuclear ribonucleoprotein particles. After fractionation of nuclear proteins by extraction with solutions of different ionic strength, these polypeptides are present in the fraction of nuclear sap proteins soluble in 0.1 M Tris - HCl buffer, pH 7.6, and in the fraction of non-histone chromosomal proteins which are soluble in 0.35 M NaCl. The increase in synthesis of non-histone nuclear proteins with an approximate molecular weight of 40 000, observed in the infected cells, represents an increase in the synthesis of proteins concerned with post-transcriptional events and, therefore, is the result and not the cause of gene activation.


Influence of dibutyryl cyclic AMP on thymidine uptake by herpes simplex virus infected cells and the intracellular level of cyclic AMP. Dibutyryl cyclic AMP inhibits the increase of dThd and BrdUrd transport normally observed after infection with Herpesvirus hominis, type I and II. Incorporation is also reduced. Inhibition of uptake is non-competitive as analysed by the Lineweaver-Burk plot. Addition of this drug to infected cells also reduces the activity of the thymidine kinase (EC 2.7.1.75). Transport of dUrd, dCyd and dAdo is not reduced. 4--8 h after infection with thymidine kinase (+) herpes strains the level of cAMP increases. On infection with a thymidine kinase (-) virus, only a small elevation  of cAMP can be shown. It was also found that early addition of actinomycin D or of cycloheximide prevents the increase of the cAMP level. This increase seems to depend on the activity of the herpes genome, because ultraviolet irradiation of infective particles destroys this ability.


Mitochondrial biogenesis in cultured animal cells. I. Effect of chloramphenicol on morphology and mitochondrial respiratory enzymes. The effects of chloramphenicol on the morphology and respiratory enzymes of BHK-21 cells in spinner culture have been examined with time. Cells treated with chloramphenicol double twice before growth ceases; these cells have increased size as measured by several techniques. Mitochondria are enlarged and appear to degenerate with prolonged treatment. Cytochrome c oxidase and succinate cytochrome c reductase activities are reduced while there is no decrease in the activities of monoamine oxidase, glutamate dehydrogenase or NADPH-cytochrome c reductase. Cytochromes aa3 and b disappear on treatment while cytochromes c + c1 appears to be unaffected. All these effects are reversible if chloramphenicol is removed within a limited period of time.


The effect of cyclic GMP on phosphofructokinase from rat tissues. In view of the recently proposed hypothesis of biologic regulation through opposing influences of cyclic AMP and cyclic GMP, and since cyclic AMP is a well-known allosteric activator of phosphofructokinase (ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11), the effect of cyclic GMP on the activity of this enzyme from several rat tissues was investigated. It was found that cyclic GMP exerted an inhibitory effect on the activity of rat heart and skeletal muscle phosphofructokinase. This effect was most pronounced under conditions in which the enzyme was partially inhibited by ATP or by citrate. Cyclic GMP also antagonized the deinhibitory action of cyclic AMP and other allosteric activators, such as glucose 1,6-bisphosphate or AMP, on the ATP or citrate-inhibited heart or muscle phosphofructokinase. In contrast to the heart and skeletal muscle phosphofructokinase, the adipose-tissue enzyme was not affected by cyclic GMP to any significant degree. The antagonistic action of cyclic GMP to the activation of heart-phosphofructokinase, may suggest a mechanism by which the activity of phosphofructokinase is synchronized with the activity of glycogen phosphorylase, as a result of acetylcholine action in heart, to achieve a decrease in total glycogenolysis and glycolysis.


A continuous fluorometric assay for flavokinase. Properties of flavokinase from Peptostreptococcus elsdenii. A continuous fluorometric assay that utilizes apoflavodoxin as a trapping agent for riboflavin 5'-phosphate (FMN) has been developed for flavokinase (ATP:riboflavin 5'-phosphotransferase, EC 2.7.1.26). Use of this assay is illustrated in a procedure for the partial purification of flavokinase from the strict anaerobe Peptostreptococcus elsdenii. The purified enzyme catalyzed the formation of 8.3 nmol FMN - min-1 - mg-1 at 37 degrees C and had apparent Km values for riboflavin and ATP of 10 and 4.7 micronM, respectively. ATP could be replaced by ADP (22% of the rate observed with ATP) but not by GTP. The enzyme also phosphorylated 5-deaza- and 8-bromoriboflavin with activities of 15 and 70%, respectively, of that with riboflavin; it was inactive with iso riboflavin and deoxyriboflavin.


Kynureninase-type enzymes and the evolution of the aerobic tryptophan-to-nicotinamide adenine dinucleotide pathway. Kynureninase-type (L-kynurenine hydrolase, EC 3.7.1.3) activity has been found to be present in the livers of fish, amphibia, reptiles, and birds. In addition to past information concerning this enzyme activity in mammalian liver, it is now clear that all the major classes of vertebrates carry a highly specialized kynureninase-type enzyme, which we have termed a hydroxykynureninase. To compare the reactivities of these enzymes with L-kynurenine and L-3-hydroxykynurenine, ratios of tau values (Km/V) were used. Based on this comparison, the bacterium Pseudomonas fluorescens carries the most efficient kynureninase, whereas the amphibian Xenopus laevis has the most efficient hydroxykynureniase. In these two cases, the ratio of tau values differs by a factor of 38 000. It is hypothesized that the tryptophan-to-nicotinamide adenine dinucleotide biosynthetic pathway evolved from a catabolic system of enzymes, and that the differences observed in the kynureninase-type enzymes between lower and higher organisms reflect the specialization of the function of these enzymes from a strictly catabolic role to an anabolic one during the course of evolution.


In vitro regulation of cholesterol metabolism by low density lipoproteins in skin fibroblasts from hypo-and hyperresponding squirrel monkeys. When squirrel monkeys (Saimiri sciureus) are fed diets containing cholesterol, some individuals (hyperresponders) become hypercholesterolemic, while others (hyporesponders) are able to maintain nearly normal plasma cholesterol concentrations. Skin fibroblasts were grown from three hyperresponder and threehyporesponder squirrel monkeys, previously characterized on the basis of their plasma cholesterol response to two cholesterol-containing diets and the pheno-type of their parents. The rates of cholesterol synthesis and esterification were determined in the cultured fibroblasts incubated with low density lipoproteins isolated from normocholesterolemic squirrel monkeys or hypercholesterolemic rhesus monkeys. Both lipoprotein preparations influenced the metabolic parameters measured in a similar manner in cells from both hypo- and hyperresponder animals. Exposure of skin fibroblasts to low density lipoproteins resultd in a stimulation of cholesterol esterification and a suppression of cholesterol synthesis in cells from both hypo- and hyperresponder animals. When incubated with increasing concentrations of low density lipoprotein cholesterol, up to 50 microgram/ml, fibroblasts from both hypo-and hyperresponding animals responded with a similar maximum percentage suppression of sterol synthesis. Thus, hyperresponsiveness to dietary cholesterol in squirrel monkeys, although a heritable characteristic, is not associated with an inability of low density lipoprotein to suppress cholesterol synthesis or stimulate cholesterol esterification as occurs in familial hypercholesterolemia in man.


Effects of cyclic nucleotides on the conformational states of the alpha core of the cyclic AMP receptor protein. The alpha core gragment produced by limited proteolysis contains the cyclic AMP binding domain and the two buried sulfhydryl groups of the cyclic AMP receptor protein. The buried sulfhydryl groups of the alpha core react with 5,5'-dithio-bis(2-nitrobenzoic acid) after denaturation by 3 M urea or digestion with subtilisin. The rate of sulfhydryl modification in the presence of 3 M urea or subtilisin is markedly decreased in the presence of cyclic nucleotides which are proposed to tighten the conformation of the alpha core. Incubation of the alpha core in 3 M urea or dithionitrobenzoic acid does not affect cyclic AMP binding while dithionitrobenzoic acid plus 3 M urea inhibits cyclic AMP binding suggesting a role for the buried sulfhydryls in cyclic AMP binding or their proximity to the cyclic AMP binding domain of the alpha core. The data are consistent with a ligand-induced conformational change in the alpha region of the native cyclic AMP receptor protein that is required for DNA binding.


Particle distribution of human serum high density lipoproteins. Density gradient ultracentrifugation of human serum high density lipoproteins (HDL) from both normolipemic males and females results in a distribution of HDL concentration versus subfraction hydrated density which has three maxima. Gradient gel electrophoresis of total HDL is characterized by three banding maxima, the positions of which suggest the presence of three particle size ranges: I. 10.8-12.0 nm, II. 9.7-10.7 nm, and III. 8.5-9.6 nm. Gradient gel electrophoresis of density gradient subfractions established an inverse relationship between particle size and particle hydrated density which was corroborated by electron microscopy and analytic ultracentrifugation. Comparison of male HDL from size ranges I, II, and III with female HDL from the same size ranges showed only small differences in the mean value of the peak F degrees 1.20 rate, size, molecular weight, protein weight percent, and weight protein/weight phospholipid. Major differences between males and females were seen in the relative amounts of HDL in density gradient subfractions 1-3 (size range I material) and 11-12 (size range III material); the percent total HDL in the group of subfractions 1-3 was greatly increased in female HDL while that of the group of subfractions 11-12 was increased in the male HDL. These studies indicate the presence of at least three major components in HDL instead of two (HDL2 and HDL3) and that peak F degrees 1.20 rate differences in HDL schlieren patterns between males and females are a function of the relative levels of these three components.


Dimeric ("big") human placental lactogen. Immunological and biological activity. Dimeric ("big") human placental lactogen has been isolated in near homogeneous form from placental tissue. It consists of a disulfide-linked (stable) form and a noncovalently associated (unstable) form of the native hormone. The two forms were separated by exposure to denaturing conditions and resolution by gel exclusion chromatography. Both forms retained immunological activity, ability to bind mammary membranes, and ability to induce mammary N-acetyllactosamine synthetase in vitro. On a molar basis, stable dimeric placental lactogen was more active than placental lactogen in the radioimmunoassay indicating that the immunological determinants on both monomeric units could bind to antibody. On a molar basis, stable dimeric placental lactogen was equally active with monomeric placental lactogen in competing for mammary gland membrane binding sites, indicating that only one active site in the molecule could interact with the membrane at a time. Stable dimeric placental lactogen was also active in an in vitro bioassay using the induction of N-acetyllactosamine synthetase. It is concluded that dimer formation does not alter the biologically active portion of the placental lactogen molecule. Since the carboxyl-terminal region (residues 182-191) is involved in the interchain disulfide bonds of dimeric placental lactogen, this portion of the molecule is probably not necessary for its biological activity.


Properties of human erythrocyte phosphatidylinositol kinase and inhibition by adenosine, ADP and related compounds. 1. An improved assay for the enzyme phosphatidylinositol kinase is described. The kinase activity is increased more than 10-fold by the addition of mercaptoethanol and exogenous phosphatidylinositol in the presence of Triton X-100. The enzyme is solubilized by non-ionic detergents. 2. Phosphatidylinositol kinase is inhibited by physiological concentrations of ADP and by similar levels of adenosine. Cyclic AMP, AMP and theophylline inhibit at higher concentrations. Caffeine is much less effective than theophylline as an inhibitor. Guanine nucleotides do not appreciably inhibit the kinase. All the inhibitors tested seemed to compete with ATP. Theophylline also reduced the rate of polyphosphoinositide synthesis in intact cells. 3. Possible roles of polyphosphoinositides in energy charge maintenance and inversion and resealing are discussed.


Effects of polyamines and polyanions on a cyclic nucleotide-independent and a cyclic AMP-dependent protein kinase. The phosphorylation of phosvitin in vitro by a cyclic nucleotide-independent protein kinase (phosvitin kinase) derived from rooster liver is markedly stimulated by the divalent cation, Mg2+. In addition, the activity is further stimulated by low concentrations of the polyamines putrescine, spermidine and spermine leading to higher rates of phosphate incorporation than could be obtained at any concentration of Mg2+. Spermine is inhibitory at higher concentrations. The polyamines shift the Mg2+ requirement for maximal activity to lower concentrations. The activity of a cyclic AMP-dependent histone kinase from beef heart is not altered by the presence of polyamines. Heparin is a potent inhibitor of phosvitin kinase but has no effect on histone kinase. Polyribonucleotides (polyadenylic acid and transfer RNA) inhibit both types of kinases, but the degree of inhibition of phosvitin kinase is variable and depends upon the type of the polyanion present. Sermidine and spermine, but not Mg2+, efficiently counteract the inhibitory action of heparin and tRNA. The results suggest that, also in vivo, naturally occurring polyamines and polyanions such as tRNA may have a regulatory function on protein kinases.


Elevation of guanosine 3',5'-monophosphate level by adenosine in cerebellar slices of guinea pig. Effect of adenosine on the level of guanosine 3',5'-monophosphate in guinea pig cerebellar slices was investigated. Adenosine increased the concentration of guanosine 3',5'-monophosphate in the slices 3--4 fold. Upon removal of adenosine from the medium, the concentration of guanosine 3',5'-monophosphate returned to the initial level. AMP, ADP or ATP also increased the guanosine 3',5'-monophosphate level to the same extent as adenosine, while adenine or other nucleosides were not effective. In the absence of Ca2+ in the incubation medium, adenosine did not increase the concentration of guanosine 3',5'-monophosphate in cerebellar slices although level of adenosine 3',5'-monophosphate was elevated by adenosine. Anticholinergic agents, adrenergic blocking agents or antihistaminics did not prevent the increase of guanosine 3',5'-monophosphate by adenosine indicating that the effect of adenosine was not mediated by the release of neurotransmitters. The combination of adenosine with depolarizing agents showed an additive effect on the level of guanosine 3',5'-monophosphate indicating that adenosine increased the level of guanosine 3',5'-monophosphate by a different mechanism from the depolarization.


Comparative EPR study on high-spin ferric porphine complexes and cytochrome P-450 having rhombic character. Comparative EPR studies were made on two high-spin Fe(III) porphine model systems and mammalian liver microsomal cytochromes P-450, all of which exhibit approximately the same degrees of rhombicity in their EPR spectra. Comparison of g values and linewidths as a function of temperature, and of the microwave power saturation demonstrated that EPR characteristics of P-450 are more similar to the Fe(III) porphines having the thiolate axial ligand than in the other model systems, the mixed crystals of Fe(III) porphine with the corresponding free base porphine, in which no thiolate ligand is involved. There is, however, a discrepancy between P-450 and the model thiolates with respect to the size of the zero-field parameter D. These observations indicate that P-450 heme has essential structural features in common with thiolates but the Fe-S bond of P-450 may be modified from its normal orientation in model thiolates, probably as a result of the constraints imposed by the protein structure.


Preparation and metabolic characterization of isolated rat lung cells. Collagenase digestion of minced lung tissue yielded isolated cells, functionally viable as judged by several metabolic and morphological criteria, representative of all the cell species normally present in the tissue. The efficiency of the isolation procedure was about 25 per cent. Aerobic metabolism was not affected by most of the substrates tested except by succinate which increased oxygen utilization, and glucose, fructose and octanoate which significantly decreased oxygen uptake. Since no significant changes have been observed in the cellular adenine nucleotide content during glucose depression of aerobic metabolism it is concluded that the glycolytic flux had to be sufficient as to account for the decrease in the mitochondrial energy production. The mechanism responsible for these effects as well as their physiological significance are discussed herewith.


Sequential enzymolysis of human aorta and resultant stress-strain behavior. The human aorta has five major components from which the aortic walls can be characterized: mucopolysaccharides, smooth muscle, collagen, micro-fibrilar glycoprotein (associated with the elastic fiber), and elastin. Enzymes were employed to remove four of the components sequentially without destroying the mechanical characteristics of the remaining components in order to elucidate the structure-property relationship in the human aorta. Before treatment the initial mechanical behavior was recorded on an Instron Tensile testing machine. After enzymolysis the samples and controls were again tested and these results compared to their prior characteristics. Stress-strain characteristics after a sequence of enzyme treatments indicate that two of the components share the major part of the stress in the circumferential direction. These components, elastin and collagen, contribute as if they were in parallel to each other with the collagen in a crimped state.


Electron paramagnetic resonance of Cu2+:Mb single crystal. Conformational changes. Copper introduced into met-myoglobin crystals occupies various sites as indicated by electron paramagnetic resonance parameters. Cu2+ (A) is probably liganded to histidine A10, lysine A14, and asparagine GH4 (Banaszak et al., 1965) and shows superhyperfine interaction with a single (imidazole) nitrogen. Cu2+ (B) and Cu2+ (C) correspond to other anisotropic sites described in less detail. Cu2+ (A) exhibits a transition to an isotropic form with a transition temperature of 40.5 degrees C. This transition indicates a conformational change in myoglobin and could correspond to a motion of A helix away from the GH section. The transition temperature is 7 degrees C higher than the one previously reported (Atanasov, 1971) for myoglobin in solution.


Collective behaviour of hemoglobin in dense solutions. The formation of radical pairs of spin-labelled hemoglobin molecules (beta-93 cysteine) in a mixture with unlabelled hemoglobin of concentrations higher than the critical one [(5.7 less than c less than 6.3) mM/sigma per heme] is due to some kind of long-range order characterizing these solutions. Labelled molecules induce pertubation in an unlabelled hemoglobin solution of concentration higher than the critical one. The collective behaviour of unlabelled molecules gives rise to the formation of radical pairs of spin-labelled hemoglobin molecules. Such behaviour is dependent on the ligand bound at the heme iron (carbonmonoxide, oxygen, or water.)


[Participation of the adenylate cyclase system in inductive synthesis of acetylcholinesterase in the brain]. The enhancement of the brain acetyl cholinesterase (ACHE) activity in rats by the intraventricular injection of adrenaline or dibutiryl analogue cyclic adenosine-3',5'-monophosphate (cAMP) was shown to be due to the enzyme inductive synthesis. ACHE induction manifests itself more in the subcortical white matter than in the cortex. The stimulating effect of adrenaline on the ACHE activity is suppressed under the beta-adrenoreceptor block, while the cAMP effect remains unchanged. On the contrary, the block of the alpha-adrenoreceptors stimulates the enzyme synthesis induction. The effects of adrenaline and cAMP are of the same direction and are realized through the beta-adrenoreceptors. The enhancement of ACHE activity during the block of the alpha-adrenoreceptors is accounted for by the elimination of their inhibitory influence on the beta-adrenoreceptors.


[Possible participation of prostaglandins A1 and B1 in the development of internal carotid artery spasm]. Experiments were carried out on dogs with the internal carotid artery circulatory isolated and continuously perfused in situ with oxygenated Ringer-Krebs bicarbonate solution. Prostaglandins A1 and B1 (PG-A1 and PG-B1) exerted a constrictory effect on the artery. The effect of both prostaglandins was significantly less, but more prolonged than that of the same doses of serotonin. A possible role of PG-A1 and PG-B1 in the development of vasospasm is testified to by the following experimental results: a) during repeated action of PG-A1 AND PG-B1 their effect did not change, and therefore it should not decrease under prolonged influence of these prostaglandins on the vascular wall;) b) the mentioned prostaglandins caused potentiation of the constrictory effect of both serotonin and norepinephrine.


[Factors influencing the serum levels of cholesterol and beta-lipoproteins in starving rabbits]. The influence of starvation on the lipid metabolism was studied on male rabbits under usual conditions and in the presence of pyroxidine deficiency (4-deoxypyridoxine administration) and of thiamine (oxythiamine administration), and also in administration of neurotropic preparations (phenamine, seduxen). Starvation for 7 to 10 days led to increase of cholesterol and beta-lipoproteins level in the serum. Pyridoxine deficiency and phenamine administration caused a greater increase of cholesterol and especially or beta-lipoproteins. On the other hand, thiamine deficiency and seduxen administration limited an increase of cholesterol and beta-lipoproteins during hungry stress. Administration of aerovit for prophylactic purpose promoted a decrease of the metabolic shifts. The amount of cholesterol increased in the liver of hungry animals, especially after the phenamine administration and in the presence of pyridoxine deficiency; aerovit administration prevented increased cholesterol accumulation in the liver. The differences in the cholesterol level in the serum and and the liver can be explained by the changes of its biosynthesis during hungry stress.


[Inhibition of rat heart and brain cyclic adenosine-3',5'-monophosphate phosphodiesterase in vitro under the influence of neurohormone C]. The work was devoted to one of the new neurohormones (neurohormone "C") obtained from the bovine hypothalamus which produced a pronounced relaxation of the coronary vessels. A study of the effect of "C" on cyclic 3',5'-adenosine monophosphate (cAMP) phosphodiesterase (PDE) showed "C" to be a potent inhibitor of PDE in the crude PDE-preparations of the rat heart and brain. Experiments were carried out on the 2000 g supernatant obtained from the homogenized tissues. It is supposed that the relaxant effect of "C" on the coronary vessels is due to its inhibitory action on PDE mediated by the accumulated cAMP.


[Functioning of the lactose operon of E. coli K-12 under the influence of non-specific regulators]. A possible role played by cAMP in the stimulating action of ACTH and hydrocortisone on lactose E. coli K-12 operon was studied. It was shown that ACTH caused no effect in the E. coli WZ-78/F'lac (cya855) and E. coli CA8001 (L1) strains with destroyed positive cAMP control system of the lactose operon function, at the same time producing a stimulating effect on the lactose operon in the strains of wild type, i.e. E coli 200PS/F'lac and E. coli 3000. Hydrocortisone stimulated the lactose operon function both in E. coli 3000 and in the mutant E. coli CA8001 (L1). It was supposed that the accelerating effect of ACTH on the lactose operon was mediated through cAMP; as to hydrocortisone--it stimulated the lactose operon function independently of cAMP.


Evidence for intracellular amyloid formation in myeloma. Evidence for intracellular formation of amyloid fibrils in a patient with kappa light-chain myeloma is described. Amyloid fibrils were seen as intracytoplasmic inclusions within plasma cells, histiocytes, renal tubule cells, and possibly in hepatocytes. Extracellular amyloid was also present. The electron microscopic studies suggest that amyloid fibrils may form in the Golgi apparatus and within lysosomes.


Ultrastructural, cytochemical, and membrane surface marker characteristics of the atypical lymphocytes in infectious mononucleosis. Atypical lymphocytes from nine young adults with acute infectious mononucleosis (IM) were studied for morphologic, ultrastructural, cytochemical, and membrane surface marker characteristics. There was an absolute increase in T lymphocytes in the patients. Atypical lymphocytes accounted for 83%-96% of the lymphocyte population. These lymphocytes contained cytoplasmic inclusions which ranged in size from 1000 to 6000 A, were usually membrane bound, and consisted of parallel arrays of microtubulelike structures. The inclusions, which have been referred to as parallel tubular arrays (PTA), were found in 15%-75% of the lymphocytes from the IM patients. Ultrastructural cytochemical methods demonstrated acid phosphatase activity within many of the membrane-bound PTA. The function of the PTA is unknown. Since they were observed only in the lymphocytes which appeared to correspond to the atypical lymphocytes on light microscopy, the majority of which typed as T cells, there appears to be an association between PTA and T lymphocytes. It is possible that PTA identify a specific subset of T lymphocytes which is expanded in IM. Alternatively, PTA may be a transient finding in lymphocytes appearing only in certain biologic states of the cell such as during T-lymphocyte activation.


Synovial sarcoma of the soft parts in the 1st toe: recurrence after a 35 year interval. A female patient with a recurrent synovial sarcoma of right 1st toe (35 years) after the first operation is presented. The histology of the tumor 35 years ago showed evidence of the biphasic structure, while the recurrent tumor contained only the fibrous elements. The patient has been examined 3 years after the last operation. She has no pain and there is no evidence of any recurrence. The patient refuses any radiation or additional surgery at this time.


The generation of nerve and muscle repetivie activity in the rat phrenic nerve-diaphragm preparation following inhibition of cholinesterase by ecothiopate. 1 Simultaneous extracellular recordings were made from two end-plate zones of the isolated diaphragm and from the phrenic nerve of the rat in response to stimulation of the nerve. The contractions of the diaphragm were also recorded.2 In the curarized diaphragm, the introduction of ecothiopate, a non-competitive inhibitor of cholinesterase, caused a threefold increase in the amplitude of the end-plate current and an eightfold increase in the duration at half the peak amplitude.3 In the non-curarized diaphragm, the introduction of ecothiopate caused the generation of repetitive activity (RA) in first the phrenic nerve: this was then followed by RA in the diaphragm. At that stage, nerve RA possessed a shorter latency than muscle RA. The generation time for nerve RA was 1.6 ms and for mRA, it was 2.7 milliseconds.4 Nerve RA was more labile than muscle RA; it was readily abolished by increasing the frequency of stimulation, by magnesium, by tubocurarine or by high concentrations of ecothiopate, whereas muscle RA was still generated. Steady exposure to acetylcholine abolished both forms of RA.5 Two competitive inhibitors of cholinesterase, neostigmine and ambenonium, were also shown to evoke RA in nerve and muscle. The generation times for nerve RA and muscle RA were similar to those following ecothiopate.6 It was concluded that nerve RA and muscle RA were generated after the inhibition of cholinesterase by ecothiopate as a result of the prolonged action of acetylcholine upon cholinoceptive sites on the nerve terminal and motor endplate respectively. A direct excitatory action of ecothiopate upon the phrenic nerve terminals was excluded.


Cyclic adenosine 3',5'-monophosphate and leucocyte chemotaxis in vivo. 1 The effect of local elevation of leucocyte cyclic adenosine 3',5'-monophosphate (cyclic AMP) content on the continued migration of leucocytes to a site of acute inflammation was studied in the pleural cavity of rats. 2 Leucocyte cyclic AMP levels were elevated by injecting with the irritant into the pleural cavity dibutyryl cyclic AMP alone or with theophylline. 3 The treatments both produced a marked reduction in leucocyte migration into a pleural reaction induced by immediate hypersensitivity, but had no effect on cell numbers in pleurisy induced either by pyrophosphate or urate crystals.


The role of beta-adrenoceptors in the responses of the hepatic arterial vascular bed of the dog to phenylephrine, isoprenaline, noradrenaline and adrenaline. 1 The sympathetically-innervated hepatic arterial vascular bed of the dog was perfused from a femoral artery. Hepatic arterial blood flow and perfusion pressure were recorded continuously, and the hepatic arterial vascular resistance (HAVR) calculated from these measurements.2 Intra-arterial injections of phenylephrine caused dose-dependent rises in HAVR, indicating hepatic arterial vasoconstriction, at all doses above threshold. No secondary reductions in HAVR followed these responses.3 Intra-arterial injections of isoprenaline caused only dose-dependent reductions in HAVR at doses above threshold.4 Intra-arterial injections of noradrenaline typically caused an initial increase in HAVR which was followed at all but the highest doses by a secondary, delayed, reduction in HAVR.5 Intra-arterial injections of adrenaline, like those of noradrenaline, resulted in hepatic arterial vasoconstriction followed by hepatic arterial vasodilatation.6 On a molar basis, the most potent hepatic arterial vasoconstrictor was noradrenaline, followed by adrenaline and phenylephrine.7 The maximum reductions in HAVR caused by adrenaline (mean reduction = 21.9%) and noradrenaline (16.9%) were significantly smaller than those due to isoprenaline ((P) < 0.001).8 Propranolol attenuated the hepatic arterial vasodilator responses due to isoprenaline, and the secondary falls in HAVR following intra-arterial adrenaline and noradrenaline.9 Propranolol did not modify the vasoconstrictor responses to phenylephrine.10 Both adrenaline and noradrenaline were more potent hepatic arterial vasoconstrictors after propranolol than in the absence of beta-adrenoceptor blockade. The potentiation of the vasoconstrictor effects of adrenaline was statistically significant.11 After propranolol, adrenaline was a more potent hepatic arterial vasoconstrictor than noradrenaline.12 Since the beta-adrenoceptors in the hepatic arterial vasculature were not blocked by atenolol, but were stimulated by salbutamol, it is concluded that they are predominantly of the beta(2)-type.13 The vasoconstrictor actions of phenylephrine, noradrenaline and adrenaline were all antagonized by the systemic administration of phentolamine, all three dose-response curves being shifted to the right.14 The results are discussed with regard to the possible control of the hepatic arterial vasculature by naturally-occurring catecholamines.


Neuroticism and dreaming sleep: a case for interactionism in personality research. The value of a trait approach to sleep and dreaming characteristics is evaluated by reviewing recent research on differential patterns and interactions manifest by high and low neuroticism subjects. Differential patterning of dream content and REM sleep characteristics under positive and negative presleep conditions suggest a qualitative difference in experience and behavior, and a quantitative difference in "importance" attributable to the dreaming sleep process. It is hypothesized that, under "normal" conditions, low neuroticism individuals make more exclusive use of the dreaming period, while under "artificial" conditions, they demonstrate a greater motivation to compensate for restricted REM by intensification of the REM process (shortened REM onset latencies, greater REM rebound, intensified dream experiences). Finally, it is suggested that theories of dreaming sleep should recognize the importance of traits, as well as situational factors to the psychology of dreaming sleep.


Metyrapone in long-term management of Cushing's disease. Metyrapone was used in the long-term management of 13 patients with pituitary-dependent bilateral adrenal hyperplasia (Cushing's disease). The total length of treatment ranged from two to 66 months, with a mean of 21 months. The clinical features of the disease rapidly improved on metyrapone and this improvement was maintained. Although plasma ACTH concentrations rose in all patients, the increase was insufficient to overcome the adrenal blockade induced by the drug. Eight of the 13 patients had additional external pituitary irradiation as definitive treatment of their disease and one underwent a transfrontal hypophysectomy. Radiotherapy cured one patient, and after three years metyrapone was withdrawn. Slight hirsuties was noted in four of the seven women who received the drug for six months or more. A fifth woman had more severe hirsuties and this led to bilateral adrenalectomy. Other than hirsuties, side effects were few and the routine use of metyrapone is recommended as an adjunct to more definitive treatment in all patients who present with Cushing's syndrome, irrespective of aetiology.


BK antibody and virus-specific IgM responses in renal transplant recipients, patients with malignant disease, and healthy people. Haemagglutination-inhibition (HAI) antibodies to BK virus, including BK-virus-specific IgM, were determined before and after renal transplantation in 20 patients, in 57 patients with malignant disease, and in 66 healthy controls, Before transplantation 11 of the renal transplant recipients were seronegative, but eight later serocconverted, two before and six after transplantation. Twenty of the patients with malignant disease and 22 controls were also seronegative. The geometric mean titre of BK HAI antibodies was significantly higher among transplanted patients (1/180) than among controls (1/90). BK-virus-specific IgM antibody was detected in seven renal transplant recipients, six patients with malignant disease, and 13 healthy controls. In transplant recipients BK-virus-specific IgM antibody usually persisted throughout the duration of the study, and studies on controls from whom second serum samples were available suggested that they too had persistent BK-virus-specific IgM responses. The geometric mean titre of BK-virus-specific IgM HAI antibody was significantly greater in post-transplantation sera (1/223) than in control sera (1/28). The specificity of the detection of BK-virus-specific IgM HAI antibody was confirmed by direct visualisation of antibody by immune electron microscopy. The persistence of BK-virus-specific IgM suggested that BK virus continued to provide an antigenic stimulus. Nevertheless, there was no obvious association between the serological findings and any clinical features, and prospective studies will be needed to elucidate any such association.


Effects of vagal volleys on units of intralaminar and juxtalaminar thalamic nuclei in monkeys. As part of an attempt to clarify the nature of inputs to the limbic cortex, the thalamic intralaminar and juxtalaminar nuclei were explored for unit responses to vagal volleys in awake, sitting squirrel monkeys. Vagal shocks elicited responses of a large percentage of units in the anterior medial, paracentral, lateral dorsal, and lateral, and medial dorsal nuclei, as well as in part of the ventral lateral nucleus adjacent to the paracentral. Responsive units showed either initial excitation or initial inhibition. As in the preceding study on the cingulate and supracingulate cortex, there were two main types of initially excited units: type 1 responded with a discharge of 1-3 spikes at relatively short and constant latencies, while type 2 units were characterized by a burst of 3-14 spikes at longer and more variable latencies. Although the findings were compatible with the hypothesis that the anterior and paracentral nuclei transmit vagal impulses to the cingulate and supracingulate cortex, an analysis of latencies suggested that a more rapidly conducting pathway(s) accounts for latencies as short as 12 msec of some cingulate units. Twenty-eight percent of 367 units in the medial dorsal nucleus responded to vagal volleys. This finding gives substantial support to the traditional view that the medial dorsal nucleus transmits interoceptive information to limbic and neocortical areas of the orbitofrontal region.


Specific uptake of a behaviorally potent [3H]ACTH4-9 analog in the septal area after intraventricular injection in rats. Distribution within the brain of a behaviorally potent [3H]ACTH4-9 analog 2 h after intraventricular injection in rats was studied in the presence and absence of behaviorally and structurally similar peptides, to explore the significance of earlier found preferential uptake of the [3H]ACTH4-9 analog in the septal area. Hypophysectomy resulted in significantly enhanced uptake of radioactivity in the septum as compared to normal rats. No increase in this brain area of hypophysectomized rats was observed after intraventricular injection of [3H]Phe. Elevated circulating ACTH levels after adrenalectomy seemed too low to compete with the septal uptake of the ACTH4-9 analong. Subcutaneous substitution of hypophysectomized rats with sustained release zinc phosphate preparations of the behaviorally equipotent peptides ACTH1-24 and ACTH4-10 decreased the accumulation of the [3H]ACTH4-9 analog in the septum, whereas treatment with the behaviorally inactive fragment ACTH11-24 is not effective. Retreatment of hypophysectomized rats with neuropeptides, differing structurally from natural ACTH peptides (7-D-Phe-ACTH4-10, BETA-LPH61-76 and 9-desglycinamide, 8-Lys-vasopressin), did not change the uptake of the ACTH4-9 analog in any of the investigated brain areas. These results give evidence for specific uptake of the ACTH4-9 analog in the septal region, because competitive displacement occurs only with peptides which both behaviorally and structurally are closely related to the ACTH4-9 analog.


Sleep and activity rhythms in mice: a description of circadian patterns and unexpected disruptions in sleep. Studies on daily and circadian rhythms in wheel running and electrographically defined wakefulness, NREM sleep, and REM sleep in M. musculus were done to gather data on the temporal distribution of activity and sleep. Generally, peaks in NREM and sleep tended to coincide and to alternate with the coincident peaks of wakefulness and wheel running. However, during the active phase of the circadian wheel running cycle some NREM and REM sleep did occur; conversely, during its rest phase, wakefulness was often present. The most striking finding was that in mice with clearly entrained or free-running activity onsets, the circadian peak-through patterns in wakefulness, NREM, and REM sleep were not always distinct--they could be damped and/or polyphasic. Several explanations of these phenomena are considered.


ACTH release during passive avoidance behavior. ACTH was measured by radioimmunoassay during learning and retention of a passive avoidance response. Electric footshock, used during the learning trial, appeared to be a weaker stimulus for ACTH release than was the retention test. Moreover high latency scores during retention were associated with high plasma ACTH levels, whereas shorter latencies were associated with lower levels. The results indicate that psychological mechanisms organizing behavioral coping are important in the response of the pituitary-adrenocortical system to stimuli which are related to a previous adversive experience.