Cadmium and zinc flux in wild-type and cadmium-resistant CHO cells

Cellular flux of cadmium-109 and zinc-65 is characterized in cultured Chinese hamster ovary cells. The transport of cadmium is primarily unidirectional and, following uptake, cadmium is strongly retained. Zinc transport is bidirectional and intracellular zinc continuously leaches out into the medium. Nonradioactive cadmium or zinc enhances the efflux of⁶⁵Zn from prelabeled cells. Transport of these metals into wild-type cells is not affected by azide, ouabain, cycloheximide, or actinomycin D. A cadmium-resistant mutant was isolated that exhibited altered sensitivities to certain inhibitors of macromolecular synthesis as well as quantitative differences in metal transport and accumulation. Although the mutant accumulates less cadmium than the wild-type cell, that which is retained is bound much more tightly. In addition, this lower rate of cadmium uptake is significantly decreased by either cycloheximide or actinomycin D. This suggests that thede novo synthesis of a protein or proteins is required for much of the net cadmium retention by the cadmium-resistant cells.     

DEVELOPMENTAL CHANGES IN Na+, K+ AND ATP AND IN THE LEVELS AND TRANSPORT OF AMINO ACIDS IN INCUBATED SLICES OF RAT BRAIN

Abstract—

• 1 Upon incubation, slices of brain tissue took up fluid; the degree of swelling increased with increasing age. No sweiling occurred in slices from foetal brain. Since this swelling was associated with increases in the inulin space, the percentage of inulin space in slices at the end of incubation increased during brain development.
• 2 Most of the capacity for ion transport seemed to be absent from foetal brain. In vivo and in slices, Na⁺ was very high and K⁺ was very low in comparison to levels at other ages. There was a rapid change around birth, but no significant change at later ages. Upon incubation, Na⁺ levels increased in other slices, but not in slices of foetal brain.
• 3 Upon incubation of the slices, ATP levels were restored to levels close to those in the living brain; there were no significant alterations in available energy during development to explain changes in amino acid transport.
• 4 The composition of the free pool of cerebral amino acids in vivo changed with development, with some compounds (glutamic acid and related compounds) increasing, others (mostly‘essential’amino acids) decreasing, with age. These changes were not linear with time, and the level of a compound might exhibit several peaks during development.
• 5 The uptake (influx) of taurine, glutamate and glycine into brain slices increased rapidly during the foetal and early neonatal periods, reached a maximum between 2 and 3 weeks of postnatal age and then declined to adult levels. The levels of steady-state uptake with glycine also exhibited a maximal peak at 2-3 weeks of postnatal age. Steady-state uptake of taurine and glutamate reached adult levels by about 3 weeks of age.
• 6 The pattern of inhibition of amino acid transport by two specific amino acid analogues changed during development for some amino acids (GABA, glycine and glutamate), indicating an alteration in substrate specificity.
• 7 The results demonstrate complex changes in cerebral amino acid transport during development, with several maxima or minima and with changes in specificity for at least some compounds.

     

Osmotic alterations of the lysosomal system during rat liver perfusion: Reversible suppression by insulin and amino acids

Livers from nonfasted rats were perfused in situ under conditions known from previous studies in this laboratory to increase or decrease overall endogenous proteolysis. At the termination of the experiments, lysosomal alterations were evaluated by the increase in free acid phosphatase or N-acetyl-β-D-glucosaminidase that occurred when tissue homogenates were subjected to osmotic shock in hypotonic sucrose. In control perfusions, osmotic sensitivity increased spontaneously over unperfused values, reaching maximum by 60 min or earlier. Additions of insulin, amino acid mixtures, or cycloheximide in amounts known to suppress proteolysis prevented this spontaneous perfusion effect or, when added at 60 min, rapidly reversed it. Glucagon alone during perfusion did not increase osmotic sensitivity further; however, stimulation with glucagon was observed when the perfusion effect was suppressed by insulin or cycloheximide. Anoxia, induced by gassing with nitrogen instead of oxygen, markedly reduced the perfusion effect and also doubled the amount of free acid phosphatase in the initial isotonic homogenates. Total acid phosphatase activities in the perfusion experiments were not significantly different from unperfused values and, with the exception of the anoxia perfusions, the amounts of free enzyme present in the initial isotonic sucrose homogenates did not change.     

Ribonucleic Acid Synthesis of Vesicular Stomatitis Virus: I. Species of Ribonucleic Acid Found in Chinese Hamster Ovary Cells Infected with Plaque-forming and Defective Particles

Plaque-forming B particles of vesicular stomatitis virus (VSV) induce the synthesis of virus-specific ribonucleic acid (RNA) in Chinese hamster ovary cells, whereas defective T particles do not. Infection with low input multiplicities of B results in the formation of four species of RNA. During infection with high multiplicities, RNA synthesis begins with mainly these four species of RNA but gradually shifts to a new pattern of RNA synthesis involving five other species of RNA. The change can also be induced by superinfection with T at 2.5 hr after infection with a low multiplicity of B. T added at the same time as B prevents virtually all RNA synthesis. Synthesis of the first group of RNA species correlates with the formation of B particles, whereas synthesis of the second group correlates with the formation of T particles. The various species of RNA formed after infection with VSV particles include single-stranded RNA, a completely double-stranded RNA, and RNA with partially double-stranded regions. These observations begin to establish a molecular basis for understanding the ability of T particles to interfere with the growth of B particles.     

Rapid Fluorescent-Antibody Staining Technique

The rapid fluorescent-antibody staining technique described by Kellogg and Deacon for staining Neisseria gonorrhoeae and Treponema pallidum was applied to fluorescent-antibody tests for group A streptococci and enteropathogenic Escherichia coli. Results obtained with this staining technique were compared with results using the conventional staining procedure; excellent correlation was obtained. Considerable time and materials were saved by using the rapid method; it was also found completely satisfactory.     

BIOSYNTHESIS OF RIBONUCLEIC ACID IN RAT BRAIN SLICES

The incorporation of uridine into RNA in brain slices was studied. Optimal conditions for uridine incorporation were determined. The characteristics of the product suggest that de novo DNA-directcd synthesis of fairly high molecular weight material takes place. Incorporation into RNA of several areas of brain was studied. The incorporation was also studied as a function of the age of the animal. Finally, an apparent correlation was observed between the decrease in uridine incorporation with age and the increase of the enzyme uridine nucleosidase which hydrolyses uridine to uracil, a material which cannot be incorporated into RNA.     

The Photosynthetic Electron Transport Chain of Chlamydomonas reinhardi. VII. Photosynthetic Phosphorylation by a Mutant Strain of Chlamydomonas reinhardi Deficient in Active P700

Electron transport activity and absorbance changes associated with P700 were investigated in a mutant strain of Chlamydomonas reinhardi with impaired photosynthesis. This mutant strain, ac-8oa, cannot reduce NADP with electrons from either water or dye and ascorbate, but it has considerable Hill activity. The mutant strain shows none of the absorbance changes characteristic of P700. Although unable to carry out cyclic photosynthetic phosphorylation, ac-8oa is able to synthesize ATP when ferricyanide is provided as an electron acceptor.

These observations lead to the conclusion that a site for the coupling of photosynthetic phosphorylation with electron transport must exist between the 2 photochemical systems.