Division competence in Tetrahymena: determination of minimum cell volume and rate of nutrient uptake

Cell volume and doubling time have been determined for exponentially growing Tetrahymena pyriformis cells in broth medium with and without glucose and in media made from these media by dilution with water. The cells tolerate media with dry weights from 105 down to 0.06 g/L. In the diluted media the cells have small volumes and the doubling time is increased. When the cell volume increase per time per cell in a given medium is expressed as a function of the cell volume in this same medium, a direct proportionality is found. From this equation the minimum cell volume of division competence (MVDC) can be found. It is 2,100 microns 3 for T. pyriformis at 28 degrees C. The lag period resulting from an upshift of exponentially growing cells from diluted media to more concentrated media is a function of the initial and resulting cell volumes and MVDC. The increase in cell volume per unit of time for a given cell depends on the dry weight of the medium. This parameter can be transformed to mass increase per cell surface area per time, which represents rate of nutrient uptake. When plotted against the dry weight of the media, a Michaelis-Menten-like curve is obtained with two Km values of 3.8 and 0.08 g/L with corresponding Vmax values of 20 and 4 ng/cm2.s. The low Km value (0.08 g/L) indicates that Tetrahymena is able to take up nutrients from highly diluted media. The high value of Vmax (20 ng/cm2.s) increases the ability of growth in more concentrated media.(ABSTRACT TRUNCATED AT 250 WORDS)     

Energetics of gramicidin hybrid channel formation as a test for structural equivalence. Side-chain substitutions in the native sequence.

To determine whether amino acid side-chain substitutions in linear gramicidins after the structure of membrane-spanning channels formed by the modified peptides, we have developed a quantitative measure of structural equivalence of the peptide backbone among gramicidin channels based on functional (single-channel) measurements. The experiments exploit the fact that gramicidin channels are symmetrical dimers, and that channels formed by different gramicidin analogues can be distinguished on the basis of their single-channel current amplitudes or durations. It is thereby possible to determine whether hybrid channels can form between chemically dissimilar peptides, i.e. whether the peptides can adapt to each other. Further, since the relative rates of channel formation as well as the relative concentrations of pure and hybrid channel types can be measured in the same membrane, these experiments provide a quantitative measure of the energetic cost of hybrid channel formation relative to the formation of the pure channels. For a wide variety of different side-chains, we find that substitutions as extreme as glycine to phenylalanine at position 1, at the join between the two monomers in a membrane-spanning dimer, incur no energetic cost for channel formation, which implies that channels formed by each of the modified peptides are structurally equivalent. In addition, the average durations of the hybrid channels (except those having tyrosine or hexafluorovaline at position 1) are intermediate to the average durations of the respective pure channel types, thus providing further evidence for structural equivalence among channels formed by sequence-substituted gramicidins.     

Glutathione Turnover in Cultured Astrocytes: Studies with [15N]Glutamate

The incorporation of [15N]glutamic acid into glutathione was studied in primary cultures of astrocytes. Turnover of the intracellular glutathione pool was rapid, attaining a steady state value of 30.0 atom% excess in 180 min. The intracellular glutathione concentration was high (20-40 nmol/mg protein) and the tripeptide was released rapidly into the incubation medium. Although labeling of glutathione (atom% excess) with [15N]glutamate occurred rapidly, little accumulation of 15N in glutathione was noted during the incubation compared with 15N in aspartate, glutamine, and alanine. Glutathione turnover was stimulated by incubating the astrocytes with diethylmaleate, an electrophile that caused a partial depletion of the glutathione pool(s). Diethylmaleate treatment also was associated with significant reductions of intraastrocytic glutamate, glycine, and cysteine, i.e., the constituents of glutathione. Glutathione synthesis could be stimulated by supplementing the steady-state incubation medium with 0.05 mM L-cysteine, such treatment again partially depleting intraastrocytic glutamate and causing significant reductions of 15N labeling of both alanine and glutamine, suggesting that glutamate had been diverted from the synthesis of these amino acids and toward the formation of glutathione. The current study underscores both the intensity of glutathione turnover in astrocytes and the relationship of this turnover to the metabolism of glutamate and other amino acids.     

Evidence of a Monoaminergic-Cholinergic Imbalance Related to Visual Hallucinations in Lewy Body Dementia

Senile dementia of Lewy body type is characterized clinically by a relatively acute onset of fluctuating memory loss and confusion, frequently accompanied by visual hallucinations. Neurochemical analyses of temporal cortex has revealed a distinction between hallucinating and nonhallucinating patients in both cholinergic and monaminergic transmitter activities. In contrast with the cholinergic enzyme choline acetyltransferase, which was more extensively reduced in hallucinating individuals, serotonergic S2 receptor binding and both dopamine and serotonin metabolites were significantly decreased in nonhallucinating cases. These results suggest that an imbalance between monaminergic and cholinergic transmitters is involved in hallucinogenesis in the human brain.