Stimulation of Rb+ transport by glucagon in isolated rat hepatocytes.

Glucagon caused a 2-2.5-fold increase in 86Rb+ (a K+ tracer) uptake in primary monolayers of rat hepatocytes. Removal of the hormone led to a slow reversal of the effect over a period of hours. Glucagon acted by increasing the maximum velocity of influx. Ouabain inhibited Rb+ uptake in both hormone-induced and control cells by 85-90%. Hormone stimulation and ouabain inhibition did not affect exit. The induction by glucagon required 3-4 h for full expression and was dependent on protein synthesis and changes in cAMP. Half-maximum stimulation occurred at a concentration of 0.4 nM of the hormone. The ionophores monensin and gramicidin, which lead to Na+ influx, stimulated ouabain-sensitive Rb+ transport. This suggested that the (Na,K)-pump rate is limited by the internal Na+ concentration in both glucagon-stimulated and control cells.

Interaction of the alkylating antitumor agent 2,3,5-tris(ethyleneimino)benzoquinone with the plasma membrane of Ehrlich ascites tumor cells.

The effect of the alkylating agent 2,3,5-tris(ethyleneimino)benzoquinone (Trenimon) on the uptake of 2-aminoisobutyric acid, 1-aminocyclopentane-1-carboxylic acid (cycloleucine), 3-O-methyl-D-glucose, and 86Rb was studied. All transport studies were performed at nonsaturating conditions where the specific transport system was rate limiting for the uptake. The activities of all systems are reduced after treatment with the alkylating agent. The impairment of the plasma membrane is expressed 30 sec after exposure to the drug, as measured by the 86Rb uptake, and lasts for at least 12 hr according to the reduced 3-O-methyl-D-glucose uptake. Inhibition of protein synthesis by cycloheximide for 2 hr does not affect the uptake of 86Rb. The short interval which is necessary before an inhibition of 86Rb uptake can be registered and the resistance of the 86Rb transport system to an inhibition of protein synthesis are considered as indicative for a direct alkylation of a membrane constituent. Treatment with the alkylating agent increases the cyclic adenosine 3':5'-monophosphate of the cells. This effect is not due to an effect on adenylate cyclase or the membrane-bound cyclic adenosine 3':5'-monophosphate phosphodiesterase. In view of the known correlations between plasma membrane functions and the regulation of cell division, it is proposed that the growth inhibition by Trenimon of Ehrlich ascites tumor cells may be caused by its interaction with the plasma membrane.

CO2 fixation and its regulation in Anacystis nidulans (Synechococcus).

Anacystis nidulans (Synechococcus) had a minimal doubling time of 5 hrs at 30 degrees C at saturating light intensity and carbon dioxide concentration. Half maximal growth rates in saturating CO2 occured at a light intensity of 0.54 mW per cm2, and there was an apparent threshold intensity of 0.13 mW per cm2 below which no growth occurred. Growth rate in saturating light was dependent on the concentration of CO2+H2CO3 in the medium, rather than on total dissolved CO2; half maximal rates were estimated at 0.1 mM CO2+H2CO3. Under saturating conditions of light and CO2, 14CO2 was fixed primarily into 3-PGA, and subsequently moved into sugar phosphates and amino acids. Incorporation into aspartate was relatively slow. CO2 fixation was strictly light-dependent. The changes in adenylate and pyridine nucleotide pools were followed in light/dark and dark/light transitions. Whereas adenylates relaxed slowly over 15-20 min to the concentrations characteristic of illuminated cells following the abrupt changes induced by darkening, the sharp drop in intracellular NADPH showed little dark recovery although rapid restoration occurred on reillumination. Other pyridine nucleotides showed no changes during these transitions. The nucleotide specificity and Km of partially purfied GAP dehydrogenase suggest a role for this enzyme in the regulation of CO2 fixation.

Phosphate utilization and alkaline phosphatase activity in Anacystis nidulans (Synechococcus).

Anacystis nidulans (Synechococcus) was maintained in a medium of low phosphate concentration (0.1 mM) and grew with a normal doubling time of 5 hrs at 30 degrees C. Such cultures had a normal pigment composition and alkaline phosphatase was detectable at low specific activities only. The onset of phosphate-limited growth occurred when the phosphate concentration in the medium fell to a value below 4 muM (the limit of accurate determination by the assay method used ) and resulted in increases in alkaline phosphatase activity, reaching a final 10 to 15 fold increase in specific activity after a period of several hours. Marked changes in the overall pigment composition occurred in this period of growth restriction. The addition of phosphate to such cultures resulted in a halt in synthesis of the enzyme and the restoration of normal pigmentation before growth resumed at the normal rate. Several oraganic phosphate esters could replace inorganic phosphate for growth and were also hydrolyzed by the partially purified enzyme, but growth rates were characteristically lower and the specific activity only 3 to 4 fold higher than in cultures grown in phosphate excess. Studies with the partially purified enzyme suggested that it differed in some of its properties from other alkaline phosphatases described in the literature.