Experimental Biology and Medicine

Isotopic cholesterol was administered per os for 30 days to cholesterol-fed White Carneau pigeons. At weekly intervals and for 100 days after cessation of isotope administration, subgroups were killed and their aortas were divided into normal tissue, fatty streaks, and atherosclerotic plaques. Cryostat sections from each plaque were examined for morphologic features, and from the different areas of the aorta cholesterol concentration and radioactivity were determined. From the values obtained, rates of influx and efflux of plasma cholesterol were calculated. The results indicated that in normal aorta the influx of free cholesterol greatly exceeds that of cholesteryl esters. There appeared to be no net positive cholesterol balance. In fatty streaks, the influx of both free and esterified cholesterol was greater than for normal tissue, and influx rates exceeded those for efflux. In plaques, still greater rates of influx and efflux were seen, and the influx of cholesteryl esters frequently exceeded that of free cholesterol, and like fatty streaks, plaques appeared to be in positive cholesterol balance. No relationship was observed between the morphologic and metabolic characteristics of the plaques.

The protein of low density rat serum lipoproteins contained 5.2% carbohydrate, consisting of galactose (33%), mannose (31%), glucose (9%), glucosamine (20%), and N-acetyl neuraminic acid (8%). The protein of high density rat serum lipoproteins contained 2.2% carbohydrate, consisting of galactose (23%), mannose (24%), glucose (8%), glucosamine (26%), and N-acetyl neuraminic acid (18%).

The Na⁺, K⁺, and Cl^- content of aqueous humor of the bullfrog is 100, 2.4, and 82 meq/kg of water, respectively. The [Na⁺] and [K⁺] of the corneal stroma are significantly higher than those of the aqueous humor while the Cl^- content of the stroma is essentially the same as that of the aqueous humor.

Studies were carried out to measure the effect of a synthetic double-stranded RNA, poly I:C, on SV₄₀ transplant tumor growth and SV₄₀ viral oncogenesis in hamsters. Single or multiple doses of poly I:C given intraperitoneally before and/or after subcutaneous inoculation of SV₄₀ virus into newborn hamsters had little or no effect on the percentage of animals developing subcutaneous tumors later in life. Likewise, single or multiple doses of varying amounts of poly I:C administered intraperitoneally before and /or after subcutaneous inoculation of 1000 to 500,000 virus-free SV₄₀ tumor cells had little or no effect on the rate of tumor appearance in adult male hamsters.

The authors are indebted to W. R. Clark, W. S. Collom, and T. A. Napier for technical assistance. Statistical computations were done by A. McLean.

RSV stocks in common laboratory use, BH-RSV and its pseudotype designated as RSV (RAV-1), were examined to demonstrate occult xenogenetic RAV by using genetically different host cells and antisera specific to subgroups A and B avian tumor viruses. The virus stocks contained a phenotypically mixed virus, i.e., subgroup A RAV genome enveloped in subgroups B RAV protein coat. The virus replicated in C/A cells and yields the progeny of subgroup A RAV. The progeny replicated in C/O cells serially but not in C/A cells. The phenotypically mixed virus could not infect C/AB cells and was neutralized by an antiserum specific to subgroup B viruses. The possible origin and the significance of the virus were discussed.

The authors are indebted to Dr. William H. Murphy, the University of Michigan, for his helpful comments and criticisms during the preparation of this manuscript.

The location of a descending vesicopressor pathway in the spinal cord was investigated in Macaca mulatta, lightly anesthetized (pentobarbital) and immobilized in a stereotaxic frame. The lateral columns of the spinal cords were explored with stimulating microelectrodes moved 250 to 500 μ between stimuli using dorsoventral penetration. Detrusor activity was monitored using a pressure transducer connected to an intravesical catheter. A descending vesicopressor pathway was outlined in the lateral funiculus of spinal cord segments C₆ through T₁ by mapping the points of maximum detrusor response. These points of maximum detrusor response were found in an area near the junction of the dorsal and ventral halves of the lateral columns extending from the region of the denticulate ligament laterally, to the spinal cord gray medially.

The author wishes to thank Dr. Edward W. Lauer and Dr. Charles L. Votaw in whose laboratories and under whose guidance much of this work was done. My appreciation goes also to Mr. William L. Brudon who prepared the drawings and photographs.

The effect of azaserine on liver lipid composition, on lipogenesis in vitro in liver slices and on serum lipoproteins, on liver content of ATP, biotin, and nicotinamide nucleotides in the rat is reported. The effect of ATP or adenine in azaserine-treated rats is also presented. The neutral fats in the liver of azaserine-treated rats significantly increased, while the incorporation in vitro of acetate-¹⁴C into total lipids decreased. In the same experimental conditions the liver concentration of biotin and nicotinamide nucleotides fell. A marked decrease of the levels of serum lipoprotein also was observed. The administration of ATP or adenine to azaserine-treated rats did not prevent either the development of fatty liver, the decrease of liver biotin and of serum lipoprotein, or the decrease of acetate-¹⁴C incorporation into liver lipids. On the contrary, the administration of ATP or adenine partially reversed the azaserine effects on the liver content of nicotinamide nucleotides and increased the liver concentration of ATP.

Mice infected with Candida albicans exhibited enhanced RES phagocytosis as measured by the clearance of colloidal carbon from the blood stream. Production of accelerated carbon clearance required living organisms. The time of onset of acceleration seemed to be dependent upon the size of the infecting inoculum and return to normal of the clearance occurred as the infection terminated. While accelerated clearance of colloidal carbon was produced by the infection, no change was seen at any time in the rate of clearance of microaggregated human serum albumin.

We thank Mr. Joseph Edelin for his expert technical assistance.

Asparaginase, injected in conventional therapeutic doses into mice previously inoculated with the lactate dehydrogenase-elevating virus (LDH-virus), disappeared from the peripheral blood at a significantly slower rate than in corresponding virus-free mice. Direct testing of the transplanted tumors and leukemias employed in the earlier studies on the antitumor properties of asparaginase have indicated that the virus, was unknowingly present. The evidence indicates that the virus, through its impairment of host enzyme clearance and possibly other factors, played a significant role in the observed therapeutic effects of EC-2 l-asparaginase against sensitive mouse leukemia and lymphosarcoma.

We thank J. D. Loveless, M. A. Fitzmaurice, and Edith R. Shapiro for assistance.

A method for the isolation of rat 7S gamma globulin based on multiple adsorptions with DEAE-Sephadex A-50 was presented. The separation of rat 7S globulin requires a procedure modified from the one used for humans in terms of the buffer pH, and the number of adsorptions. Ultracentrifugal analysis indicates the resulting fraction is a single component. In addition, immunoelectrophoresis of the fraction when tested against antirat serum confirms the homogeneity of this 7S fraction. A monospecific antiserum to rat 7S gamma globulin was produced. This antiserum is free of reaction with all other serum components when subjected to immunoelectrophoresis against normal rat serum.

A comparative study of serum β-lysin and muramidase activity in germfree, ex-germfree and conventional rats showed no significant differences in β-lysin levels between the three groups; however, upon separation and purification of β-lysin from germfree serum, unusual differences in the bactericidal activity of the different fractions obtained were noted. The CV rats showed significantly higher levels of muramidase than exGF or GF rats and these differences were closely correlated with differences in blood granulocyte counts. Higher levels of both β-lysin and muramidase were found in exGF male rats in contrast to exGF females.

We thank Dr. David W. Ailing, NIAID, NIH, for the statistical analyses and Drs. John E. Tobie and Phillip J. Baker for their criticisms of this manuscript.

The clearance of C. albicans organisms from the blood in Lewis rats is not altered in neonatally thymectomized rats despite the presence of profound immunological disturbances. The selective clearance of C. albicans by the lung is also unimpaired in these rats.

The rates of recovery of mouse spleen and marrow cells after sublethal irradiation were studied. Cells from donors given 500 R were transferred to lethally-irradiated recipients at various times after donor irradiation. Donor spleen cells were given with sheep erythrocyte antigen (SRBC), while donor marrow cells were given with normal thymus cells and SRBC. The recovery rates of cellular proliferative ability, hematopoiesis (by ⁵⁹Fe incorporation) and immunocompetence (by antibody formation to SRBC) were measured. The cellular proliferative ability and hematopoietic recovery were parallel; marrow cells had recovered by day 11 after irradiation (following an “overshoot”) while spleen cells had also recovered by day 11 but without overshoot. The marrow and spleen recovery rates of immunocompetence were markedly different, however. By day 11, marrow immunocompetence had reached normal levels while spleen immunocompetence was only 3.4% of normal.

Rhinovirus-infected HeLa suspension cultures did not increase in cell number and lost viability late in the infectious cycle. Maximum adsorption occurred within 30 min with approximately 40% of the virus being adsorbed. Yields of intracellular virus of about 8-10 PFU/cell were obtained at 12 hr after infection. Comparable levels of extracellular virus were not obtained until 30 hr after infection. The suspended cell culture system should have practical value in studies on replication and for obtaining concentrated preparations of virions.

The ability of soterenol and isoproterenol to reduce endotoxin lethality in chick embryos and mice was compared. Soterenol was significantly more potent than isoproterenol at lower concentrations in both models. At higher concentrations (10 mg/kg in mice of 0.075 μg/egg) the difference between the drugs was not significant. The role of β-adrenergic stimulators in combating endotoxin lethality was discussed.

The authors wish to acknowledge the technical assistance of R. L. Stratman and the advice of K. W. Dungan.

The influence of various cardio-depressant and beta adrenergic blocking drugs on the daunomycin induced bidirectional arrhythmia in the hamster was determined. Diphenylhydantoin reverted the arrhythmia to sinus rhythm with minimal decreases in heart rate and blood pressure. Quinidine exerted some antiarrhythmic activity but the resulting ECG patterns were not normal in most cases. In addition, the higher doses produced a persistant decrease in heart rate and blood pressure. These results support the evidence that the antiarrhythmic action of diphenylhydantoin is different from quinidine. Other quinidine-like agents such as lidocaine and procainamide exerted minimal antiarrhythmic activity. The most effective agent tested racemic propranolol antagonized the arrhythmia at lower doses than dexpropranolol although each isomer decreased heart rate. Both d(—)-INPEA and dl-MJ 1999 exerted almost no antiarrhythmic activity. These results tend to indicate that antagonism of the daunomycin arrhythmia is due to actions other than beta receptor blockade but in the case of racemic propranolol these actions are enhanced if simulataneous beta adrenergic receptor blockade also occurs.

The authors are grateful to the following suppliers of compounds: Dr. W. Murmann of Selvi and Company (Milano) for isomers of INPEA and Dr. Alex Sahagian Edwards of Ayerst Laboratories (New York) for isomers of propranolol.

Immune globulin synthesis by sensitized lymph nodes, obtained from mice treated with cyclophosphamide, following the application of subcutaneous grafts of sarcoma I was studied by the technique of radi-oimmunoelectrophoresis. There was complete suppression of labeling of 7S γ₁ during the homograft reaction as well as almost complete suppression of immunoglobulin synthesis during the first 7 days after grafting. Labeling of 7S γ₂ and IgM appeared on the 10th day after the application of the graft. These findings differed from those obtained with untreated tumor-bearing murine hosts.

Intravenous infusion of histamine phosphate in intact lightly anesthetized dogs produced a consistent increase in pressure in small pulmonary veins, slight reduction in stroke volume and decrease or lack of change in left atrial pressure. The rise in pulmonary venous pressure was due to an active increase in tone of these veins without change in their caliber, since the pulmonary blood volume did not change significantly.

Rats were prepared with chronic ig and iv catheters and fed ad libitum. The effect of ig loading of hypertonic glucose or NaCl on food intake was studied with and without concomitant iv infusion of distilled water. With water infusion, the depression of food intake induced by ig glucose was not affected, while the depression induced by ig NaCl was significantly attenuated. It was concluded that the depression of food intake following ig glucose is not the result of osmotic stress.

Absorption of ⁵⁹Fe given with antibody to gastric mucosa, was studied in rats with and without induced gastric atrophy. Whereas mild but significant depression of absorption occurred in rats without gastric atrophy, profound depression occurred in rats with gastric atrophy. This reduction in absorption could be largely corrected by concurrent administration of normal gastric juice.

Plasminogen activator activity (bilokinase) was demonstrated for the first time in fistula bile of the hamster as well as in the bile of isolated hamster liver. Sodium taurocholate was shown to stimulate the release of bilokinase into bile. Bilokinase in bile is analogous to urokinase in urine, but its role in coagulation homeostasis or in gallstone formation remains to be determined.

Acknowledgment is made of the expert assistance of Mr. Emery Van Hook in preparing the isolated liver system and of Miss Linda L. Livingston in laboratory determinations.

The generation time of marrow nucleated erythroid cells was estimated with a colchicine technic in adult and 6-day-old Sprague-Dawley rats. Extrapolation of metaphase accumulation to 100% provided comparable estimates for these age groups. Cell cycle parameters were: generation time (t_G )≈ 10.5 hr: mitotic interval (t_M ) 40 min adult vs 60 min 6 day old; metaphase interval (t _Metaphase) 31 min adult vs 53 min 6 day old. The higher erythroid mitotic index (MI) in the neonatal animal appears best explained by a longer duration of mitosis. Neonatal animals were more sensitive to the stathmokinetic effects of colchicine. The change in sensitivity to colchicine appeared to parallel the transition of neonatal to adult erythropoiesis.

Nephrotoxic sera were prepared in rabbits with rat kidney cortex homogenate in complete Freund adjuvant. The γ-globulins were precipitated with (NH₄)₂SO₄ and fractionated through gel filtration in Sephadex G-200. Antibodies responsible for the production of pulmonary edema in rat and capable of fixing complement were consistently associated with the 19S form of immunoglobulins; both these activities were destroyed by reduction with 2-mercaptoethanol followed by alkylation with iodoacetamide. The antibodies associated with the 7S immunoglobulins were revealed by gel double-diffusion analysis, possessed low capacity to fix complement, and only occasionally produced slight symptoms of pulmonary edema. The 19S antibodies were present in rabbit serum 8 days after the immunization and reached peak titers by days 15–18. The 7S antibodies, on the other hand, only were present 1 week after the booster.

At 5, 7, 9, and 11 ATA of 100% O₂: Cysteine + succinate offers the greatest protection against O₂ toxicity. Succinate is the most effective single compound for protection against O₂ toxicity. The failure of GSH to sustain its degree of protection at 7, 9, and 11 ATA is thought to be due to insufficient time available to permit conversion of GSH to succinate due to the increased severity of stress. The acid-base buffer, Tris, was less effective at 7, 9, and 11 ATA than -SH group protectants. Malate, a NAD-linked substrate offers no protection against O₂ toxicity.

We thank Marvin and Julie Nunn for valuable technical assistance.

Fourteen percent of guinea pig uteri survived freezing to a core temperature of —14° but none to —16°. Dose-response studies utilizing DMSO, dextran, and MgCl₂ as cryoprotectants were conducted on guinea pig uteri frozen to —16°. Uteri treated with MgCl₂ did not survive freezing. Uteri treated with dextran in concentrations as high as 20% survived freezing to —14° but not to —16°. The optimal dextran concentration, 10%, allowed a 62% survival at —14°. Uteri treated with 2 M DMSO survived freezing to —16°; and when treated with 5 M DMSO, survived cooling to —50°.

An isotope equilibrium technique was used to measure mitochondrial thyroid hormone concentration in thyroidectomized, partially thyroidectomized, and intact rats in order to correlate this with the P:O ratios in the mitochondria from these animals. The data suggest that thyroidectomized rats have P:O ratios higher than normal rats. Furthermore, there appears to be an inverse linear relationship between mitochondrial thyroid hormone content and the P:O ratio.

The two beta-adrenergic receptor blocking drugs, propranolol and sotalol (MJ-1999), were studied for their effects on myocardial oxygen consumption (MVO₂) and hemodynamic functions in anesthetized, open chest dogs. Propranolol (0.5 and 1.0 mg/kg) and sotalol (5.0 mg/kg) iv produced a significant decrease in MVO₂, although the effect of the latter drug was greater in magnitude and persisted longer. No significant effects on arterial or left ventricular blood pressure were produced by either drug.

The decreases in MVO₂ were associated with a negative chronotropic and inotropic effect, the former change being more pronounced with sotalol. Coronary sinus blood flow (CSBF) also was decreased by both drugs, with sotalol having a greater effect than either of the doses of propranolol. The lowering of MVO₂ by sotalol can be attributed largely to its negative chronotropic effect. This is supported by studies with the (+) -isomer (relatively weaker as a beta-blocker) showing significantly less change in MVO₂ compared to the recemic drug. In addition, racemic sotalol failed to significantly lower MVO₂ in heart rate-controlled experiments. The decrease in CSBF could be explained by the lowering of the oxygen demands of the heart accompanying heart rate and contractility changes, although a direct coronary vasoconstriction mechanism may also contribute to the observed decrease in CSBF. These results are discussed in terms of the possible application of these drugs to the treatment of ischemic heart disease.

Data and statistical computations were provided by the Research Data Facility, Pacific Medical Center (Dir., R. Abbott). Sotatol was donated by the Mead Johnson Research Center, Evansville, Ind.

Hemagglutination titers of cholera antitoxin (rabbit sera) determined with sheep erythrocytes sensitized with crude or purified cholera toxin correlated with the antitoxin levels as determined by the rabbit skin permeability factor neutralization test. Tannic acid treatment of erythrocytes was unnecessary. Both fresh and formalinized erythrocytes were satisfactory, and the latter, sensitized with toxin, could be stored without loss of sensitivity for at least 60 days. The presence of “somatic” antigen in crude toxin did not interfere with the specificity of the test.

In rats with chronic gastric fistulas, secretin had no effect on histamine-stimulated acid secretion. This finding, together with our earlier studies showing that secretin inhibited pentagastrin-stimulated acid secretion but not the increase in histidine decarboxylase activity, cannot be reconciled with the hypothesis that gastrin stimulates acid secretion by releasing histamine.

We are grateful to Dr. Morton I. Grossman for advice and support and to Dr. Lawrence W. Way for help with surgery. This study was supported by Veterans Administration Research Funds. Dr. Johnson was supported by NIH Training Grant No. 5 TO1 AM05470-05. Dr. Johnson's current address is the Department of Physiology and Biophysics, University of Oklahoma Medical Center, Oklahoma City, Oklahoma 73104.

The endogenous pyrogen from rabbit polymorphonuclear leukocytes was partially purified by butanol-methanol and elution from Sephadex G-200. This protein in addition to increasing the body temperature of the rabbit also lowered the plasma iron concentration in endotoxin-tolerant rats.

The effects of phenoxybenzamine and propranolol on norepinephrine anti-ADH action were studied in water loaded rats. Norepinephrine blocked the ADH anti-diuresis. This norepinephrine effect was blocked by phenoxybenzamine but not by propranolol. These observations suggest that different adrenergic receptors may have differing functions in the kidney with the alpha receptors supporting an anti-ADH effect.

EB, P, and R significantly reduced milk yield in the day 14 lactating rat. Relaxin acts synergistically with the steroid hormones to reduce milk yield below that observed with EB and P alone. It is suggested the ovarian hormones act by inhibiting the normal action of oxytocin on the mammary gland during milk-ejection reflex.

The effect of cardiac glycosides on rat skeletal muscle intracellular action potenials was studied and correlated with changes in serum and muscle electrolytes and water. In the early minutes after subcutaneous administration of ouabain, 25-200 μg, there was a significant shortening of depolarization and repolarization times. After 15-min repolarization lengthened and after 50 min depolarization also becomes prolonged. The early phase of shortening is more easily demonstrated with 25-50 μg doses, and the latter prolongation of the action potential is more marked and consistent with the larger doses.

There is also a biphasic action on muscle water content. This is apparently an increase in intracellular water as no change was seen in inulin space. The time relationship to this effect is unrelated to the time of changes in action potentials. Muscle water was increased for the first hour after injection of ouabain, returned to normal 1 to 2 hr after huge doses of ouabain, 400-600 μg, and was decreased 5 hr after the last of 3 doses of digitoxin over a 24-hr period. Muscle sodium increased significantly and progressively from 15 to 120 min after ouabain and was normal 5 hr after 3 doses of digitoxin. Muscle potassium was decreased after 15 min and became progressively lower over the next 105 min. It was also low 5 hr after 3 doses of digitoxin. Serum and muscle electrolyte and water changes are not demonstrable during the phase of shortening of the stimulated action potential. Prolongation of the action potential correlates in time only with diminution of muscle potassium content. Shortening of the action potential is thought to represent the therapeutic effect of cardiac glycosides, prolongation the toxic effect. Possible correlations with increased active sodium transport and with inhibition of Na and K dependent ATPase are discussed.

1. Marrow cells and thymus cells which had been exposed in vivo to SRBC antigen interact synergistically to produce hemolysin plaque-forming cells (Table II) (2).

2. The possibility of similar thymus-marrow cell interaction in producing cell mediated responses was tested in a variety of systems.

3. Synergistic interactions were not observed: (a) in the restoration of homograft reactivity of sublethally irradiated mice (Fig. 1); (b) in the production of graft-vs.-host responses as measured by splenomegaly (Figs. 3, 4, and 5) or of secondary disease (Table III) (cf. 4); and (c) in the restoration of the homograft response in sublethally irradiated, thymectomized recipients injected with marrow cells and with spleen cells from irradiated mice which had been injected previously with thymus cells and “weak” isoantigen (non-H-2).

4. A nonspecific stimulation of the homograft response occurred under conditions similar to 3c above but where the isoantigenic disparity was strong.

5. This nonspecific stimulation and the resuits of Globerson (9) which show a synergism between thymus and marrow cells in the production of in vitro GVH responses, offer perhaps the only known evidence that thymus–marrow cell interactions are involved in cell-mediated immunological responses.

6. Specific syngergistic interactions so evident in the initiation of the hemolysin response do not occur when cell-mediated immunological responses are tested under similar conditions. This does not definitely rule out the possibility that thymus–marrow cell interactions are involved in cell-mediated immunological responses.

7. Under certain conditions, thymus and bone marrow cell populations, like lymph node cell populations are each capable of conferring cell-mediated immunological competence. Hence these cell populations may have either one cell type capable of responding to isoantigen, or two or more cell types which interact to produce the response.

Blood levels of PGA and cyano-cobalamin and the urinary excretion of PGA and CF were estimated in scorbutic and paired-fed normal guinea pigs. The urinary excretion of Figlu after administration of histidine also was determined. No significant changes in the blood levels of PGA and cyanocobalamin or in the urinary excretion of PGA, CF, and Figlu were observed in the vitamin C-deficient guinea pigs. The intermediary metabolism of PGA did not seem to be affected as a result of vitamin C deficiency. It seems that in guinea pigs, unlike rats, ascorbic acid has a limited role in the conversion of PGA to CF.

The intraperitoneal toxicity of nicotine was studied in bats (Myotis lucifugus). The LD₅₀ was 39.7 mg/kg (33.1–47.7 mg/kg; 95% confidence limits). Thus, nicotine appears to be less toxic to the bat than to the mouse, rat, or rabbit.

Direct quantitative observations of vascular diameters in unanesthetized bats were made with a microscope combined with a closed circuit television system. The diameter of innervated minute arteries markedly increased with intraperitoneal nicotine administration. Minute vein diameter slightly increased while minute vein vasomotion slightly decreased. Sympathetic denervation abolished the increase in minute artery diameter when nicotine was administered. However, minute vein diameter decreased and was also accompanied by decreased minute vein vasomotion. Nicotine caused an increase in heart rate which was not found to be significant. These data suggest that nicotine may produce vasoconstriction by direct action on the peripheral vasculature, and that minute vein dilatation may have been mediated via the sympathetic system. However, we cannot presently exclude the possibility that a passive dilation of the minute vein occurs for the innervated series because minute artery dilatation may have increased minute vein pressure and flow. Since a decrease in vasomotion has usually been found to accompany an increase in minute vein diameter, our data suggests that minute vein diameter and vasomotion can be manipulated independently.

Nicotinate is readily converted to NAD in animal tissues. The pathway involves nicotinate mononucleotide and nicotinate adenine dinucleotide as intermediates (1, 2). Animal tissue also contains enzyme(s) having NAD glycohydrolase activity that breaks NAD down to nicotinamide and adenosine diphosphoribose (3). The administration of low levels of nicotinate to mice results in a marked increase in hepatic nicotinamide levels (4) presumably from NAD via NAD glycohydrolase. This observation that nicotinate appears to be a more efficient precursor of hepatic nicotinamide than nicotinamide itself motivated us to explore the matter further. The results presented here indicate that hyperphysiological levels of hexestrol cause a marked alteration in the means whereby nicotinate is converted to nicotinamide in mice.

Materials and Methods. The C57 black male mice (25-35 g) received hexestrol or sesame oil intramuscularly as described in the tables. The ¹⁴C-nicotinate was administered by intravenous injection at the level indicated. The animals were sacrificed by cervical rupture at the time indicated, the livers immediately removed and dropped into a dry ice-acetone freezing mixture. Individual frozen livers were dropped into 3 ml of boiling 0.04 M potassium phosphate buffer, pH 5.4. After 1 min in a boiling water bath the samples were homogenized for 30 sec, cooled in an ice bath and centrifuged at 21,000g for 15 min. Carrier NAD, nicotinamide, and nicotinate where then added to the supernatant fluid and aliquots were applied to sheets of Whatman 3MM paper. The sheets were developed in either the isobutyrate-NH³ solvent of Magasanik et al. (4), or in butanol saturated with water containing solid NH₄HCO₃ (5). The spots corresponding to NAD, nicotinamide, and nicotinate were cut out and quantitated for radioactivity (6).

Result and Discussion. The intravenous administration of tracer amount of ¹⁴C—nictinate (1.5-7.5 μg) (Table I, Fig. 1) results in the following events: (i) The intrahepatic levels of labeled nicotinate decreases rapidly during the first few minutes after intravenous injection.

The litters of lactating rats were adjusted to 8 pups on the day of parturition (day 1 of lactation) and various methods of cervical stimuli were performed. The litters were weaned at regular intervals and vaginal smears were examined for the return of estrous cycles. Thirty percent of mothers who either were not suckled or were suckled for 48 hr showed diestrus smears for 10 days or more. Mothers whose litters were removed on days 6, 9, and 12 of lactation did not exhibit an extended diestrum. Mechanical stimulation of the cervix of mothers weaned on day 3 of lactation did not increase the incidence of extended diestrus from the controls; electrical stimulation, however, almost doubled the number responding (50 vs. 30%). If the initiation of vaginal smearing was delayed from the first day of lactation until the third, none of the animals had pseudopregnancy-like vaginal smears. Estradiol administration on day 1 of lactation resulted in 60% of the animals exhibiting an extended diestrum. Combining both estradiol injection and mechanical cervical stimulation resulted in a slightly lower percentage (40%). The suckling stimulus is believed to induce an acute pattern of prolactin secretion while cervical stimulation induces a chronic pattern. The summation of these two secretory patterns may be observed early in lactation.

The author wishes to acknowledge the technical assistance of Mr. Jon Maguire in these studies.

The amino acid composition of the perfusate of an isolated rat liver varies markedly, and highly individually for given amino acids, over a 6-hr period. Glutamic acid, valine, isoleucine, and leucine concentrations increased whereas the concentrations of aspartic acid, proline, glycine, alanine, methionine, phenylalanine, lysine, and arginine decreased. In livers bathed in warfarin (10 μg/ml), vitamin K₁ added at 2 hr in a concentration of 25 μg/ml caused a prompt and linear increase in prothrombin. The specific activity of L-leucine-1-¹⁴C added to the perfusate at zero time declined exponentially to 4% of initial values as the concentration of total leucine increased 5-fold. The prothrombin synthesized under these conditions was highly radioactive, suggesting that vitamin K stimulates de novo synthesis of this coagulation protein.

The authors wish to thank Miss Lourdes Maglasang and Miss Katherine Compagno for able technical assistance.

Pyran copolymer, administered with sheep red blood cells to mice, enhances the early rate of appearance of antibody-forming spleen cells. Although the cytotoxicity and pyrogenicity of pyran and bacterial endotoxins are similar, pyran, in contrast to endotoxin, does not trigger a nonspecific initiation of antibody formation; it only enhances specific responses. The major effect of pyran appears to be on macrophages.

Human heart muscle cells are diploid at birth. In agreement with other data, they become tetraploid in normal adults and develop greater polyploidy in adult hypertrophy. Some cases of infantile hypertrophy appear to have mostly tetraploid nuclei. In keeping with the enlargement of muscle cells which occurs during growth, the DNA/protein ratio is high at birth, progressively decreasing until adulthood, after which it does not change in hypertrophied hearts except in clinically unusual conditions. This stability of the DNA/protein ratio in hypertrophy thus appears due to a concomitant increase in DNA and protein in the muscle cells.

Thanks are due to Jacqueline Cerniak and Hellen Ellis for skilled technical assistance and Dr. George M. Hass for encouragement and support.

Rat thymus cells were used as target cells in Jerne agar plaque technique to detect and enumerate antibody-forming cells of mouse stimulated with this antigen. ⁵¹Cr-labeled rat thymus cells were also used in these experiments to carry out plaque assay autoradiographically. The techniques demonstrated complement-dependent cytolytic plaques caused by antibodies released from immune mouse spleen cells and could be used to investigate kinetics of antibody-forming cells.

Achylia gastrica induced in growing rats by X-irradiation of the exposed stomach led to iron depletion despite a normal diet. This implied that gastric secretion or acid was essential for the optimal assimilation of dietary iron by rats.

Use of either incomplete Freund, alhydrogel, or pertussis vaccine as adjuvants together with rat thyroid extract (RATE) elicited neither antibodies nor thyroiditis when injected into Lewis rats. A complete Freund adjuvant emulsion with RATE gave inconsistent results. Multiple, spaced injections of such an emulsion elicited a severe thyroid reaction. Combination of pertussis vaccine with either incomplete Freund adjuvant or alhydrogel elicited moderate thyroiditis. The most marked thyroiditis was obtained using a combination of CFA and RATE emulsion and a separate injection of pertussis vaccine.

The authors thank Mr. Steven Buck for his expert technical assistance. The pertussis vaccine was donated by Dr. H. B. Devlin of Parke, Davis & Co., Detroit, Mich. The alhydrogel was donated by Dr. Niels Neergaard, Superfos Export Co., Copenhagen, Denmark.

Cyclophosphamide given in sublethal doses for 1 year reduced the numbers of oocytes, follicles, and corpora lutea in mouse ovaries. The mechanism, unknown at present, is presumably related to that causing ovarian suppression in humans.

Further studies on the mode of action of streptozotocin and the prevention of its effects by nicotinamide have yielded the following results: 1. The injection of nicotinamide at varying time intervals after streptozotocin afforded partial protection against its β-cytotoxic effects, decreasing with the length of the intervals but still significant when the interval was 2 hr. 2. Manno-heptulose failed to suppress the insulin release occurring between 6 and 10 hr after the injection of streptozotocin. 3. Nicotinic acid administered by gastric tube failed to protect against the diabetogenic effect of streptozotocin. 4. Nicotinamide injected 10 min after alloxan afforded no protection against the diabetogenic effect of the latter. It is concluded (1) that the β-cytotoxic effect of streptozotocin is not immediately irreversible or that it does not affect all B-cells simultaneously; (2) that the insulin release observed between 6 and 10 hr after the injection of streptozotocin occurs by way of leakage from damaged B-cells; (3) that the ineffectiveness of nicotinic acid in preventing the β-cytotoxic effect of streptozotocin is probably not the consequence of its insufficient conversion to nicotinamide after parenteral administration; and (4) that streptozotocin and alloxan differ in their mechanism of action on the pancreatic B-cell.

The Snell-Bagg dwarf mouse was immunologically deficient as shown by severe lymphopenia, decreased agglutinating antibody response to sheep erythrocytes and Brucella antigen, and decreased graft-versus-host reactivity of dwarf spleen cells. The lymphoid organs of the dwarf were small, atrophic, and generally depleted of small lymphocytes. Serum immunoglobulins appeared to be within normal ranges. These findings suggested a selective deficiency of the thymus-dependent lymphoid system of the dwarf, caused by a defective thymotropic pituitary control.

The cytotoxic effect of an antiserum to neutral salt-soluble chicken collagen on tissue culture of chicken fibroblasts was studied. The cytotoxic action was present up to a final dilution of the antiserum of 1:768, and was complement-dependent. No cytotoxicity was observed when tissue cultures of liver or mesonephros of chick embryo or mouse embryo fibroblasts were incubated with the antiserum. The action was, therefore, tissue- and species-specific. The damage observed consisted of irregularities in the cell membrane followed by vesiculation and swelling of the cytoplasm. Nuclear changes were observed only when high concentrations of the antiserum were used.

These experiments indicate that the toxic effects shown by deoxycholic acid in vitro do not occur in vivo, apparently because transfer of free bile acids into the mesenteric venous system prevents them from reaching dangerous intracellular concentrations. This adds further evidence to the concept that in the blind loop syndrome, unconjugated bile salts are not responsible for the production of steatorrhea, but that decrease in the concentration of conjugated bile salts is the more likely mechanism. The importance of an intact mesenteric circulation in keeping intestinal mucosal free bile acid concentrations from reaching damaging levels is emphasized by these experimental findings.

Net transtubular movement of urea and water was studied in rats made diuretic by infusion of a volume of isotonic saline solution equivalent to 10% of their body weight. The major site of urea reabsorption was the proximal tubule and there was little urea loss from more distal parts of the nephron. There was no evidence of addition of urea to tubular fluid flowing through the loop of Henle. Throughout the nephron the percentage of filtered water remaining varied exponentially with transit time.

An assay using the L-929 mouse fibroblast cell culture line was described to evaluate the toxicity of Arlacel A. The toxic lots of AA produced a characteristic cytotoxic effect consisting of intracytoplasmic refractile bodies. The results obtained by the cell culture and mouse assays were similar for 18 of 19 lots tested; one lot was moderately toxic by the cell culture assay only. A correlation was seen between the amount of FFA and the toxicity of AA. The cell culture assay provides a sensitive and reproducible assay to evaluate AA toxicity.

We thank Dr. B. S. Berlin for his critical review of the manuscript, the Cell Biology Section (DBS) for providing the cells used in these studies, and acknowledge the excellent assistance of Mr. Donald Riggs, Mrs. Norma Duffin, Mrs. Marion Grabowski, Mr. Ethan Summers, and Mrs. Hazel Young.

In vivo and in vitro studies revealed that incorporation of thymidine-³H into DNA was inhibited in Ehrlich ascites cells treated with cortisone 24 hr before. Incorporations of other DNA precursors were not inhibited. Thymidine kinase was markedly inhibited by pretreatment with cortisone. Other nucleoside kinases (except deoxyuridine kinase), thymidylate kinase, and DNA polymerase were not appreciably affected by the cortisone treatment.

Feeding 5 and 8% excess L-histidine to rabbits caused significantly elevated plasma cholesterol levels. Phospholipid levels were slightly but not significantly elevated. The 5% histidine-supplemented diet produced plasma levels of 20–50 mg/100 ml of histidine. After 17 weeks on diet, no effects on brain cerebrum and cerebellum or liver cholesterol concentrations were found. Brain and liver total lipid and phospholipid and cerebrum galactolipid concentrations were also normal. The effect of excess dietary histidine on lipid components is apparently localized in the circulatory system. The effects noted in rabbits are much less than those previously observed in monkeys.

The technical assistance of Robert Colwell and Robert Jaeger was greatly appreciated.

Purified myelin was prepared in high yield by a single centrifugation in the L-4 zonal centrifuge. A whole rabbit brain (8 g) homogenate in 7% sucrose was layered over a 10–35% sucrose gradient in the B-IV rotor and sedimented for 2 hr at 40,000 rpm. The gradient was removed by pumping 50% sucrose into the rotor with the recovery of 120 fractions of 13 ml each. Every fifth to tenth fraction was examined under the electron microscope and was assayed for RNA, DNA, sialic acids, and 10 enzymes. The latter included cytochrome oxidase (CyOx) to monitor for mitochondria, K-activated ATPase (K∗ATP) and acetylcholinesterase (AChE) for synaptic membranes, leucylglycinase for soluble and microsomes, acid phosphatase for lysosomes, and glucose-yields in fractions 90–110, in keeping with the presence of all subcellular constituents in these fractions.

We gratefully acknowledge the support of this project through NIH, Grant NB 08278.

Pregnant rats on two levels of dietary Na (1.1, 28.7 meq/100 g) were studied along with their nonpregnant counterparts. At the end of gestation, adrenals were removed and studied histochemically for the demonstration of G6PD and sudanophilic lipids. Plasma Na and K concentrations and hematocrit values were taken as supporting data to aid in the interpretation of the histochemical findings.

Either pregnancy or a restricted intake of dietary Na led to slight hypertrophy and hyperplasia of the zona glomerulosa. The combination of Na restriction and pregnancy resulted in extreme hypertrophy and hyperplasia of the zona glomerulosa.

G6PD in the zona glomerulosa increased by 54% due to pregnancy and by 68% due to dietary Na restriction. However, the combined stress of pregnancy plus Na restriction caused a 656% increase in G6PD.

Alterations in the patterns of sudanophilic lipids were noted. Dietary Na restriction or the stress of pregnancy resulted in a slight increase in zona glomerulosa lipid as compared to the nonpregnant controls. However, the combined stress of Na restriction and pregnancy led to lipid depletion in an hypertrophied zona glomerulosa.

It is suggested that the increased activity of G6PD in the zona glomerulosa and the changes in the patterns of lipid distribution are due to an increased stimulation to aldosterone synthesis. The results support the suggestion that Na restriction during pregnancy exhausts the aldosterone-producing capacity of the cells of the zona glomerulosa.

Mammary tumor virus (MTV) did not induce detectable circulating interferon in female C57BL or male Swiss mice upon intravenous injection nor did it induce resistance in mice to subsequent challenge with Columbia SK virus. Numerous other viral and nonviral inducers of interferon did protect mice from challenge with Columbia SK virus. Neither milk obtained from mice during their third to fifth lactation, which contained very high concentrations of MTV (10¹² particles by electron microscopy), nor blood from tumor-bearing RIII mice, contained any detectable interferon or interferon-like inhibitor. MTV did not exhibit any specific hemagglutinin (HA) activity with the erythrocytes from 20 species of vertebrates. The data indicate that MTV, which shares certain properties with the myxoviruses, lacks the HA activity and interferon-inducing capacity frequently associated with many myxoviruses.

We are indebted to M. Lieberman and A. Pascale for conducting the mouse protection tests and the interferon assays.

Determination of the electrophoretic mobility of PSL particles in increasing concentration of PLG showed an unexpectedly high positive mobility at a coverage corresponding to one monolayer. This elevated mobility was tentatively ascribed to a particular orientation of the adsorbing PLG molecules due to interaction with the PSL surface. Other surfaces examined suggest that the effect is produced by hydrophobic surfaces. The mobility changes were shown to be correlated with changes in the organ distribution of PSL particles injected into rats.

It is a pleasure to acknowledge the enthusiastic technical help of Mr. Craig A. Warren. The work was supported in part by grants from The John A. Hartford Foundation, Inc., the National Institutes of Health (Grant No. AL 07161-02), and the Office of Naval Research. [Grant No. Nonr-4170 (00, NR 301-740)].

The effects of thermal changes on the organization of connective tissue of rat tail tendon was studied by polarization microscopy. Loss of birefringence in the temperature range 48–60° occurs most rapidly in young animals (1–3.5 months). After 3.5 months the trend continues but is less significant. The thermal effect could be due to rupture of low energy bonds in the collagen or collagen-ground substance aggregate. The method may be valuable for the study of changes of state of connective tissues in physiologic and pathologic conditions.

A bacterial carbohydrate-containing group-specific antigen of N. meningitidis has been identified to be of a high molecular weight (mean 140,000) by sedimentation equilibration and greater than 200, 000 by Sephadex G-200. This antigen was proved to be of serological importance for the measurement of antibody by PHA technique. Enzymatic digestion, with RNase, DNase, and trypsin, along with selective methods of isolating polysaccharides, was performed without appreciably diminishing the HA activity. Satisfactory levels of carbohydrate for sensitization of RBC were from 6 to 10 μg/ml (A, C, and 29E), 25 μg/ml (B) and 0.08 μg/ml (Bo). The concentration requirements of the group-specific carbohydrates were discussed along with the possibility of a multivalent molecule with both CF and HA activity and the likely difference in size of antigenic determinants.

Appreciation is expressed to G. Barlow, Abbott Laboratories, North Chicago, Illinois, for his technical assistance, and to Mrs. Lula Duggan for the preparation of this manuscript.

The subcutaneous implantation of Spirometra mansonoides spargana into hypophysectomized rats produced a striking increase in body weight and tibial cartilage width comparable to that noted with growth hormone. Other tissue responses confirmed this similarity. Injection of plasma from implanted rats also produced a growth hormone-like response including a stimulation of the uptake of ³⁵SO₄ ^2- by costal cartilage.

The authors are indebted to Dr. Justus Mueller for his valuable guidance in the initial studies. Dr. William C. Campbell and Dr. Dan Ostlind of the Merck Institute for Therapeutic Research greatly assisted in the work by growing and maintaining the SMS. Edward Chapin, Clinton Cope, Darlene Dotson, Roseann Leonzi, Barbara Oakley, Margaret Pastor and Bonni Yaeger provided valuable technical assistance in the studies reported.

Seven healthy subjects were exposed to imaginary food ingestion under hypnotic stimulus and plasma glucose, immunoreactive insulin and NEFA were measured during the experiment. Three subjects showed elevation in plasma insulin either after the induction of “hunger” or after the beginning of the “meal.” Fall in NEFA was seen in four subjects. No changes in blood glucose were detected.

Total acid-labile CO₂ was measured in aliquots of dog renal cortex and medulla frozen in liquid nitrogen. A higher (15.87 μmoles/g wet weight vs 13.55 μmoles/g wet weight, p <.05) acid-labile CO₂ content was found in the renal medulla. It is suggested that the higher medullary total acid-labile CO₂ content is primarily due to a high [HCO₃-] in the medullary intracellular fluid rather than to a high medullary Pco₂.

Inhibition of antibody formation to heterologous red cells in mice was studied using combinations of cortisone acetate with cyclophosphamide. The two drugs mutually potentiated or antagonized each other. Interaction between the compounds depended on the dose of antigen used for immunization. Pretreatment with cortisone prevented the development of cyclophos-phamide-induced tolerance. It is suggested that in the case of large particulate antigens steroids may block a common pathway leading both to tolerance and to immunity.

The skilled technical assistance of Miss D. Nitsche, Miss U. Petzoldt, and Mrs. T. Glanzmann is gratefully acknowledged.

An antibody in human sera that reacts with Burkitt's lymphoma biopsy cells and with leukemic lymphoblasts was detected by use of a complement-dependent cytotoxicity test. This antibody was detected in the sera of patients with Burkitt's lymphoma and also in the sera of other indivdiuals. The studies indicate that the antibody reacted with an antigen that is not tumor-specific but is also present on lymphoid cell lines and on normal peripheral lymphocytes.

Neonatally castrated male rats (some bearing ovarian transplants) and female rats given testicular grafts at birth were injected chronically with ¹³¹I-labeled L-T₄ and carrier T₄ as adults; the radioactivity in plasma and kidneys was measured and compared with that in normal adult males and females. Both renal and plasma radioactivity were significantly higher in normal male rats than in females. Neonatally castrated males as well as females given testicular transplants at birth exhibited an increase in renal T₄ to levels of normal males whereas neonatally castrated males bearing sc ovarian transplants showed a significant reduction in renal retention of T₄. A sex difference in the capacity of the kidney to concentrate radiothyroxine was demonstrated. This was not solely related to adult androgen levels and was influenced by neonatal hormonal manipulations which are known to alter hypothalamic-pituitary relationships in the adult rat.

The skilled technical assistance of Mr. Phil Cooper and Mrs. Joy Ewing is most gratefully acknowledged. Statistical analyses were done with the program EXBIOL on file at the Common Research Computer Facility, Texas Medical Center, Houston, Texas. The authors are indebted to Dr. Edvart Sakiz for this program.

Corynebacterium rubrum–a gram-postive nonpathogenic bacterium–induces severe arthritis a week to 10 days after its injection in rats. Production of arthritis is species-specific and dose-dependent, the optimal dose being 600 gamma per animal. Morphologically, the arthritis–similar to adjuvant arthritis induced by mycobacteria–is characterized by granulation tissue invading synovia, periarticular tissue, tendons, articular cartilage, periosteum, and bone spaces. The discovery of a nonpathogenic and easily cultivated substitute for mycobacteria may help to identify the responsible antigen and the mechanism in adjuvant arthritis.

A number of organisms occurring on blue crabs do not seem to affect their normal growth and survival.

The authors wish to acknowledge their indebtedness to Professor W. Van Engel from the College of William and Mary and the Virginia Institute of Marine Science, Gloucester Point, Virginia where this work was started. The authors also wish to express their gratitude to Dr. Ross Nigrelli, Director of the New York Aquarium for his valuable counsel during the performance of this work.

Obesity was induced in 45 and 36% of mice by intracerebral injection with the carcinogens 4-nitroquinoline 1-oxide (NQO) and 4-hydroxyaminoquinoline 1-oxide (HAQO), respectively, but not with the related noncarcinogens, 4-aminoquinoline 1-oxide, 3-methyl-4-nitroquinoline 1-oxide, and 4-nitropyridine 1-oxide. Food and water intake were markedly increased in the obese mice. While dietary restriction decreased the body weight, release of the restriction caused a conspicuous increase of food intake followed by a gain in body weight. No obesity could be induced with the following 4 local carcinogens: 3, 4-benzpyrene, β-propiolactone, N-ni-trosomethylurea and N-hydroxy-N-2-fluorenylactamide.

The authors thank Mr. T. Yamamoto and Mr. K. Ohno for their technical assistance. We are indebted to Dr. A. Ohta, this institute, Dr. Y. Kawazoe, National Cancer Center Research Institute, and Drs. John and Elizabeth Weisburger, National Cancer Institute, U. S. A., for their generous supply of the carcinogens and related compounds. We are also grateful to Professor Emeritus E. Ochiai, Tokyo University, and Dr. W. Nakahara, National Cancer Center Research Institute, for their interest and encouragement.

Mouse blood labeled with ⁵¹Cr was used to determine the blood volume of the obese and the diabetic mouse. While mutant mice were twice as heavy as normal controls, their blood volume was only 25% greater than normal. This finding suggests the possibility that blood volume rather than body weight should be used as the basis for in vivo dosage calculations.

The authors thank Dr. Robert S. Gordee for his valuable consultation and Mrs. Jean Pieper and her associates for radioactivity determinations.

Rabbits infected subcutaneously with E. coli developed hyperlipemia related to increased serum levels of free fatty acids early in the infection and later to increased levels of triglycerides. Rabbits challenged subcutaneously with S. aureus had normal levels of serum lipids during the acute febrile period but after 24 hr developed hyperlipemia related to hypertriglyceridemia.

Prostaglandin F_2α was luteolytic in rhesus monkeys when injected 30 mg/day subcutaneously b.i.d. for 5 days, if the injections were initiated on day 11, 12, or 13 postovulation of fertile cycles. At this time circulating progestin levels are elevated, presumably due to a trophic stimulus from the blastocyst. Such an effect was not obtained by injection earlier in the luteal phase of the reproductive cycle. This indicates that the corpus luteum of this species is more vulnerable to luteolysis in the later stages of the reproductive cycle.

Using column chromatography with Amberlite XAD-2 and silica gel and countercurrent analysis, a metabolite of naloxone was isolated from human urine. The metabolite was identical to naloxone-3-glucuronide isolated earlier from rabbit urine.

The author is grateful to Drs. Ralph Jacobsen and Harold Blumberg of Endo Laboratories for arranging this study with Drs. Max Fink and Arthur Zaks of the Department of Psychiatry, New York Medical College.

The administration of glutamate to rats decreases the rate of reaction between labeled cystine and the sulfhydryl groups of plasma globulins to form mixed disulfides. The rupture of these mixed disul-fides bridges also appears to be inhibited by glutamate.

Mouse L cells infected with nonreplicating Newcastle disease virus first developed antiviral resistance and then produced the interferon protein. Such infected cells, during inhibition of protein synthesis by cycloheximide, did not produce viral components or interferon and did not develop antiviral resistance. However, after reversal of protein inhibition and the addition of actinomycin D interferon was produced but antiviral resistance failed to develop. The production of interferon but not antiviral resistance after reversal of inhibition of protein synthesis was interpreted to indicate that the mRNA for interferon, but not the mRNA for antiviral resistance, was transcribed during infection in the presence of inhibitors of protein synthesis. The failure of development of the proposed mRNA for antiviral resistance in the absence of production of interferon suggests that induction of resistance by this virus is mediated by interferon. Finally, the production of the mRNA for interferon in the absence of even partial replication of Newcastle disease virus indicates that the stimulus for induction of interferon in this system is provided by a component of the input virus or an associated physical event.

The authors express their gratitude to Dr. Hilton B. Levy, Dr. Robert M. Friedman, and Prof. Claudio Pellegrino for helpful suggestion and criticism. The excellent technical assistance by Mr. G. Eutropi is gratefully acknowledged.

Seventeen of eighteen type 22 rhinoviruses isolated in this laboratory were shown to be antigenic variants of the prototype strain (127-CV22). The variant viruses were neutralized poorly or not at all by immune serum to the prototype strain. However, immune serum prepared to one of the variant viruses (Chase) neutralized both the prototype and homologous strains to similar titer and also neutralized all of the other isolates. The neutralizing reactivity of the Chase virus immune serum for the prototype and homologous strains was found to be essentially unaltered by multiple absorptions with human liver powder, a procedure capable of removing cytoxic factors or nonspecific inhibitors found in certain rhinovirus antisera. The Chase virus thus appears to represent a “prime strain” of rhinovirus type 22.

The authors acknowledge the excellent technical assistance of Mrs. Dana Gallo and Mrs. Florence Lester.

Polyriboinosinic–polyribocytidilic acid stimulated formation of hemolytic antibodies in mice immunized with sheep erythrocytes. Similarly, poly I_r:C_r treatment of mice resulted in a marked reduction in survival of isografts. Temporarily, administration of poly I_r:C_r at the time of exposure of mice to antigen resulted in optimal enhancement of humoral and cellular antibody formation. Optimal enhancement of both humoral and cellular antibody formation was induced by 100-200 μg of poly I_r:C_r, while significant stimulation of hemolytic antibody formation was induced by a lower dose of 10 μg of poly I_r:C_r.

We wish to thank Mr. Hugh Pettigrew of the Biometry Branch, National Cancer Institute, for the statistical evaluation of the experimental data.

Pituitary glands from mice bearing pituitary isografts in the renal capsule for 6-74 weeks were incubated with ¹⁴C- labeled amino acids, and the incorporation into prolactin and growth hormone was determined. Mice with isografts for 23 weeks synthesized 60% less prolactin and growth hormone than did normal intact mice. Implantation of isografts for periods up to 74 weeks further decreased the ability of the pituitary gland to synthesize the prolactin hormone; however, at 74 weeks, the extent of this inhibition in growth hormone synthesis was some-what decreased. It is proposed that the decrease in prolactin and growth hormone synthesis is mediated by hormone(s) secreted by the tumor acting at the hypothalamo-pituitary level through an autofeedback process.

The author thanks Prof. O. Muhlbock and Drs. L. M. Boot and H. W. Kwa for providing the laboratory facilities and fruitful discussions during his visit to the Netherlands Cancer Institute.

Plasma 17-hydroxycorticosteroids (17-OHCS) exhibited a significant circadian rhythm in rhesus monkeys (M. mulatta) in the first week of life. In two experiments, plasma 17-OHCS levels averaged 35 μg/100 ml at 8-9:00 am, and declined to levels of 29 μg/100 ml at 9:00 pm (Expt. I) and 24 μg/100 ml at 12:00 midnight (Expt. II). These changes were about half the magnitude reported by others in the adult monkey. Correlated with this steroid rhythm, the infant monkey, although having a polyphasic 24-hr sleep pattern, also exhibited a tendency to sleep more at 1:00 and 3:00 am than at other times of the day. This suggested a relationship between sleep–activity cycles and steroid cycles in the infant monkey similar to that in the adult monkey.

Rough strains of gram-negative bacteria are not necessarily more sensitive than smooth strains to the bactericidal action of normal serum or C. The resistance of certain smooth strains to C may merely reflect a lack of normal antibody in serum. Other smooth organisms may be refractory to C even when sensitized with homologous antiserum. In such instances, it is believed that the C-sensitive substance may be too far removed or inaccessible to the C enzymes activated by antigen–antibody complexes on the cell surface (6).

Cell size is smaller and mitotic rate and DNA content of parotid glands of rats maintained on liquid diet (Metrecal) from weaning on are lower than those of litter mates maintained on chow. At 32 days of age, when mitotic rate is 4 per 1000 acinar cells in parotid of chow-fed rats, it is nearly zero in the Metrecal rats. Substitution of a solid for liquid diet at 32 days brings about a marked increase in mitotic acitvity and DNA content, and increase in cell size, that are evident with 24 hr. The change is most marked at 1 day with regard to cell size, 2 days for mitotic rate (60 per 1000 acinar cells) and 5 days for DNA. Removal of the postganglionic innervation prevents the increase in mitosis and DNA that occurs with introduction of solid food. Neurally-mediated normal phisiological activity of the gland apparently evokes the excessive mitotic activity.