Sheep erythrocyte rosettes in pigs, sheep, cattle and goats demonstrated in the presence of dextran. Formation of rosettes  with sheep RBC by blood lymphocytes from young pigs was increased from 24.4% +/- 2.2 (mean +/- S.E.) in PBS to 54.1% +/- 1.9 in the presence of dextran. This increase was achieved without inducing appreciable rosette formation with other RBC which do not form rosettes in PBS. Lymphocytes which rosette only in dextran are predominant in pigs between 20 and 160 days old, when the peripheral lymphocyte pool is increasing very rapidly. The use of dextran revealed major populations of blood lymphocytes rosetting with SRBC in adult sheep (30.7% +/- 2.0), adult cattle (37.3% +/- 4.2) and adult goats (13.3% +/- 1.2). Proportions of rosette-forming lymphocytes tended to increase with age. In calf lymphoid tissues the distribution of rosette-forming lymphocytes suggested that these were T cells. In the improvement of rosette formation with SRBC, dextran was more effective than foetal calf serum, papain treatment of the SRBC or combinations of these treatments.


Kinetics of immunoglobulin production by cultured human peripheral blood lymphocytes. A study of immunoglobulin synthesis by human peripheral blood lymphocytes cultured in Marbrook flasks is described. Maximal amounts of immunoglobulin were produced by 3 weeks and most of this was synthesised between 6 and 21 days. In the region of 90% of the immunoglobulin was secreted into the medium. Pokeweed mitogen (PWM) stimulated cultures produced about 10 times as much immunoglobulin as cells cultured in medium only. After 21 days stimulated cultures produced about 150 microgram of IgG, 125 microgram of IgM and 50 microgram of IgA per 10(7) cells.


In vitro stimulation of goat peripheral blood lymphocytes: optimization and kinetics of the response to mitogens and to allogeneic lymphocytes. Experiments were performed to establish the optimal concentrations for specific and non-specific lymphocyte transformation in vitro, in cultures containing 10(5)-10(6) caprine lymphocytes. A microculture system was used in conjunction with a semiautomatic sample harvester. The assay was optimized for mitogen concentration (PHA-P, Con A, PWM and LPS) and allogeneic cell number, number of responding cells, incubation time and amount of tracer. The effect of addition of serum and the cytotoxic effect of phytomitogens on cultured cells is discussed.


Lymphocyte response to phytohemagglutinin: temporal variation in normal dogs. The lymphocyte response to mitogens is frequently used as a screening method for monitoring general immunocompetence. In practice, however, the technique is potentially misleading. The results presented here suggest that temporal variation in lymphocyte response is the norm. Considerable variation in response to phytohemagglutinin was found for several dogs tested over different time periods. The response for any individual dog also fluctuated whether testing was at 2-week intervals, daily, or even every 4 h. This temporal variation remained whether separated lymphocytes or whole blood was cultured and when the results were expressed on a per lymphocyte basis. This suggests that there is a real difference in the response capability of individual lymphocytes. Factors affecting the numbers of leukocytes and the proportions of different types of lymphocytes in the peripheral blood are suggested as a possible source of the variation. Because of the inherent variation in lymphocyte responsiveness, it is necessary to test at different intervals to obtain an accurate indication of the general status of an individual's immunocompetence and to determine the time of peak response capability for that individual.


Quantitation of secretory protein levels by radioimmunoassay. A radioimmunoassay was designed for the detection of secretory protein, a component of secretory immunoglobulin A, in human serum. The assay uses free secretory protein isolated from human colostrum, and antisera raised in rabbits to the purified antigen. The mean level of secretory protein in the control group was 2.34 +/- 0.41 microgram/ml (mean +/- S.E.M.). The level in cord blood was slightly lower (0.74 +/- 0.26 microgram/ml), while the level in patients with ovarian carcinoma was significantly increased (12.67 +/- 1.43 microgram/ml). Pregnant women have increasing secretory protein levels with increasing length of gestation (5.86 +/- 2.02, 11.55 +/- 1.30 and 17.00 +/- 1.16 microgram/ml for the first, second and third trimesters, respectively.


Purification of human thyroglobulin for radioimmunoassay and testing by ultracentrifugal analysis and immunoelectrophoresis. The development of sensitive radioimmunoassays for the measurement of thyroglobulin (Tg) in human serum has demanded a high degree of purity of the Tg preparation. A procedure for purification of Tg including immunological methods for both purification and control of purity was therefore used. Extract of human thyroid glandular tissue from a patient with Grave's disease was chromatographed on Sepharose CL6B and subsequently on an affinity column containing antibody to whole human serum. Control of both purification steps was by fused rocket electrophoresis of fractions and crossed immunoelectrophoresis of the concentrated solutions. It could thus be shown that traces of contaminating serum proteins, present after column chromatography, were removed by affinity chromatography. Ultracentrifugation of 125I-labelled Tg indicated that it had a sedimentation rate corresponding to 19 S.


Change of electrophoretic mobility of purified alternative pathway factor D. The electrophoretic mobility of factor D (D) of the alternative pathway of human complement activation was examined by the method of lysoelectrophoresis. Purified D was found to have beta-mobility, while D in fresh serum showed alpha-mobility. Addition of D-depleted serum to D induced a change of electrophoretic mobility from beta to alpha. Addition of guinea pig serum to D did not produce this change. The change of electrophoretic mobility of D was not due to complex formation between D and other known alternative factors, P, C3NeF, B and C3. The Factor(s), which mediated the change in the electrophoretic mobility of D, had pseudoglobulin properties and was distributed around the third peak on Sephadex G-200 gel filtration of human serum with a mol. wt. of 50,000-80,000. This phenomenon might be restricted to a semi-solid state reaction, because complexes containing D were not observed upon analyses by sucrose density gradient ultracentrifugation and isoelectrofocusing.


Isolation and characterization of highly stable rabbit C1q. Rabbit C1q has been isolated by techniques which retain elements of previous isolation methods along with novel procedures incorporated to yield stable, non-aggregating, highly purified and biologically active C1q. The procedure involves euglobulin preparation by dialysis of serum and two column chromatographies, DEAE-Sephadex and Ultragel ACA 34. The method results in yields of C1q from rabbit sera of 20-30%. Ouchterlony analysis, sedimentation velocity experiments, equilibrium ultracentrifuge molecular weight analysis, SDS, polyacrylamide gel electrophoresis and isoelectric focusing experiments indicate the presence of only a single component, which has all the physical characteristics previously reported for C1q. The preparation has all the biological characteristics of C1q such as the ability to agglutinate IgG-coated latex particles and to restore hemolytic activity to RC1q serum. Preparations obtained by this procedure are stable to freezing (-70 degrees C) for extended periods of time and show no evidence of aggregation upon thawing following periods of storage of up to 6 months. Mild acetylation and handling of preparations for extended periods at 37 degrees C and 45 degrees C indicate no evidence of aggregation or loss of agglutinating and hemolytic activities. Preparations so isolated meet all the criteria required for critical physical-chemical studies of this protein.


Evaluation of various filter membranes for Boyden-type leukocyte migration studies. Various types and makes of membranes were assessed for suitability in Boyden-type chemotaxis studies. Pooled exudate neutrophils and macrophages from the rat were used as indicator cells. Sartorius filters were found to be superior to other cellulose ester filters both in efficiency and consistency of cell migration. Variations between and within Sartorius filter lots were small and filter inconsistency could be ruled out as a factor in day-to day variation in efficiency of cell migration. In our hands, Millipore membranes were unsatisfactory for chemotaxis studies. With the exception of the shortened incubation times, polycarbonate filters did not offer significant advantages over Sartorius filters when the vigorously responding rat exudate cells were tested. The orientation of the cellulose ester filters in the chemotaxis chambers was found to be a critical factor when the lowermost level counting method was used. The observed polarity could not readily be washed out by acid, alkali or ethanol treatment nor by boiling the filters in distilled water and remains unexplained.


Microsurgical cannulation of the thoracic duct in cats. Technical note. The authors describe a new technique for the microsurgical cannulation of the thoracic duct. A small silastic tube is utilized which can be joined to a subcutaneously implanted one-way flow, CSF, Ommaya-type reservoir. Extended drainage is avoided. No deletion or alteration of major venous channels occurs. Lymph collection is accomplished by tapping the subcutaneous reservoir with a small-bore needle.


In vitro assessment of delayed hypersensitivity in the human. Inhibition of cell migration from agarose microdroplets. The in vitro migration inhibition responses of peripheral blood leukocytes from tuberculin skin test positive and negative donors were tested to validate and determine optimal conditions for the agarose droplet method in the human. In vitro migration inhibition was observed in skin test positive donors in the presence of 25 microgram PPD/ml of medium using both unfractionated leukocytes and a mixture of immune lymphocytes with allogeneic polymorphonuclear leukocyte indicators. The supernatants of tuberculin positive lymphocytes cultured with PPD also inhibited the migration of human polymorphonuclear leukocytes and guinea pig peritoneal exudate cells but did not alter that of murine peritoneal exudate cells. These studies establish that the agarose droplet method is an efficient approach to the in vitro assessment of cell-mediated immunity in the human and define suitable indicator cell populations for assay of human lymphokines.


Monolayer immune complex (MIC) fractionation of Fc receptor bearing human spontaneous killer cells. A simple high recovery method is described for depletion and enrichment of Fc receptor bearing human spontaneous killer cells. The technique is called monolayer immune complex (MIC) fractionation and is based on the fact that Fc receptor positive cells will bind to immune complexes bound to plastic surface and that bound cells can be partially recovered by protein A elution.


Reversed immunosorbents: a simple method for specific antibody immobilization. A method is presented for permanently converting an antigen immunosorbent into a purified antibody immunosorbent which retains specific reactivity for antigen. Following immunological reaction of the antibodies in an antiserum with the antigen adsorbent, the specific immunoglobulin is chemically bound to the antigen by means of the divalent crosslinking agent. Various such agents, including glutaraldehyde, suberimidate, a diazide, and isocyanates, were employed. The conditions of preparation using the first two were optimized, and the characteristics of the resulting immobilized antibodies were investigated. Applications in radioimmunoassay, preparative chromatography, and affinity-constant studies discussed in view of the unique attributes of these reagents.


Radioimmunoassay for hepatitis B surface antigen and antibody using a two-component immunoadsorbent. A solid-phase radioimmunoassay is described for detection of hepatitis B surface antigen and antibody. The assay uses a two-site (antigen and antibody) immunoadsorbent capable of binding antigen and antibody. Antigen is detected by enhanced binding of [125U]antibody through sandwich formations of antibody--antigens--[125I]antibody on the immunoadsorbent. Antibody presence is revealed by inhibition of [125I]antibody binding.


The preparation of latex particles with covalently bound polyamines, IgG and measles agglutinins and their use in visual agglutination tests. Carboxylated latex particles were substituted with side arms terminating in primary amine and hydrazine groups. The particles were coupled to aldehyde groups generated on glycoproteins which were treated with sodium periodate. Particles having the alipathic primary amine putrescine hapten as the sole substituent and particles linked to glycoproteins such as measles agglutinins and IgG were used to detect the presence of the corresponding antibodies or antigens in biological fluids by agglutination tests.


The application of complement fixation to antibody weight estimates I. To soluble antigens. A method is described for estimating early mouse IgG antibodies to the dinitrophenyl (DNP) determinant in absolute weight units. The antibodies to be measured are adsorbed quantitatively onto an appropriate immunoadsorbent such as DNP polyacrylamide. The particulate immune complexes are then incubated with a standard dilution of a potent rabbit antiserum to mouse IgG. Removal of the rabbit anti-mouse IgG (RaM) immunoglobulins by the complexes decreases the C fixing ability of the RaM supernatants reacted with C and mouse IgG. The decrements in C fixation are linearly related to the logarithm of the weight of mouse IgG. The decrements in C fixation are linearly related to the logarithm of the weight of mouse IgG  anti DNP added to the immunoadsorbent. A calibration curve is constructed with known amounts of mouse IgG )anti-NDP as measured by quantitative precipitation. Concurrent assays with the early mouse anti-DNP sera permit conversion of the losses in C fixing potencies to weight units of mouse IgG anti-DNP by interpolation on the calibration curve.


The use of [125I]deoxyuridine for semi-microevaluation of human lymphocyte response to phytohemagglutinin in vitro. [125I]5-iodo-2'-deoxyuridine ([125I]UdR) was compared with [6-3H]thymidine ([3H]TdR) in a semi-micromethod used to evaluate the response of human lymphocytes to phytohemagglutinin (PHA) in vitro. 5-Fluoro-2'-dexoyuridine (FUdR) was added to [125I]UdR in order to increase its incorporation, by preventing endogenous thymidine synthesis. The relationship between PHA concentration and the isotope uptake by lymphocytes was studied. Similar dose-response curves were found for the two precursors, with a peak at 15 microgram/ml of PHA. Nevertheless the recovery of the labeled compounds was higher when [125I]UdR together with FUdR was used.


Studies on the microcytotoxicity test. III. Comparison of  [75Se]selenomethionine with [3H]proline, Na2, 51CrO4 and [125I]iododeoxyuridine for pre-labelling target cells in long-term cytotoxicity tests. Four intracellular radioisotope labels, [3H]proline, Na2 51CrO4, [75Se]selenomethionine and [125I]iododeoxyuridine, were evaluated for use in a pre-labelling long-term microcytotoxicity assay for cell-mediated immunity. Adherent rat tumour cells established in tissue culture were used as targets and the basic variables studied were labelling efficiency, toxicity and spontaneous release rates. [125I]Iododeoxyuridine was found unsuitable on account of its high toxicity and correspondingly high spontaneous release rate, and Na2 51CrO4 for its toxicity and low labelling efficiency. Of the two other radiolabels, [75Se]selenomethionine had the advantage over [3H]proline of higher labelling efficiency (especially in Ham's F10 medium), lower toxicity, and being a gamma-emitter. Furthermore, released 75Se was shown to be non-reutilisable and its retention by target cells provided an accurate measure of cell survival in an alloimmune system. Methods of calculating the results of pre-labelling cytotoxicity tests based on the total radioactivity in target cells at the beginning of the assay were found to be invalid.


I. An ultrastructural investigation on erythrocytes and lymphocytes coated with proteins: use of CrCl3 as an electron-dense marker. Proteins (human immunoglobulin G and protein A of Staphylococcus aureus) coupled to the surface of erythrocytes (sheep, mice, man and chicken) and to mouse spleen lymphocytes by chromic chloride technique acquire electron-dense properties and may be visualized in electron microscopy. Sheep red blood cells coated with rabbit anti-erythrocyte antibodies and treated with chromic chloride showed continuous electron-opaque labelling of the cell membrane due to chelation of rabbit IgG with chromium atoms.


II. Ultrastructural differences in attachment patterns of indicator cells to target cells in the rosette-forming process. A comparison was made of the electron microscopic appearances of rosettes formed between mouse spleen lymphocytes and indicator cells with the ligands exposed on their surfaces either in clusters or in continuity. Sheep red blood cells (SRBC) coated with protein A of Staphylococcus aureus or human IgG by the chromium chloride technique were used as indicator cells with cluster distribution of the ligand. RBC coated with rabbit IgG anti-sheep red blood cells or staphylococci were used as indicator cells with uniform distribution of the ligand. Results show that the appearance of the zones of contact (point attachment or broad zone attachment) between lymphocytes and indicator cells are mainly influenced by the distribution of the specific ligands on the surface of the indicator cells and do not reflect the pattern of distribution of receptors on lymphocytes.


Individual serum bile acid concentrations in normo- and hyperlipoproteinemia as determined by mass fragmentography: relation to bile acid pool size. Combined gas-liquid chromatography-mass spectrometry with specific ion monitoring (mass fragmentography) has been used for assay of cholic acic (C), chenodeoxycholic acid (CD), and deoxycholic acid (D) in human serum. Deuterium-labeled C and D were used as internal standards. The relative standard deviation of duplicate samples was 3, 4, and 7% for C, CD, and D, respectively. The variation within the same individual in the fasting state was small, while the day-to-day variation was greater, especially for the dihydroxy bile acids. In normal control subjects (n = 24), the fasting serum concentration of C averaged 184 +/- 24 ng/ml (mean +/- SEM), and that of CD and D 526 +/- 62 and 407 +/- 44 ng/ml, respectively. Patients with type IIa hyperlipoproteinemia (n = 32) displayed low values of serum bile acids, with a C concentration of 121 +/- 11 ng/ml (P less than 0.01 vs. controls). A similar pattern was seen in patients with a type IIb lipoprotein pattern (n = 10). Subjects with type IV hyperlipoproteinemia (n = 32) showed serum bile acid levels within the normal limits. No relationship to age, sex, or body weight was seen in any of the patient subgroups. Bile acid kinetics were determined with an isotope dilution technique using 14C-labeled C and CD under steady state conditions in control subjects and patients with type IIa and type IV hyperlipoproteinemia. The serum concentration of C correlated significantly to its pool size in control subjects and in patients with type IIa hyperlipoproteinemia but not in patients with type IV. The serum level of CD was not related to CD pool size in any of the subgroups. The data obtained are discussed in relation to present concepts of the enterohepatic circulation. It is suggested that the intestinal content of C in the fasting state is proportional to the total C pool size. The possibility of a defective intestinal uptake of C in some patients with type IV hyperlipoproteinemia is raised.


Isolation and characterization of human liver hematoside. The monosialoganglioside hematoside (GM3) is an important precursor in the synthesis of the more complex gangliosides. To obtain large quantities of GM3 for use in ganglioside biosynthetic experiments, we have devised a column chromatographic procedure for the isolation and purification of GM3 from human liver. A total ganglioside mixture was obtained from a 550 g sample of normal human liver. Quantitative analysis by gas-liquid chromatography indicated about 66 microgram of lipid-bound N-acetylneuraminic acid per gram of fresh tissue. Hematoside appeared as double bands on thin-layer plates and represented 86.4% of the total sialic acid content. Additional ganglioside species, notably GD3, were also seen. Hematoside was separated from the other gangliosides by GM3 obtained was 120 mg, which represented a 90% recovery. Although the long-chain base and sugar compositions of the upper and lower GM3 fractions were similar, striking fatty acid differences were detected. The upper fraction contained predominantly unsubstituted fatty acids, while the lower fraction showed a preponderance of alpha-hydroxy fatty acids. The unsubstituted fatty acid and hydroxy fatty acid composition of the total GM3 fraction was calculated to be 56.9% and 43.1%, respectively.


Age-related changes in cholesterol and bile acid metabolism in rats. Age-related changes in serum and liver cholesterol, phospholipid and triglyceride levels, serum lipoproteins, biliary secretion of cholesterol, phospholipid and bile acids, and fecal excretion of sterols and bile acids were examined in Sprague-Dawley and Wistar strain male rats, 7 to 106 weeks in age. Serum and liver lipid levels increased with age in both strains but the liver phospholipid level remained unchanged. The proportion of serum alpha-lipoprotein increases and that of beta- and prebeta-lipoproteins slightly decreased. Cholesterol and phospholipids in low density lipoprotein and cholesterol in high density lipoprotein fractions also increased with age. Bile flow and biliary secretion of cholesterol and bile acids decreased in aged rats, but when they were expressed in terms of units per rat they were almost constant without regard to age. Pool size, synthesis, secretion, and turnover frequency of bile acids also did not change when they were expressed per rat, though 7-week-old rats showed a low value for turnover frequency. Biliary secretion of phospholipid, however, increased in aged rats. Biliary secretion of chenodeoxycholic and alpha-muricholic acids decreased but that of cholic and hyodeoxycholic acids increased. Daily excretion of feces and fecal neutral sterols decreases in aged rats but the excretion of bile acids remained constant regardless of age. The ratio of coprostanol and cholesterol in the total sterols was not affected. Fecal lithocholic, beta-muricholic and P10 (probably omega-muricholic) acids were decreased with age but the other bile acid components were not changed or were slightly increased.


On the quantitative importance of omega-oxidation of fatty acids. The relative importance of omega-oxidation of fatty acids in livers of starved rats was studied by determination of the ratio between 14C at C1 and at C2, C3, C4 in acetoacetate formed from [16-14C]palmitic acid in vivo and in vitro and from [10(-14)C]decanoic acid in vitro. The results are consistent with the possibility that at least a small fraction of the fatty acids may be subject to primary omega-oxidation prior to beta-oxidation in the ketotic state. The maximal figure for this pathway was calculated to be of the magnitude 5--10%. The metabolic consequences of a pathway involving primary omega-oxidation of fatty acids are discussed.


Purification of calciferol-binding proteins from kidney: physicochemical and immunological properties. The calciferol-binding system of rat kidney cytosol has been purified and is shown to consist of two proteins, each capable of binding either 25-hydroxy-vitamin D3 (25-OH-D3) or 1,25-dihydroxyvitamin D3  (1,25-(OH)2D3). The two proteins, designated A and B, have similar sedimentation coefficients (S20w) of 5.2 S. Component A binds 25-OH-D3 with a dissociation constant (Kd) of 10(-7) M while component B binds 1,25-(OH)2D3 with a Kd of 1.6 x 10(-8) M. The estimated molecular weights (Mr) of the two proteins are 105,000 for component A and 250,000 for component B. Amino acid analyses revealed that glutamic acid is the most abundant residue in both proteins, comprising 12% of the total number of amino acid residues. Immunodiffusion test using commercial anti-human serum group-specific protein antiserum gave a precipitin reaction when purified rat serum calciferol-binding protein was used as an antigen, but no reactions could be detected  with proteins A and B. This result significantly eliminated the possibility of the presence of the rat serum binding protein in either of the purified kidney proteins. In contrast, anti-rat serum calciferol-binding protein antiserum prepared in rabbits interacted with the rat serum and kidney proteins. This result suggests that the antigenic determinants recognized by the antiserum against the rat serum calciferol-binding protein appear to be similar to those recognized in the kidney proteins A and B. Immunoelectrophoresis of the three rat proteins demonstrated dissimilar electrophoretic mobilities with the serum protein showing the least mobility, a property consistent with its higher lysine content relative to proteins A and B.


Identification and characterization of mannosyl retinyl phosphate occurring in rat liver and intestine invivo. A study was conducted to determine whether mannosyl retinyl phosphate occurred in rat liver and intestine in vivo, and, if so, to partially purify it and investigate its properties. After injection of [(3)H]retinol and [(14)C]mannose, a chloroform-methanol 2:1 extract of rat liver and small intestinal mucosa yielded two (3)H/(14)C-labeled peaks on DEAE-cellulose column chromatography: peak I eluted with 10 mM and peak II eluted with 29 mM ammonium acetate. Peak II, subjected to silicic acid column chromatography, gave principally two (3)H/(14)C-labeled fractions, one eluted with chloroform-methanol 2:1 and the other with chloroform-methanol 1:1. The latter showed, on thin-layer chromatography in a chloroform-methanol-water 60:25:4 system, an R(f) of 0.25 (with coincidence of the (3)H and (14)C radioactivity), which is identical to the R(f) of authentic mannosyl retinyl phosphate. The chloroform-methanol 1:1 peak, on mild acid hydrolysis, yielded [(3)H]retinol (identified by two thin-layer chromatography systems), [(14)C]mannose, and [(14)C]-mannose phosphate (identified by paper chromatography). On mild alkali hydrolysis, the peak yielded [(3)H]retinol and [(14)C]mannose phosphate. The substance eluted in the chloroform-methanol 1:1 peak from silicic acid was therefore concluded to be mannosyl retinyl phosphate. When chromatographed on silicic acid, peak I from the DEAE-cellulose column primarily showed a fraction eluted with chloroform-methanol 2:1. When chromatographed on thin-layer plates in the above solvent, this fraction showed an R(f) of 0.3, with coincidence of (3)H and (14)C radioactivity; it was resistant to mild acid hydrolysis, mild and strong alkali hydrolysis, and glucuronidase action. Mannosyl retinyl phosphate occurs, therefore, in vivo in liver and intestinal mucosa, and it is accompanied by a closely similar, though slightly less polar, compound that remains unidentified.


Rates of removal and degradation of chylomicron remnants by isolated perfused rat liver. Chylomicron remnants are removed intact by isolated perfused rat livers and their lipid components are metabolized by the liver (Biochim. Biophys. Acta 488: 464, 1977). The present study provides quantitative information regarding these processes. When the lipoprotein concentration of the perfusate was constant, the removal of chylomicron remnants increases lineraly for 17 min. The rate of remnant removal was a hyperbolic function of the perfusate's remnant concentration. The removal rate had aV max of 28microgram cholesterol per g liver per min and an apparent Km of 64 microgram cholesterol per ml perfusate. Feeding the liver donors a diet containing 1% cholesterol or 4% cholesterol and 1% cholic acid failed to alter the hepatic removal rate. The cholesteryl ester removed from the remnants was hydrolyzed at a rate that was a small fraction of the removal rate (about 0.5% of removed cholesteryl ester per min). The rate of cholesteryl ester hydrolysis did not appear to approach saturation in the range studied. Studies of the lysosomal cholesteryl ester hydrolase suggested that this enzyme was not responsible for limiting the initial rate of hydrolysis, raising the possibility that the degradation rate is determined by the movement of the removed remnant to the site of hydrolysis.


Monoacylmonoalkylglycerol as a substrate for diacylglycerol hydrolase activity in adipose tissue. The synthesis and use of 1(3)-[3H]oleoyl-2-0-oleylglycerol as a substrate for the assay of diacylglycerol hydrolase activity in adipose tissue is described. Neither the compound nor its reaction product are hydrolyzed by purified adipose tissue monoacylglycerol lipase.


d-amphetamine and fixed-interval performance: effects of operant history. Sixteen rats were initially exposed for 50 sessions to either a fixed-ratio 40 or an interresponse-time-greater-than-11-second food reinforcement schedule, then shifted to a fixed-interval 15-second food reinforcement schedule. Animals with fixed-ratio 40 histories lever pressed at much higher rates under the fixed-interval schedule than did animals with inter-response-time-greater-than-11-second histories. This difference persisted across 93 sessions of fixed-interval exposure. The effects of d=amphetamine were assessed after 15 and 59 sessions of fixed-interval exposure. On both occasions, the low-rate responding of animals with interresponse-time-greater-than-11-second histories was typically increased by all doses of the drug, while the high-rate responding of animals with fixed-ratio 40 histories was typically decreased by all doses of the drug. These results suggest that control response rate under the fixed-interval schedule, which may be affected by a history of responding under another schedule, is the primary determinant of the relative effects of d-amphetamine.


Attentional changes during discrimination learning by retarded children. Eight moderately retarded children were trained on a simultaneous two-choice discrimination problem and a series of discrimination-shift problems. The procedure required the subjects to perform overt observing responses to produce elements of the discriminative stimuli, making it possible to measure directly changes in attention to different aspects of stimuli during learning. The patterns of change in observing responses were generally in line with descriptions of attentional changes derived from two-process theories of discrimination learning; for example, the frequency of irrelevant observing responses was high during the presolution period during extradimensional shifts but was low during intradimensional shifts. Contrary to current theories, extradimensional shifts caused an immediate increase in irrelevant observing responses, and intradimensional shifts usually caused an increase in relevant observing responses. Subjects responded to later shift problems by initially increasing both relevant and irrelevant observing responses, then withholding irrelevant observing responses.


Ultrastructural observations on the chorionic epithelium, parathyroid glands and bones from chick embryos developed in shell-less culture. Three-day-old chick embryos were cultured in Petri dishes until they reached developmental stages 37 or 39 (Hamburger & Hamilton, 1951). The electron microscopical study of the chorio-allantoic membranes showed that, even in the absence of the shell and the corresponding calcium supply, 'capillary-covering' and 'villus-cavity' cells differentiated well. The parathyroid glands from the cultured embryos showed ultrastructural signs indicative of active synthesis and secretion of the parathyroid hormone. This correlates well with the significant hypocalcemia (5.4 mg/100 ml) observed in these embryos. In all cultured embryos mineralization of bones was greatly reduced as shown by alizarin bulk staining and confirmed by histological and electron microscopical analysis. The ultrastructural characteristics of osteoblasts and osteocytes, as well as of the bone matrix, appeared normal. The defect in mineralization appeared thus to be due to a deficiency in the availability of calcium and not to a delay in bone differentiation. This implies that the yolk sac appears to lack calcium regulatory capacity since it cannot compensate for the absence of shell by increasing its own contribution in order to assure an adequate mineralization of the embryonic bones.


Ultrastructural analysis of preimplantation lethal yellow (Ay/Ay) mouse embryos. Embryos recovered at 62 and 80 h post coitum from reproductive tracts of yellow (Ay/a) and black (a/a) female mice mated to Ay/a males were examined ultrastructurally to define the developmental defect of lethal Ay homozygotes. No abnormalities were observed in five embryos from control matings (female a/a x male Ay/a). Of 24 morulae and blastocysts from Ay/a x Ay/a matings, six were observed to possess some morphological aberration. Two of the six abnormal embryos were morulae and contained isolated blastomeres which had developmental features typical of younger embryos; remaining cells of these embryos were normal. The third, an early blastocyst, contained a degenerating trophoblast cell' other cells of this embryo were also abnormal but not in an advanced stage of degeneration. The fourth abnormal embryo (late cleavage stage) was in an advanced stage of degeneration affecting all blastomeres. Finally, the remaining two abnormal morulae had a unique nucleolar morphology and an unusual abundance of intra-cisternal A particles. Presumably, one or more of the six abnormal embryos from Ay/a x Ay/a matings were Ay homozygotes. However, no single ultrastructural alteration characteristic of Ay/Ay embryos was found.


The paths and destinations of the induced ipsilateral retinal projection in goldfish. Adult goldfish had one tectal lobe removed surgically, and several months later, the eye contralateral to the missing tectum was injected with radioactive proline. Radioautographs of the brains were studied to trace the paths and termination sites of the optic fibers. The optic tract decussated at the chiasm, as normally, but then ran caudally in a large neuroma on the tectum-less side of the brain. Substantial numbers of fibers left this neuroma to enter two or more of five commissures, through which they recrossed the midline. These commissures: transverse, minor, horizontal, posterior and ansate, ordinarily contain few or no optic fibers. All are normally linked with the tectum. Negligible numbers of aberrant optic fibers recrossed the midline elsewhere. On the intact side of the brain, ipsilateral to the injected eye, the optic fibers innervated some or all of the nuclei and areas normally served by contralateral retinal fibers. An earlier behavioral study of these same fish had shown that some of them made reversed optokinetic nystagmus in response to stripe movement seen by the eye projecting ipsilaterally; others failed to respond to stimuli through this eye. In all the reversed responders, a caudal group of retinal projection sites was labeled ipsilaterally. This included the basal optic nucleus and the caudal portions of nucleus dorsolateralis thalami and area pretectalis. In the non-responders, these targets were not labeled ipsilaterally. Together, these results suggest that one or more of these three sites is or are responsible for optokinetic nystagmus in normal goldfish.


Control of somite number in normal and amputated mutant mouse embryos: an experimental and a theoretical analysis. A regulation is shown for size and number of serially repeated axial structures, the somites, in a mammalian embryo. The mammalian embryo is normally inaccessible to operation at post-implantation stages. This problem is resolved by the quantitative analysis of somite size, number and development in a recessive mutant of the mouse, amputated, whose axial length is greatly reduced. The effect of the gene simulates an experiment ablating part of the embryonic tissue available for somitic segmentation. Regulation occurs at the time when the somite is first formed, by control of the quantity of cells included in each new somite. A model is devised for the control of somitic segmentation which explains most of the features observed and which can be simulated on a computer.


Catch-up growth in the rat skull after retardation during the suckling period. In rats whose growth was restricted by rearing in large litters the skull was less affected than the spine and the brain case less than the face. No sex difference was apparent during undernutrition but on subsequent rehabilitation females had caught up to controls in four out of five skull measurements within 4 weeks and by the end of the experiment the skulls of the neonatally undernourished females were the same size and shape as the controls. In the case of males, however, the skull of the undernourished rats never attained the same width as the controls.


Anatomical mapping of retino-tectal connections in developing and metamorphosed Xenopus: evidence for changing connections. Neural connections between the eye and optic tectum in Xenopus laevis were anatomically traced by observing the tectal location of Wallerian degeneration after discrete retinal lesion. These retinotectal connections were mapped in postmetamorphic frogs and tadpoles at stage 51, the stage at which retinal axons have grown into about the rostral one-half of the tectum. The course of the experimental degeneration was the same in frogs and tadpoles, but degeneration proceeded faster in the younger animals. In the frogs, connections were ordered, with nasal retina mapping to the caudal part of the tectum and temporal retina mapping to the rostral tectum. In the tadpoles, within the innervated area at the rostral tectum, the retino-tectal connections were generally organized as in the adults, with the temporal retina mapping to the rostral part of the innervated tectum and nasal retina mapping primarily to the caudal part. But a portion of the nasal fibers consistently mapped to the far rostral tectum as well. Electron microscopic observations showed degenerating synaptic terminals at both rostral and caudal portions of the innervated tectum after lesion of just the nasal retina. Degeneration was not seen in control animals. These results indicate that some fibers (particularly from nasal retina) may shift their terminals caudally on the tectum to match tectal growth and produce the adult pattern of connections. If there is such connection readjustment, the 'aberrant' connections from nasal retina in tadpoles may be an indication of this process.


Serum-dependent avian skin differentiation in vitro: time sequence of induced events. Information concerning the developmental mechanisms involved in skin differentiation have been mainly derived from in vitro experiments. We have previously observed that 6-day chick embryonic thigh skin keratinizes in vitro in chicken serum-containing medium, but does not do so in chick embryo extract-containing medium. This system seems adequate in many respects for investigating the mutual regulative relationship between epithelium and mesenchyme (so called epithelio-mesenchymal interactions). We have therefore attempted better to define the time course of differentiative events and their dependence upon the time of serum administration. Skin explants have been sequentially supplemented with differentiation-stimulating or non-stimulating nutrient, removed at different intervals and examined by histological and histochemical procedures. Different epithelial and mesenchymal behaviour results according to serum supplementation time. Serum administered in the first 48 h of in vitro maintenance is unable to stimulate either the subsequent epidermal keratinization or the correlated changes in dermal histochemical pattern. On the other hand, serum-containing medium induces keratinization and changes in dermal intercellular composition (glycoproteins accumulate to a greater extent than glycosoaminoglycans, as does hyaluronic acid relative to chondroitin sulphuric acids), if added for only the second two days. In the last 48 h, serum may promote epidermal keratinization provided that in vitro incubation has been prolonged. Administration of actinomycin D with serum in the second 48 h prevents epidermal keratinization and modifies dermal histochemical reactivity, but is ineffective if added in the last 48 h. The above findings demonstrate that serum factor(s) stimulating epidermal differentiation act(s) on skin explants in the central incubation period and that correlatively mesenchyme acquires a characteristic histochemical pattern, supporting the possibility that the effect of serum may be mediated by changes in the composition of dermal ground substance.


Somite abnormalities caused by short heat shocks to pre-neurula stages of Xenopus laevis. This paper describes the small disturbances, in the regular pattern of the somites and the fissures between them, that are seen following short (around 300 s) heat shocks at 37.5 degrees C delivered to pre-neurula stages of Xenopus laevis. Affected groups of cells still finally differentiate as somite muscle, but the normally precise spatio-temporal sequence in which they move beforehand to give rise to the actual pattern of somite blocks, is disrupted. Examination of the position and sizes of patches of disrupted morphogenesis, in relation to the precise embryonic stage at shock, leads to certain conclusions about the nature of the disturbance induced by a brief period at high temperature, in cells due to form somites. The pattern of results is compared with that produced by similar temperature shocks given to tail-bud (later) staged embryos. The discussion includes a brief consideration of how the various results of heat shocks, given at different embryonic stages, might be understood in terms of one particular model (Cooke & Zeeman, 1976) for the spatio-temporal control of the developing somite pattern.


Changes in responsiveness of preimplantation mouse embryos to serum. Changes in uptake of radioactive uridine and its incorporation into RNA were determined in preimplantation mouse embryos, from the 2-cell to the blastocyst stage, as a measure of their responsiveness to extracellular conditions. Two media were tested, one contained serum and the other contained bovine serum albumen as a control. An increase in the acid-soluble pool occurred at the 8-cell stage and a marked increase in RNA synthesis occurred at the early blastocyst stage when the embryos were incubated with serum.


Postembryonic development of the visual system of the locust, Schistocerca gregaria. I. Patterns of growth and developmental interactions in the retina and optic lobe. The visual system of the locust, Schistocerca gregaria, has a highly ordered and predictable arrangement of neurons. The retina and the outermost layer, or lamina, of the optic lobe are each composed of repeating units, ommatidia and cartridges respectively. Each ommatidium has eight photoreceptor cells, which send axons directly to a group of five neurons in the lamina to form the cartridge. The importance, for the development of this precise pattern, of the mode of growth of the two arrays and of interactions between them was investigated. The spatial and temporal sequences of cell proliferation, differentiation and death in the developing retina and optic lobe were examined quantitatively under normal and experimental conditions. The retina grows from its anterior margin by addition of new ommatidia formed from recruited epidermal cells. The lamina also grows by addition of new neurons to its anterior margin, but these neurons are derived from a stem cell population. The parallel pattern of growth of the retina and lamina may be important for the formation of neuronal connexions between them. The retina grows and differentiates even when deprived of the underlying lamina. In laminae deprived of the ingrowth of new axons from the retina, the production of new neurons is also autonomous, but these neurons do not differentiate, but degenerate. A limited amount of cell death occurs in the laminae of control insects. These two observations suggest that a plausible mechanism for coordinating the sizes of the two arrays during normal development might be production of lamina neurons in excess of requirements and death of those remaining non-innervated.


Chromosome analysis of early postimplantation presumptive haploid parthenogenetic mouse embryos. The chromosome constitution of early postimplantation presumptive haploid parthenogenetic mouse embryos was examined. All the embryos isolated were at the egg-cylinder stage and seven contained dividing cells. In two of the apparently healthy embryos only haploid mitoses were seen, whereas in five others an approximately equal proportion of haploid and diploid mitoses was observed. Out of 52 cells in which unequivocal counts could be made, only one contained more than the euploid number of chromosomes (mouse, n = 20). Possible reasons for the poorer development of haploid compared to diploid parthenogenetic embryos are discussed.


Respiratory and hydrostatic functions of the intestine of the catfishes Hoplosternum thoracatum and Brochis splendens (Callichthyidae). 1. The air-breathing behaviour of Hoplosternum thoracatum and Brochis splendens has been studied and their strategy of coordinating the respiratory and hydrostatic functions of the accessory respiratory organ has been examined. 2. H. thoracatum and B. splendens are continuous but not obligate air-breathers and individuals of the former breathe air in synchrony with each other. 3. Frequency of air-breathing increased with increased activity in H. thoracatum. 4. Aquatic respiration (Vo2) in H. thoracatum decreased in hypoxic water but aerial Vo2 maintained a fairly constant total Vo2 independent of aquatic O2. Total Vo2 is higher when fish breathe both air and water but aerial Vo2 did not exceed 75% of total Vo2. 5. The accessory respiratory organ provides about 75% of the lift required to attain neutral buoyancy whereas the swimbladder provides less than 5%. The mean decreases in volume of the accessory respiratory organ in the period between breaths of B. splendens and H. thoracatum were 13.2 and 7.8% respectively. 6. With a gas phase of O2, B. splendens maintained a similar frequency of air breathing and showed a slightly greater reduction in buoyancy between air breaths when compared to breathing air. With a gas phase of N2, air breathing was less frequent and decreases in buoyancy between air breaths were much less than when breathing air. 7. The respiratory and hydrostatic functions of the accessory respiratory organ are compatible. Buoyancy is maintained by frequent air breaths taken in part in response to a decrease in volume of the accessory respiratory organ. This reservoir of O2 could increase metabolic scope during bursts of activity.


The effect of training on the swimming muscles of the goldfish (Carassius auratus). Goldfish (Carassius auratus) were exercised continuously for periods of 28 days at swimming speeds of 1.5, 3.0 and 4.5 body lengths per second and their rates of growth were determined. Changes in muscle fibre size were examined, as were changes which occurred in the concentrations of the major chemical constituents of these cells. These fish, typical of the carp family in that they are found only in still or slowly moving water, did not adapt well to the flowing water environment at any swimming speed. They often grew less than the controls, although consuming much more food. Changes in the composition of the muscle fibres indicated that excess food was not being stored, and also indicated that the major fuel for swimming at all speeds was glycogen. The fish survived well at high speeds and it was suggested that this was due to the ability of the species to metabolize glycogen anaerobically without the production of lactic acid.


Influence of water temperature on the electric organ discharge (EOD) of the weakly electric fish Marcusenius cyprinoides (Mormyridae). 1. The influence of different water temperatures on the electric organ discharge (EOD) of a mormyrid fish Marcusenius cyprinoïdes was studied. The range of the water temperatures was fixed according to the seasonal temperature variations of the rivers in Central Africa, the natural habitat of this species. 2. The EOD activity was characterized using the following parameters: mean EOD rate, EOD pattern in the form of Interpulse Interval Histograms (IIH), IIH range, and shortest pulse interval. These parameters remained constant during control experiments at constant temperature (27 degrees C) for 4 days. 3. The mean EOD rate increases with increasing water temperatures. The lowest mean EOD rate is always found at 17 degrees C, the highest between 26 and 33 degrees C. The characteristics of the IIH are modified by stepwise temperature increases. These IIH show during high temperatures (26-33 degrees C) similar patterns to those previously observed during high level motor activity and excitement. The IIH range diminishes with stepwise temperature increases. The shortest pulse interval has a negative, linear correlation with water temperature. 4. The possible role of water temperature in the reproduction of the mormyrids is discussed. The high discharge rate of M. cyprinoïdes produced by high water temperatures during the rainy season could serve to improve the resolution of the electroreceptors during this period.


The effect of hypertonic solutions on the rate of relaxation of contracture tension in Mytilus smooth muscle. 1. The effect of the bathing solution tonicity on the mechanical properties of the anterior byssal retractor muscle (ABRM) of Mytilus edulis was examined. 2. The rate of relaxation of contracture tension produced by acetylcholine (ACh) was greatly reduced when the bathing solution tonicity was increased by adding NaCl, KCl or LiCl after the removal of ACh, whereas a decreased tonicity increased the rate of relaxation. 3. The contracted ABRM in hypertonic solutions showed no active shortening after an isotonic release and barely redeveloped active tension after a quick release. 4. The resistance to stretch increased with increasing tonicity of the bathing solution. 5. The wet weight of the ABRM decreased much more markedly in sucrose-hypertonic solution than in Na-, K- or Li-hypertonic solutions, but the decrease in the rate of relaxation was less marked in the former, indicating that there may be little relation between the rate of relaxation and the degree of osmotic deformation of the ABRM fibres. 6. It is suggested that the elevated ionic strength in the myoplasm may be related to a reduction in the rate of detachment of the cross-linkages between the thick and thin filaments.


Kinetics of sodium and lithium movements across the blood-brain barrier of an insect. The electrical responses of axons were used to monitor the time-course of a change in the concentration of an ion species in the fluid bathing the axons in connectives of isolated cockroach nerve cords. Initial exposure of the connectives to sodium-deficient Ringer resulted in a depletion of extra-axonal sodium which was much slower than the restoration observed on return of the sodium Ringer. It is suggested that this asymmetry could result from a sodium reservoir which delays the initial decline. Subsequent net inward and outward movements of sodium ions were rapid and symmetrical. Unlike sodium ions, lithium ions were apparently unable to reach the axon surfaces following sodium depletion. In view of the similar properties of sodium and lithium ions in many biological systems it is therefore unlikely that the sodium movements were passive. Instead, the results support the idea of net sodium transport by the perineurial and/or glial elements.


Rebound resilience of tendons in the feet of sheep (Ovis aries). The interosseous muscle of the fore foot of the sheep is multipennate, with large tendons but extremely short muscle fibres. It is believed to save energy in locomotion by elastic storage in its tendons and its sheath. Its rebound resilience (at 5-7 Hz and 20-37 degrees C) is about 0.62.


Auditory masking patterns in the goldfish (Carassius auratus): psychophysical tuning curves. The masking effects of tones on the detection auditory signals were studied in goldfish using the psychophysical tuning-curve paradigm. For signals below 350 Hz, masking is an inverse function of the frequency separation between masker and signal; a finding consistent with previous masking studies on fishes, birds and mammals. For signals above 350 Hz, masking peaks occur both in the 350 Hz region and at the frequency of the signal. Quantitative comparisons with recent neural tuning curves for goldfish saccular neurones suggest that the filtering observed may be determined by mechanical frequency selectivity below 350 Hz, but by a neural analysis of temporal patterns above this range.


The origin, kinetics, and characteristics of the Kupffer cells in the normal steady state. Enzymatic digestion with pronase and DNAase was used to isolate Kupffer cells from mouse liver. The characteristics of these cells were found to be similar to those of peritoneal macrophages, except that in the initial suspension the percentage of Kupffer cells with Fc receptors was low, C receptors were absent and the ingestion of opsenized bacteria was very poor, because of the effect of pronase on the cell membrane. After 24 h incubation in vitro all these characteristics return. The in vitro and 1 h-pulse [(3)H]thymidine labeling of the Kupffer cells is low (0.8 and 1 percent, respectively) indicating that in essence these cells do not divide. It was also shown that the small percentage of in vitro labeled Kupffer cells was recently derived from the circulation. After an intravenous injection of zymosan the in vitro labeling index of the Kupffer cells increased 16-fold, but it was proven that these dividing cells were immature mononuclear phagocytes very recently recruited from the bone marrow. The labeling of Kupffer cells aider one or four injections of [(3)H]thymidine reached a peak of 10.4 percent at 48 h or 24.1 percent at 60 h, respectively, indicating that these cells are derived from labeled monocytes. Further evidence for this conclusion was obtained by the absence of an increase of labeled Kupffer cells during treatment with hydrocortisone, which causes a monocytopenia during which no circulating monocytes are available to migrate to the tissues. Labeling studies in animals X-irradiated with hind-limb shielding gave a Kupffer cell labeling index of 5-10 percent of the normal values, which confirms their bone marrow origin. A quantitative study on the production of labeled monocytes in the bone marrow and their transit through the circulation showed that in the normal steady state at least 56.4 percent of the monocytes leaving the circulation become Kupffer cells. Considering the Kupffer cells as kinetically homogeneous this gives a mean turnover time of the total population of Kupffer cells of 21 days.


The role of antigen form and function in the primary and secondary intestinal immune responses to cholera toxin and toxoid in rats. This report describes studies of the mucosal antitoxic response in rats after enteric administration of several forms of cholera toxin or toxoid, proteins which differ primarily in their ability to bind to cell membranes and activate cellular adenyl cyclase. These two characteristics appeared to markedly enhance the local primary response to these antigens. A single dose of toxoid lacking these features was ineffective in local priming even though it was absorbed and induced a systemic immune response. Single dose mucosal priming occurred only with preparations which bind to cell membranes and was enhanced by those which also activate cellular adenyl cyclase. In contrast, single-dose mucosal boosting was best accomplished by materials with these properties but was also seen with a toxoid lacking both of these functions. The property of membrane binding appears to be most advantageous in mucosal priming, perhaps by increasing effective trapping of absorbed antigen in unprimed mucosal lymphoid tissue, whereas the ability to activate adenyl cyclase appears to enhance primary and secondary type responses about equally. Combinations of crude toxoid and toxin were also more effective in mucosal priming than purified materials, a finding which is unexplained. A single dose of this combination induced mucosal priming which was fully developed in 2 wk, undiminished after 4 too, and only modestly diminished after 8 mo, thus demonstrating relatively prolonged memory in the IgA mucosal immune system. Effective two-dose local immunizing regimens were developed, and it was shown that there was no correlation between the mucosal and systemic secondary antitoxin responses provoked by these regimens.


Human C4-binding protein. I. Isolation and characterization. C4-binding protein (C4-bp), a new component of the complement system, was isolated from human plasma by precipitation with polyethyleneglycol, followed by chromatography on ion exchangers. C4-bp was identified on sodium dodecyl- sulfate polyacrylamide gel electrophoresis (SDS-PAGE) by two independent criteria: its ability to bind to C4b, and immunoprecipitation with a monospecificantiserum. Purified C4-bp is a 10.7 s glycoprotein. It consists of several disulfide bonded subunits of mol wt 70,000 daltons. Under nonreducing conditions, its mol wt has been estimated on SDS-PAGE as 540- 590,000 daltons. C4-bp moves as a slow B-globulin at pH 8.6 in the absence of free divalent cations, but when the buffers contain Ca(++)-lactate, C4-bp is a gamma globulin. Purified C4-bp binds to purified C4b. The reaction proceeds in the presence or absence of divalent cations and is not inhibited by diisopropylfluorophosphate. The C4b/C4-bp complexes have sedimentation coefficients between 15 and 17 s on sucrose gradient ultracentrifugation, and can be readily identified by crossed immunoelectrophoresis (CIE). The complexes move faster toward the anode than either protein. C4-bp is multivalent. Saturation is reached at molecular ratios of C4b/C4- bp of between 4 and 5. The interaction between C4b and C4-bp may complicate the electrophoretic patterns of these proteins in normal human serum, if the complement system is activated before or during the run. However, in EDTA-plasma, native C4 and C4-bp do not form stable complexes and can be identified in separate peaks after CIE.


Habituation of the forelimb-withdrawal response in neonatal rats. The purpose of the present research was to examine the ontogeny of habituation in the neonatal rat, using the forelimb-withdrawal response. Thresholds and latencies of the response, changing patterns of responding to shock stimuli, and habituation of the response were studied in rats 3 to 15 days of age. It was found that although response thresholds do not change during this period of development, response latencies decrease and amplitudes increase. Three- and 6-day old pups remain active much longer following shock than do 10- and 15day-olds. When compared for habituation to different frequencies of stimulation, 3-day-old rat pups were found to be much more susceptible to habituation at low frequencies than 15-day-olds. Insertion of a simple intense shock in the habituation series produced marked dishabituation in the 3-day-olds but little or none in the 15-day-olds. This pattern of results was obtained regardless of the locus of the dishabituating shock. Analysis of response latencies showed that although 15-day-olds responded 2 to 3 times more quickly than 3-day-olds, both age groups responded more rapidly to a strong shock than to a weak one. During habituation, response amplitudes of both 3- and 15-day-old pups declined with no change in latency. Age-related differences in habituation were shown to be independent of differences in reactivity to shock and more likely due to the emergence of response sensitization.


Mononuclear phagocytes from carrageenan-induce granulomas. Isolation, cultivation, and characterization. Stable cultures of mononuclear phagocytes from carrageenan-induced granulomas in mice have been established after enzymatic dispersion of these lesions. The cells can be maintained for up to 3 wk without division in serum-free media. The mononuclear phagocytes were identified by several criteria. The cells are adherent, phagocytic, contain lysosomal acid hydrolases at high specific activities, secrete lysozyme, and bind soluble aggregates of IgG. The activities of 5'-nucleotidase and leucine aminopeptidase in the cultured granuloma cells showed that they resembled macrophages from thioglycollate-stimulated mice but not unstimulated macrophages in these respects. Supernates from the cultured granuloma cells contain factor(s) which induce the proliferation of thymocytes; the release of such factors by the cells is stimulated by lipopolysaccharide.


Frustration odor of rats receiving small numbers of prior rewarded running trials. Three related experiments were performed to examine trial-to-trial differences in production of frustration odor during runway extinction by rats that had received 0, 2, 4, or 6 prior rewards. Differences in odor production were indexed by the subsequent speed of escape from the runway goal box by conspecifics. The results indicated that (a) subjects will perform a hurdle-jump response to escape from frustration odor; (b) escape performance even after several prior escape trials accurately reflects trial-to-trial changes in production of frustration odor; and (c) production of frustration odor requires only 4 prior rewards. Implications of the present data for frustration analyses of the limited-trial partial reinforcement effect and future investigations of frustration odor are discussed.


Reinforcement and the organization of behavior in golden hamsters: Pavlovian conditioning with food and shock unconditioned stimuli. The effects of Pavlovian conditioned stimuli (CSs) for food or shock on a variety of behaviors of golden hamsters were observed in three experiments. The aim was to see whether previously reported differences among the behaviors produced by food reinforcement and punishment procedures could be accounted for by differential effects of Pavlovian conditioning on the behaviors. There was some correspondence between the behaviors observed to the CSs and the previously reported effects of instrumental training. However, the Pavlovian conditioned responses (CRs) alone would not have predicted the effects of instrumental training. Moreover, CRs depended to some extent on the context in which training and testing occurred. These findings, together with others in the literature, suggest that the results of Pavlovian conditioning procedures may not unambiguously predict what system of behaviors will be most readily modified by instrumental training with a given reinforcer.


The role of predrug signals in morphine analgesic tolerance: support for a Pavlovian conditioning model of tolerance. According to a model of morphine tolerance, which emphasizes Pavlovian conditioning principles, tolerance results from an association between predrug environmental cues and the systemic effects of the drug. To assess this model, groups of rats were administered morphine on either three or nine occasions, with a complex environmental stimulus either paired or not paired with each injection. Control groups had equivalent experience with the environmental cue and injection procedure, but the injected substance was physiological saline. Subsequently, the analgesic effect of the opiate was tested in all subjects following administration of the drug in conjunction with the environmental cue. As expected on the basis of the conditioning model of tolerance, subjects with a pretest history of paired morphine administrations displayed analgesic tolerance, but subjects with a pretest history of unpaired administration displayed no evidence of such tolerance. The results suggest that prior demonstrations that the display of morphine tolerance is specific to the drug administration environment may be readily interpreted by a conditioning analysis of tolerance.


Conditioned fear and startle magnitude: effects of different footshock or backshock intensities used in training. In Experiment 1 four groups of rats received 30 light-shock pairings using footshock intensities of either .2, .4, .8, or 1.6 mA. One day later all rats were tested for startle by presenting tones in the presence or absence of the light CS. Potentiated startle (the difference between startle on light-tone vs tone-alone trials) was nonmonotonically related to the shock intensity used in training, with the greatest potentiation at intermediate shock levels. Experiment 3 demonstrated a similar relationship when backshocks instead of footshocks were used. In Experiment 2 rats were trained with either a moderate or high shock and then given an extended extinction-test session 1 day later. The moderate-shock group showed a gradual decline in potentiated startle over extinction. The high-shock group showed a nonmonotonic extinction curve where potentiation progressively increased toward the middle of extinction and dissipated thereafter. The results suggest that acoustic startle bears an inverted U-shaped relationship to fear and are discussed in relation to other studies concerned with this issue.


Radioimmunoassay of 17-alpha-alkylated anabolic steroids. A method for the detection of anabolic 17-alpha-alkylated androstane derivatives in both plasma and urine is described and evaluated. The goat and rabbit antisera against 17-alpha-Methyltestosterone-3-carboxymethyloxim-Rinder-serum albumin were raised and compared using [3H]methandrostenolone as a tracer. 22 Steroids including 10 potent synthetic anabolics were tested for their cross-reaction with these antisera.


[Short programs for the determination of amino acids in physiological fluids (author's transl)]. Short programs for the determination of a few amino acids are described. The studies were performed with two commercially available ion exchange resins (Durrum DC6A, Phoenix XX 907 OPKU). All the programs are constructed so that they are suitable for urine analysis with unfavourable concentration ratios. The economic aspects and the areas of application of these methods are discussed.


Evaluation of the enzymatic ammonia method for urine on the Du Pont automatic clinical analyzer. The enzymatic method for ammonia with the Du Pont Automatic Clinical Analyzer was tested in our laboratory. The method is very suitable for urine samples although the method was released by Du Pont only for plasma samples. The results correlated well with those obtained by a method based on the Berthelot indophenol reaction and by a method based on the use of an ammonium electrode, which has an ammonium ion selective membrane. The day-to-day coefficient of variation is about 2 and 4% at concentration levels of 100 and 50 mumol/1 respectively. The measured and relative ammonia concentration are linearly related. Recovery of added known amounts of ammonium is complete and no interference from glucose, urea, creatinine and uric acid could be demonstrated. Recommendations are given for handling blood and urine samples before analysis.


[Electrolyte and collagen content of rat heart in chronic Mg-deficiency and stress (author's transl)]. In chronic Mg-deficiency, there is a decrease in the content of Mg and K+ in the heart muscle. The contents of Ca, Na+ and hydroxyproline increase. There is also an increased urinary excretion of adrenaline and especially noradrenaline. With simultaneous noise stress, these changes are even greater. The magnitude of the changes increases with the degree of Mg-deficiency. The changes in the contents of Na+, K+, Ca, Mg and hydroxyproline correlate with the excretion of noradrenaline. The increase in the collagen content can be verified by electron microscopy, and it is due to a stimulation of the fibrocytes.


[Studies on the interaction of Phenprocoumon with Diazepam and with Nitrazepam (author's transl)]. The influence of Diazepam and Nitrazepam on the anticoagulating activity of Phenprocoumon was studied in rats. It was found that the repeated oral application of Diazepam or Nitrazepam during the treatment with Phenprocoumon did not change the adjusted prothrombin time. The prothrombin time was considerably shortened when the application of Diazepam or Nitrazepam was stopped although Phenprocoumon was still applied. Furthermore, the influence of Chlordiazepoxid, Diazepam and Nitrazepam on the normal prothrombin time was studied. It was found that the treatment with benzodiazepine derivatives had no influence upon the prothrombin time, not only during the treatment, but also after the treatment was stopped.


[Determination of serum iron; a comparison of two methods: atomic absorption and bathophenanthroline without deproteinisation (author's transl)]. Two methods for the assay of serum iron are compared; determination by atomic absorption after deproteinisation, and spectrophotometric determination after liberation of the iron by a detergent, reduction with dithionite, and chelation with bathophenanthroline disulfonate, without deproteinisation. Regression: y = 0.996 x --0.41 (s: atomic absorption; y: bathophenanthroline method; mumol/1 Fe; r = 0.986. Precision (CV) for a series of identical samples with 14.3 mumol/1: 2.0% for atomic absorption and 1.3% for bathophenanthroline.


Comparative studies on the radioimmunological determination of carcinoembryonic antigen in tumour tissue. Human carcinoembryonic antigen (CEA) was measured in perchloric acid extracts from 9 malignant gastrointestinal tumours, using two different radioimmunological methods (Hoffmann-La Roche and CEA-IRE-SORIN). When standard curves were set up, quantitative differences in the degree of inhibition were found between the standards from the two manufacturers. In the Hoffmann-La Roche assay the CEA binding curves of all tumour extracts ran parallel with the standard. However, when the CEA-IRE-SORIN assay was applied two of the nine tumours exhibited different slopes. In one case, using the CEA-IRE-SORIN method, various parts of the tumour exhibited binding curves with non-parallel slopes. The present results suggest that radioimmunologically heterogeneous CEA occurs in different gastrointestinal tumours as well as in various parts of the same tumour.


Diagnostic value of lysozyme activity estimation in the feces of infants with acute diarrhoea. The activity of lysozyme in feces was estimated in a control group of 50 healthy infants and in a group of 152 infants with acute diarrhoea. All infants investigated were artificially nourished. In the latter group the activity of lysozyme was estimated twice: a) at the beginning of clinically active phase of the disease and (b) in the convalescence period immediately after withdrawal of clinical symptoms. The range of normal values was 14.9--77.0 (average 44.0) of egg-white lysozyme units/g dry feces. In acute diarrhoea the activity of lysozyme in feces was found to be elevated in 72.4% of cases in the first determinations and in an additional 7.6% of cases in the second determination (i.e. a total of 80% of cases in both determinations). The average elevations of lysozyme activity in the feces and the dynamics of their normalization after withdrawal of clinical symptoms were generally related to the severity of the disease.


A rapid screening test for lactic aciduria. A rapid test for the determination of lactate in urine is described, which differentiates adequately between normal and elevated urine lactate levels. The method depends on the enzymatic decolourization of the blue dye dichlorophenolindophenol by lactate in the presence of the enzyme L-lactate cytochrome c oxidoreductase (EC 1.1.2.3). Interference by vitamin C is prevented by pre-treatment with charcoal. The test is simple and inexpensive, and a compact testing unit, which is stable and convenient for most laboratories, can be assembled in a glass capillary.


[Endogenous 3-O-methyldopa in man: normal level and effect of an inhibitor of aromatic amino-acid decarboxylase (Benserazid) (author's transl)]. The fasting level of 3-O-methyldopa in human plasma is 120 +/- 18 nmol/1. Four hours after a single dose of 425 mumol benserazid, the level increased to 313 +/- 69 nmol/1.


[A model for the adsorptive distribution of thyroid hormone in the cell (author's transl)]. Mitochondria from rat livers were suspended in solutions of cytoplasmic proteins ("cytosol") containing 0.5--4.7 g total protein per liter suspension. In these suspensions the adsorptive distribution of radioactive 3,5,3'-L-triiodothyronine was studied in the concentration range between 0.04 and 0.50 nmol/1. The experiments show that the distribution of the hormone between the liquid phase of the cytosol and the solid phase of the mitochondria depends mainly on the protein concentrations in both phases. The mathematical formulation of this distribution equilibria which is also applicable to thyroxine corresponded closely with the experimental findings. The processes of the adsorptive binding of thyroid hormones to cell proteins observed in vitro are thought to represent part of a transport chain that exists between different compartments in vivo. The consequences of these considerations on the transport of thyroid hormones within the cell are discussed.


[The reaction of cyclohexylsulphamic acid (Cyclamate) with nitrous acid in aqueous HCl-solution, isolated human gastric juice and urine (author's transl)]. The pH dependence of the formation and decomposition of the N-nitroso derivative, which is produced by the reaction of cyclohexylsulphamic acid with nitrous acid, was investigated by means of continuous photometric measurement of the changes in absorbance. Reaction at pH 2.45 resulted in an optimal yield of the N-nitroso derivative. The half-life of the decomposition reaction (the reactants cyclohexylsulphamic acid/sodium nitrite being in a molar ratio of 1:1.11) at pH 2.61 was 13.6 min. The N-nitroso derivative showed a relatively high stability at pH 4.5. The reaction in isolated human gastric juice and urine showed the same pH dependency as in aqueous HCl solution.


Variations in cerebral organization as a function of handedness, hand posture in writing, and sex. During the past century, it has become increasingly apparent that there is a great deal of variation in the direction and degree of cerebral lateralization, a plurality of people having language and related functions strongly specialized to the left hemisphere and visuospatial functions strongly specialized to the right, with substantial minorities manifesting various deviations from this pattern. In particular, in 35%-50% of sinistrals and 1%-10% of dextrals, the right hemisphere is specialized for linguistic skills, and in some unknown fraction of the two handedness groups, verbal and/or spatial abilities are, to varying extents, bilateralized. Levy (1973) suggested that the hand posture adopted during writing might be an index of the lateral relationship between the dominant writing hand and the language hemisphere, a normal posture indicating contralateral language specialization, and an inverted posture indicating ipsilateral language specialization. In the present investigation, two tachistoscopic tests of cerebral lateralization, one measuring spatial functions and one measuring verbal function, were administered to 73 subjects classified by handedness, hand posture during writing, and sex. Among both dextral and sinistral subjects with a normal writing posture, language and spatial functions were specialized to the contralateral and ipsilateral hemispheres, respectively, and lateral differentiation of the brain was strong. The reverse was seen in subjects having an inverted writing posture. In all groups, females were less laterally differentiated than males. In 70 out of 73 subjects, the direction of cerebral laterization was accurately predicted by handedness and hand posture. The 3 subjects (2 females and 1 male) who failed to manifest the predicted relations were all left-handers having an inverted hand posture . In this group, lateral differentiation was so weak that the reliability of the tachistoscopic tests was reduced, and we attribute these three predictive failures to this cause. Thus, almost all of the variation in the lateral organization of the brain was accounted for by handedness, hand posture, and sex.


An alternative interpretation of the stimulus suffix effect. The influence of a stimulus suffix was examined in yes-no recognition and probe-recall tasks. As with previous findings with serial recall, reliable suffix effects were obtained in probe recall. However, no suffix effects were found in the recognition task. Both serial and probe-recall tasks require the retention of order and item information, while only item information is required in recognition. An interpretation of the suffix effect based on interference with order information is presented.


Inositol pentaphosphate in fish red blood cells. Inositol pentaphosphate (IP5) has long been characteristic of avian erythrocytes. We now report that this compound is also present in the red cells of two species of elasmobranch fishes, Squalus acanthias (spiny dogfish) and Narcacion nobiliana (torpedo ray). The mean concentration of I5 is 0.36 and 0.24 micromoles per ml of red cells respectively. ATP is the major organic phosphate in both fishes. 2.3-diphosphoglycerate is absent and GTP levels correspond approximately to the levels of I5.


Estrogen localization in the brain of the lizard, Anolis carolinensis (1). Estrogen induces reproductive behavior in lizards by acting on specific areas of the brain. Neural areas that selectively concentrate radioactivity after 3-h-estradiol administration were mapped using thawmount autoradiography. Major accumulations of hormone-concentrating cells occur in the medial preoptic area, basal hypothalamus, amygdala (n. vetromedialis telencephali) and in and around the torus semicircularis. Numerous smaller accumulations are also described and the topographical distribution is compared with that of avian and mammalian species.


The effects of lumicolchicine, colchicine and vinblastine on pigment migration in fish chromatophores. The effects of lumicolchicine, colchicine, vinblastine and cold temperature on the pigment migration in melanophores and xanthophores of Fundulus heteroclitus and Oryzias latipes were examined by light and electron microscopy. Xanthophores of both species which contain numerous microfilaments and a poorly developed microtubule system were extremely sensitive to the alkaloids. Lumicolchicine and colchicine induced irreversible dispersion while vinblastine caused permanent aggregation of the pigments. Treatment in lumicolchicine or colchicine at 5 mM for 60 minutes did not disrupt microtubules of melanophores to an appreciable degree, however, melanosome aggregation was partially inhibited by these drug in Oryzias. When melanophores were kept in the cold in the presence of colchicine at 1 mM, almost all microtubules were disrupted and their repolymerization at room temperature was nearly completely inhibited by colchicine. These melanophores lacking in microtubules responded to epinephrine with slow aggregation. Vinblastine at 0.1 mM induced partial aggregation of melanosomes and disruption of microtubules but most melanophores were still able to respond with pigment migration. Vinblastine at 1 mM made all melanophores punctate and immobile. Large vinblastine-induced crystals were frequently seen in the dendritic processes. The results of the present investigation suggest that cytoplasmic microtubules in fish melanophores facilitate melanosome migration only in directional orientation and appear not be responsible for the motive force.


Inhibition in the Limulus lateral eye in situ. Inhibition in the Limulus lateral eye in situ is qualitatively similar to that in the excised eye. In both preparations ommatidia mutually inhibit one another, and the magnitude of the inhibitory effects are linear functions of the response rate of individual ommatidia. The strength of inhibition exerted between single ommatidia is also about the same for both preparations; however, stronger effects can converge on a single ommatidium in situ. At high levels of illumination of the retina in situ the inhibitory effects are often strong enough to produce sustained oscillations in the discharge of optic nerve fibers. The weaker inhibitory influences at low levels of illumination do not produce oscillations but decrease the variance of the optic nerve discharge. Thresholds for the inhibitory effects appear to be determined by both presynaptic and postsynaptic cellular processes. Our results are consistent with the idea that a single ommatidium can be inhibited by more of its neighbors in an eye in situ than in an excised eye. Leaving intact the blood supply to the eye appears to preserve the functional integrity of the retinal pathways which mediate inhibition.


Effects of bicarbonate on lithium transport in human red cells. Lithium influx into human erythrocytes increased 12-fold, when chloride was replaced with bicarbonate in a 150 mM lithium medium (38 degrees C. pH 7.4). The increase was linearly related to both lithium- and bicarbonate concentration, and was completely eliminated by the amino reagent 4, 4'- diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). DIDS binds to an integral membrane protein (mol wt approximately 10(5) dalton) involved in anion exchange. Inhibition of both anion exchange and of bicarbonate-stimulated lithium influx was linearly related to DIDS binding. 1.1 X 10(6) DIDS molecules per cell caused complete inhibition of both processes. Both Cl- and Li+ can apparently be transported by the anion transport mechanism. The results support our previous proposal that bicarbonate-induced lithium permeability is due to transport of lithium-carbonate ion pairs (LiCO-3). DIDS-sensitive lithium influx had a high activation energy (24 kcal/mol), compatible with transport by the anion exchange mechanism. We have examined how variations of passive lithium permeability, induced by bicarbonate, affect the sodium-driven lithium counter-transport in human erythrocytes. The ability of the counter-transport system to establish a lithium gradient across the membrane decrease linearly with bicarbonate concentration in the medium. The counter-transport system was unaffected by DIDS treatement. At a plasma bicarbonate concentration of 24 mM, two-thirds of the lithium influx is mediated by the bicarbonate-stimulated pathway, and the fraction will increase significantly in metabolic alkalosis.


A polysaccharide antigen from the Gram-positive organism Eubacterium saburreum containing dideoxyhexose as the immunodominant sugar. A highly active surface antigen, reacting by precipitation and complement fixation, has been isolated from Eubacterium saburreum strain L32. The antigen is a polysaccharide polymer built up of galactose, ribose and dideoxyhexose. The dideoxyhexose is the immunodominant sugar.


Mutants of Acinetobacter calcoaceticus NCIB 8250 constitutive for the mandelate enzymes. Methods for isolating mutants of Acinetobacter calcoaceticus NCIB 8250 constitutive for the mandelate enzymes were compared. Non-inducing substrates were not available. Continuous culture gave no stable constitutive mutants. Alternate culture in inducing (L-mandelate) and non-inducing (L-glutamate) media led to the isolation of mutants mesoconstitutive for L-mandelate dehydrogenase and phenylgloxylate carboxy-lyase, and hyperinducible for benzaldehyde dehydrogenase I. Screening of possible anti-inducers showed that 2-phenyl-propionate was very effective; it was a competitive inhibitor of gratuitous induction by thiophenoxyacetate, and was used for the isolation of constitutive mutants. Some of the mutants were magnoconstitutive for L-mandelate dehydrogenase, phenylglyoxylate carboxylyase and benzaldehyde dehydrogenase I and could not be further induced; others had lower specific activities which could be increased by induction with phenylglyoxylate or thiophenoxyacetate. Similar constitutive mutants were derived from a mutant which lacked L-mandelate dehydrogenase.


Origins of fermentation products formed during growth of Bacteroides ruminicola on glucose. Bacteriodes ruminicola grown on complex medium with glucose as carbon source gave acetate, CO2, formate and succinate as main fermentation products. No evidence was found for significant glucose catabolism by pathways other than the Embden-Meyerhof sequence. However, [U-14C]glucose fermentation gave products whose specific radioactivities were much lower than expected. There appear to be two main causes. Firstly, a rapid exchange occurred between metabolic intermediates and CO2, probably due to reversibility of the pathway between phosphoenolpyruvate and fumarate. Secondly, non-glucose precursors, mainly peptides and acetate, added to the medium as growth factors, also gave rise to the above end-products. The distortions that such reactions introduce into measurements of ATP molar growth yields based on product analyses and measurements of carbon flux based on radioactivity recovered in products are discussed.


Properties of a temperature-sensitive mutant of Staphylococcus aureus defective in DNA replication and cell division and replication of plasmids in the mutant. The properties of a temperature-sensitive mutant (ts39) of Staphylococcus aureus NCTC 8235 are described. After transfer to the restrictive temperature (42 degrees C), absorbance increased 10-to 20-fold but DNA content did not increase beyond 150 to 200% and cell division continued at a greatly reduced rate. On transfer back to the permissive temperature, both cell division and DNA synthesis resumed if the transfer occurred after less than 120 min at 42 degrees C. Resumption of DNA replication was blocked by chloramphenicol (100 microgram ml-1). The results are discussed with reference to possible defects in DNA replication. Replication of the plasmids pI258 and pT10501 and the chromosome were affected to a similar extent in ts39. Growth at 42 degrees C resulted in the appearance of an increased amount of pI258 DNA in a form that sedimented slowly in a sucrose gradient.


[Haemoglobinosis C/beta-thalassemia double heterozygosity in an Algerian patient with total suppression of haemoglobin A synthesis (author's transl)]. The authors describe the case of a young Algerian, aged 32, suffering from mild icterus, accompanied by a marked splenomegaly. The blood count revealed a moderate degree of anaemia with reticulocytosis, pronounced anisocytosis, micro-spherocytes, bulls eye cells, folded cells, hypochrome cells, a marked polychromasia and a mild erythroblastosis. Present also were hyperbilirubinaemia, raised plasma haemoglobin, zero haptoglobin, a reduced osmotic fragility and half-life of erythrocytes. Haemoglobin electrophoresis showed 17.25% haemoglobin F, 62.8% haemoglobin C+A2 and no haemoglobin A. The genetic study indicated that the patient was a double heterozygote C/beta thalassaemia, his mother and his son both suffering from this disease. This thalassemic gene of type beta (0) totally inhibited the synthesis of haemoglobin A, the defect found in our patient.


[Sclero-atrophic keratodermal genodermatosis of the extremities (sclerotylosis) (author's transl)]. The authors report a new case of sclero-atrophic keratodermal genodermatosis of the extremities, which is frequently degenerative. This condition, described by Huriez et al. in 1967, is characterized by symmetric lesions of the hands and feet, sclerodactyly, keratodermia, onychopathy and hypohidrosis. This disease, which is of dominant autosomal transmission, is associated with the MNSs trait, the two genes being probably located on chromosome No. 2.


[Constitutional stereotyped gap in human chromosomes (author's transl)]. The stereotyped break with gap of a chromosomal variant is rarely observed. This anomaly is transmitted according to autosomal dominant rule. The distal part of the broken chromosome may either be still bound to the sister chromatid through mitotic non-disjunction, forming a triradial figure, or take a moniliform and pulverized appearance, evoking premature chromosome condensation.


Ovarian differentiation in Turner's syndrome. The authors have studied the gonadal histogenesis and the sexual chromosome influence on the gonads of 17 patients with the following complements : 45,X/46,XXqi (1 case); 45,X (4 cases); 45,X/46,XXP--(1 cases); 45,Xq--  (1 case); 45,X/46,XX (8 cases); 45,X/46,XX/46,XXqi (1case); and 45,X/46,XXqi/47,XXqiXqi (1 case).  The presence of sexual differentiation structures was investigated : coelomic epithelium, stromal characteristics, follicles sexual cords, medullary tubules, rete ovarii, hilar cells, mesonephric remnants and coelomic epithelium inclusions. All gonads were constituted by rudimentary ovarian stroma with different states of hyalinization. Primordial follicles were noted in two patients with respectively 45,X/46,XX and 45,X/46,XXqi/47,XXqiXqi karyotypes, and a cystic follicle was present in one patient 45,X/46,XXp--. Sexual cords were seen in 6 patients and medullary tubules in 9. Different amounts of hilar cells were found as well. The authors conclude that in Turner's syndrome there exists an ovarian dysgenesis which is probably caused by early involution before reaching the maturation, conditioned by the genetic incapacity of the oogonia to complete the meiotic prophase.


Prevalence of Klinefelter's syndrome (47,XXY) in a general male population. In a representative sample of 3,840 males examined for military service chromosome examination was made in those with testes equal to or less than 12 ml and those with a stature equal to or greater than 181 cm, as well as in males not recruited because of physical or mental disability. Testes equal to or less than 12 ml were found in 59 patients (1.45%). Three of these males had a 47,XXY karyotype (5.1%), the prevalence among the total sample of 3,840 being 0.78 per 1,000. Hypogonadal signs, except for gynaecomastia, which was only present in one patient, were found in the saem proportion as in 47,XXY males ascertained in institutions and clinics. The results of EEG investigations were alos similar to those found in psychiatric institutions. The intelligence level was comparatively low; none had an IQ above 100. The personality traits corresponded to those found in institutionalized Klinefelter males.


Apparent non-penetrance for dystopia in Waardenburg syndrome type I, with some hints on the diagnosis of dystopia canthorum. Two large pedigrees with Waardenburg syndrome type I (W--I), i.e. with dystopia canthorum and blepharophimosis, are described to show both the variable expressivity of dystopia canthorum, which may be confused with non-penetrance of this sign, and the possibility to firmly diagnosis it with the new biometric index W, which differentiates a dystopic from a non-dystopic or a non-apparent dystopic subject,  the latter within a defined biometric range. A general discussion of the relative value of blepharophimosis and dystopia canthorum as diagnostic features in W--I is presented, to conclude on the greater value of dystopia canthorum, which can be identified with confidence in more than 96% of carriers. Empirical probabilities are given for dystopia canthorum and blepharophimosis in the general populations, based on data from the world literature, useful for all ethnic groups.


ABO blood groups and fertility in an Indian population. A total of 589 compatible mating couples could be investigated against 432 incompatible mating couples in order to determine the selective mechanism operating on ABO blood groups. There appears to be no striking difference in the proportion of childless couples between the two groups. The mean number of living children presents a significant difference. There is 21% deficiency of 'A' children in the two groups. Similarly, there is 16% deficiency of 'B' children in the two groups. It appears that there is 31.9% fetal wastage in incompatible matings as compared with 17.15% in compatible matings.


[Do all cases of trisomy 18 with long survival (beyond 10 years) show mosaicism in fibroblasts? (author's transl)]. Description of a young girl aged 16 1/2 years who presents the typical features of trisomy 18 : small height, microdolichocephaly, anti-mongoloid palpebral fissures, micrognathia, microstomia, arched palate, malformed ears, atrophy of thenars and hypothenars, clinodactyly of fifth fingers and abortive cutaneous syndactyly between IV and V. At the lower limbs, there is a shortening of the right leg, an atrophy of the calves, as well as genua valga and bilateral pes excavatus with dorsiflexion of the toes. The gait is rigid with enlarged basis of sustentation. The results of the cardiac examination point to a minor ventricular septal defect. The development of secondary sexual characters (breasts and body hair) corresponds to the puberal age; the large pudendal lips are hypoplastic. The X-rays show a double left kidney. There is a very severe oligophrenia (I.Q. = 20). Cytogenetic examinations showed a typical trisomy 18 in 100% of observed lymphocytes, while the analysis of cutaneous fibroblasts revealed a mosaicism with 87% of trisomic cells. Out of 11 cases of trisomy 18 with long survival from the literature only 3 cases were of mosaic type. The authors assume that this small number of mosaic cases in trisomy is probably due to the fact that no examination of fibroblasts has been carried out in the seven other cases.


[Partial trisomy 13 (13q14 leads to 13qter) in mosaic (author's transl)]. A case of partial trisomy 13 with mosaicism in a 6 1/2 year-old boy is reported. Concerning this case and according to previous data in the literature, the authors try to give a precise topology for trisomy 13; it is possible to outline phenotypes specific for a trisomy of both the proximal third and the distal two thirds of the chromosome, but it appears difficult to suggest a more precise picture for the topology of trisomy 13. This difficulty is related to the notion of phenotypic variability encountered in complete trisomy 13.


Familial transmission of a 3q;22p translocation, with partial trisomy of chromosome 3 in the propositus. A family, carrying a balanced 3q;22p translocation, was detected through a propositus who showed multiple congenital malformations. As there are no previous references of similar cases where identification techniques were performed, the authors present this material for consideration in the delineation of clinical syndromes associated with specific chromosomal anomalies.


[Karyotype 46,XY,22p+ in a male patient (author's transl)]. The authors report the case of a 2-month-old infant with psychomotor retardation and several physical stigmata. Cytogenetic studies of the patient using the normal technique show in all the cells a karyotype 46,XY with a G group chromosome substituted by an F-like mediocentric element with satellites. The R, G and C-banding methods reveal that it is the 22 with too developed short arms (22p+). This element was found in the mother's and maternal grandfather's karyotypes although they both present normal phenotypes. The authors advance two hypotheses concerning the origin of the alteration but cannot exclude a possible connection between this particular chromosome and the proband's anomalies. The difficulties of genetic counselling in this case are evident.


[Trisomy of chromosome No. 10 in mosaic (author's transl)]. A case of trisomy of chromosome No. 10 in mosaic is described in a boy who died at the age of 6 months. The frequency of pathological cells is less than 30% (28% from lymphocytes, 20% from fibroblasts); it is possible, anyway, to rule out the hypothesis of a cellular cloning in vitro, since the trisomy 10 was observed in two different cultures, terminated after 48 and 72 hours. The parents' karyotype was normal, except for a litte number (6%) of cells with trisomy 10 from cultured lymphocytes of the mother. The morphological features of the present case are compared with those of the boy described by Higurashi et al. in 1969 (mosaic of trisomy 10, with a higher frequency of pathological cells).


Evaluation of a simplified sucrose gradient method for the detection of rubella-specific IgM in routine diagnostic practice. Rubella-specific IgM was measured in a single fraction of serum from a sucrose density gradient. Haemagglutination inhibition (HAI) tests were performed on paired aliquots of the fraction untreated and after treatment with 2- mercaptoethanol, dilutions of the aliquots being incubated over night with rubella antigen before the addition of red cells. Of 822 sera tested, specific IgM was found in 249, but not in 492. When first tested, the remaining 81 sera gave unsatisfactory results because of contamination of the IgM fraction with IgG (6.0%), probable aggregation of IgG (3.5%), or the persistence of chick red cell agglutinins (0.4%). Tests were performed on 134 patients with rubella confirmed by a rise of HAI antibodies. Rubella-specific IgM was found at a titre of more than eight in the sera taken from 62 of 64 patients between 10 and 29 days after the onset of the rash but in only one of the sera taken between 80 and 119 days, and in none taken later. However, specific IgM was still to be found at lower titre in the sera of 13 patients collected between 80 and 162 days after the onset of the illness. In routine diagnostic tests over three years on the serum from 479 patients with suspected acquired rubella, specific IgM was found at a titre of more than eight in 51 patients and in only 10 instances (2.1%) did a lower level pose a problem in interpretation.