Regulation of Renal Na-HCO3 Cotransporter: VIII. Mechanism of Stimulatory Effect of Respiratory Acidosis

We examined the effect of respiratory acidosis on the Na-HCO₃ cotransporter activity in primary cultures of the proximal tubule of the rabbit exposed to 10% CO₂ for 5 min, 2, 4, 24 and 48 hr. Cells exposed to 10% CO₂ showed a significant increase in Na-HCO₃ cotransporter activity (expressed as % of control levels, 5 min: 142 ± 6, 2 hr: 144 ± 13, 4 hr: 145 ± 11, 24 hr: 150 ± 15, 48 hr: 162 ± 24). The increase in activity was reversible after 48 hr. The role of protein kinase C (PKC) on the stimulatory effect of respiratory acidosis on the cotransporter was examined in presence of PKC inhibitor calphostin C or in presence of PKC depletion. Both calphostin C and PKC depletion prevented the effect of 10% CO₂ for 5 min or 4 hr to increase the activity of the cotransporter. 10% CO₂ for 5 min or 4 hr increased total and particulate fraction PKC activity. To examine the role of phosphotyrosine kinase (PTK) on the increase in cotransporter activity we studied the effect of two different inhibitors, 2-hydroxy-5-(2,5-dihydroxylbenzyl) aminobenzoic acid (HAC) and methyl 2,5-dihydroxycinnamate (DHC) which inhibit phosphotyrosine kinase in basolateral membranes. Cells were pretreated either with vehicle or HAC or DHC and then exposed to 10% CO₂ for 5 min or 4 hr. In cells treated with vehicle, 10% CO₂ significantly increased cotransporter activity as compared to control cells exposed to 5% CO₂. This stimulation by 10% CO₂ was completely prevented by HAC or DHC at 5 min (5% CO₂: 1.8 ± 0.2, 10% CO₂: 2.6 ± 0.2, 10% CO₂+ HAC: 1.6 ± 0.2, 10% CO₂: +DHC: 2.0 ± 0.3 pH unit/min) and also at 4 hr. The protein synthesis inhibitors actinomycin D and cycloheximide appear to prevent the effect of 10% CO₂ for 4 hr on the cotransporter. Our results show that early respiratory acidosis stimulates the Na-HCO₃ cotransporter through PKC and PTK-dependent mechanisms and the late effect appears to be mediated through protein synthesis. Received: 28 March 1997/Revised: 22 December 1997     

Activation of yeast mannan synthetase by alpha factor pheromone

The first mannose of O-linked oligomannose chains in S. cerevisiae is transferred to Ser/Thr residues via dolichylphosphate mannose. Only this reaction (and not the subsequent reactions requiring GDP-Man) proceeds at the endoplasmic reticulum.     

Protective Effect of Adonitol on Lactic Acid Bacteria Subjected to Freeze-Drying

The protective effects of glycerol, adonitol, and four other related polyhydric alcohols on lactic acid bacteria subjected to freeze-drying were examined. The presence of adonitol in the suspending medium markedly protected the viabilities of the 12 stains tested. Dulcitol, mannitol, m-inositol, and sorbitol were found to provide little or no protection.     

Diurnal variations of rat liver enzymes catalyzing cholesterol ester hydrolysis

Cholesterol ester hydrolase activity was determined at 3 h time intervals over 24 h in lysosomes, cytosol and microsomes from ad libitum-fed and 24 h food-deprived female rat liver. Diurnal rhythms were identified for the acid and neutral esterases, which were strikingly changed by fasting. In fed animals, lysosomal esterase specific activity exhibited a peak at noon and a sustained medium rate at early darkness, whereas total esterase was maximal at midnight. The circadian patterns of the cytosolic and the microsomal esterases paralleled each other, though the amplitude of rhythms differed, showing higher activities around midnight. After fasting, cholesterol esterase activity from all cell fractions reached a maximum near dark onset. These results are the first to indicate that cholesteryl ester hydrolysis may play a role in generating the diurnal rhythm of hepatic cholesterol.     

Circadian and seasonal variations in pineal gland intercellular canaliculi in the white rat.

Seventy Wistar rats are used to study the changes in pineal intercellular canaliculi over a 21-hour period and for two different photoperiods (pre-autumn, first week of September, and winter, first week of February). The study considers these changes at pineal body, cortical and medullar level separately, and compares the values obtained. The results show variations in canalicular surface at different point times (10:00, 14:00, 18:00) and for both photoperiods. The variations are found to favour the cortical layer, and are also observed between nocturnal and diurnal hours. Canalicular surface to greater during the diurnal hours of both photoperiods. Interesting histological findings are described that suggest an important function of the intercellular canaliculi in pineal gland metabolic exchange.     

Kinetics of cellular proliferation and hormonal receptors in EVSA-T breast cancer cell line.

The hormone-independent human breast cancer cell line EVSA-T, originally described as negative estrogen and progesterone receptors is shown to become positive hormone receptors when the cellular proliferation rate is slowed down. The experimental procedure included the following steps: 1) EVSA-T cells were seeded in minimum essential medium supplemented with 10% fetal bovine serum and kept undisturbed for 2 days; 2) culture medium was replaced with Dulbecco's solution and Ham's F-12 and cells were incubated in serum-free media for another 24 h; 3) then, cells were "rescued" with 10% FBS supplemented medium and estrogen (ER) and progesterone receptors (PgR) were measured immediately, time 0, and 6, 12, 18, 24 and 30 h after the media were changed. Cell yield was quantified at the same times. Experimental data indicate that changing the proliferation kinetics makes it possible to detect estradiol and progesterone receptors on EVSA-T cells. Estrogen receptor appeared at 18 h after rescue, 6 hours before progesterone receptor could be detected. Immunohistochemical analysis of ER content confirmed this observation, showing maximal positive stain at 18 h. Furthermore, ER disappeared when cells recovered their normal proliferation rate.     

Birefringence of macromolecules. Wiener''s theory revisited, with applications to DNA and tobacco mosaic virus.

We summarize Wiener's theory of the dielectric constant of heterogeneous systems and extend its application to suspensions of particles with corrugated surfaces and interstitial solvent. We retain a simple geometrical shape for the particles and account specifically for the solvent associated with the particles. We calculate the birefringence of the rodshaped Tobacco Mosaic Virus (TMV) particle and of DNA and find excellent agreement between our numerical results and experimental values from the literature.     

Effect of alpha factor pheromone on the activity of enzymes which catalyze the synthesis and hydrolysis of cell wall structural polymers from Saccharomyces cerevisiae

Cells of Saccharomyces cerevisiae of the a mating type treated with alpha factor contain an increased amount of structural polymers (beta-glucans and chitin) in their cell walls and, consequently, exhibit higher glucan synthetase and chitin synthetase activities than untreated cells. However, alpha factor has no detectable effect on the activities of these enzymes when they are assayed, "in vitro", in the presence of the pheromone. On the other hand, the activity of beta-glucanases remains constant during the time that growth of a cells is kept arrested by alpha factor at the G1 phase of the cell division cycle and starts to increase when budding of the cells is reinitiated.     

R1881 regulation of steroidogenesis in cultured testicular cells

The influence of a synthetic androgen R1881 upon hCG stimulated steroidogenesis in cultured rat testicular cells was investigated. Testicular cells were cultured for 8 days in medium alone and thereafter reincubated for 48 h with appropriate treatments before the collection of media for steroid RIA. Addition of R1881 (10(-6) M) resulted in an overall decrease of hCG (0.3-10 ng/ml) stimulated pregnenolone and progesterone production by cultured cells. The conversion of exogenous steroids of the delta 4 pathway (progesterone,17 alpha-OH-P and delta 4-A) was also studied in cultures supplemented with cyanoketone (10(-5) M) and/or spironolactone (10(-5) M) to prevent endogenous testosterone production. R1881 inhibited progesterone and 17 alpha-OH-P conversion to testosterone (T) and was ineffective when delta 4-A served as precursor for T biosynthesis. The inhibitory effect of R1881 upon Testosterone production was prevented by concomitant treatment with CPA. These observations suggest that R1881 decreases the hCG stimulated testosterone production via inhibition of CSCCE,3 beta-HSD,C17-20 Lyase and likely 17 alpha-Hydroxylase, whereas no effect on 17 beta-HSD could be observed.     

Role ofβ-oxidation in inhibitingLactobacillus leichmanii by fatty acids

The effect of saturated fatty acids from 6∶0 to 16∶0 and oleic acid onLactobacillus leichmanii ATCC 4797 growing in non-skim-milk media was determined. The inhibition by lauric acid was higher than that obtained with any other fatty acid. A mutant (MC12) resistant to the fatty acid inhibition with high β-oxidation activity was also studied. A positive correlation between the ability ofL. leichmanii ATCC 4797 and its derivative MC12 to degrade fatty acids and their resistance to the fatty acid inhibition is shown in this report.