Experimental Biology and Medicine

A rapid and convenient method for the standardization of I¹³¹ solutions is described. A statistical evaluation of the method showed that the standard deviation of the mean for 10 samples was approximately 1%.

Pregnancy has been maintained in rats hypophysectomized on day 6 of their first pregnancy and injected daily subcutaneously for 6 days with placentae from rats, 12 days pregnant. Five placentae or approximately 275 mg daily accomplished this in the majority of the test rats. One 12-day placenta plus 0.5 μg of estrone proved equally efficacious. Pregnancy was not maintained by similar treatment if the rats were also oöphorectomized.

Administration of leukotaxine, a bacterial polysaccharide or an exposure to ionizing radiation induces increased capillary fragility. Injury to the capillary wall, whether it is caused by ionizing radiation, by leukotaxine, or by a bacterial polysaccharide, can be prevented to a considerable degree by the administration of a vitamin‘P'compound composed of flavonoids naturally present in citrus fruit.

Observations have been reported in which after the creation of large arteriovenous fistulae in dogs endocarditis occurred without intentional introduction of bacteria. This result occurred in about 8 out of 10 dogs in which sufficiently large shunts existed for more than 4 weeks. Adrenal gland enlargement occurred following creation of a large fistula. Other concomitant findings have been reported and discussed.

The plasma and blood volumes of mice and of various mouse organs were determined by injecting mice with a protein iodinated with iodine containing tracer quantities of radioactive iodine, and determining the amount of radioactivity present in the various organs. The average plasma and blood volumes of the mice were found to be 6.7 ml and 12.7 ml per 100 g of body weight, respectively. The average plasma volume in ml per 100 g of wet tissue for the brain was 1.6; kidney, 19.1; liver, 20.2; lung, 23.9; small intestine, 5.0; spleen, 9.2; submaxillary gland, 5.9; testes, 3.4.

Comparison of the average weight gains made by young, growing rats (offspring of B₁₂-depleted animals) on diets of varying fat and carbohydrate content, with and without B₁₂, shows that subcutaneous administration of B₁₂ produces increased weight gains in both sexes on all four of the rations studied. It appears that when the calories are derived primarily from carbohydrate the vitamin exerts a greater effect on weight gain in young, growing, female rats than in their littermate males. The weight stimulating effect of B₁₂ is more marked in females on high carbohydrate diets than in littermate sisters on diets of like protein content but with the calories provided by fat or a mixture of fat and carbohydrate

The administration of L-hydrazinophthalozine results in inhibition of sympathetic vasoconstrictor reflexes in man. In addition it has epinephrine-like effects on the heart. C-5968 differs from other “sympatholytic” drugs studied thus far in man.

An apparatus has been described permitting intracavitary radiation of hollow viscera accessible to intubation. In preliminary trials the technic has been used to induce achlorhydria in the stomach of dogs. This technic may have additional practical applications for lesions in the rectum, bladder, bronchi and other organs or tracts accessible to intubation.

The valuable suggestions of Dr. A. C. Corcoran and Dr. Otto Glasser, and the technical assistance of Dr. C. Ballinger are gratefully acknowledged. Assistance in fabrication of the intragastric bag was provided by the American Anode, Inc., Akron, Ohio.

Microbiological determinations of plasma values for arginine, glycine, histidine, lysine, phenylalanine, serine, and threonine have been determined in a group of normal individuals and compared with the values obtained from patients with rheumatoid arthritis.

1. The values obtained for arginine, histidine, and threonine in patients with rheumatoid arthritis were lower than those obtained from the normal group. These differences were highly significant 11 . 2. The values for glycine, lysine, phenylalanine and serine were not significantly different in the group with rheumatoid arthritis from those found in the group of normal individuals.

The free amino acids of sea-urchin (Strongylocentrotus purpuratus) eggs and embryos were extracted with 75% ethanol and identified by means of paper chromatography. Extracts of unfertilized eggs contain relatively large amounts of glycine and small amounts of alanine, glutamic acid, valine, threonine, lysine, glutamine, and leucine (or isoleucine) as well as several peptides and unknown ninhydrin-positive substances. The patterns of amino acid and peptide spots on chromatograms of extracts of cleavage stages, blastulae, gastrulae and prism larvae are dissimilar to that of unfertilized eggs.

The author is indebted to Mr. Arthur Karler from the Division of Medical Physics for advice on chromatographic technics.

Several free amino acids and one amide were identified by paper chromatography in alcoholic extracts of eggs and early developmental stages of Rana pipiens and Hyla regilla. Other ninhydrin-positive substances, including several peptides, will be studied later. One experiment on Hyla revealed no major chromatographic difference between explants of dorsal lip and ventral ectoderm.

Dogs, digitalized to cardiac toxicity will ouabaine (0.07 or 0.08 mg/kg) or with digitoxin 0.3 mg/kg) were anesthetized with cyclopropane, ether and thiopental sodium.

Cyclopropane improved or abolished the arrhythmias produced by digitalis preparations. Ether caused a reversion to normal rhythm. Thiopental sodium exerted no effect upon the electrocardiographic abnormalities of over-digitalized dogs.

The egg-shell membranes of the chicken, duck, and turkey contain 0.66 to 1.00% hydroxyproline depending upon species. The remainder of the contents of the egg contains no detectable amount of hydroxyproline. Hydroxyproline appears in detectable amounts in the chick embryo after the fourth day and increases to 1.16% of the dry weight on the nineteenth day. The serosa contains hydroxyproline after the fifth day which increases to a maximum of 2.1% on the fourteenth day. Most of the hydroxyproline of the embryo and serosa is present in a form extractable by autoclaving, probably as collagen.

When a solution of crystalline vit. B₁₂ was subjected to a series of reducing agents between pH 4 and 7, at which range this vitamin is stable, a marked loss of the microbiological activity occurred. This loss of microbiological activity can be attributed to the reducing power of the agents, but is not necessarily related to the disappearance of the intensity of the red color. Our preliminary data also demonstrate that the destruction of the microbiological activity was not accompanied by the destruction to an equal extent of the APF activity.

The authors are indebted to Dr. A. Langlykke of E. R. Squibb and Sons for some of the microbiological assays of vit. B₁₂.

1. Prostatic tissue obtained surgically from 100 consecutive males, who had lived for many years in an endemic area of brucellosis in Minnesota, were cultured for brucella and in no instance were brucella isolated. 2. On the basis of historical evidence, as well as positive intradermal tests and the presence of brucella agglutinins, many of the individuals had been exposed to brucellosis.

Concentrates of the Leuconostoc citrovorum factor (CF) and of “folinic acid” partially counteract the inhibitory action of aminopterin for the chick embryo. Folic acid and formylfolic acid were ineffective or only slightly effective under the same conditions. The magnitude of the effects obtained with CF were similar to those obtained previously with mixtures of thymidine and hypoxanthine desoxyriboside 1). The viewpoint is developed that conversion of folic acid to CF is a necessary preliminary to the catalytic action of the former compound, that synthesis of thymidine and one or more of the purine desoxyribosides are among the synthetic reactions for which CF or folic acid) is required, and that these synthetic reactions are those inhibited in the chick embryo by small amounts of aminopterin. The great variation that exists in the ability of CF to counteract aminopterin inhibition in various organisms is pointed out.

The intracellular distribution of nitrogen and nucleic acids in the livers of rats fed a diet free of protein for 5 weeks was found to be similar to that previously observed in rats subsisting on the same diet for 3 weeks, although the changes from control values were of greater magnitude in the case of the animals depleted for the longer period of time.

The inclusion of 1% DL-methionine in the protein-free diet had no appreciable effect in protecting the liver from the observed changes due to protein depletion except in partially preventing the increase of PNA in the residual cytoplasmic fraction. It is noted that the changes encountered in the intracellular distribution of nitrogen and nucleic acids in the livers of protein-deficient animals fall into the same pattern as the changes which have been found to obtain in the livers of precancerous and cancerous animals.

The authors are indebted to Dr. Maurice Ogur of Brooklyn College foro making available to them the details of the method for the extraction of nucleic acids prior to publication.

Premedication with moderate doses of dihydroergocornine greatly diminished the renal hyperemia produced in the dog by a bacterial pyrogen. No significant change of glomerular filtration rate was observed following administration of either the pyrogen or the alkaloid.

The feeding of 2% of phthalylsulfathiazole to rats maintained on a purified ration containing a commercial soybean protein and DL-methionine as the only source of amino acids produced no depression of growth; it supported better reproductive performance and no cases of acute uremia of the newborn were observed among about 200 rats. on the basal ration, acute uremia of the newborn occurred in 35% of the litters born.

An analysis of the phosphoryl-ated intermediates of the liver and brain of hypophysectomized rats with and without ACTH was made. The decreased ability of hypophysectomized rats to synthesize glycogen from glucose was restored to normal by ACTH. Phosphoprotein phosphorus levels also returned to normal with ACTH. The anaerobic glycolysis of the brain of hypophysectomized rats was reduced 20% below that of normal rats but this was restored to normal by the administration of ACTH.

The injection of non-epinephrine in the rat is more effective in causing an augmentation of water diuresis than is adrenalin. Pure 1-epinephrine has no stimulating action on diuresis.

Dibenamine causes a water retention in intact rats but fails to do so in adrenal medullectomized rats.

Dibenamine blocks and reverses the diuretic activity of adrenalin in intact rats but it less effectively blocks such activity of norepinephrine. Dibenamine, in the dosage used, was incapable of blocking the action of norepinephrine in medullectomized animals.

These results indicate that the diuretic action of adrenal medullary hormone is not due to epinephrine but to the presence of norepinephrine.

A study of the blood volume of the dog as measured simultaneously by the Dye T-1824, by carbon monoxide and by radioactive iron, Fe₅₅, has shown that the values measured by the cell-tagging methods are distinctly lower than the value given by the dye method whenever the donor blood tagged with CO or Fe₅₅ shows incompatibility with the blood of the recipient animal. However, when the bloods of the donor and the recipient animals show no incompatibility, the results by the three methods show improved agreement. These results suggest that the cell-dumpings occurring in the reaction of incompatibility, by making the smaller vessels inaccessible to the clumped cells, reduce the volume measured by the tagged cells.

The experiments herein reported corroborate the observations of other investigators that the mercury of an organic mercurial diuretic may be boud by the protdeins of the plasma. The extent of this binding is a function of mercurial concentration; at dilutions of the diuretic of 1: 1000 or higher the mercury is over 90% bound; at dilutions of 1:100 or lower, the mercury is over 50% free. This may well explain certan phenomena observed in the tracer studies of the mercurial diuretics.

It has been shown that the plasma iron of man undergoes a regular diurnal variation. These results have been compared with those of other investigators.

It was shown that dilute homogenates of rate organs catalyze in vitro the conversion of 3-phosphoglycreate to phosphoenol-pyruvate as measured by the formation of iodine labile phosphorus. A marked sex difference was observed between the rates of phosphopyruvate formation by male and female organs.

The methanol extract factor required by the mink has been found to be soluble in 90% phenol and insoluble in acetone, n-butanol and 3/4 saturated ammonium sulfate.

Fish solubles contain this factor, but dried distillers solubles, dried whey, pork spleen, and hog intestinal mucosa were found to be inactive.

The relationship between the methanol extract factor and vitamin B₁₂ is discussed.

The original observation that the mink requires a methanol insoluble factor as well as a soluble factor has been confirmed by the data presented.

We wish to acknowledge our indebtedness to Merck and Co., Rahway, N. J., for the crystalline vitamins; to Lederle Laboratories, Pearl River, N. Y., for the pteroylglutamic acid.

The utilization of the optically active and racemic forms of asparagine and aspartic acid by Leuconostoc mesenteroides has been studied under a variety of test conditions. D-asparagine was found to be entirely inactive, and DL-asparagine was found to have 50% the activity of L-asparagine. Conditions were described under which D-aspartic acid was essentially inactive and under which DL-aspartic acid had 50% the activity of L-aspartic acid. Under other conditions acid with Leuconostoc mesenteroides were suggested

Phlorhizin has been found to inhibit the intestinal absorption by rats of glucose, galactose, and possibly of mannose and sorbose, but not of fructose. This may indicate the presence in the intestinal mucosa of a specific enzyme for fructose phosphorylation which is not inhibited by phlorhizin.

The mouse toxic factor.of influenza virus, PR₈ strain, originally reported by the Henles was reinvestigated, and similarities and differences between their observations and ours are reported and discussed. The toxin also was shown to affect ferrets.

Quantitative data are presented on the urinary excretion of phenylalanine, phenylpyruvic acid and phenyllactic acid in twenty patients affected with phenylpyruvic oligophrenia. The values, expressed in mg per g of total nitrogen, show inconspicuous differences from patient to patient kept on similar diets. High protein diet or the ingestion of phenylalanine and phenylpyruvic acid result in an increased daily output of phenyl compounds. Data on sweat excretion of phenyl compounds are also presented.

The serum level of free phenylalanine and phenyllactic acid was determined in 18 patients suffering from phenylpyruvic oligophrenia. No significant amounts of the hydroxy acid were found. The concentration range of the amino acid was between 19 and 38 mg per 100 ml of serum with larger inter- than intra-individual variations. No correlation exists between the degree of mental deficiency and the concentration of phenylalanine in serum. Phenylalanine concentration in the spinal fluid varied from 6.1 to 8.2 mg/100 cc.

It is suggested that although the increased amount of phenylalanine is caused by the metabolic error, its blood plasma level is determined by the ability of the kidney tubuli to reabsorb the amino acid.

Cross serological reactions indicated that egg strains of influenza B virus obtained in different years were dissimilar in antigenic composition. The extent of the differences in antigenic pattern was analyzed. All strains studied were antigenically different from the mouse adapted Lee strain. The implication of these findings on the problem of immunization in man was discussed.

Ryanodine causes a high oxygen consumption in muscles which do not shorten. In those muscles that shorten the oxygen consumption falls off as the muscle shortens. The length of the muscle seems to regulate the oxygen consumption of the Ryanodine-treated muscle. Complete cutting off of the extra metabolism due to Ryanodine occurs at 30% of the rest length.

We are indebted to Dr. Roeder and to Merck and Co. for supplying us with the purified Ryanodine powder.

We wish to thank Dr. Robert W. Ramsey for his helpful advice and interest in this problem. We are also indebted to him for making many of the microscopic observations on the structure of the muscle at the conclusion of the experiments.

The antihistaminic, anticholinergic and musculotropic spasmolytic properties of the para-halogen, para-methoxy and some para-alkyl derivatives of 2-benzhydryloxy-dimethylethyl amine have been investigated. Para-substitution with methyl, ethyl and halogen atom results in an enhancement of antihistaminic activity, a decrease in acute toxicity, or both. It lowers the atropine-like action but does not produce a significant change in musculotropic spasmolytic activity.

Considerable evidence has accumulated that both the sodium ion and the adrenal cortex are concerned to some extent with the maintenance of blood pressure levels in hypertensive patients ( 1 , 2 , 3 ). Although the rigid restriction of salt masks the pressor response of hypertensives to desoxycorticosterone( 4 ), this type of observation does not establish the fact that alterations in sodium metabolism modify the arterial tension through an adrenal mechanism. It has been noted that the adrenals of rats on a reduced sodium intake may be smaller and different in color( 5 ), temporarily depleted of ascorbic acid( 6 ), and may show secretory changes( 7 ) and (in nephritic animals) subcapsular hyperplasia ( 8 ). Furthermore, excessive salt, or liberal amounts in animals receiving desoxycorticosterone, has resulted in progressive structural and functional atrophy of the glomerular zone of the adrenal cortex( 9 , 10 ). Finally, chromatographic patterns of urinary steroid excretion may be influenced by the salt intake( 11 ).

In order to investigate further the possibility that sodium influences the blood pressure through steroidal action, studies were undertaken in a patient with uncomplicated hypertensive vascular disease and Addison's disease who also had mild diabetes mellitus and rheumatoid arthritis.

Case report and methods. G.S., a 55-year-old, white housewife, was admitted to the metabolism ward of the Presbyterian Hospital for research study. Hypertension was first recorded 10 years before admission, with repeatedly elevated blood pressure readings up to 210/110 mm of mercury. Addison's disease developed one year before entry, as demonstrated by weakness, skin and buccal mucous membrane pigmentation and the disappearance of hypertension. The diagnosis was clinically apparent and was substantiated by the finding of repeated serum sodium values which were markedly below normal limits following salt withdrawal, calcification in the adrenal areas by x-ray and abnormally low 17-ketosteroid (2.9 mg) and neutral reducing lipid values (0.9 mg)(12)(†) in 24-hour urine specimens.

A synthetic ration in which all of the nitrogen is furnished as free amino acids is not as palatable to the mouse as one containing casein. The inferior palatability results in less food consumption during the first 3 to 7 days and consequently poorer growth. When the initial lag period is diminished or eliminated by the addition of 1% of monosodium glutamate to the ration, the final weights of the mice are more consistent and approach more closely those of the animals fed a ration containing casein.

A simple method is described for the purification of the egg-white inhibitor of influenza virus hemagglutination by filtration of diluted egg-white through rapid-flow filter papers and extraction of the unfilterable residue. The inhibitor has been purified about 80-fold in this way. The method has proved useful also for the further purification of acid-purified inhibitor. The advantages and disadvantages of the new method are discussed in relation to those of other methods.

The serums of 90 patients in various phases of viral hepatitis were tested for the presence of C-reactive protein, and the results were uniformly negative. The significance of this is undetermined but the question is raised as to whether it may be an expression of a fundamental difference in the pathogenesis of certain viral infections.

1. Erythrocytes from 2 individuals with sickle cell anemia, when injected into rabbits, produced antisera which agglutinated erythrocytes of 19 individuals with sickle cell anemia, but not those of 21 individuals with sickle cell trait nor those of 124 normal individuals.

2. Such sera may be useful in the differential diagnosis between sickle cell anemia and sickle cell trait.

3. The presence of agglutinin antibody has been demonstrated in the sera of all of 13 patients with sickle cell anemia. This finding may explain the hemolysis which is characteristic of this disease.

4. Of 11 patients with sickle cell trait, definite agglutinin antibody was demonstrated in the sera of 4, and doubtful agglutinin antibody in 2.

The inactivation of renal and intestinal alkaline phosphatases by HCl was studied by histochemical and by quantitative methods. In the mouse, guinea pig and dog the renal enzyme is inactivated at a distinctly higher pH than is the intestinal enzyme. These observations support the view that renal and intestinal alkaline phosphatases are not identical.

1. The addition of supplementary methionine, choline or cystine to an “adequate” basal diet failed to protect the livers of rats against damage produced by the chronic oral administration of carbon tetrachloride. 2. A supplement of methionine, but not of cystine or of choline, showed a protective effect against damage produced by feeding sodium selenate (selenium 20 p.p.m.). 3. This action was demonstrated only in the presence of alpha-tocopherol. The relationship is discussed briefly.

In a series of experiments it has been shown that male dba line 1 mice are relatively resistant to intravenous injection of the yeast phase of Histoplasma capsulatum. Intraperitoneal injection of enormous numbers of the organisms suspended in saline produced occasional deaths. Intraperitoneal injection of approximately the same numbers of viable organisms suspended in 5% mucin increased the death rate. Intracerebral injection, however, was far superior to any of the other routes employed, both with respect to the numbers of organisms necessary to produce death 30 days after injection and the percentage of fatalities obtained within this period of time.

1. The distribution of total ferritin in the intestine and mesenteric lymph nodes in a control horse was compared with the corresponding distributions in 2 horses fed iron, using an immunochemical technic for determining the ferritin in extracts of the tissues in question.

2. Ferritin was found to be present in the first 11.5 feet of intestine of the control horse as well as in the last 3 feet of intestine. Small amounts of ferritin were detected in the mesenteric lymph nodes of the control animal.

3. Twenty-four hours after the oral administration of an amount of ferrous ammonium sulfate containing 30 g of iron to a second horse the total ferritin content was appreciably increased in the first 2.5 feet of intestine while 48 hours after feeding the same amount of ferrous ammonium sulfate to a third horse, there were notable increases in the amounts of total ferritin in the first 11.5 feet and in the last 3 feet of intestine.

4. Twenty-four hours after feeding iron to a second horse there was approximately a 4-fold increase in the amount of total ferritin in the mesenteric lymph nodes based on the value for the control horse, while 48 hours after feeding iron to the third horse, more than a 5-fold increase was noted, as judged by the value for the control horse.

5. The data indicate that ferritin is involved in the phenomenon of iron absorption through the intestine of the horse, and that the lymphatic system is concerned with iron absorption by some process in which ferritin also plays a role.

1. Chemical measurements were made of the collagen content of the lungs, liver, kidneys, spleen, heart and skeletal muscle of 58 guinea pigs weighing between 60 and 1000 g. 2. The concentration of connective tissue components varied with different tissues; the lungs contained the greatest proportion, and the liver the least proportion, of collagen. 3. Tissues from older animals usually contained a higher per cent collagen than comparable younger tissues, except for the spleen where the converse was true. 4. The values obtained for guinea pig tissues were in essential agreement with previously reported figures for other species.

The administration of sulfasuxidine eliminates the growth response produced by vitamin C in the chick, on a semi-purified ration. No effect is observed on the growth response produced by vitamin B₁₂, if the levels of the other B vitamins are adequate.

The administration of sulfasuxidine to a semi-purified ration eliminates the increase of the PGA liver storage produced by vitamin C. Both vitamin C and vitamin B₁₂ raise the caecal concentration of PGA in the chick. The administration of sulfasuxidine lowers the caecal levels of PGA to a minimum in all cases. Sulfasuxidine has no effect on the caecal level of vitamin B₁₂ in the chick.

Vitamin C, when injected, has no effect on growth, liver storage or caecal concentration of PGA in the chick. The injection of PGA markedly increased the liver storage and caecal levels of both PGA and vitamin B₁₂.

Widely different degrees of stromatolysis were observed electron microscopically in erythrocytes which occurred in preparations of Treponema carateum taken from tissue fluid of skin lesions. A comparison of electron images with light microscopic images of cells under controlled conditions of stromatolysis showed striking similarities. Isolated appearances in the electron micrographs similar to all the stages of disintegration of hemolyzed erythrocytes reported by Furchgott from light microscopy are represented and observed.

Vitamin B₁₂-deficient chicks were fed vitamin B₁₂-deficient basal diets supplemented with 0, 1 and 4% glycine in combination with 0, 3, and 30 μg of vitamin B₁₂ (supplied by Merck's APF Supplement No. 3) per kilogram. The growth of the chicks that received either 1 or 4% added glycine in the vitamin B₁₂-deficient basal diets was depressed. Those chicks which received 4% added glycine without vitamin B₁₂ suffered excessive mortality. The addition of as little as 3 micrograms of vitamin B₁₂ per kilo of diet overcame the inhibitory action of both levels of glycine. These results indicate that vitamin B₁₂ functions in the metabolism of glycine.

Calcification in vitro of the hypertrophic epiphyseal cartilage was found to be inhibited by 4.8 × 10⁻⁴ M sodium desoxycholate, 1.9 × 10⁻³ M sodium cholate, 1.4 × 10⁻³ M Duponal C, and .08% Tween−20. By preliminary treatment of rachitic sections with higher concentrations of these surface active agents for a short time interval, it was possible to demonstrate inhibition only with the bile salts and Duponal C.

These experiments corroborate Hektoen's original classic findings that antibody formation is suppressed by total-body X radiation. It has been demonstrated, in addition, that if the spleen or appendix of the rabbit is protected by lead shielding during total-body irradiation, the capacity to produce antibodies to an injected particulate antigen is retained to a marked degree even though lymphatic tissue elsewhere in the body is temporarily destroyed.

1. The rate of serum protein formation as indicated by the incorporation of methionine labeled with S35 was determined in 4 patients with lipoid nephrosis and in two control subjects. 2. The rate of serum protein formation thus determined would appear to be greater in the nephritic patients than in the control subjects.

The effects of experimentally induced hypo-and hyperthyroidism on the ovarian response to pregnant mares' serum were determined in immature rats and mice by administering thiouracil for 4 to 20 days and thyroxine or thyroprotein for 10 days. A constant dose of PMS was injected into each animal during the last 4 days of each experiment and the increase in ovarian weight was measured. It was found in rats that all doses of thyroprotein or thyroxine significantly decreased the action of the gonado-trophin on the ovaries, while in mice thyroprotein significantly increased the ovarian response to PMS. When thiouracil was fed to rats for 4, 7, 10 and 15 day periods, there were significant increases in ovarian response to PMS, but when fed for 20 days thiouracil reduced the ovarian response to PMS. These data on young female rats and mice corroborate the results previously obtained in young male rats and mice. It is suggested that insofar as the ovarian response to PMS is concerned, young rats of both sexes secrete more and young mice less than an optimal amount of thyroid hormone.

The guanine analogue, 5-amino-7 - hydroxy - lH - v - triazolo (d) pyrimidine (guanazolo), which has been reported to be relatively non-toxic for adult mice, wras found to be toxic for the developing chick embryo.

This was manifested by a high mortality rate, inhibition of growth, and the occurrence of developmental abnormalities. The toxic effects of guanazolo were inhibited by the simultaneous injection of guanine HCl.

The intravenous injection of a purified preparation of the hyperglycemic factor of the pancreas produces an increase in the venous blood sugar of man. This response is variable in quantity and is not increased by increased dosage. The production of anesthesia enhances the increase in venous blood sugar produced by the hyperglycemic factor. This effect of anesthesia is attributed to an increase in peripheral blood flow rather than to an augmented hyperglycemic effect.

The yeast cell phase of Histoplasma capsulatum, when injected intravenously in suitable numbers, has been shown to yield a consistently reproducible fatal infection in the rat. The necessity for standardizing the inoculum on the basis of the number of viable organisms, rather than the absolute number of organisms, is emphasized.

1. Chick embryo-adapted mumps virus has been shown to infect the tissues of the guinea pig's eye after intraocular inoculation.

2. Mumps virus has been carried through 7 successive guinea pig passages by intraocular injection with the subsequent production of complement-fixing antibodies in the sera of guinea pigs in passages I, II and VII. A virus culturally and immunologically indistinguishable from the original virus has been recovered from ocular tissue of passages V, VI and VII by inoculation into chick embryos.

3. When doses of virus 6 to 10 times as large as those necessary to infect the anterior chamber were introduced into the plantar spaces, peritoneal cavity, or heart, antibody formation was low or absent as shown by titers of the complement fixing antibody in the convalescent sera.

4. The guinea pig appears to be a suitable animal for the study of mumps virus infections when inoculations are made into the anterior chamber of the eye.

1. Albino rats and rabbits were injected intraperitoneally with sterile mineral oil to produce peritoneal exudates.

2. Peritoneal exudates of rats contained greater numbers of white blood cells than rabbit exudates.

3. There was a significantly greater mononuclear phagocyte response to the same stimulus in rats than in rabbits. It can be postulated that this response may be related to the native resistance of the rat to tuberculosis.

We wish to thank Miss Mary Louise Wilcox for her technical assistance throughout this study.

Blood urethane levels using the method of Archer et al. were performed on patients receiving enteric-coated urethane tablets for the treatment of leukemia and multiple myeloma. Blood levels ranging from 0 to 27 mg per 100 cc of blood were obtained. The correlation between dosage of the drug, blood level and clinical response was poor. This negates the value of such determination in the regulation of oral urethane therapy.

It was recently reported that ACTH is capable of suppressing the phenomenon of local tissue reactivity, when administered intramuscularly 2 hours prior to the provocative injection of meningococcus culture filtrate ( 1 ). The experiments embodied in the following paper serve to extend these observations.

Experimental. Substances failing to inhibit or modify the phenomenon of local tissue reactivity to bacterial filtrates. The interpretation of the results of the present studies is facilitated by considering the failure of a variety of substances to inhibit or modify the phenomenon of local tissue reactivity.

In published( 2 ) and unpublished experiments by one of the authors (G.S.), the substances employed were administered repeatedly by various routes. The substances which failed to suppress the phenomenon were acetylcholine, adenosine-5-phosphoric acid, alypin, antiplatelet serum, amino acids in various combinations, ascorbic acid, atropine; benadryl, biotin, calcium chloride, calcium gluconate, casein hydrolysate, choline, cocaine, congo red, curare, DCA, dicumarol, distilled water, estradiol, ether general anesthesia, folic acid, glucose, heparin, hesperidin, histaminase, histamine, india ink, inositol, lemon “citrin,” milk, Niagara sky blue, nicotinic acid, pantothenic acid, paraminobenzoic acid, physostigmine hydrochloride, pilocarpine, progesterone, pyribenzamine? pyridin (in doses reducing significantly the blood platelet count), pyridoxal, pyridoxamine, pyridoxine, rat liver extract, rat spleen extract, riboflavin, sodium oxalate, sulfadiazine, sulfanilamide, sulfathiazole, thiamine, a-toco-pheroi, trypan blue, trypsin, urethane, vitamin K (water soluble and oil soluble preparations), wheat germ oil, and yeast extract. Becker( 3 ) showed that BAL, mapharsen, partial exsanguination and thyroidectomy failed to inhibit the phenomenon, while Smith and Humphrey observed no effect with anthi-san( 4 ).

Substances capable of suppressing the phenomenon of local tissue reactivity. The bacterial preparations used for the preparatory and provocative injections wrere meningococcus, 44B, “agar washings'” filtrates(2). Rabbits were prepared by a single intradermal injection of 0.25 ml of the filtrate diluted 1 : 2 and injected intravenously 24 hours later with 125 to 150 multiples of the minimal provocative dose.

Evidence of an increased rate of production of gonadotrophins in castrated mice with intrasplenic ovarian grafts has been obtained by means of parabiotic experiments. The ovaries of normal mice placed in parabiosis with female littermates that had been castrated and had an ovary grafted into the spleen from 30 to 50 days previously averaged 37% heavier than the corresponding ovaries when parabolic was performed simultaneously with the other operations. The duration of parabolic ranged from 9 to 13 and from 11 to 35 days, in the 2 groups, averaging 11 days in the former and 17 days in the latter.

The administration of Cortisone to 42 sensitized mice 18 hours before the intravenous injection of a challenging dose of antigen prevented fatal anaphylactic shock in 38 animals. In contrast, 2S of 34 sensitized but unprotected control mice died in anaphylactic shock.

The authors with to thank Mr. Bernard K. Friedman and Mrs. Jacqueline Isola for their technical assistance in this work.

The Cortisone used in this investigation was purchased by funds from The United States Public Health Service.

The subcutaneous administration of testosterone propionate depressed the rise in urinary nitrogen excretion following burns in male and female rats. The effect of testosterone propionate in reducing the total urinary nitrogen excretion of male and female adult rats is essentially unchanged by a thermal burn for a period of 8 days after the burn or onset of the testosterone medication. For 2 to 4 days after this period the effect of the drug post-burn is no longer evident. In normal female animals the effectiveness of testosterone propionate continues for at least 18 days whereas in normal males it disappears at 12 to 14 days. Possible reasons for this wearing-off of the effectiveness of testosterone propionate after burns are considered. Various problems connected with the possible use of testosterone. propionate in the damaged patient are discussed.

A part of this report is based on work done by Capt. Braasch in the Department of Physiology of the University of Illinois School of Medicine in partial fulfillment of the requirements for the degree of Master of Science in Physiology. The authors wish to thank the Schering Corporation for a generous supply of testosterone propionate.

A shock-like state was produced in rats by applying rubber band tourniquets to the limbs for a certain length of time and then releasing them. A progressive increase in blood amino acid and urea nitrogen occurred after the release of the tourniquets. Application of the rubber band per se had no effect on these blood constituents but was found to cause a definite stimulation of the adrenal cortex as indicated by a lowering of the adrenal cholesterol content. Release of the tourniquets produced a further decline in the adrenal cholesterol content. The “sparing” effect of 0.9% NaCl administration in lowering mortality of rats during “tourniquet shock” was found to be associated with a decrease in the elevation of blood amino acid concentration and in the fall of the adrenal cholesterol content when compared with untreated animals.

It has been demonstrated that slices of rat liver have caused the in vitro incorporation of radioactive orotic acid into the pyrimidine nucleotides of nucleic acid and not into the purines.

Three milligrams of orotic acid (labeled in the 4 position) were incubated with about 6 g of slices of rat liver for 5 hours at 37°C in a Krebs saline-phosphate buffer of pH 7.4. The solution was saturated with a 95% C0₂: 5% C0₂ mixture at the beginning and nothing was used to diminish bacterial action. The slices were homogenized after preliminary washing and the homogenate precipitated with 10% trichloroacetic acid. The extraction of lipids was then carried out with a 3:1 alcohol and ether solution. The nucleic acid was extracted from the protein with hot 10% NaCl and precipitated with alcohol. All of the dried impure nucleic acid (35 mg containing both r.n.a. and d.n.a.) was hydrolyzed with 0.75 cc of N HCl in a boiling water bath for one hour. This hydrolyzed the Purina nucleotides and left the pyramiding nucleotides unchanged. 0.04-0.06 cc were placed on each of 4 filter paper strips 12 cm wide and run with ascending columns of tertiary butyl alcohol (70% made 0.8 N with HCl, 30% water) (1). After drying, the papers were viewed with an ultraviolet Mineralite. The separated bands on paper were eluted with 0.01 N HO, and the solutions read with a Beckman ultraviolet spectrophotometer. The maximum absorptions and the ratios of densities (278-262 mμ for cytidylic and uridvlic acids; 248-262 for guanine and adenine) were determined in order to identify and quantitate the bases. The separate solutions were evaporated on plates and radioactivity determined with a windowless counter.

Approximately three-fourths of the hydrolysate from the above experiment containing Purina bases and pyrimidine nucleotides was adjusted to pH 8 and the guanine precipitate removed by centrifugation.

The effects of crystalline vit. B₁₂ on thiouracil action was determined in immature female rats for a 30-day period. The vitamin completely counteracted the growth-inhibiting action of thiouracil, and this was accompanied by a considerable increase in food consumption. Although vit. B₁₂ decreased the thyroid hypertrophy induced by thiouracil, the uptake of radioactive iodine (I¹³¹) by the thyroids was even less than in the rats which received thiouracil only. It is suggested that vit. B₁₂ may be able to induce normal growth in hypothyroid rats.

Crystalline vit. B₁₂ was fed to immature rats of both sexes in order to determine whether the vitamin could alter thyroid function in normal or thyroprotein-treated rats. The growth rate of the rats supplemented with the vitamin was increased above that of the normal or thyroprotein-treated controls, but there was no significant effect on thyroid weight or uptake of I¹³¹. It is concluded that vit. B₁₂ does not alter normal thyroid activity in rats.

1. In the oxygen unsaturated state the abnormal sickle cell hemoglobin molecules undergo orderly orientation, forming-by specific linkage of the individual molecules-long chains of hemoglobin elements. Subsequent parallel alignment of these elements results in birefringent tactoids.

2. The birefringent sickled erythrocyte is in all probability a membrane-covered hemoglobin tactoid.

3. The clinical and pathologic manifestations of sickle cell disease apparently follow as a consequence of the effects of the abnormal hemoglobin molecules upon the physical behavior of the erythrocytes.

We are grateful to Dr. W. B. Castle and Dr. T. H. Ham for valuable counsel during this work and the preparation of this manuscript. David F. Waugh, Associate Professor of Physical Biology at the Massachusetts Institute of Technology, aided in demonstrating the anisotropy of the sickled cells and of the hemoglobin tactoids and gave advice concerning the interpretation of the meaning of the toxoid form.

Guinea pigs were used throughout this study. The number of eosinophils in the blood of males was significantly lower than that in females. Cortisone reduced the number of eosinophils in the blood of males and females. ACTH produced a pronounced eosinopenia in males and nonpregnant females. Pregnancy abolished the eosinopenic effect of ACTH. Neither cortisone nor ACTH had any effect upon the degree of anaphylactic shock produced in guinea pigs by the intravenous injection of the agent to which the animals had been sensitized.

When growing bacteria, S. marcescens, are placed in contact with their specific antibody and with normal serum or gamma globulin there is an increased O₂ consumption by the former over the latter, particularly when C0₂ is absorbed.

Nutritional. experiments with Leptospira conicola have been described A semisynthetic medium containing dialyzed rabbit serum, salts, vitamins, amino acids, and purine and pyridine bases was developed and appeared to be adequate as a basis for determining amino acid and vitamin requirements of this organism.

1. The rate of respiration of a given volume of embryonic blood gradually decreases from the 14th to the 20th day of incubation. From this time until 29 days after the onset of incubation (the greatest age investigated) it remains virtually constant. 2. It is suggested that the decrease in respiratory rate is due to a decrease in the number of young cells having a high respiratory rate and a hemoglobin with a lesser affinity for oxygen. 3. Methylene blue shows no catalytic effect on the rate of respiration of embryonic blood of the chick. It will, however, counteract the inhibitory effect of KCN when present.

Yeast autolysates have been found to contain enzymes which catalyze the phosphorylation of ribose and of adenosine, when ATP is added as a phosphate donor. The enzymes, designated ribokinase and adenosine phosphokinase, have been partially purified, but the best preparations still contain large amounts of interfering enzymes, making it impossible to isolate and to identify the products of the reactions. The possible physiological significance of the reactions is discussed.

Protein digestibility of raw and autoclaved legume seeds were determined in growing rats and the trypsin inhibitor content of the same seeds was measured in vitro. The proteins of kidney beans, soy beans, lima beans, and hyacinth beans were 12-25% more completely digested when autoclaved as compared with the raw seeds. These samples had also the highest trypsin inhibitor activities.

Serum glycoproteins are increased in mice bearing transplantable tumors. Water soluble glycoproteins are increased in the connective tissue bordering and abutting on the tumor. These findings support the thesis that increased circulating glycoproteins arise from the ground substance of the connective tissue at the site of invasive growth by a process of depolymerization, whereby smaller, water soluble and diffusible glycoprotein moieties are produced.

A method for the quantitative estimation of Ac-globulin in plasma has been described. The method is based upon the use of three variables-conversion rate, thrombin yield, and onset of thrombin formation. With the use of all three variables combined, a greater degree of accuracy can be obtained.

A marked retardation in body and gonadal weight was observed in immature male rats fed a purified ration under conditions of low environmental temperature. Supplements of desiccated whole liver, water-insoluble liver residue or water-soluble liver extract resulted in a marked increase in both body and gonadal weight. A supplement of the known B vitamins was similarly effective. It is suggested that the protective effects of the various supplements were due, at least in part, to their vitamin B₁₂ content.

ω-Methylpantothenic acid has been shown to exhibit low acute toxicity toward rats, mice and chicks. Growth inhibition of Tetrahymena geleii H at high levels of this analog was reversed by pantothenic acid. Plasmodium lophurae infections in chicks on the other hand were unaffected by ω-methylpantothenic acid administered intraperitoneally at a level of 2 g/kg twice daily for 4 days or orally at 5 g/kg daily for 5 days. In chicks on a pantothenic acid-deficient diet, the simultaneous administration of ω-methylpantothenic acid did not alter the course of the infection with P.lophurae.

The feeding of 100 mg of aureomycin daily to lambs resulted in markedly decreased feed consumption and weight loss. Bacterial counts of rumen contents of lambs fed aureomycin and aureomycin plus vitamins was much higher than control animals, indicating that perhaps the aureomycin had destroyed certain strains of bacteria, thereby, eliminating a normal competitive environment and permitting less desirable strain of bacteria to multiply.

A vit. B₁₂ requirement was not demonstrated for the Syrian hamster when fed either a corn-soybean oil meal ration with or without iodinated casein, or a casein basal ration with or without iodinated casein. No increase in the rate of gain was observed when 2% whole liver substance was added to these rations. Parallel experiments were conducted with rats fed the corn-soybean oil meal ration plus iodinated casein in which a vit. B₁₂ requirement was demonstrated. A reduction in food efficiency was observed in rats attributable to the vit. B₁₂ deficiency.

Benadryl, Pyribenzamine and Dilantin were tested in rats for toxicity and for ability to modify maximal electroshock seizure pattern and to prevent Metrazol convulsions. The data indicate that all three drugs modify maximal electroshock seizure pattern but do not prevent Metrazol convulsions. The significance of the data is discussed.

1. Methadone (0.0005M) increases the oxygen uptake of liver slices rich in glycogen and depresses it in slices poor in glycogen. 2. The increased oxygen uptake is accompanied by an increased rate of glycogenolysis as demonstrated by biochemical and histologic methods. 3. A possible explanation for the findings is discussed.

1. l-norepinephrine and l-epinephrine produce vasodilation of the isolated surviving coronary arteries of swine.

2. 1-norepinephrine produces about 2^4 times the degree of vasodilation produced by l-epinephrine in the same artery in the same dose. This difference exceeds the anticipated difference attributable to differing molecular weights.

Excessive amounts of niacinamide, niacin or the alcohol of niacin were given to 21 diabetic rats. The latter two compounds caused a marked increase in the excretion of ketone bodies in 10 of these animals. The 11 remaining rats were fed a diet containing 20 to 30% fat in order to make them mildly ketonuric or bring them to the borderline of ketosis. When excessive amounts of the niacin compounds were then added to the high fat diets, 7 of the rats showed an increase in acetone body excretion, niacin next and niacinamide the least ketogenic

1. The optimal growth in tissue culture of macrophages and the exoerythrocytic forms of Plasmodium gallinaceum occurred between 7 and 11 atmospheres of pressure. Growth was reduced to 50% at 5 and 13 atmospheres.

2. High concentrations of serum (over 50%) were inhibitory to both the malarial parasites and the host cell, the macrophage.

Purified prothrombin can be dried from the frozen state without immediate loss of activity. Thereafter progressive loss of prothrombin activity occurs, characterized by refractivity to the action of calcium plus thromboplastin plus Ac-globulin. In about one year most of the prothrombin is altered and in addition some is insoluble in aqueous solution. Prothrombin altered as the result of the freeze drying technic can be activated autocatalytically in 25% sodium citrate solution. The altered prothrombin has essentially the same electrophoretic properties as purified prothrombin and represents a derivative of prothrombin not previously described.

The injection S. typhosa endotoxin appears to stimulate the liberation of adrenalin or adrenalin-like substances, bringing about peripheral vasoconstriction. Dibenamin, a sympatho-adrenal blocking substance, antagonizes this effect.

When the rate of growth of chicks is used to measure the nutritional efficiency of carbohydrates they fall in the following order: dextrin, cerelose, sucrose, and lactose. Cellulose, sulfasuxidine, reticulogen, fish solubles, or vit. B12 do not change the significant differences observed when these carbohydrates are fed. However, the lactose-fed birds do give the greatest response to reticulogen, fish solubles and the higher levels of vit. B₁₂. The excretion times of chickens fed these different carbohydrates are in the following decreasing order: dextrin, sucrose, and lactose. The possibility of these nutritional differences being explained by the synthesis of known and/or unknown factors is discussed.

The L.E. cell phenomenon has been produced experimentally in the skin of two normal human volunteers following inoculation of windows with the plasma of a patient with acute disseminated lupus erythematosus. In vivo, under the conditions of our experiments, the neutrophilic L.E. cell usually developed as the result of ingestion of other neutrophilic nuclear lobes or nuclei which had undergone a previous partial peculiar lysis. Less commonly the L.E. cell may represent the originally affected neutrophilic leukocyte in which lobar lysis is out of step in the individual nuclear lobes. The experiments described should provide a means of testing the proposed identity of the L.E. cells and the “hematoxylin staining bodies” of the diseased tissues in acute disseminated lupus erythematosus, an identity recently suggested by several workers (11,12).

In a controlled study, cortisone has been found to be antipyretic in rabbits given pneumococcal vaccine and a pseudomonas pyrogen. The mechanism for this effect is not known.

Virgin female mice receiving long term progesterone treatment attained an appreciably greater body weight than their untreated littermate controls. Male mice, both intact and castrate, receiving the same progesterone treatment failed to attain a greater body weight than their untreated controls. Progesterone exerted a protective action against the body weight suppressing effect of estrogen.

1. Ten samples of human milk collected from young, healthy, nursing women, whose sisters, mothers, or grandmothers had breast cancer, were examined with the aid of an electron microscope. Spherical particles, of a smooth surface and a high density† to the electron beam, varying in diameter from 20 to 200 mμ, in some instances grouped in pairs, or clusters, were found in all samples; they were particularly numerous, however, in 5 of the 10 samples examined. These spherical particles appeared to be similar to those previously observed in mouse milk known to contain the mouse mammary carcinoma agent.

2. Thirty-two control human milk samples, collected from young, healthy, nursing women having a family record apparently free from any malignant tumors for 2 preceding generations, were also examined with the aid of an electron microscope. Eleven of them were found to contain spherical particles essentially similar to those described above. Of the remaining 21 control milk samples, 17 were found to contain only occasional, isolated, single particles in some of the electron micro-scopic fields; the other 4 samples appeared to be free from spherical particles, but contained some unidentified debris.

3. No definite conclusions can be reached at this time as to the nature of the spherical particles revealed with the aid of the electron microscope in human milk.

1. The stock from which the host is taken is apparently important for the outcome of attempts to induce in mice experimental disseminated encephalomyelitis, for certain stocks of albino mice are susceptible and others, even though of related lines, are relatively resistant. 2. In 19% of the mice which reacted to homologous brain tissue-Freund type adjuvant mixtures, lesions characteristic of disseminated encephalomyelitis can be found in the central nervous system in the absence of any demonstrable objective symptoms of illness. 3. A diagnosis of the experimental encephalomyelitis should there-fore be based on the results of histological examination of the nervous system as well as on typical symptomatology. The question is discussed whether reactors might not have been overlooked among animals of other species hitherto employed, when the basis for the diagnosis of the encephalomyelitis was only the presence of outward signs of illness.

It is shown that the inhibitor present in allantoic fluid for hemagglutination by and absorption of mumps virus is more active when human erythrocytes rather than when chicken red blood cells are used. It appears that the species of erythrocyte present influences the reaction between mumps virus and inhibitor in the direction of more or less combined (non-hemagglutinating) virus. A similar influence, though to a less striking degree, is also shown for the red blood cell in the influenza virus-inhibitor reaction.

The development of fatty livers has been noted in 8-week-old fasted CBA male mice bearing a transplantable lymphosarcoma of varying age. Similar mice free of the tumor fail to develop fatty livers following a 48-hour fast under comparable conditions, nor does the growth of the tumor in fed mice result in fatty livers. Adrenalectomy has no inhibitory effect on the development of a fatty liver under these conditions.

Rheumatoid arthritis patients treated with ACTH showed a highly significant increase in urinary free threonine, lysine, and tyrosine as determined as the average and maximum 24-hour excretion. Cortisone-treated patients excreted a highly significant amount of threonine and tyrosine at the maximum, but lysine was not increased significantly. Arginine excretion was not significantly affected by either ACTH or Cortisone. Supplementary medication did not affect significantly the responses of the patients to ACTH or Cortisone, chemically or clinically. Clinical improvement in all patients was both subjective and objective. The cause of the increase in urinary excretion of the amino acids herein reported is not known and may or may not be associated directly with the metabolic changes brought about by the remission of rheumatoid arthritis.

Two sows which consumed synthetic diets and were given injections of approximately 400 μg each of vit. B12 reared their first litters. The weights of the pigs when weaned at 8 weeks were subnormal, 20 and 26 lbs., and there were other signs of mild nutritional deficiency. One of the sows received by injection 2,380 μg of the vitamin during a second gestation and lactation and reared a litter of 7 pigs with the unusual average weaning weight of 47 lbs. The other sow consumed during her second gestation and lactation a diet that contained a water extract of liver and weaned a litter of 7 with an average weight of 35 lbs. The inclusion of a liver extract in the synthetic diet was not more effective than was the injection of vit. B12 and the evidence indicates that swine can complete a normal life cycle on diets that contain no unrecognized nutrients.

Particles having the known dimensions of the influenza virus are observed in electron micrographs of thin sections cut from infected chorioallantoic membranes and mouse lungs. These particles are in groups and clusters apparently developing from the borders of membrane cells and from the walls of the alveoli.

The orotic acid requirement of Lactobacillus bulgaricus 09 can be met by ureidosuccinic acid (10-20% as active as orotic acid) or by 5-(carboxy-methylidine)-hydantoin (30-40% as active as orotic acid but not by any one of a large group of lated or unrelated compounds studied. The data are taken as microbiological evidence that ureidosuccinic acid and possibly ureidofumaric acid (derived from 5-(carboxy-methylidine)-hydantoin) are precursors of the pyrimidine ring.

Data on the distribution of orotic acid in nature are presented.

Various natural substances especially whey were found to contain a substance required for growth of Lactobacillus bulgaricus 09. In the survey of known compounds orotic acid was found to satisfy the requirement of this organism for natural material. The microbiologically active material was isolated from one of the natural sources, whey, and identified with orotic acid.

When blood fibrin (Armour) was the protein in the ration, on controlled food intake, increasing the concentration of mixtures of various components of the vitamin B complex, resulted in marked increases in the protein efficiency ratio, expressed as gains in body weight per gram of protein intake Summary. When blood fibrin

The hyperglobulinemia accompanying hypercholesterolemia produced by cholesterol feeding in the rabbit is predominantly due to beta-globulin.

The administration of a single dose of a combination of Vit. C and p-aminobenzoic acid to nephrectomized rats significantly increases the survival time.