Effect of defolliculation and 17α-hydroxy, 20β-dihydroprogesterone on cyclic AMP level in full-grown oocytes of the rainbow trout,Salmo gairdneri

The changes in cAMP were followed in trout oocytes incubated in vitro after defolliculation performed by either enzymatic or manual dissection. Both defolliculation methods induced a highly significant rise in oocyte cAMP level (4.5 times the basal level of control [follicle-enclosed oocytes], after 6 h). Treatment of defolliculated oocytes with 17α-hydroxy,20β-dihydroprogesterone (17α,20β-OH-P) (10^?6 M), which induced oocyte maturation (germinal vesicle breakdown [GVBD]) was able, first, to interrupt the increase of oocyte cAMP level promoted by defolliculation and then to lower this level significantly down to values that still remained higher than folliculated controls. Very low concentrations of 17α,20β-OH-P (1.38–55.6 10^?9 M), or physiological doses of testosterone (0.35 10^?6 M, in the range found in vivo before ovulation) were able to induce a similar decrease of oocyte cAMP level without inducing GVBD. Under the same experimental conditions estradiol (0.35 10^?6 M) exhibited no action. These results suggest that some factor(s) originating in the follicle (FIF), inhibit the oocytes' tendency to accumulate cAMP before the final surge of 17α,20β-OH-P. This factor might be a follicular steroid such as testosterone or nonmaturing concentrations of 17α,20β-OH-P. Moreover our data favour the hypothesis that the final surge of 17α,20β-OH-P could induce distinct intraoocyte mechanisms: the first induces an irreversible blockage of cAMP level before the inhibitory action of the FIF is suppressed by ovulation, and the second mechanism leads to GVBD.     

Isolation and characterization of plasma membrane fromAcanthamoeba culbertsoni

A rapid method for preparation of plasma membrane fromAcanthamoeba culbertsoni involving toluene treatment followed by lithium bromide extraction is described. In the plasma membrane preparation, 5′-nucleotidase, Na⁺ + K⁺ -ATPase, Mg²⁺ -ATPase and glucose-6-phosphatase activities were enriched. The membrane preparation was free from nucleic acid, cytochrome P-450 and cytochrome b5. Amino acid (¹⁴C-Ieucine) was not incorporated in the plasma membrane in 2 min. Succinic dehydrogenase was not detectable in the plasma membrane preparation. The molar ratio of cholesterol and phospholipids was 0.95 which is characteristics for plasma membranes. Under electronmicroscopy the preparation was homogenous without any other component of the cell. Plasma membrane proteins and glycoproteins were separated on acrylamide gel electrophoresis     

Physiology of a temperature-sensitive mutant of Saccharomyces cerevisiae defective in phosphofructokinase activity.

In this paper, we describe a temperature-sensitive mutant of the yeast Saccharomyces cerevisiae (P5-9) which at a restrictive temperature (36 degrees C) shows a pleiotropic defect for transport of many different metabolites. The temperature sensitivity of the mutant is closely related to a reduction in phosphofructokinase activity. This conclusion is based on the following criteria. (i) Both the primary isolate, designated P5-9 (ts [rho-] Ino-), which is an inositol auxotroph and respiration deficient, and a purified derivative, SB4 (ts [rho+] Ino+ ), which is respiration competent and capable of growing in the absence of inositol, are temperature sensitive for growth and ethanol production in media containing glucose or fructose as the sole carbon source. (ii) The respiration-competent derivative SB4 is not temperature sensitive in media containing glycerol or glycerol-pyruvate; glucose inhibits its growth at 36 degrees C in these media. (iii) Assays of glycolytic enzymes in P5-9 and SB4 extracts, prepared from cells incubated for 1 to 2 h at 36 degrees C before harvesting, show selective reduction in phosphofructokinase activity. Analysis of tetrads derived from the cross of mutant and nonmutant haploids indicates that temperature sensitivity for growth is due to a single gene or to two closely linked genes. The biochemical analysis of spores from seven such tetrads revealed a uniform cosegregation of temperature sensitivity for growth and phosphofructokinase activity. Transport and ATP levels were drastically reduced in SB4 cells incubated at 36 degrees C for 1 to 2 h with glucose as the carbon source, but not when glycerol-pyruvate or lactate was the energy source. Therefore, depletion of energy as a result of phosphofructokinase inactivation appears to be the cause of the pleiotropic transport defect observed in the mutant.     

Nitrate reductase,nitrite reductase activities and leaf diffusive resistance in wheat under water deficit

The flag and second leaves of wheat showed physiological and biochemical differences in the normal and water stressed plants. Although the flag leaf had lower enzyme activities in the control plants, in stressed plants it could continue nitrate assimilation better than the second and other leaves. The flag leaf also exhibited a higher tendency to resume normal metabolic activity after the release of stress as is indicated by higher NRA, NiRa, lower r_ad and r_ab.     

Purification and properties of indole 2,3-dioxygenase from maize leaves

An indole 2,3-dioxygenase was purified ca 38-fold from maize leaves. The enzyme had an MW of about 98000, an optimum pH of 5.0 and the energy of activation was 9.1 kcal/mol. The K_max for indole was 1.4 × 10^?4 M. The enzyme was inhibited by diethyldithiocarbamate, salicylaldoxime and sodium dithionite. The inhibition by diethyldithiocarbamate was specifically reversed by Cu²⁺. The dialysed enzyme was stimulated by Cu²⁺. Four atoms of oxygen were utilized in the disappearance of 1 mole of indole. Inhibition of the enzyme by -SH compounds and -SH group inhibitors, and their partial removal by Cu²⁺ only, suggested the involvement of -SH groups in binding of Cu²⁺ at the catalytic site.     

Studies on the C-24 configurations of Δ7-sterols in theseeds of Cucurbita maxima

Uncertainties surrounding the structures of the Δ⁷-sterols in the seeds of Cucurbita maxima have been resolved. Seven components were found by TLC, GLC, HPLC, mass spectrometry and ¹H NMR. They were 24β-ethyl-5α-cholesta-7,22,25(27)-trien-3β-ol, 24β-ethyl-5α-cholesta-7,25(27)-dien-3gb-ol, avenasterol, spinasterol, 24-dihydrospinasterol, 24ζ-methyllathosterol and 25(27)-dehydrofungisterol. The ¹H NMR spectra indicated that the sterols with an ethyl substituent at C-24 occurred in the absence of their C-24 epimers. This seems to be the first instance of the detection of 25(27)-dehydrofungisterol in a higher plant.     

Kinetics of reabsorption of nutrients in renal brush border membrane vesicles from rats with experimental ascending pyelonephritis

The uptake of nutrients was investigated in the renal cortical brush border membrane (BBM) vesicles at different stages of ascending pyelonephritis. There was significant difference (p less than 0.05) in the uptake of D-glucose, L-alanine, L-aspartate, L-lysine and L-proline 3 days postinfection and onwards in both right unobstructed and left obstructed experimental kidneys as compared to the sham operated control. The uptake of D-glucose, L-lysine and L-proline was found to be significantly decreased (p less than 0.05) during the course of infection. While uptake of L-alanine and L-aspartate increased (p less than 0.05) in early stages and decreased (p less than 0.05) in later stages of infection. The differential effect was attributed to the compensatory measure and different kinds of transport systems for different types of amino acids.     

Suppression of ouabain-insensitive K-ATPase activity in rabbit nephron segments during chronic hyperkalemia

Recently, we demonstrated that an ATPase stimulated by K (and not inhibited by ouabain, Na-K-ATPase inhibitor) is present in the connecting tubule (CNT) and collecting duct segments of the rabbit. In this study, we determined the effects of high- and low-K diet on K-ATPase activity in the CNT and collecting duct segments of rabbit. One group of animals was given a low-K diet (34 mEq/kg diet) and the other group was given a high-K diet (700 mEq/kg diet) for 1 week. K-ATPase activity was measured by a microfluorometric assay in which ATP hydrolysis is coupled to oxidation of NADH. Low-K animals had plasma K = 3.1 +/- 0.2 as compared with 5.5 +/- 0.5 mEq/l in high-K animals. Low-K animals had significant K-ATPase activity in CNT, CCD (cortical collecting duct) and MCD (medullary collecting duct). On the other hand, K-ATPase activity in all 3 segments from high-K animals was not significantly different from zero. These results support a hypothesis that chronic K loading suppresses the ouabain-insensitive K-ATPase in the distal nephron.