Two color light scattering identifies physical differences between lymphocyte subpopulations

Forward angle light scattering of two different wavelengths by cells in a flow cytometer was used to investigate physical differences between lymphocytes of different lineage, functional subclass and developmental stage. Correlation of the ultraviolet (UV: 351 nm and 364 nm) and 488 nm light scattering signals produced by lymphoid cells demonstrated that the two signals were not equivalent and that they placed different emphasis on the physical parameters characterizing lymphocytes. Both small T and B lymphocytes from peripheral lymphoid tissues and mitogenically activated large T and B lymphocyte blasts were discriminated by both wavelengths. Differences between the Lyt-2 negative and Lyt-2 positive T lymphocyte subsets were also apparent. Two color light scattering could also discriminate between immature thymocytes and mature peripheral T cells and between small bone marrow cells and mature peripheral B cells. In bone marrow an increase in UV light scattering coincided with the appearance of cell surface immunoglobulin on small cells. These data establish that two color light scattering is a sensitive probe for distinguishing cells of apparently similar morphology and that it can be used to study the physical changes that occur during lymphoid cell differentiation.     

Influence of oxytocin on human memory processes: Validation by a control study

An earlier report (1) of an adverse effect of high doses of oxytocin on human memory included results of studies on women receiving oxytocin as part of the treatment to induce 2nd trimester therapeutic abortion. These women served as their own controls. We have now been able to study a group of women who have been treated in all ways like the original group, with the exception that they did not receive oxytocin. The results from this external control corroborate the finding that oxytocin affected memory.     

Isolation of zymogen granules from rat pancreas and characterization of their membrane proteins

A zymogen granule fraction has been isolated from rat pancreas, and its purity has been assessed by biochemical and morphological criteria. Specific activities of two marker enzymes, amylase and chymotrypsin, are increased by 4.6 and 5.4-fold, respectively, as compared to the homogenate. The purified fraction is devoid of detectable RNA, DNA and 5'-nucleotidase, glucose-6-phosphatase, and cytochrome c oxidase activities. Electron micrographs confirm the absence of mitochondria, lysosomes, and rough endoplasmic reticulum fragments. Zymogen granule membranes were isolated from this fraction on a sucrose gradient following lysis in alkaline buffer. Secretory contaminants were efficiently removed from the membranes as indicated by experiments in which labeled secretory proteins were added during the isolation procedure and secondly by measuring residual levels of amylase and chymotrypsin. Three enzyme activities were found in the membranes: thiamine pyrophosphatase, ATP-diphosphohydrolase, and low levels of acid phosphatase. Membrane proteins were solubilized by urea-Triton X-100 and separated in double-dimension (isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Isoelectric point and molecular weight of each protein band were determined.     

The influence of hormones and other substances on lens regeneration in vitro

Culturing the dorsal iris epithelium of a newt with a pituitary gland in organ culture greatly enhances the ability of the iris epithelium to produce advanced lens regenerates in vitro. In an attempt to elucidate the mechanism by which the pituitary enhances lens regeneration irido-corneal complexes from adult newts were cultured in medium to which various substances had been added either singly or in numerous combinations. Prolactin, insulin, hydrocortisone, and thyroxine failed to enhance the production of advanced lens regenerates in any of the doses or combinations tested. Similarly, addition of 50 microgram/ml of sodium or calcium ascorbate had no effect on the progress of lens regeneration in vitro. Addition of dibutyryl cyclic-AMP caused an inhibition of depigmentation and regeneration at high doses. The results of these experiments show that the effects of the pituitary cannot be duplicated by hormones which other authors have asserted to be beneficial to limb or tail regenerates in vitro. The results with cyclic AMP suggest that prolonged exposure to high doses of cyclic AMP inhibit regeneration and indicate that further studies on the fluctations in cyclic AMP levels throughout the process of lens regeneration must be done.     

Cat satellite DNA. Isolation using netropsin with CsCl gradients

The absence of centromeric bands in the karyotype of Felis catus is confirmed. It is also confirmed that no satellite band is visible in CsCl density gradients. However, a satellite is observed both by recentrifuging the fraction of the DNA that bands at high density in CsCl and by using netropsin to enhance the resolution of a CsCl gradient containing total F. catus DNA. The satellite, about 0.5% of total DNA, was isolated by repeated centrifugation in CsCl alone and in CsCl with netropsin. Netropsin was removed and a pure satellite DNA obtained. The reassociation kinetics (C0t1/2 less than 10(-3) M . s) show that the satellite is of the simple sequence type and hence a candidate for centromeric heterochromatin. Its cytological localisation awaits in situ hybridisation experiments.     

Structure—activity relationships in the mutagenicity of quinone methides of 7-hydroxyflavylium salts for Salmonella typhimurium

Several synthetic 7-hydroxyflavylium salts related to apigeninidin, a natural 3-deoxyanthocyanidin, have been studied in the Ames mutagenicity test using strain TA1537 of Salmonella typhimurium. Under the neutral pH conditions of the test, these flavylium salts are deprotonated through ionization of the C7-OH (pK_a′ = 4.2–4.4) to form quinone methides. Only the quinone methides of 4-methyl-7-hydroxyflavylium chloride and 4′-methoxy-4-methyl-7-hydroxy-flavylium chloride showed mutagenicity. Responses of 4–8 times the background were observed at the higher doses (1000 μg/plate), both with and without metabolic activation. It was concluded that the induction of frameshift mutagenicity by this group of compounds is caused by those quinone methides that have non-ionic, stable polycyclic structures at neutral pH.     

Mechanism of action of ryanodine on cardiac sarcoplasmic reticulum

Ryanodine was found to initially inhibit calcium uptake by cardiac sarcoplasmic reticulum. This initial depression was followed by a later marked stimulation of calcium uptake. These effects were noted when calcium uptake was measured in the presence or absence of oxalate. The requirement for preincubation with ryanodine was highly dependent on ryanodine concentration and temperature. The mechanism of action of ryanodine clearly was not an effect on oxalate entry or calcium oxalate precipitation because the effects were also observed in the absence of oxalate. Ryanodine also had no effect on passive calcium efflux from actively loaded vesicles. Because ryanodine had no effect on Ca2+-ATPase activity under defined conditions of an ATP-regenerating system and no calcium gradient, we suggest ryanodine does not change the stoichiometry of the pump. Our results are consistent with the hypothesis that ryanodine closes a calcium channel in a subpopulation of the vesicles.