Radiation damage to bull sperm motility. III. Further x-ray studies.

The results of previous radiation experiments, which indicated that the centriole serves as a control center for bull sperm motility, appear to be in conflict with experiments showing that the bull sperm flagellum is an autonomous oscillator. To resolve this conflict experiments were conducted to calibrate absolutely the dose-response curves for the radiation damage, and to measure the force production and the mechanochemical energy conversion after irradiation in bull sperm. The results indicate that the centriole acts as a mechanical anchor for the contractile fibers.


Immunodeficiency and autoimmunity in aging.

In general, both cell-mediated and humoral immune activities decline with advancing age and, associated with this decline, is an increase in the incidence of certain types of autoimmune disease, cancer and infection. Not only may the relationship be causal, but the former may be a significant factor involved in the expression of the latter. Discussion is focused primarily on the relative contribution of extrinsic and intrinsic factors responsible for the immunodeficiency of aging individuals.


[Effect of the mitochondrial ionophore for divalent cations on bilayer phospholipid membranes].

It has been shown that ionophore of bivalent cations (IBC) isolated from fatless, subjected to partial triptic hydrolysis cattle heart or liver mitochondria decreases BPM resistance inducing Ca2+ conductivity. Ions of lanthane in micromolar concentrations decrease calcium conductivity induced with IBC. When ten-fold gradient in Ca2+ was created on BPM the intitiation of the membrane potential fo 9-11 mV was observed. The role fo IBC and water soluble factors binding Ca2+ with high affinity, in the mitochrondial mechanism of Ca2+ translocation is discussed.


Biosynthesis and secretion of immunoglobulins by peripheral blood lymphocytes in severe hypogammaglobulinemia and by cultured lymphoblast cells.

Peripheral blood lymphocytes of two patients with infantile X-linked agammaglobulinemia could neither synthesize nor secrete immunoglobulins in detectable amounts. Peripheral blood lymphocytes of patients with common variable type of hypogammaglobulinemia synthesize but do not secrete immunoglobulins. Serologic precipitation with antiimmunoglobulin serum and acrylamide-gel analysis of immunoglobulins synthesized by lymphocytes of these patients showed predominant mu-chain, as compared to immunoglobulins synthesized by normal lymphocytes. Biosynthesis and secretion of immunoglobulins by cultured lymphoblast cells from normal persons was very much analogous to that of lymphocytes of hypogammaglobulinemic patients. These findings suggest that lymphocytes of patients with common variable type of hypogammaglobulinemia are immature B cells like cultured lymphoblast cells, and that a block in cytodifferentiation of IgM to IgG-producing cells is present in their B cells.


Variation in activity of monoamine metabolizing enzymes in rat liver during pregnancy.

1. Catechol-0-methyltransferase (COMT) and monoamine oxidase (MAO) activities in rat liver were measured during pregnancy, parturition and postpartum. Compared with activity in non-pregnant controls, both enzymes showed a significant decrease in activity which was most pronounced at day 18. 2. The metabolism of intravenously infused [3H]-adrenaline to [3H]-metanephrine and to [3H]-acidic metabolites was also significantly depressed during pregnancy but had returned to control values by the 21st day. 3. The effects of reserpine and/or nialamide on hepatic COMT and MAO were studied in control and 20-day-pregnant rats. Their action on COMT activity differed in the two groups. MAO was inhibited to a similar extent in these groups whether the drugs were given separately or in combination. 4. It seems possible that the changes in endocrine function which occur during pregnancy are responsible for the observed alterations in enzyme activity.


An abnormality of oestrogen feedback in amenorrhoea-galactorrhoea.

Fourteen patients with amenorrhoea and hyperprolactin-anemia but no evidence of pituitary tumours were each given an intramuscular injection of 1 mg oestradiol benzoate. Thirteen patients failed to release luteinizing hormone in response to the oestrogen. This hypothalamic abnormality may help to explain the menstrual disturbances in subjects with hyperprolactinaemia.


Tuberoinfundibular neurons: dopaminergic and norepinephrinergic sensitivity.

The techniques of microelectrophoresis and antidromic identification have been utilized in the female rat anesthetized with urethane to localize the tuberoinfundibular neurons and to determine their pharmacological sensitivity to dopamine (DA), norepinephrine (NE) and glutamate (GLUT). Extracellular recordings were made of 149 single units in the arcuate (ARC) nucleus; 79 of which were antidromically identified (43 were spontaneously active and 36 were silent) and the remaining 70 cells were determined to be spontaneously active but could not be antidromically identified (unidentified ARC neurons). The mean latency of the antidromically activated potentials was 9,5 msec, which gave a conduction velocity of 0.05 mm/msec. The spontaneous firing rate of antidromically identified ARC neurons varied from less than 1/sec to 8/sec while unidentified cells discharged at rates from less than 3/sec to 10/sec. These data indicate that ARC neurons exist which are capable of conducting impulses and send their axons into the external layer of the ME. Successful drug applications were made on 104 ARC neurons; of the 43 antidromically identified units, 37 were spontaneously active and 6 were silent. There were 61 spontaneously active, unidentified neurons. Two distinct pools of antidromically identified ARC neurons were found based on their sensitivity to NE and DA. Neurons displaying excitation to NE(N = 23) were either inhibited (N = 10) or non-responsive (N = 13) to DA applied electrophoretically; no NE-sensitive ARC neurons were excited by DA. The second pool of neurons (N = 14) was excited by DA. These neurons when tested with NE were either reproducibly inhibited (N = 3) or nonresponsive (N = 11); no DA-sensitive neurons were excited by NE. In the unidentified ARC neurons, iontophoretically applied NE and DA gave reproducible effects on individual neurons, so that some neurons were excited by both NE and DA, some were inhibited by both, and still others were excited by one and inhibited by the other chemical. Glutamate (GLUT) had a powerful excitant action on nearly all the neurons tested, increasing the electrical activity of the spontaneously active neurons as well as initiating activity in quiescent ARC neurons. These data are suggestive of a tuberoinfundibular neuronal system whose activity is modulated by either DA or NE. This evidence is certainly compatible with a functional role for both DA and NE in regulating pituitary function by altering the activity of presumed neurosecretory neurons which release hypophysiotropic hormones.


Effects of chronic administration of sodium diphenylhydantoin ('Dilantin') on bones and teeth of the rat and hamster: a preliminary study.

Male Charles River rats, 31-days old, received i.p. injection of sodium diphenylhydantoin (DPH), 100 mg/kg in 0.9% NaCl, once daily for 26-27 days before death. Male Syrian hamsters, 40-days-old, received similar injections of DPH, 25 mg/kg for 46 days, no treatment for 39 dyas, then DPH FOR A FURTHER 17 DYAS BEfore sacrifice. All rats receiving DPH gained less weight than the controls, and more than 50% displayed acute neurotoxic reactions to the drug; hamsters were not so affected. Morphology and composition of caudal vertebrae, teeth, and jaws from control and DPH-rats were compared on the basis of measurements on radiographs and gross specimens, histological investigation, and determined of % dry volumes of ash, volatile inorganic component, lipid, and organic matrix. DPH-bertebrae were smaller and showed impaired osteogenesis; but chondrogenesis was similar to controls. Overall tail length was similar in both animal groups because caudal intervertebral spaces were wider in DPH-rats, compensating for reduced longitudinal growth of corresponding vertebrae. Incisors were smaller and third molar roots shorter in DPH-rats. In DPH-hamsters the attachment of the periodontal ligament to maxillary incisors was deranged. DPH administration did not change the composition of rat bone or teeth. Densities of dry bones and teeth were in accord with their composition. Possible modes of action of DPH are discussed. Species differences in response of mineralized tissues to DPH administration are emphasized in relation to reports of rickets and osteomalacia in patients on long term DPH therapy.


The effect of a low-calcium diet in lactating rats; observations on the rapid development and repair of osteoporosis.

Female rats were given a low-calcium diet (0.05%) during the last three weeks of the lactating period, followed by a normal diet (1.03% calcium) during the first three weeks after lactation. The resulting bone loss and its recovery were studied by means of microradiography, tetracycline-uptake, quantitative estimation of the cortical area of cross-sections from the femoral midshaft, and estimation of total body calcium. The cortical area in the femoral midshaft fell to 46% of its original value during depletion, and then rose to 78% during the first three weeks after weaning. Total body calcium fell from 1.12% to 0.60% of body weight and then increased to 0.89%. Removal of bone occurred mainly in the spongiosa and on the endosteal side of the cortex. Subsequently, new bone was laid down on the endosteal side, but also to some extent on the periosteal side of the cortex. The mineral density of this new bone was low. During the recovery phase resorption cavities within the cortex were filled in a concentric manner as in Haversian remodeling. Neither this feature nor the low mineral density of bone are normally present in the rat.


Effect of modifiers on the hydrolysis of basic and neutral peptides by carboxypeptidase B1.

The initial rates of hydrolysis of Bz-Gly-Lys and Bz-Gly-Phe by carboxypeptidase B (CPB) are increased in the presence of the modifiers beta-phenylpropionic acid, cyclohexanol, Bz-Gly, and Bz-Gly-Gly. The hydrolysis of the tripeptide Bz-Gly-Gly-Phe is also activated by Bz-Gly and Bz-Gly-Gly, but none of these modifiers activate the hydrolysis of Bz-Gly-Gly-Lys, Z-Leu-Ala-Phe, or Bz-Gly-phenyllactic acid by CPB. All modifiers except cyclohexanol display inhibitory modes of binding when present in high concentration. Examination of Lineweaver-Burk plots in the presence of fixed concentrations of Bz-Gly has shown that activation of the hydrolysis of neutral and basic peptides by CPB, as reflected in the values of the extrapolated parameters, Km(app) and kcat, occurs by different mechanisms. For Bz-Gly-Gly-Phe, activation occurs because the enzyme-modifier complex has a higher affinity than the free enzyme for the substrate, whereas activation of the hydrolysis of Bz-Gly-Lys derives from an increase in the rate of breakdown of the enzyme-substrate complex to give products. Cyclohexanol differs from Bz-Gly and Bz-Gly-Gly in that it displays no inhibitory mode of binding with any of the substrates examined, activates only the hydrolysis of dipeptides by CPB, and has a greater effect on the hydrolysis of the basic dipeptide than on the neurtal dipeptide. Moreover, when Bz-Gly-Lys is the substrate, cyclohexanol activates its hydrolysis by CPB by increasing both the enzyme-substrate binding affinity and the rate of the catalytic step, an effect different from that observed when Bz-Gly is the modifier. The anomalous kinetic behavior of CPB is remarkably similar to that of carboxypeptidase A, and is a good indication that both enzymes have very similar structures in and around their respective active sites. A binding site for activator molecules down the cleft of the active site is proposed for CPB to explain the observed kinetic behavior.


[Effect of neonatal thymectomy on the content of magnesium in the plasma and in various tissues of the rat].

White rats were thymectomized neonatally. After 60 days, no alteration was noted in the plasma or testis magnesium level. On the other hand, a marked decrease was observed in the magnesium content of muscle (-23%) and of bone (-19%). The injection of thymosin reversed these changes. These findings suggest that a correlation between the thymus and the muscle is exerted by a protein factor (thymosin), and that the thymus may act on sex glands through an indirect pathway.


Isolation and identification of rumen bacteria capable of anaerobic phloroglucinol degradation.

Eight strains of rumen bacteria capable of degrading phloroglucinol (1,3,5-trihydroxybenzene) under anaerobic conditions were isolated from enrichment cultures of the bovine rumen microflora established in a prereduced medium containing 0.02 M phloroglucinol. Five of the strains were facultatively anaerobic Gram-positive streptococci which were identified as Streptococcus bovis. Three strains of obligately anaerobic Gram-positive cocci were assigned to the genus Coprococcus. Anaerobic cultures of the Streptococcus bovis strains in a 40% rumen fluid medium initially containing 0.02 M phloroglucinol degraded 50-80% of the substrate within 2 days, whereas cultures of the Coprococcus strains degraded more than 80% of the substrate under the same conditions. The Streptococcus bovis strains were incapable of degrading phloroglucinol in brain heart infusion or in the medium of de Man, Rogosa, and Sharpe (MRS broth) incubated aerobically.


Changes in pattern of respiration and glucose utilisation in Candida utilis during the cell cycle: some variations with growth rate.

The release of 14CO2 from 14C-labelled glucose(G-1(-14)C, G-3,4(-14)C, G-6(-14)C) was followed in phased cultures of Candida utilis grown in a glucose- mineral salts medium under altered conditions of carbon:nitrogen limitation at doubling times of 2,4 and 6, h.  Changes in oxygen uptake and CO2 evolution were observed and respirometric studies showed that the relative contributions of the Embden-Meyerhof-Parnas and hexose monophosphate pathways varied over the cell cycle and changed with growth rate.  The results are discussed in relation to the growth metabolism of the cells.


The effect of glucose and lactose on beta-D-galactosidase activity and formation of sclerotia in Sclerotium rolfsii.

Addition of 0.5% (w/v) lactose to a glucose-mineral mdeium (SM) induced formation of sclerotia and beta-D-galactosidase (beta-D-galactoside galactohydrolase)(EC 3.2.1.23) synthesis in Sclerotium rolfsii types A and R; These effects as well as lactose uptake were inversely related to glucose concentration within the tested range of 0.5 to 2.5% (w/v).  Transfer of lactose-grown colonies to a glucose-supplemented medium nullified the inducible effect of lactose on formation of sclerotia, whereas transfer to water agar did not. It is concluded that glucose nullifies the effect of lactose on S. rolfsii by interfering with its active uptake.


The development and structure of thick-walled, multicellular, aerial spores in Diheterospora chlamydosporia (=Verticillium chlamydosporium).

The aerial, thick-walled spores in Diheterospara chlamydosporia arose as terminal swellings on erect hyphae.  Repeated septation of the continuously swelling spore resulted in a multicellular structure.  Immediately after the onset of septation secondary wall material was laid down between the two-layered primary wall and the plasmalemma.  The presence of secondary wall material indicates that the multicellular spore is a dictyochlamydospore and not an aleuriospore.  The relationship between chlamydospores and aleuriospores in other fungi is discussed.


Prospects for para-aortic irradiation in treatment of cancer of the cervix.

The incidence of para-aortic node metastasis in cancer of the cervix ranges from 5%to 50% in clinical Stages i-iv.  Extended field irradiation is being administered more frequently because of high incidence of extrapelvic disease.  The cancer of the cervix material at Walter Reed General Hospital has been analyzed as to site and time of failure to determine who might have benefited from extended field irradiation.  Of patients who fail, the majority will have uncontrolled pelvic disease regardless of other involvement.  Diagnostic procedures including staging laparotomy are unable to predict pelvic failure or the presence of occult distant disease in lung and bone. The maximum number of patients who could benefit from pari-aortic irradiation is lessthan 3% of a total series.  It is concluded that the salvage of patients with para-aortic metastasis will be small, but for those few long-term survivors, the benefit will be great.


Carcinoma of the endolarynx: results of irradiation.

Irradiation at the University of Michgan in the 1960's was carried out in 192patients with endolaryngeal carcinoma.  Thirty-two and 66% of the cases were located inthe supreaglottic and glottic regions respectively.  Cases were classified in stages appropiate TNM GROUPING.  Results were analyzed with respect to tumor location, size, anddose-time relationship.  Early lesions were controlled more often than advanced ones.  Radical surgery to the primary and/or neck improved overall survival, which was 89% at 5years for 127 glottic carcinoma cases and 61% for 61 patients with supraglottic tumors.  No significant complications were found following radiation therapy.


Lymphangigraphic accuracy in the staging of testicular tumors.

Since 1965, 103 patients with testicular neoplasms have been evaluated with lymphangiography for evidence of retroperitoneal disease.  Thirty-five patients with testicular carcinomas had lymphadenectomies; corroboration of their initial lymphangiographic interpretaions with the surgical specimens was possible.  There was an overall 79% (26 of 33) correlation.  We feel that even with relatively inexperienced examiners, lymphangiography can be a useful means of evaluating the retroperitoneum.


Effect of methotrexate on thymidylate synthetase in cultured parenchymal cells isolated from regenerating rat liver.

The effect of methotrexate (MTX) on thymidylate synthetase activity during liver regeneration was examined with parenchymal cells isolated 22 and 44 hr after partial hepatectomy and cultured as a monolayer. The synthetase activity in these cells decreased with a half-life of 18 to 24 hr, but if MTX (1.5 X 10(-6) to 1.5 x 10(-5) M) was present in the culture media, this decline could be delayed for at least 48 hr. In contrast, thymidine kinase activity decreased at a rate which we unaffected by MTX. Dihydrofolate reductase was inhibited at all concentrations of MTX used to block the decrease in synthetase activity. Folic acid at 10(-4) M, although less effective than MTX, also delayed the decrease in synthetase activity. The addition of cycloheximide, puromycin, or antinomycin D to the culture media did not alter the response of the synthetase to MTX. The latter studies, coupled with those indicating that the rapid loss of synthetase activity in crude extracts could be prevented by MTX or, more effectively, by MTX plus deoxyuridine 5'-monophosphate, suggest that the primary effect of MTX on thymidylate synthetase in vivo is that of enzyme stabilization. Similar stabilizing effects were obtained in liver cell extracts with 10(-5) M deoxyuridine 5'-monophosphate in combination with 10(-4) M folate or 10(-4) M dihydrofolate.


Toxicity of amphotericin B and its methyl ester toward normal and tumor cell lines.

The toxicity of amphotericin B, amphotericin B methyl ester (AME), and Fungizone toward three "normal" and three tumor-derived human and mouse cell lines was evaluated in monolayer culture. AME was less toxic than amphotericin B and Fungizone to all cell lines, but the sensitivity of the normal and tumor lines was different. The human (HEL-8 and WISH) and mouse (L-M) cells derived from normal tissue were more resistant to AME than the tumor-derived human (KB and HeLa) and mouse (RAG) cell as indicated by: (a) increased 24-hr survival, (b) increased 72-hr viability, and (c) growth rates at higher AME concentrations. In contrast, no pattern of differential sensitivity was observed with amphotericin B and Fungizone.


A human gastric choriocarcinoma cell line with human chorionic gonadotropin and placental alkaline phosphatase production.

A gastric choriocarcinoma cell line synthesizing human chorionic gonadotropin (HCG) was established in 1971 by Oboshi et al. and was found to possess human placental alkaline phosphatase. The present paper also deals with the relationship between the cell growth and HCG secretion and with cellular localization of HCG and human placental alkaline phosphatase by cytochemical and ultrastructural methods. This cell line was found to secrete HCG during cellular proliferation, with the maximum secretion in the stationary phase (about 1 muIU/cell/48 hr), and the hormone could be detected in a small proportion of mono- and/or multinuclear cells in both logarithmic and stationary phases. The organ-specific, heat-stable, L-phenylalanine-sensitive, immunoreactive human placental alkaline phosphatase was localized on the cell membrane of many cells. Ultrastructurally, the line consisted mainly of cytotrophoblastic and intermediate cells in the process of syncytial formation, with more or less squamous metaplasia. From these findings it was concluded that the cell line maintained the properties of trophoblastic cells from morphological and functional aspects, i.e., it was a cell line with two distinct marker substances.


Direct determination of polyamines in human serum by radioimmunoassay.

Antipolyamine antibodies were produced following immunization of New Zealand white rabbits with thyroglobulin spermine conjugate, using multiple-site injection and macrophage-harvesting techniques. The antispermine antibody in a radioimmunoassay system cross-reacted with spermidine (22%), putrescine (1%), diaminopropane (6%), and cadaverine (0.16%). No reaction with L-lysine, ornithine, or histamine was noted. Using this antibody, the radioimmunoassay for direct determination of serum polyamines was developed. A sensitivity of 200 pg was achieved on human serum samples. This radioimmunoassay would appear to be a useful tool for the detection and monitoring of a variety of pathophysiological states associated with abnormal polyamine metabolism, i.e., cancer.


Progression from hormone dependence to autonomy in mammary tumors as an in vivo manifestation of sequential clonal selection.

The natural history of breast cancer with respect to its progression from hormone dependence to autonomy was studied by successively transplanting the earliest possible form of a rat mammary adenocarcinoma in syngeneic rats and isolating sublines of this tumor manifesting new endocrinological and other biological characteristics over a period of 15 years. The original, fully mannotropin-dependent tumor MT-W9, whose growth is dependent entirely on pharmacological levels of mannotropin supplied by an extraneous source, gave rise to an estrogen-dependent variant, MT-W9A. This MT-W9A growns only in normal adult female hosts and regresses promptly upon oophorectomy. Subsequently, this tumor produced a subline, the fully autonomous MT-W9B, which grows well in any syngeneic rats regardless of their hormonal status. The third subline derived from the autonomous tumor was designated MT-W9C, the androgen-responsive line, because it growns better in male than in female rats. Having reversed its original female preference, it might also be characterized as reversely responsive. Chromosomal analysis of these 4 tumors disclosed 4 distinct stem cell lines, but each tumor also contained small numbers of cells from other stem lines. The progression from mammotropin dependence to androgen responsiveness in this mammary tumor system seems to have been accomplished by merely shifting from 1 stem line to another in an orderly sequence, and it appears to be an irreversible process. In addition to the hormone dependency, the progression of the tumor is accompanied by a gradual increase of cachexia-producing effects on the host that may be related to the immunogenicity of tumor cells. These 4 distinct hormonal characteristics encompass the entire known spectrum of breast cancer in man and animals with the exception of the metastasizing property. These tumors are being studied by many investigators and are maintained vertically and horizontally in syngeneic rats by us. The hormonal and cytogenetic characteristics of each stem cell line have been stable for over a decade.


Different agglutinability of fibroblasts underlying various precursor lesions of human uterine cervical carcinoma.

Fibroblasts underlying human uterine cervical dysplasia, carcinoma in situ, and invasive carcinoma are agglutinable by concanavalin A (Con A) but not by wheat germ agglutinin, except at very high concentration. Studies with low levels of Con A show that maximal agglutination is obtained with fibroblasts from invasive carcinoma, while the fibroblasts underlying dysplasia give minimal agglutination reactions. Fibroblasts underlying carcinoma in situ give agglutination reactions halfway between those obtained with fibroblasts underlying dysplasia and invasive carcinoma. An epithelial-like cell line obtained from a case of dysplasia shows agglutinability by Con A very similar to that obtained with fibroblasts underlying dysplasia. These epithelial-like cells are also not agglutinable by wheat germ agglutinin. Treatment of the cervical cells, both epithelial and fibroblasts, with neuraminidase leads to slight increase in agglutination by both Con A and wheat germ agglutinin. Marked increase in agglutination is not obtained even after treatment with high concentration of neuraminidase (10 units/10(6) cells). Marked agglutinability, however, is observed after trypsin treatment. The results suggest that, while the fibroblasts obtained from normal cervix are not agglutinable by Con A, surface alterations necessary for Con A-specific agglutination exist in fibroblasts during the early stage of development of uterine cervical epithelial neoplasia (dysplasia) and increase with the progression through carcinoma in situ to invasive carcinoma. Loss of cell surface sialic acids may result in a slight increase in agglutinability, but some other mechanism(s) is likely to be involved in alteration of surface properties that lead to marked agglutinability of the human uterine cervical cells obtained from cancer precursor lesions.


A solid-phase radioimmunoassay for human prostatic acid phosphatase.

A solid-phase technique for radioimmunoassay of human prostatic acid phosphatase (EC 3.1.3.2) is described. Human prostatic acid phosphatase was purified from prostatic fluid. Monospecific antisera to the purified acid phosphatase were produced in rabbits. Disposable polypropylene tubes were coated with antiserum and used for radioimmunoassay with 125I-acid phosphatase. The nonspecific binding was minimized by saturating the binding sites of the tubes with bovine serum albumin. The working range of the technique was 1 to 30 ng of antigen. The solid-phase radioimmunoassay is rapid, sensitive, and efficient. In preliminary clinical trials it was shown that (a) patients with advanced prostatic cancer had elevated prostatic acid phosphatase levels by both enzymatic assay and radioimmunoassay assays, and (b) patients with other cancers were in the normal range for prostatic acid phosphatase.


The production of acidic polysaccharides by 5-bromodeoxyuridine-treated B16 mouse melanoma cells.

B16 melanoma cells were treated in culture with 5-bromo-2-deoxyuridine. The cell-associated and released proteoglycans and sialoglycopeptides were compared to those of control cultures treated with thymidine. The 5-bromo-2-deoxyuridine-treated cultures showed a marked reduction in the proportion of cell-associated proteoglycans and sialoglycopeptides, an increase in the synthesis of hyaluronic acid, the absence of high-molecular-weight chondroitin sulfate, and the presence of increased amounts of heparan sulfate in the media. In addition, the 5-bromo-2-deoxyuridine-treated cells had a higher DNA content and were larger than controls.


The failure of methotrexate to inhibit chicken fibroblast proliferation in a serum-containing culture medium.

Methotrexate (1 muM) abolishes the proliferation of chicken fibroblasts in a culture medium containing defibrinogenated chicken plasma but does not affect proliferation in a medium containing chicken serum.


Matrix simulation of duodenal crypt cell kinetics. I. The steady state.

Steady state crypt cell kinetics have been simulated using matrix algebra. The model crypt cell population is distributed through two proliferation compartments (P1 and P2) and a quiescent state (Q). Under steady state conditions half the daughter cells produced on completion of P1 enter G1 of P2 and half enter G1 of P1. Both P2 daughter cells enter Q. Cells in Q are non-dividing but retain the potential to divide. On completion of Q, cells lose the potential to divide and move up onto the villi. The model has been developed by simultaneously simulating the following biological data: (1) the per cent labeled mitosis (PML) curve, (2) the number of labeled cells per crypt as a function of time following an injection of 3H-thymidine, and (3) the total number of cells per crypt.


Matrix simulation of duodenal crypt cell kinetics. II. Cell kinetics following hydroxyurea.

The perturbed cellular kinetics of the duodenal crypt following a single injection of hydroxyurea (HU) have been simulated using matrix algebra. Following the direct effects of HU (S-phase cytotoxicity and a G1/S block) the crypt cell kinetics undergo several alterations. Previously documented alterations include: (1) a temporary partial synchronization of the surviving cells, (2) a shortening of the cell-cycle transit time, and (3) recruitment of normally non-proliferating cells into active proliferation. These conclusions have been extended by constructing several different complex but theoretically possible recovery models and the validity of each of these models has been evaluated by simulating the following biological data: the number of cells in the S and M-phase of the cell cycle, total viable cells per crypt, and the per cent labeled mitosis and the number of labeled cells following 3H-TdR injections at 9 and 21 hr after HU treatment. The model which showed visually the best overall agreement with all sets of the data was chosen as "most probable' and leads to the following interpretations. Immediately after the end of the HU block (i.e. 5 hr after HU injection) the modal cell-cycle transit time is reduced to 8 hr. By 17 hr after HU, the modal transit time is increased to 10 hr. Repopulation of the proliferating compartment, i.e. restoration of the proliferating compartment back to the control value, occurs between 12 and 17 hr after HU injection and probably consists of both recycling of the proliferating cells (i.e. they do not progress up into the non-proliferating compartment) and recruitment of the non-proliferating cells into active proliferation. Also, the rate at which the non-proliferating cells move onto the villi is reduced temporarily. The overall recovery process results in a crypt which temporarily is larger than control and produces villi cells at a rate which is faster than the control. The time when the crypt size and villus cell production rate return to normal cannot be established using the available data.


Variation of cell kinetic parameters in relation to the growth rate of the Ehrlich ascites tumor.

A multicompartmental model of the cell cycle and proliferation kinetics was used to analyse the time-course behavior of the cell cycle time, the growth fraction, and the cell loss rate during Ehrlich ascites tumor growth. The growth rate of Ehrlich ascites tumor cells as the tumor aged was significantly influenced by change in the cell cycle time.


Distribution of microtubules in prolactin cells of lactating rats.

The intracellular distribution of microtubules was studied using serial sections of prolactin cells in anterior pituitary glands from lactating rats. Numerous microtubules were present in these cells following fixation with glutaraldehyde and osmium tetroxide. The greatest number of microtubules were present in the Golgi complex, situated around the perimeter and in association with the cisternae, vesicles and developing secretory granules. Microtubules were found in channels between groups of parallel cisternae of rough surfaced endoplasmic reticulum and in close proximity to small vesicles. They were also located adjacent to mitochondria, the plasmalemma, the nuclear envelope, and among mature secretory granules. Due to their orientation within the cell, it is suggested that the microtubules may act to direct the movement of organelles from one region of the cell to another and to give internal support to the cell.


Cell differentiation during early development of Nassarius reticulatus L. (Gastropoda Prosobranchia). I. Zygote to 16-cell stage.

The ultrastructure of the zygote and of early segmentation stages, up to the 16-cell stage, was investigated in normal Nassarius reticulatus from Roscoff (France). This study deals predominantly with structures that remain morphologically unchanged throughout this period, such as yolk granules, lipid droplets, and multimembranous vesicles. These organelles do not change in position, fine structure, or quantity from the egg to the 16-cell stage. The cortex and the vitelline layer also remain almost unchanged until the 16-cell stage. Cortical granules could not be observed and thus the thin vitelline layer is not transformed to a thick fertilization membrane. This phenomenon seems to be related to the facts that the eggs are protected by a tough capsule and that the cells of the embryo have to incorporate extraembryonic nutrient substances. From the 2-cell stage until the 16-cell stage micropinocytotic vesicles 500-2000 A in diameter are associated with the plasmalemma. Perhaps they help to incorporate the extraembryonic nutrient reserves. Desmosomes first appear at the 4-cell stage and are common at the 8-cell stage, when a small transitory blastocoel appears. Normally the cell borders are in close proximity during interphase. Syncytial connections between the blastomeres were not observed. Beginning at the trefoil stage the mitochondria increase in number and many apparent division stages are observed. The increase in mitochondria occurs in the perinuclear region throughout the embryo and does not result in a polar lobe especially rich or poor in mitochondria. From the 4-cell stage onwards, the number of ribosomes increases differentially in different blastomers.


Influence of light and darkness on the ultrastructure of the pineal organ in the blind cave fish, Astyanax mexicanus.

Ultrastructural changes of the pineal organ were investigated in the blind cave fish, Astyanax mexicanus, kept under continous artificial light (5000 lux), in continuous darkness, and under natural light conditions. The pineal end-vesicle of the fish kept under natural photoperiod consisted of photoreceptor cells and supporting cells mixed with a few ganglion cells. The photoreceptor cells possessed well-developed outer segments with regularly arranged lamellar membranes. The supporting cells contained a number of lipid droplets and large globular cisternae filled with fine granules. In the fish kept under continuous light or in darkness, the pineal end-vesicle displayed a dilated lumen, and the outer segments of the receptors showed signs of degeneration. Furthermore, alterations of cell organelles were observed in the photoreceptor and supporting cells.


Polyploidization and endomitosis in giant cells of rabbit trophoblast.

Morphological and cytophotometric investigations have been performed on giant cells of the rabbit trophoblast to reveal a mechanism of nuclei polyploidization and define the level of polyploidy. The character of endomitotic chromosomes is found to differ and depend largely on the degree of nuclei polyploidy. Small chromosomes were found in nuclei with low levels of polyploidy. For highly polyploid nuclei, two stages are distinguished. In the first case condensed chromosomes join into bundles resembling Riesenchromosomen in plants, whereas in the second, decondensed chromosomal threads separate and disperse in the karyoplasm. The splitting does not involve nuclei-forming chromosomes in the region of the nucleolar organiser. The degree of polyploidy was determined on the 15th day of development. It was found that giant cell nuclei contain DNA in amounts corresponding to 32-512 chromosomal sets. Most of the nuclei have levels of 128c and 256c. Highly-polyploid nuclei disintegrate into small nuclei with the degree of polyploidy varying from 1c to 32c. Di- tri- and tetraploid nuclei predominate.


Metabolism of the polyadenylate sequence of nuclear RNA and messenger RNA in mammalian cells.

The poly(A) sequences at the 3' end of mRNA and nuclear RNA molecules of mouse sarcoma and Chinese hamster cells are subject to an elongation process distinct from de novo synthesis. This process continues in cells treated with a high level of actinomycin D to block transcription. This results in the labeling of the steady-state poly(A) population in the cytoplasm and of unusually long poly(A) segments in the nucleus. In cells incubated with 3H-adenosine in the absence of drug treatment, cytoplasmic steady-state poly(A) segments with short labeled sequences at the 3' end can be detected by their heterogenous size distribution and by measurements of adenosine and AMP released by alkaline hydrolysis. These measurements indicate an average size of 8 residues for the labeled sequences. In the nucleus, a slow elongation of preexisting poly(A) chains can also be detected through measurements of AMP and adenosine. The cytoplasmic elongation process leads to turnover of the 3' end of the poly(A) sequence on mRNA, because of concomitant removal of AMP residues. It is apparently not linked to mRNA translation. The Chinese hamster and mouse sarcoma cells appear to differ markedly with respect to relative extents of poly(A) chain extension and de novo synthesis.


Sequence complexity of heterogeneous nuclear RNA in sea urchin embryos.

The sequence complexity of heterogeneous nuclear RNA is sea urchin gastrulas was measured by RNA-driven hybridization reactions with nonrepetitive sea urchin DNA. 28.5% of the sequence complexity of the genome is represented in the nuclear RNA. This amounts to 1.74 X 10(8) nucleotides of diverse sequence, more than 10 times the nucleotide complexity of the polysomal messenger RNA extracted from sea urchin embryos at the same stage. The complex set of nuclear RNA sequences driving this hybridization reaction was shown to be the same as the rapidly labeled hnRNA, using pulse-labeled nuclear RNA as driver.


Studies on the persistence of differentiated functions in rat hepatocytes set into primary tissue cultures. Specific binding of 3H-bilirubin after eight days of staying in vitro.

Preliminary experiments were performed which indicate that even after 8 days of staying in primary in vitro tissue culture rat hepatocytes are still able to take up and bind tritiated, unconjugated bilirubin in a specific fashion and much more intensely than do connective tissue cells present in the same cultures. The data are suggestive of the maintenance in 8-day cultured hepatocytes of at least some of the specific pathways bound to bilirubin metabolism.


High-speed quantitative karyotyping by flow microfluorometry.

We describe a new flow-system technique for high-speed quantitative karyotyping. Metaphase chromosomes are isolated from cells, stained with a DNA-specific  fluorochrome, and individually measured for stain content at rates up to 10(5)/min in a flow microfluorometer. The resulting distribution contains quantitative information about the relative frequency of occurrence and the chromosomal DNA content of the measured chromosomes. Such analyses of Chinese hamster chromosomes are in good agreement with independent results from scanning cytophotometry and with cytogenetic observations of the banded chromosomes. Analyses of two karyotypically different clones of Chinese hamseter M3-1 lines provide the quantitative characterization of two stable translocations. Initial data on human chromosomes are presented and agree well with results of scanning cytophotometry and suggest potential clinical applications as preparative and instrumental techniques improve.


Adaptation of a T3-uptake test and of radioimmunoassays for serum digoxin, thyroxine, and triiodothyronine to an automated radioimmunoassay system--"Centria".

We report the adaptation of four radioassays to the prototype of an automated radioimmunoassay system ("Centria," Union Carbide). The system consists of three integrated modules: (a) an automated pipettor, which dispenses samples and reagents; (b) the key module, an incubator/separator, in which centrifugal force is used to initiate and terminate multiple radioassay incubations and separations simultaneously; and (c) a gamma-counter/computer, which counts three tubes simultaneously and converts counts into concentration units. Radioimmunoassays for thyroxine, triiodothyronine, and digoxin were developed with use of well-characterized antibodies and of prepackaged Sephadex-containing columns to separate bound and free radioactive ligand. A triiodothyronine-uptake test in which the same kind of columns were used was also adapted to the instrument. Results for clinical samples compared favorably with those obtained by manual procedures. We report data on correlation between different methods and preliminary data on precision of the prototype system.


Effect of extraction methods on two pregnancy related, antigenic components of human chorionic gonadotropin preparations.

One of the pregnancy related, antigenic components of HCG preparations is not readily detected in kaolin extracts of pregnancy urines; it is however, readily detected in these urines when concentrated by lyophilization or in their benzoic acid extracts.


In vivo activation of C3 revealed by crossed immunoelectrophoresis as a parameter of immunological activity in disease.

A technique suitable for the demonstration of in vivo activation of the third component of complement (C3) is described, and the results of detailed methodological studies are presented. The technique is a useful tool for monitoring the activity of the humoral immune system in disease states. The serum levels of complement components reflect a balance between synthesis and catabolic rates, while the present method gives a more dynamic picture of the state of the complement system. The results of the application of the technique to different groups of patients are presented.


The measurement of urinary digoxin and dihydrodigoxin by radioimmunoassay and by mass spectroscopy.

A radioimmunoassay for urinary digoxin is described which includes an initial solvent extraction to remove factors in urine which cause non-specific interference in the assay. The recoveries obtained using different solvents are compared and the non-specific factors influencing the assay investigated further. These effects were overcome by the use of a small urine volume (10 mul) in a direct, unextracted, urine assay and the results obtained correlated closely with those from the assay using prior extraction (r=0.99). No false positive results were obtained with unextracted urine samples from hospitalised patients not receiving digoxin. The specificity was also determined with regard to the natural steroids, spironolactone and the metabolites of digoxin including dihydrodigoxin. The metabolite dihydrodigoxin, with a saturated lactone ring, was not detected whereas the mono-, and bis-digitoxo-sides and digoxigenin metabolites did cross react in the assay. It was not possible to separate dihydrodigoxin and digoxin by thin-layer chromatography or solvent extraction due to their similar structures, however, mass spectroscopy was successful in this respect and was employed to obtain the ratio of dihydrodigoxin to digoxin in extracted urine samples. Levels of urinary digoxin excreted by patients maintained on different oral doses of the drug were measured. The percentage excreted in the urine as digoxin correlated closely with the oral dose (r = 0.96) but was found to be lower than that reported in most previous studies. Mass spectroscopy measurements showed that an average of 16.4% (range 12.2-19.7%) of the total oral dose was excreted as dihydrodigoxin in the urine of nine patients investigated.


Specific determination of testosterone in female plasma by radioimmunoassay: a rapid and reliable procedure for the routine clinical laboratory.

A rapid and reliable procedure for the specific determination of testosterone in female plasma is described. The assay entails a simple biphasic solvent partitioning of plasma extracts in order to remove cross-reacting steroids before quantitation of testosterone by radioimmunoassay using an antiserum raised against testosterone-11alpha-hemisuccinyl-bovine serum albumin. "Testosterone' values obtained in this way are in excellent agreeement with those obtained following thin-layer chromatographic purification of plasma extracts. However, the present procedure is easier, faster and cheaper to perform, and enables a single technician to assay 32 females plasma samples (in duplicate) in one working day.


Dominance and reproduction in Baboons (Papio cynocephalus).

This monograph reports on a 14 month study of yellow baboons (Papio cynocephalus) in the Masai-Amboseli Game Reserve, Kenya. The study was an attempt to determine the relationship between agonistic dominance and reproductive success in male baboons and centered around testing a priority-of-access model of mating behavior. Explicit criteria for determining dominance in baboons are presented and the consistency of dominance relationships through time is analyzed for all classes of individuals. Related data on reproductive cycle length, perineal and behavioral indications of the optimal day for mating, changes in female behavior during estrus, and effects of the presence of estrous females on group organization are also included. This work constitutes the first comprehensive field study of baboon mating systems and social organization and emphasizes the use of systematic behavior sampling techniques in the field and quantitative models in the study of primate social behavior.


Idiopathic hypertrophic subaortic stenosis with ostium secundum atrial septal defect: successful surgical correction.

The first report of coincident IHSS and ostium secundum ASD is corroborated and successful septal myotomy and myectomy with patch closure of the ASD is described.


Idiopathic hypertrophic subaortic stenosis and Wolff-Parkinson-White syndrome. Changes of obstruction in left ventricular outflow depending on the type of ventricular activation.

A case of idiopathic hypertrophic subaortic stenosis (IHSS) associated with Wolff-Parkinson-White (WPW) syndrome is studied. On three occasions preexcitation was suppressed with an intravenous injection of ajmaline. The left ventricular outflow tract obstruction (LVOTO) was analyzed; when WPW syndrome activation disappeared, the obstruction decreased. The ventricular depolarization pathway is noteworthy as a factor in the severity of LVOTO in IHSS.


Idiopathic hypertrophic subaortic stenosis presenting as cough syncope.

In a patient with idiopathic hypertrophic subaortic stenosis, syncope developed as a result of a sustained decrease in aortic pressure induced by severe cough paroxysms. Treatment with propranolol was effective in abolishing the syncopal episodes, by reducing the post-tussive gradient and facilitating a more rapid return to normal of aortic pressure. Post-tussive syncope in IHSS may result from both an unusually strong cough paroxysm and augmented left ventricular outflow obstruction consequent to reflex sympathetic stimulation.


Efficacy of standard anticonvulsants in monkey model with spontaneous motor seizures.

Monkeys were rendered chronically epileptic by injection of alumina gel into the pre- and postcentral gyrus. To test the validity of this primate model, the effects of diphenylhydantoin (DPH), phenobarbital, and primidone on spontaneous seizures evaluated for 8 months with a Latin-Squar experimental design. All three drugs were effective, the frequency of seizures being reduced by at least one-half during 6 weeks with treatment as compared with 6 weeks without. In most monkeys the frequency and severity of seizures were correlated to the number of interictal spikes in the EEG, and were inversely related to the level of drug in plasma. During withdrawal of phenobarbital and primidone, epileptic activity increased over that during control periods. Side effects were minimal with all three drugs. Patterns of behavior, although they differed from one monkey to the next, exhibited trends specific to each drug but particularly DPH. The seizures of some animals seemed to be related to the sleep-waking cycle.


Comparative electron-microscope studies on type-III and type-I collagens.

Long-spacing-segment crystallites prepared from type III collagen with the chain composition [alpha1(III)]3 and type I collagen with the chain composition [alpha(I)]i12alpha2 have been compared in the electron microscope after positive. staining with phosphotungstic acid and uranyl acetate. The comparison revealed several differences in intensities of the cross-striation bands as well as significant differences in band positions. The latter occur most prominently in three distinct regions of the crystallites. Further, crystallites prepared from type III collagen contain an additional intensely staining band in an area corresponding to the carboxy-terminal end of the molecule. The latter band is still observed following limited pepsin digestion and presumably represents a slight elongation of the helical portion of the type III molecule when compared to the type I molecule. In spite of the somewhat altered distribution of charged groups as indicated in studies on the long-spacing-segment crystallites, type III molecules are capable of forming fibrils of the native type with a cross-striation pattern and periodicity virtually identical to that observed when type I molecules are precipitated as native fibers.


Isohormones: molecular forms of the human chorionic somatomammotropic hormone.

The chorionic somatommaotropic hormone extracted from the human placenta exists in several molecular forms. Analytical electrophoresis in polyacrylamide gel permits separation of a highly anodic migration form, form 1, and another form migrating slightly faster than albumin, form 2. These two forms are active as measured by radioactive immunological analysis, form 2 being about 25 times more active than form 1. The two forms are mutually interconvertible. The two forms may also be separated by filtration on Sephadex G-50. However, they do not differ in molecular weight, they have the same coefficient of sedimentation and the same coefficient of apparent diffusion, measured by analytic ultracentrifugation. Glutaraldehyde and 8 M urea do not modify their electrophoretic or chromatographic behavior. On the hand, the two forms differ in their tertiary structure, with the modification depending on the greater or lesser degree of oxidation of the intra-chain disulfide groups. The two forms also exist in placental culture media and the incorporation of tritiated leucine occurs preferably in form 1. The physiological significance of the two hormone pools is not clarified.


Renaturation of acid-denatured rabbit muscle aldolase. Existence and properties of a stable monomeric intermediate.

The effects of temperature, pH and the substrate, fructose 1,6-bisphosphate, upon the kinetics and yield of renaturation of acid-denatured rabbit muscle aldolase have been investigated. The results are discussed in terms of a sequential set of events leading from the unfolded polypeptide chain to the renatured oligomeric enzyme. One of the intermediate molecular species in this sequence has been characterized as a folded monomer with a sedimentation coefficient of 3.1 S. This monomer is shown to be much more heat-labile than the tetramer under identical conditions, thus demonstrating stabilization of the tertiary structure of the polypeptide chain by the quaternary interactions between protomers.


Substrate stereochemistry of the enoyl-CoA hydratase reaction.

1. A specimen of stereospecifically 2-tritiated 3-hydroxybutyric acid was prepared by hydroboration of ethyl crotonate. It was assumed that the hydroboration reaction took a syn course and hence that (2R,3S) plus (2S,3R)-3-hydroxy[2 minus 3H1]butyric acid was formed after oxidation and hydrolysis. 2. 3RS-3-Hydroxy[2 minus 3H2]butyric acid, symmetrically tritiated at C-2, was prepared by isotopic exchange of ethyl acetoacetate in tritiated water, followed by reduction and hydrolysis. 3. The 3R-enantiomers of the acids listed under paragraphs (1) and (2) were destroyed enzymically by use of 3R-specific 3-hydroxybutyrate dehydrogenase and the residual 3S-enantiomers were isolated. 4. The resulting specimens of 2R,3S-3-hydroxy[2 minus 3H1]butyric acid and 3S-3hydroxy[2 minus 3H2]-butyric acid were converted chemically to the acyl-CoA derivatives. These were incubated with enoyl-CoA hydratase. 5. In the presence of the enoyl-CoA hydratase symmetrically labelled 3S-3-hydroxy[2 minus 3H2]BUTYRYL-CoA lost nearly 50% of its tritium label; 2R,3S-3hydroxy [2-3-H1]butyryl-CoA lost about 78%. 6. It was concluded that the elimination of the elements of water from 3-hydroxybutyryl-CoA on the hydratase occurs stereospecifically with syn geometry.


Adrenal, plasma and urinary corticosteroids during single or repeated administration of morphine in cats.

Corticosteroids and ascorbic acid in the adrenal glands of adult cats have been investigated after single or repeated administration of morphine. Also plasma levels and urinary excretion of corticosteroids were determined. A significant increase in the content of corticosteroids in the glands and plasma was found after initial injection of morphine. After 7 days of consecutive morphine treatment a fall of corticosteroids in the glands was observed; after 2 weeks of daily injections the content of adrenal corticosteroids was significantly lower than in the control animals but no change was found in the plasma. Administration of the drug during 1 month led to a highly significant decrease of corticosteroids in the glands as well as in plasma. No significant change in adrenal ascorbic acids was found whether the adrenal corticosteroids were higher or lower than in the control cats. Urinary corticosteroids were high during the first week of morphine treatment but thereafter the excretion declined progressively and was lower than the control level after 13 days of treatment. The significant decrease of corticosteroids observed after repeated administration of morphine and the rise in adrenal corticosteroids found after the injection of nalorphine to the morphinized animals suggest that some kind of morphine dependence had been developed in the cats after repeated administration of the drug.


Effects of penfluridol and other drugs on apomorphine-induced stereotyped behavior in monkeys.

The effects of some drugs on apomorphine-induced stereotyped behavior were studied in male cynomolgus monkeys. Apomorphine produced the dose-dependent stereotyped behavior characterized mainly by continuous licking and biting, and repetitive movements of the hands, head and body in the monkeys. Penfluridol as well as haloperidol showed a clear antagonistic effect on the apomorphine-induced stereotyped behavior, while chlorpromazine was less antagonistic than haloperidol. The antagonistic effect of penfluridol lasted longer than that of haloperidol. Reserpine did not inhibit the apomorphine-induced stereotyped behavior though the drug elicited markedly the behavioral depression and alpha-methyl-p-tyrosine also did not block the stereotyped behavior. Nialamide did not depress the apomorphine-induced stereotyped behavior. In provoking the stereotyped behavior in monkeys, apomorphine probably acts directly on dopamine receptors in the extrapyramidal system, and penfluridol is suggested to act as a dopamine receptor blocker with a long action. The results indicate that protection against apomorphine-induced stereotyped behavior in monkeys may be a useful method for evaluating neuroleptic drugs.


Effect of sodium nitroprusside on myocardial performance and venous tone.

The action of sodium nitroprusside upon the mechanical performance of the isolated, working rat heart and on isolated bull veins has been studied. Cumulative concentrations of 1--1000 ng sodium nitroprusside per ml had no effect on left ventricular pressure, end-diastolic left ventricular pressure, dp/dtmax, cardiac output and calculated wall stiffness of isolated rat hearts with constant rate, preload and afterload. Isolated strips of bull veins contracted with noradrenaline showed a dose dependent relaxation by sodium nitroprusside in concentrations of 5--1000 ng per ml. This study shows that sodium nitroprusside has no direct effect on myocardial contractility and causes venous relaxation.


The behavioural effects of dopamine applied intracerebrally to areas of the mesolimbic system.

The intracerebral injection technique was used to apply dopamine directly into dopamine-containing areas of the mesolimbic system, the nucleus accumbens septi, tuberculum olfactorium and nucleus amygdaloideus centralis. 200 mug dopamine injected bilaterally into the nucleus accumbens septi caused a stereotyped sniffing behaviour and hyperactivity but only a periodic hyperactivity developed after similar injections into the tuberculum olfactorium and no change in behaviour was observed following injections into the nucleus amygdaloideus centralis. After pretreatment with nialamide, the effects of intracerebral dopamine were enhanced, doses of 1-50 mug dopamine causing consistent stereotyped sniffing and a dose-dependent hyperactivity on injection into the nucleus accumbens septi or tuberculum olfactorium. The effects initiated from the nucleus accumbens septi were most marked. In addition, 50-100 mug dopamine injected into the tuberculum olfactorium caused a periodic biting behaviour. Biting was not observed after injections into the nucleus accumbens septi and, even in the presence of nialamide, injections of dopamine into the nucleus amygdaloideus centralis failed to cause any change in motor behaviour.


Central catecholamine receptor blocking actions of BE-2254 ('heat'): comparison with chlorpromazine and haloperidol.

BE-2254, 2-[beta-(4-hydroxyphenyl)-ethylaminomethyl]-tetralone, (ED50 = 3.4 mg/kg i.p.) was about equal to chlorpromazine (ED50 = 4.4) as an antagonist of central noradrenergic receptor stimulation produced by clonidine (enhancement of the flexor reflex in spinalized rats). Haloperidol and phentolamine had essentially no effect at 9 mg/kg i.p...


Genetically determined differences in the effects of morphine on mice.

We have compared a variety of well known behavioral actions of morphine across four strains of laboratroy mice. Thes measures included locomotor activation, analgesia, tolerance, dependence and drug-seeking. We found that analgesia and drug-seeking did not vary greatly among strains, although some statistically significant differences were found. The interstrain rank ordering for locomotor activity, tolerance and dependence was different for each of these measures. This suggests that each of these effects of morphine may depend upon separate mechanisms, perhaps with each under independent genetic control. Other experiments were carried out to determine whether or not there were differences in the in vivo brain uptake of an opiate or the in vitro binding of naloxone to 'stereospecific receptors'. No significant interstrain differences could be found.


Chronic administration of morphine in cats: effects on adrenal and urinary catecholamines.

Adrenaline and noradrenaline levels in the adrenal glands and the excretion of both bioamines in urine of adult cats were investigated after chronic administration of morphine and nalorphine-induced withdrawal. After 7 days of daily consecutive morphine treatment, a significant increase in the adrenal noradrenaline content and a drop in adrenaline content were observed. After 2 weeks of daily injection of morphine, no significant changes were observed in the adrenal catecholamine level. One month of treatment with the opioid caused a significant increase in the adrenal content of both adrenaline and noradrenaline. Urinary excretion of catecholamines was significantly increased during the 4 weeks of treatment. In animals subjected to spontaneous or induced withdrawal with nalorphine, the adrenal content of catecholamines was altered and the ratio adrenaline/noradrenaline in the adrenal gland was shifted towards noradrenaline. A first injection of morphine produced an excitant manic response characterized by hyperexcitement and aggressive behaviour; animals chronically treated with the drug showed a progressively diminished response to this effect of the drug. It is concluded that physical dependence on morphine is reached by cats chronically treated with morphine and that this effect of the drug influences adrenomedulllary function in a different fashion depending on the stage of morphine treatment.


Intravenous self-administration of d- and l-amphetamine by dog.

The relative potency of d- and l-amphetamine to maintain i.v. self-administration behavior was studied. 5 dogs were trained to work for response-contingent drug infusions until a stable drug intake per 4 hr daily session was achieved. Then 2 unit doses of d-amphetamine (0.05 and 0.10 mg/kg/infusion) and 3 unit doses of l-amphetamine (0.20, 0.40 and 0.80 mg/kg/infusion) were evaluated in a parallel line bioassay. Each combination of drug and unit dose was examined separately for 5 consecutive daily sessions. Order of treatment presentation was determined by a Latin square design. By comparing the unit doses of d- and l-amphetamine which yielded the same rate of self-administration it was found that 1 mg of the l-isomer is equivalent to 0.17 mg of the d-isomer.


Response of serum prolactin to exogenous stimulation.

Previous observations by other workers indicating suppression of serum prolactin (hPRL) by water loading could not be confirmed. Sequential testing using an acute water load in patients with various clinical diagnoses did not aid conclusively in differentiating functional hyperlactinemic states from pituitary adenoma. It was of interest that the acute ingestion of water resulted in a triphasic response in serum hPRL levels. The response to breast stimulation was more marked in patients with excessive breast hypertrophy and galactorrhea. This was in contrast to patients with hypo- or hyperthyroidism, anorexia nervosa, and our normal subjects. In response to breast stimulation, there was no difference among the normal group, women with functional hyperlactinemia, and patients with proven pituitary adenoma. When they were compared with other patient groups in this study, marked responsiveness was found in patients with amenorrhea and galactorrhea, amenorrhea with elevated hPRL levels without galactorrhea, breast hypertrophy, and adrenal overactivity in response to these two stimuli. The physiologic mechanisms resulting in the increase in serum hPRL levels seen in these patients remain unclear at the present time.


Hormone-releasing silastic intrauterine devices: effect or provera-releasing intrauterine devices on two species of primates.

Silastic intrauterine contraceptive devices (IUDs) 13 mm long and 1.07 mm in diameter could be inserted easily into patas monkey uteri which, like human uteri, expelled them. Addition of 10% Provera to these devices did not reduce the expulsion rate significantly in our study. Control devices had no effect on cycle length in rhesus monkeys. After insertion, the active IUDs frequently caused a delay in onset of menstruation; however, cycles did occur with the device in situ, and normal-length cycles were resumed following removal of the device. A short period of rapid release (almost 35% of the total amount) of Provera from the device was followed by a longer period of sustained release of low levels of the hormone. Even 9 mug/day were sufficient to maintain a decidual reaction in the endometrium of the rhesus monkey. The drug could not be detected in the blood stream at 3,6, or 12 hours in patas monkeys or at 1 or 2 months in rhesus monkeys and so may never have reached the systemic circulation. Devices currently under study in baboons catain Provera or one of three other steroids to determine whether these compounds improve retention rates as well as meet the other two criteria set for the ideal IUD incorporation, for unless we meet this first criterion we can never achieve, let alone test, the others.


Mumps oophoritis: a cause of premature menopause.

One cause of secondary oligomenorrhea is ovarian infection. A rare type of infection related to the disturbance of menstrual function is mumps oophoritis. Three patients with premature menopause presumably caused by this agent were described. In one patient the symptoms coincided with a subclinical infection during the perinatal period, with subsequent infertility. Another patient seemed to have had a clinically mild oophoritis during the pubertal period, and the third patient became symptomatic following parturition. It appears that this aberration in menstrual function and fertility may be related to the time during which the infection occurs as well as to the severity of the infection. In addition, it is apparent that mumps oophoritis may be a more frequent cause of premature menopause than has heen previously suspected.


Ultrastructure of rabbit ova recovered from ovarian follicles and inseminated in vitro.

Rabbit ova were obtained from presumably preovulatory ovarian follicles of gonadotropin-treated does and inseminated with uterine sperm in vitro. Ova were serially sectioned for light and electron microscopy 0, 1 1/2, 3, 6, and 9 hours after in vitro insemination. Control ova were maintained in culture for 25 hours and examined by light microscopy for cleavage. Little difference in fertilization rate (i.e., ova fertilized/ova inseminated) was observed when criteria were based on ultrastructural evidence (56.5%) and on light microscopic evidence 25 hours post-insemination provided by cleavage of control ova (54.8%). Therefore, observations of cleavage (as in controls here) can be an accurate criterion for fertilization in the rabbit, at least under the in vitro conditions used in this study. After insemination of ova recovered from preovulatory follicles, the developmental sequence was as follows: after 1 1/2 hours, sperm penetration into the vitellus; after 3 hours, male pronuclear formation and second polar body extrusion; after 6 hours, male and female pronuclear enlargement; and after 9 hours, pronuclei in apposition. This sequence compares favorably with the normal temporal sequence of events already well known for fertilization of ovulated ova in the rabbit. Penetrating sperm cells within the matrix of the zona pellucida and the supplementary sperm cells lying in the perivitelline space had undergone the acrosome reaction. Changes associated with the acrosome reaction took place before the penetration of the zona pellucida. The presence of a number of supplementary sperm, usually five to six per ovum, within the perivitelline space of the already activated ovum indicated that the block to polyspermy in these experiments operated mainly at the level of the vitelline membrane, as is the case in the normal fertilization process. The vitelline membrane block to polyspermy occurred rapidly after penetration of the fertilizing sperm into the vitellus and was associated with cortical granule breakdown. Ultrastructural details of rabbit ova recovered from ovarian follicles and inseminated in vitro revealed no distinguishable characteristics when compared with reported observations of ovulated rabbit ova undergoing fertilization in vivo.


[Effect of MMI on the serum T3 and T4 levels in hyperthyroid patients (author's transl)].

Effect of 1-methyl-2-mercaptoimidazole (MMI) on the serum triiodothyronine (T3) and thyroxine (T4) was studied in six patients with hyperthyroidism. From 30 to 60 mg of MMI per day were administered to the patients orally. Serum T3 was measured by radioimmunoassay and T4 by radioimmunoassay of Tetrasorb Kit (Abbott Lab.). Serum T4/T3 ratios in these patients were low initially, but increased to normal range in two cases immediately after the administration of MMI. But they were unchanged at first in the other four cases. When high serum T3 values decreased abruptly to normal at a later course of medication, the serum T4/T3 ratios increased to normal range in almost all cases. Even though the serum T4/T3 ratio became normal, serum T3 and T4 values remained high in some cases. Elevated serum T4 values in all cases decreased to normal faster than serum T3 with the administration of MMI. When MMI was administered to the patents concinuously, the T4/T3 ratios, once increased to normal, then decreased again gradually in five cases. And serum T3 which decreased to normal sometimes gave high values. After the discontinuation of MMI, serum T3 increased over the normal range more rapidly than did T4.


[Effect of d,l-alpha-methyl-3, 5, 3, 5-tetraiodothyronine ethylester hydrochloride (CG-635) on experimental hypercholesterolemia and atherosclerosis in rabbits (author's transl)].

To develop the prophylactics and the curatives for atherosclerosis, thyroxine derivative, CG-635, was assayed for its physiological activities in experimental atherosclerosis in rabbits fed with cholesterol. It was found that CG-635 possessed serum TC/TP value lowering activity (total cholesterol/total phospholipid) in normal and cholesterol fed rabbits for 3 weeks, and prevented the elevation of the value of cholesterol fed rabbits by daily injection for 7 weeks. CG-635 also depressed the hyperlipemia induced by cholesterol feeding, and its inhibitory effect was shown to be more marked on the increase of cholesterol than triglyceride, phospholipid and free fatty acid in serum. CG-635 did not, however, influence GOT, GPT and G-6-Pase activities in serum with increased cholesterol intake. From the histological findings it was proved that this compound prevented to a high degree the occurrence of atherosclerosis and fatty liver of cholesterol fed rabbits. Furthermore, it was recognized that thyroid hormone and the thyroid simulating hormone-like activities of CG-635 were much weaker than thyroxine, except for the action in the lipid metabolism.


[A low response of pituitary growth hormone secretion in the patient with diabetes mellitus after intravenous administration of L-dopa (author's transl)].

Recently, L-dopa has been known as one of drugs to stimulate the secretion of growth hormone from the pituitary gland through dopamine, which is a metabolic of L-dopa, without any changes of serum insulin levels, blood sugar values, fatty acids and amino acids concentrations in the serum of human being. The present study was thus designed to assess the effect of L-dopa on the pituitary gland to secrete the growth hormone in the normal subject and the study was further extended to compare the response of the pituitary secretion of the growth hormone following the administration of L-dopa between in the patient with diabetes mellitus and in the normal subject. In the present experiment, 5 normal subjects and 11 diabetes mellitus without obesity were employed and they received a 30 min infusion of 200 mg L-dopa dissolved in 200 ml physiological saline. In normal subjects, serum growth hormone concentration measured by radio-immunoassay was started to increase within 10 min after L-dopa administration and maximum value of serum growth hormone was obtained 60 min after the drug, mean values of it, 30 ng/ml of serum. Then it was declined sharply upto the values of 2-3 ng/ml. In the patients with diabetes mellitus, on the other hand, maximum value of serum concentration of the growth hormone was only approximately 5 ng/ml of serum obtained between 45 and 75 min after the administration of L-dopa. No changes in the serum concentration of IRI (Immunoreactive insulin) and of blood sugar values were observed by the infusion of L-dopa in both normal subjects and diabetes mellitus. From the above mentioned facts, it was concluded that the ability of pituitary gland to secrete the growth hormone was considerably impaired in diabetics when it was compared with normal subjects.


Antigenic activity, in humans, of purified low molecular weight fractions of timothy pollen.

Small molecular weight fractions obtained from dialysis of crude timothy pollen extract were shown to carry immunogenic activity in monkeys. These fractions were sequentially purified through gel filtration with Sephadex G-10 and C-M cellulose. Monkeys immunized with these fractions produced almost eclusively skin-sensitizing antibodies which were shown to be of the IgE class, differing in antigenic specificity. These active fractions were assayed in patients allergic to timothy pollen exhibiting an allergic rhinitis. Direct skin tests and inhalation provocation tests were used. The results showed a discrepancy between the skin reactivity and the nasal response to the 3 active fractions. This result can be interpreted as heterogenity of IgE antibodies to small molecular weight fractions contained in crude timothy pollen extract, exhibiting different tissue sensitizing properties.


Interactions between IgE and house dust extract as studied by the radioallergosorbent test (RAST).

Several components in house dust extract can bind IgE, these interactions are not all immunologically specific. Non-immune interactions may conceivably contribute to the biological effects of house dust extract, they are actually found to interfere with the assay of allergen-specific IgE, especially when the serum under investigation has a high IgE content. By modification of the incubation medium, non-immune interactions can to a large extent be prevented. The immunological interactions between IgE and components in house dust extract have been investigated using twenty extracts, both in inhibition-type assay and in direct insolubilization assay. D. pteronyssinus-related allergens were found to be present in almost all extracts, however, not all house dust specific IgE reacted with mite-related allergens. Whether standardization of house dust extract is at all feasible, and if so, worthwhile, remains to be seen. It is clear, however, that no single figure can represent a completely reliable measure for the biological potency of a house dust extract.


Value of fluid phase complement consumption as a means of 'in vitro' allergen standardization.

Atopic allergens can readily be shown to inactivate hemolytic complement in human serum. This property has been explored as a means of in vitro allergen quantitation and standardization. The purification of house dust allergen, guided by skin tests and complement inactivation, demonstrates that the in vitro technique is clinically relevant. Since human sera differ in their sensitivity to complement inactivation by allergens, it is desirable to prepare a purified house dust allergen as a standard. By referring to this standard, other house dust extracts may then be standardized by using any given serum sample of human origin, provided that such extracts have undergone some degree of purification. Kinetic studies demonstrate that other allergens of the house dust group may be standardized in similar fashion; it was noted that the ratio of the complement inactivating powers of these respective allergens remains constant whatever the serum used. The method is not suited for the quantitation of pollen allergens.


The plasma insulin and growth hormone response to oral glucose: diurnal and seasonal observations in the Antarctic.

Plasma insulin and growth hormone levels were measured during morning and afternoon oral glucose tolerance tests performed on 12 young men at three monthly intervals in the Antarctic. No diurnal or seasonal differences in growth hormone levels were found. However there were diurnal and seasonal variations in the blood glucose/plasma insulin relationship.


Effects of buformin on the metabolsim of the isolated haemoglobin-free perfused hindlimb of normal rats.

Using the isolated, haemoglobin-free, perfused resting hindlimb of normal rats buformin neither had a direct insulin-like effect on glucose uptake by muscle tissue nor potentiated the effect of insulin on glucose uptake after oral pretreatment for one or several days with low and high doses (30 mg-350 mg/kg). Effects on glycogenolysis could not be detected. Glycerol release was inhibited after several days of pretreatment with low and high doses of buformin. The utilization of added oleate was also partly inhibited. The level of energy rich phosphates in the muscle tissue and oxygen consumption were not affected under any of the conditions used in these experiments.


Plasma levels and excretion of estrogens in urine in chronic lever disease.

Plasma levels of nonconjugated estrone (E1), estradiol-17beta (E2), and estriol (E3) and excretion of these estrogens in urine in conjugated and nonconjugated forms were measured by radioimmunoassay in 17 healthy males and 33 males with alcoholic liver disease. Patients with liver disease had significantly elevated plasma levels of E1 (74.2 +/- 8.0 SE versus 26.0 +/- 1.7 pg per ml, P less than 0.001), E2 (29.3 +/- 2.2 versus 23.3 +/- 2.0 pg per ml, P = less than 0.05), and E3 (11.5 + 1.9 versus 6.5 +/- 0.7 pg per ml, P less than 0.01). Excretion of total E1, E2, and E3 in urine was significantly increased in patients with liver disease; the increase was primarily in the nonconjugated fraction. Among those patients with chronic liver disease, the presence of ascites was associated with significantly higher plasma levels of E1 and excretion of larger amounts of E1 and E2 in urine. On the other hand, no differences were observed in the small number of patients with gynecomastia. A direct correlation between the plasma level or excretion of E1 or E2 and several liver function abnormalities was observed. The relative hyperestrogenemia we report here may account for some of the clinical observations in chronic liver disease.


Alcohol-induced testicular atrophy. An experimental model for hypogonadism occurring in chronic alcoholic men.

Elucidation of mechanisms involved in the hypogonadism and feminization observed in chronic alcoholic men requires the development of an experimental animals model system. Such an animal system should be inducible with ethanol feeding and should duplicate endocrine changes known to occur in chronic alcoholic men. We report such an animal model system. Animals fed a diet with ethanol accounting for 36% of total calories develop significant testicular, prostatic, and seminal vesicle atrophy (P less than 0.01) and greatly reduced plasma testosterone levels (P less than 0.01). Animals fed a similar diet with sucrose isocalorically substituted for ethanol do not. Testicular, prostatic, and seminal vesicular mass relative to body mass and plasma testosterone levels in these isocaloric control animals do not vary significantly from those obtained for age-matched control animals fed an ad libitum rat chow diet. These findings indicate that the caloric deprivation associated with chronic ethanol ingestion is not responsible for gonadal injury and atrophy of the sex steroid-sensitive tissues in the alcohol-fed animals. This animal model provides a useful means of directly examining perturbation in gonadal function that occurs in man as a consequence of chronic ethanol ingestion and confirms our previous data which suggest that ethanol is a primary testicular toxin.


Mutagenesis and genetic analysis with Chinese hamster auxotrophic cell markers.

Chinese hamster ovary cells were treated with various physical and chemical mutagens and subject to the bromodeoxyuridine plus visible light procedure for isolation of auxotrophic mutants. more than 200 auxotrophs have been isolated which require exogenous supplement of various nutrilites for growth, such as glycine, adenine, thymidine, inositol, etc. Fifty-five of these have been characterized by complementation tests and were shown to constitute 15 different loci. All these auxotrophs are highly stable and exhibit all-or-none growth response to the respective nutrilites. Enzyme deficiencies have been identified in several of these classes. When these auxotrophs are hybridized with human cells and grown in selective medium, the hybrids lose human chromosomes at a rapid rate and the analysis for synteny of the human genes can be successfully carried out. Moverover, in hybrids formed between an adenine-requiring auxotroph and human cells, a human esterase  activator gene has been identified which appears to regulate the expression of esterase gene activities in the Chinese hamster genome. Studies of this kind may lead to the understanding of gene regulation in mammalian cells.


Crohn's disease in Nottingham: a search for time-space clustering.

All cases of Crohn's disease in the Nottingham area were ascertained and the date and place of domicile and work at the time of onset of symptoms noted. Applying the Knox and Pike and Smith tests for clustering of patients in time and space, no significant results were observed. Further analysis of differences in time-space clustering between a group of patients and matched controls is in hand.


[Mode of action of clonidine, a central hypotensive agent, analyzed by the interaction with various sympathomimetic agents].

Injection of a low dose of clonidine showed the biphasic blood pressure response which comprises a delayed onset of long lasting hypotension and a quick onset of hypertension of short duration followed by the former, in the alpha-chloralose-urethanized rat. A second injection of low dose of clonidine administered 90 min after the first showed only two peaks of pressor response, no longer a depressor one, and tachyphylaxis occurred with repeated injections of the same dose of clonidine, together with a gradual elevation of blood pressure level. Furthermore, the depressor action was inhibited by pretreatment with cocaine or imipramine. In contrast, injection of a high dose of clonidine exhibited pressor response with only two peaks, duration of which was markedly potentiated by pretreatment with cocaine or imipramine, and also by guanethidine, but inhibited in reserpinized and spinal rats. While the blood pressure was sustained at a high level after the injection of a high dose os clonidine, blood pressure reversal was produced by tyramine, due to possible beta-mimetic action of much greater amounts of catecholamines released by tyramine. The isolated guinea-pig vas deferens contracted when treated with clonidine, and this contraction was inhibited in the reserpinized preparation. This inhibition was recovered by incubation with norepinephrine. Interaction between tyramine and clonidine was also seen in vitro, but it disappeared in the reserpinized, denervated or propranolol-pretreated guinea-pig vas deferens. The contraction of the vas deferens induced by clonidine was potentiated by pretreatment with cocaine, but not by pretreatment with guanethidine, ans was inhibited by treatment with alpha-adrenergic blocking agents phentolamine and phentolame and phenoxybenzamine, but not by tolazoline.


[Dependence on and preference for morphine (II). Comparison among morphine, phenobarbital and diazepam].

Results of a previous experiment indicated that naive rats given a choice between morphine-admixed food (0.5 similar to 1mg/g of food) and quinine-admixed food (0.5 similar to 1 mg/g of food) for 3 weeks gradually and spontaneously preferred the morphine-admixed food, and this choice behavior revealed one of the psychological aspects of morphine dependent rats. In the present work, the ability of preference formation was detected for morphine, phenobarbital and diazepam by a defferent chioce test using drug dependent rats. Rats were pretreated with morpnine, phenobarbital, and/or diazepam by drug admixed food ingestion method for 12 similar to 15 weeks, and the choice test was practiced for one week at 3 intervals (4 similar to 5 trials). Control groups of rats were given the same choice testas the naive rats. Results indicated that (a) of all the drugs employed, morphine showed the most rapid and intensive preference formation (b) Phenobarbital and diazepam had almost the same degree of preference formation. (c) Phenobarbital showed no dose-dependent intensity of preference formation in the 50-90 mg/kg/day dose range, however, a more rapid development of preference was observed dose-dependently among the 80-120 mg/kg/day dose range of diazepam. Thus these studies utilizing a choice test provide a clear demonstration of drug-seeking behavior in rats. In addition, the present method is useful for drug screening tests involving weak psychological dependence liability, and moreover, the data could be analyzed statistically.


[Effects of drugs on cell culture (I)].

Effects of drugs were studied on L-cell line: L, of mouse subcutaneous tissue origin. Results are as follows: the concentrations of 62.50 approximately 500 mug/ml for each drug were examined regarding inhibition of the growth of L-cell line. ID50 values of ibuprofen, naproxen, Y-5554, DICHLOFENAC AND AMINOPYRINE WERE FOUND TO BE 185, 185, 175, 145 AND GREATER THAN 500 MUG/ML RESPECTIVELY. Minimum concentration, caused by the detachment of the cell from the vessel wall, was as follows: ibuprofen 125 approximately 250, naproxen 250 approximately 500, Y-5554 125 approximately 250, dichlofenac 62.50 and aminopyrine 1,000 approximately 2,000 mug/ml respectively.


[Effects of drugs on cell culture (II)].

The toxicity of drugs was determined using embryonic skin and muscle from humans (Flow 1,000). The dosage causing a 50% inhibition culture growth (ID50) and minimum concentration, caused by the detachment of the cell from the vessel wall, were determined. ID50 values of ibuprofen, naproxen, Y-5554, dichlofenac and aminopyrine were found to be 150, 320,220, 110 and greater 500 mug/ml respectively. Minimum concentration, caused by the detachment of the from the vessel wall, was as follows: ibuprofen 250, naproxen 250-500, Y-5554 250-500, dichlofenac 62.50-125 and aminopyrine 1,000-2,000 mug/ml respectively.


[Simulation of cold acclimation by chronic treatment with noradrenaline, with special reference to metabolic responses to noradrenaline (author's transl)].

Rats treated with noradrenaline (NA) or L-dopa for 28 days were infused i.v. with NA for 30 min and changes in rectal temperature and plasma concentrations of nonesterified fatty acids (NEFA), glycerol, blood sugar, and lactate were observed. Results obtained were compared with those in cold-acclimated rats. The NA infusions caused a significant elevation of the rectal temperature in cold-acclimated and NA-treated rats, but not in L-dopa-treated ones. NA-induced increase in the plasma NEFA level was less in extent in cold-acclimated rats than in warm-adapted ones. Similar difference in the NEFA increase was observed between NA-treated and control rats. Changes in plasma glycerol level were the same in cold-acclimated and warm-adapted rats and in NA-treated and control rats. NA-induced increase in blood sugar level was less in extent in cold-acclimated and NA-treated rats as compared to the increase in respective control rats. No consistent changes were observed in blood lactate concentration. From the results it was inferred that the responses to exogenous NA in NA-treated rats, but not in L-dopa treated ones, were similar to those in cold-acclimated rats. It was thus suggested that the chronic treatment with NA may produce similar metabolic state as in cold-acclimation.


[Changes in blood constituents induced by noradrenaline infusion and acute cold exposure in rats treated chronically with noradrenaline and thyroxine (author's transl)].

Rats were treated with noradrenaline (NA), thyroxine (T4), NA plus T4, and NA plus ACTH for 28 days, and changes in rectal temperature and blood concentrations of nonesterified fatty acids (NEFA), sugar, and lactate were observed following NA infusion and acute cold exposure. Results obtained were compared with those found in cold-acclimated rats. NA infusion caused a marked elevation of the rectal temperature in cold-acclimated rats and NA plus T4-treated ones. No significant changes were observed in plasma NEFA level following NA infusion in cold-acclimated and NA-treated rats. Treatment with T4 tended to augment the lipolytic action of NA. Treatment with NA plus ACTH showed similar effect of augmented lipolysis. Changes in blood sugar level induced by NA infusion were small in cold-acclimated and NA plus T4 groups. Changes in blood lactate level were not significant in NA plus ACTH group. After acute cold exposure plasma NEFA level was low in cold-acclimated rats and high in NA plus T4-treated rats. Blood sugar level was slightly higher in cold-acclimated and NA plus T4-treated rats than in the other groups. Plasma lactate level showed relatively high values in T4 and NA plus T4 groups. It was thus suggested that metabolic patterns in cold-acclimated rats and in NA plus T4- as well as NA-treated rats were somewhat alike, but not always the same.


[On the toxicity of CT-1341 evoked by long-term administration. I. Subacute toxicity of the intravenous anesthetic, CT-1341 in rats (author's transl)].

CT-1341, an intravenous anesthetic was given in various daily doses in rats for a period of one month to test the subacute toxicity. The drug was administered intraperitoneally in rats. Rats tolerated to daily administration of CT-1341 at doses of less than 1.8 ml/kg without showing other particular toxic signs than anesthesia. Main pathological findings were swelling of cells in the liver and renal tubules, and perivascular cuffing in lungs. No severe patho-histological changes were observed in any organs. Mortal cases were seen in the group of rats, in which CT-1341 was given in a daily dose of 5.4 ml/kg. A paralysis of respiratory center was suggested to be cause of death, because no severe patho-histological changes were observed in any organs of mortal rats. Survivals of this group showed no particular symptom except anesthesia, but an inhibition of the growth curve was seen in male rats only.


[On the toxicity of CT-1341 evoked by long-term administration. II. Subacute and chronic toxicities of alphaxalone in rats (author's transl)].

Alphaxalone, an anesthetic steroid dissolved in 20% Cremophor solution was administered intraperitoneally to test the subacute toxicity (administration for one month) and chronic toxicity (administration for 3 months). In daily doses less than 8 mg/kg, alphaxalone did not show any particular toxic sign after administered for three months. Rats tolerated to daily administration of 20 mg/kg for three months, without showing severe toxic signs in body weight curve, blood cells and biochemical data obtained in blood and urine. However, some female rats receiving 50 mg/kg/day of alphaxalone, died by paralysis of respiratory center at the second day. Main histo-pathological changes induced by subacute and chronic administrations of the larger doses than 20 mg/kg, were swelling of cells in the liver and kidneys, but severe pathological changes were not seen in any organs.


[The lysis of proteins with cyanogen bromide (author's transl)].

Bovine beta-lactoglobulin-AB was split with cyanogen bromide, and the reaction mixture was analyzed by countercurrent distribution, gel chromatography and finally, chromatography on phosphocellulose. In addition to the previously described splitting products, we obtained three more minor products, with yields of 8, 15 and 25%. The analytical data indicate that these were formed by lysis C-terminal from the tryptophan-19 and the tryptophan-61 of beta-lactoglobulin. The result is discussed.


Constitutive heterochromatin and micronucleoli in the human oocyte at the diplotene stage.

In the diplotene stage of the human oocyte, the processes of elaboration of the nucleolar material are amplified. The principal nucleoli are more voluminous but their relations with the secondary constrictions and the satellites of the D and G chromosomes are not modified. Numerous micronucleoli, frequently to the number of 15-20 this stage. The most remarkable point is their association to various segments of constitutive heterochromatin: centromeric regions, secondary constrictions of the C9 and probably of the A1 and E16. These observations reveal that the human oocyte at the diplotene stage shows an amplification of the ribosomal cistrons. This phenomenon is homologous, to a more reduced scale, of this described from the inferior vetebrates. Besides, the role of heterochromatin in the synthesis of nucleolar material without the intervention of the classic nucleolar organizers is suggested.


Chromosome anomalies in mouse oocytes after irradiation.

We investigated the cytogenetic effects of X-rays on unfertilized mouse oocytes. NMRI females received an irradiation with 0,22.2,66.6,200, and 600 R during the preovulatory phase 3 hrs after HCG (human chorionic gonadotrophin). This is a stage during oogenesis in which the oocytes pass from late dictyotene to diakinesis. Chromosome analysis was performed after ovulation at metaphase II. From these experiments we can draw the following conclusions: 1) X-rays induced during the preovulatory phase a high number of chromosome anomalies. Among these, structural anomalies prevail. 7 out of 144 ovulated oocytes in matched controls carried such an abnormality, whereas after irradiation we observed with 22.2, 66.6, 200, and 600 R, 11 out of 72, 34 out of 108, 89 out of 102, and 122 out of 124, respectively. 2) Irradiation seems also to affect the chromosome segregation during the 1. meiotic division, as we observed after 22.2, 66.6, and 200 R a total of 6 oocytes out of 204 with a supernummary chromosome. In controls, however, no hyperploidy was found in 143 ova. This increase, however, was not significant. 3) Chromosome anomalies, e.g. breaks and deletions that go back to a one-break event increased linearly with increasing dose. Exchanges, however, going back to two-break events fittest best to the linear-quadratic dose-response model. 4) The dose of 600 R seems to represents a kind of borderline in this experiment, because nearly all (122 out 124) carried at least one structural chromosome anomaly. It is also this dose after which the highest frequency of reciprocal translocations was observed in a hump-shaped slope in spermatocytes after irradiation of spermatogonia (Preston and Brewen, 1973). With an increasing dosage up to 1200 R the frequency of translocations decrease again. The elimination of cells, crossing this borderline, might be due to genetic or non-genetic effects. 5) The frequency of radiation-induced translocations per oocyte agrees with the frequency of translocations in human lymphocytes (Dolphin and Lloyd, 1974) after in vitro irradiation. 6) Significant, lower frequencies of structural chromosome anomalies were observed irradiating earlier stages of mouse oogenesis. These stages are dictyotene from females at the age of 3, or 6 weeks and prophase I-stages in female embryos on the 17th day of gestation. This result may be due to a lower sensitivity of these stages or to modifying events during the interval between irradiation and preparations.


Evidence for two active X chromosomes in a human XXY triploid.

A human XXY chromatin negative triploid culture has been found to have both the A and B isozymes of glucose-6-phosphate dehydrogenase, as well as an intermediately migrating AB (hybrid) band. This finding indicates that the cells in this culture each possess 2 active X chromosomes. Possible mechanisms producing the abnormalities seen in this disorder are presented and discussed.


Translocation +(7p+; Bq-) associated with recurrent abortion.

A balanced translocation was found in a normal female with a history of four abortions. On the basis of the Giemsa-banding pattern the abnormality was interpreted as to be a translocation of a part of the long arm of chromosome 13 to the short arm of chromosome some 7:t(7;13)(7qter leads to 7p22::13q14 leads to 13qter;13q14 leads to 13pter::7p22 leads to 7 pter). Problems in genetic counseling are discussed with respect to this case.


Studies on the inhibition of C56-initiated lysis (reactive lysis).  V. The roleof C567-INH in the regulation of complement-dependent haemolysis initiated by cobravenom factor.

Activation of the alternative pathway of complement by a factor from cobra venom (CVF) can lead to lysis of unsensitized erythrocytes (E) of some species.  In these studies we observed that alterations in CVF-induced lysis could be produced by manipulation of C567-INH, a naturally occurring inhibitory activity which acts on fluid phase C567 complexes.  Venom lysis of sheep and guinea-pig E was markedly inhibited by serum fractions having C567-INH activity.  Microgram quantities of poly-L-lysine (PLL), molecular weight 180,000, a polycation which is a functional antagonist to C567-INH in serum, potentiated CVF lysis of sheep and guinea-pig E, and permitted the lysis of human E, which are otherwise not suscepticle to CVF lysis.  The potentiation of venom lysis by PLL seemed not to be due to alterations in the target cell membrane; furthermore, it in turn was reversed by substances with C567-INH activity.  This suggests that the generation of fluid phase C567 complexes contributes to the CVF-induced lysis of erythrocytes of these species, and that the haemolytic potential of fluid phase C567 generated during alternative pathway activation by this means is regulated by C567-INH.