Ouabain-insensitive salt and water movements in duck red cells. II. Norepinephrine stimulation of sodium plus potassium cotransport

Catecholamines induce net salt and water movements in duck red cells incubated in isotonic solutions. The rate of this response is approximately three times greater than a comparable effect observed in 400 mosmol hypertonic solutions in the absence of hormone (W.F. Schmidt and T. J. McManus. 1977 a.J. Gen. Physiol. 70:59-79. Otherwise, these two systems share a great many similarities. In both cases, net water and salt movements have a marked dependence on external cation concentrations, are sensitive to furosemide and insensitive to ouabain, and allow the substitution of rubidium for external potassium. In the presence of ouabain, but the absence of external potassium (or rubidium), a furosemide-sensitive net extrusion of sodium against a large electrochemical gradient can be demonstrated. When norepinephrine-treated cells are incubated with ouabain and sufficient external sodium, the furosemide-sensitive, unidirectional influxes of both sodium and rubidium are half- maximally saturated at similar rubidium concentrations; with saturating external rubidium, the same fluxes are half-maximal at comparable levels of external sodium. In the absence of sodium, a catecholamine-stimulated, furosemide-sensitive influx of rubidium persists. In the absence of rubidium, a similar but smaller component of sodium influx can be seen. We interpret these results in terms of a cotransport model for sodium plus potassium which is activated by hypertonicity or norepinephrine. When either ion is absent from the incubation medium, the system promotes an exchange-diffusion type of movement of the co-ion into the cells. In the absence of external potassium, net movement of potassium out of the cell leads to a coupled extrusion of sodium against its electrochemical gradient.     

Differentiation of chick embryo myoblasts is transiently sensitive to functional denervation

Clonal analysis of myoblast differentiation has been used to assess effects of denervation on developing skeletal muscle: chick embryo legs denervated by spinal cord cautery yield reduced proportions of clonable myoblasts (P. H. Bonner, 1978, Develop. Biol., 66, 207–219). The present work examines the effects on clonable myoblasts of functional denervation by d-tubocurarine. Curare treatment during the third or fourth days of embryonic development had no effect on clonable myoblasts later in development, treatment during the fifth or sixth days resulted in reduced proportions of clonable myoblasts, and treatment during the eighth or ninth days again had no effect. Clonal analysis of treated and control embryo leg muscle cells was performed between Days 10 and 18. Embryos were also permanently denervated by spinal cord cautery late in the sixth day. These embryos showed no effect of denervation on clonable myoblast proportion. It is concluded that the differentiation of skeletal muscle myoblasts is affected by interference with normal nerve-muscle relationships only during a “window” of sensitivity and that this “window” extends approximately from Hamburger and Hamilton stage 27 to stage 30.     

Changes in the structure of Nippostrongylus brasiliensis intestinal cells during development from the free-living to the parasitic stages

The ultrastructure of Nippostrongylus brasiliensis intestinal cells was examined in free-living, feeding second-stage larvae, infective, nonfeeding third-stage larvae, and parasitic, feeding third-stage larvae. The intestinal cells of second-stage larvae were characterized by a well-developed microvillar border, large numbers of ribosomes, Golgi complexes, rough endoplasmic reticulum, and nuclei with prominent nucleoli. The intestinal cells of infective, third-stage larvae had very few microvilli and the cells were extremely narrow. Few ribosomes, Golgi complexes, and little rough endoplasmic reticulum were present. Nuclei did not contain nucleoli. When worms were introduced into an in vitro culture system, development of intestinal cells began. By 36 hr, microvilli were well differentiated and the cysoplasm contained numerous ribosomes and Golgi complexes, and rough endoplasmic reticulum, mitochondria, and nucleoli were prominent. These morphological changes were related to changes in the physiology of Nippostrongylus brasiliensis which occur during development from a free-living to parasitic form.     

Experiments on the abiotic amplification of optical activity

Our earlier experiments are briefly reviewed, involving the abiotic generation of optical activity by exposure of DL-amino acids to various chiral physical forces. The enantiomeric enrichments so obtained were low, however, and additional experiments were undertaken with the objective of abiotically enhancing such small enantiomeric excesses. DL Mixtures of leucine N-carboxy anhydride gave enantiomerically enriched polymers on partial polymerization, while valine NCA mixtures behaved oppositely. Leucine polymers were also found to hydrolyze stereoselectively, providing for additional enantiomenic enhancement. A repetitive sequence of partial polymerization-hydrolysis steps is suggested as a possible mechanism for the abiotic genesis of optically enriched polypeptides on the primitive Earth.     

Kinetic determination of the genome size of the pea

Renaturation of pea (Pisum sativum) DNA has been used to estimate the size of the pea genome and the fraction of pea DNA containing repeated DNA sequences. Pea DNA renaturation and single copy tracer renaturation indicate that the size of the pea genome is 0.5 picograms. More than 70% of pea DNA sequences are repeated from 100 to 5,000 times.     

Amino acids are general growth inhibitors of Nicotiana silvestris in tissue culture

The growth of Nicotiana silvestris in suspension culture is inhibited by all of the common protein amino acids at the millimolar level, except for L-glutamine. A defined experimental system for growth/inhibition studies has been established, and growth studies were carried out with cells that had been maintained in the exponential growth phase for at least 10 generations (EE cells). The following results were obtained after particularly detailed studies with aromatic amino acids. The onset of inhibition was preceded by a duration of normal growth rate which varied within a range of 12 to 48 h. The degree of inhibition was directly proportional to amino acid concentration and inversely related to the initial cell density of the inoculum. A slowed, but still exponential rate of growth persisted during an early phase of inhibition. Under sufficiently severe conditions, this was followed by progressive diminution of growth rate and eventual lysis. The most drastic inhibitory effects caused by aromatic amino acids were in the order: phenylalanine, tryptophan and tyrosine. When EE cells cultivated under conditions of growth inhibition were diluted into fresh medium, immediate resumption of growth at the uninhibited rate occurred and persisted. On the other hand, when growth-inhibited EE cells were diluted into medium containing the same concentration of amino acid used in the first round of growth, an initial burst of uninhibited growth lasting about 24 h was followed by a drastic, progressively declining growth rate which deteriorated to cell death and lysis. When cells in stationary phase were used as an inoculum, as is done in typical growth characterizations with suspension cultures, the sensitivity to inhibition during the subsequent exponential growth phase was several-fold greater than was the case with EE cells. Hypotheses that growth inhibition might be caused by ammonia toxicity, keto-acid toxicity, or by inhibition of nitrate utilization were ruled out. Observations that provide new insight are: (i)growth-inhibited cells undergo drastic plasmolysis, (ii) L-glutamine is an effective antagonist of amino-acid inhibitors, and (iii) growth-inhibited cells exhibit a transient restoration of normal growth rate upon dilution into fresh growth medium. These results implicate a linkage of amino acids with osmotic regulation and nitrogen metabolism.     

Cytofluorometric analysis of anti-lymphocyte antibodies in AIDS

Anti-lymphocyte antibodies (ALA) have been detected in the plasma of 53.8% of HIV-positive patients tested (CD4/CD8 ratios: mean 0.265; range 0.01 to 0.5) using analytical continuous-flow cytofluorometry. IgG from the AIDS plasma was seen to bind to normal PBL in 53.8% of cases (14/26). In double labelling experiments CD4 + lymphocytes, CD8 + lymphocytes, and B lymphocytes were all bound by the ALA, but monocytes were not bound. Pre-adsorption of the diluted AIDS plasma onto an excess of mouse spleen cells did not remove lymphocyte binding activity. No evidence was found for preferential binding to phytohaemagglutinin-stimulated lymphocytes. ALA could not be detected in the plasma of normal subjects, patients with acute renal failure undergoing renal dialysis, or patients with high levels of circulating immune complexes.     

The radiolysis and racemization of leucine on proton irradiation

D- and L-Leucine have been subjected to 39–55 percent radiolysis using 0–11 MeV protons, both with the proton beam passing through the sample or being absorbed by it, and with quenching the sample immediately on completion of irradiation or after a 21-day interval. Racemization was small (1.1–1.7 percent) and comparable in all cases, suggesting that radioracemization and secondary degradative effects were not important factors in our recent unsuccessful attempts to induce optical activity in DL-leucine by partial radiolysis using 0–11 MeV longitudinally polarized protons.     

The radiolysis of tryptophan and leucine with32Pβ-radiation

Summary We have extended earlier experiments on the radiolysis of DL-tryptophan using³²P-radiation to longer reaction times, observing complete destruction of the tryptophan by secondary, non-radiolytic processes. We have also undertaken the irradiation of DL-leucine with³²P's at -196°, achieving radiolyses to the extents of ca. 20–30%, but observing no concomittant asymmetric bias. The implications of these observations are discussed with regard to the Vester-Ulbricht mechanism for the origin of optical activity.     

Computer programs for analysis of nucleic acid hybridization, thermal denaturation, and gel electrophoresis data.

Computer programs for the analysis of data from techniques frequently used in nucleic acids research are described. In addition to calculating non-linear, least-squares solutions to equations describing these systems, the programs allow for data editing, normalization, plotting and storage, and are flexible and simple to use. Typical applications of the programs are described.