Temperature Dependence of Catecholamine Secretion from Cultured Bovine Chromaffin Cells

Abstract: Secretion of both epinephrine and norepinephrine by cultured chromaffin cells was studied at temperatures ranging from 0°C to 37°C. The percentage of epinephrine secreted was always lower than that of norepinephrine when the cells were stimulated with either acetylcholine or high K⁺ at any temperature. When the cells were stimulated with acetylcholine or carbachol the percentage of catecholamine secreted at 10 min increased with temperature from 4°C to 24°C and then decreased from 24°C to 37°C. Potassium-stimulated cells secreted increasing amounts of catecholamine as the temperature was increased to 37°C. We found, however, that the initial rates of secretion increased continuously as temperature increased throughout the range for both carbachol-and K⁺-stimulated cells. The temperature maximum of acetylcholine-stimulated secretion is caused by a faster shut-off of secretion at higher temperature. The Arrhenius plots of initial rates show an inflection point at approximately 17°C for carbachol-stimulated cells. The plot for K⁺-stimulated cells is a straight line over the entire temperature range. The transition could be caused by a conformational change in the cholinergic receptor/ion channel molecule.     

Specificity and sodium dependence of the active nucleoside transport system in choroid plexus

The transport of [3H]deoxyuridine by the active nucleoside transport system into the isolated rabbit choroid plexus was measured in vitro under various conditions. Choroid plexuses were incubated in artificial CSF containing 1 microM [3H]deoxyuridine and 1 microM nitrobenzylthioinosine for 5 min under 95% O2-5% CO2 at 37 degrees C and the accumulation of [3H]deoxyuridine measured. Nitrobenzylthioinosine was added to the artificial CSF at a concentration (1 microM) that did not inhibit the active nucleoside transport system but did inhibit the separate, saturable nucleoside efflux system. The active transport of deoxyuridine into the choroid plexus depended on Na+ in the medium, as ouabain, substitution of Li+ and choline for Na+, and poly-L-lysine all inhibited deoxyuridine transport. Thiocyanate in place of chloride and penetrating sulfhydryl reagents also inhibited the active transport of deoxyuridine into choroid plexus. The active transport of deoxyuridine into choroid plexus, which is inhibited by naturally occurring ribo- and deoxyribonucleosides (IC50 = 7-21 microM), was not inhibited (IC50 much greater than 150 microM) by nucleosides with certain alterations on the 2', 3', or 5' positions in D-ribose or 2-deoxy-D-ribose (e.g., adenine arabinoside, 3'-deoxyadenosine, xylosyladenosine); or the pyrimidine or purine rings (e.g., 6-azauridine, xanthosine, 7-methylinosine, or 8-bromoadenosine). Other analogues were effective (IC50 = 8-26 microM; e.g., 5-substituted pyrimidine nucleosides, 7-deazaadenosine, 6-mercaptoguanosine) or less effective (IC50 = 46-145 microM; e.g., 5-azacytidine, 3-deazauridine) inhibitors of deoxyuridine transport into the isolated choroid plexus.     

Phylogenetic relationships in theSideritis leucantha group (Lamiaceae)

Six closely related taxa of the sect.Eusideritis of the genusSideritis (S. leucantha, S. pusilla, S. flavovirens, S. granatensis, S. biflora andS. osteoxylla) are analysed to elucidate their phylogenetic relationships and position within the sect.Eusideritis. Meiotic behaviour, karyotype features, size and fertility of pollen grains, DNA amounts and seed protein profiles are reviewed. A polyploid origin of the group (from x = 7) and the further diversification through dysploidy and chromosome repatterning is postulated.S. osteoxylla is apparently of hybrid origin.     

Trichinella spiralis: selective intestinal immune deviation in the rat

In rats, infections with 100-2000 Trichinella spiralis muscle larvae lead to a prompt immunity that is expressed in parasite expulsion within 14 days. Rats infected with more than 2000 larvae display impaired immunity with rejection delayed by 50% (7 days) or more. Suppression is selective for expulsive immunity as the antifecundity response of rats is directly proportional to dose and is expressed sooner in heavily infected subjects. Suppression of intestinal expulsive immunity was suggested by the fact that, with low doses (2000 larvae or less), worm rejection was inhibited by cortisone, whereas cortisone inhibited antifecundity but had no discernable effect on worm rejection in high-dose infections. Evidence for local immune deviation as opposed to systemic immunosuppression was obtained in experiments using parabiotic rats. When one partner was infected with 6000 worms and the other with 200, the rat infected with 200 parasites showed earlier rejection than was seen in single controls infected with 200 worms. The prolonged survival of high-dose adults was not accompanied by a change in the site of worm residence in the gut. Immunological parameters such as serum antibody levels, the number of activated cells or specific anti-T. spiralis lymphocytes in thoracic duct lymph were all increased in a dose-dependent manner. These experiments therefore demonstrate a novel autoprotective mechanism by which adult T. spiralis selectively reduce the expression of expulsive immunity in the gut.     

Sulfate influx across the rabbit ileal brush border membrane: Sodium and proton dependence,and substrate specificities

Summary In intact ileal mucosa, uptake of SO₄ across the brush border membrane requires the presence of Na and is saturable, withK1/2=1.3mm at 140mm Na (P.L. Smith, S.A. Orellana & M. Field, 1981.J. Membrane Biol. 63:199–206). The present study examines the substrate specificities and transport stoichiometry of the Na-dependent SO₄ uptake process. The effects of variations in medium anion and cation composition on lumen-to-epithelium influx of SO₄ (J _me ^SO4 ) were determined under short-circuit conditions.J _me ^SO4 is inhibited by thiosulfate, but not by phosphate, methylsulfate, vanadate or taurocholate. Cl is weakly inhibitory. Uptake of SO₄ is poorly supported by Li, and is unaffected by K, indicating a specific dependence on Na. At low SO₄ concentration (0.22mm),J _me ^SO4 is a hyperbolic function of medium Na concentration; the corresponding Hill plot is linear with a slope of 1.0, suggesting a transport stoichiometry of 1 Na: 1 SO₄. At high SO₄ concentration (6.7mm), the Na-dependent SO₄ velocity curve is sigmoidal and yields a Hill plot which is again linear but has a slope of 1.56, suggesting transport of more than 1 Na per SO₄. SO₄ uptake in presence of Na exhibits a dependence on medium pH. At 0.22mm SO₄ and 140mm Na,J _me ^SO4 was doubled by lowering pH from 7.4 to 6.8. However, at 6.7mm SO₄ and 140mm Na, changing pH had no effect onJ _me ^SO4 over the range 6.8 to 8.5. The pH dependence ofJ _me ^SO4 at 6.7mm SO₄ was restored when medium Na was lowered to 3mm, suggesting that pH sensitivity is a function of the concentration of preformed NaSO ₄ ^– ion pair. The results suggest that SO₄ influx across the ileal brush border occurs by electroneutral Na⁺/NaSO ₄ ^– or Na⁺/H⁺/SO ₄ ^2– cotransport, the former being favored by high concentrations of Na and SO₄.     

Cl− channels in basolateral renal medullary memnbranes: III. Determinants of single-channel activity

Summary We evaluated the effects of vawrying aqueous Cl^– concentrations, and of the arginyl- and lysyl-specific reagent phenylglyoxal (PGO), on the properties of Cl^– channels fused from basolaterally enriched renal medullary vesicles into planar lipid bilayers. The major channel properties studied were the anion selectivity sequence, anionic requirements for, channel activity. and the efects of varying Cl^– concentrations and/or PGO on the relation between holding voltageV _H -mV) and open-time probability (P _o).Reducingcis Cl^– concentrations, in the range 50–320mm, produced a linear reduction in fractional open time (P _v) with a half-maximal reduction inP _o atcis Cl^–170mM. Channel activity was sustained by equimolar replacement ofcis Cl^– with F^–, but not with impermeant isethionate. Fortrans solutions, the relation between Cl^– concentration andP ₀ at 10mm Cl^–. Reducingcis Cl^– had no effect on the gating charge (Z) for channel opening, but altered significantly the voltage-independent, energy (G) for channel opening.Phenylglyoxal (PGO) reducedZ and altered G for Cl^– channel activity when added tocis, but nottrans solutions, Furthermore, in the presence ofcis PGO, reducing thecis Cl^– concentration had no effect onZ but altered G. Thus we propose thatcis PGO and,cis Cl^– concentrations affect separate sites determining channel activity at the extracellular faces of, these Cl^– channels.     

Barium- and calcium-permeable channels open at negative membrane potentials in rat ventricular myocytes

Summary Ca²⁺- and Ba²⁺-permeable channel activity from adult rat ventricular myocytes, spontaneously appeared in the three single-channel recording configurations: cell-attached, and excised inside-out or outside-out membrane patches. Single-channel activity was recorded at steady-state applied membrane potentials including the entire range of physiologic values, and displayed no rundown in excised patches. This activity occurred in irregular bursts separated by quiescent periods of 5 to 20 min in cell-attached membrane patches, whereas in excised patch experiments, this period was reduced to 2 to 10 min. During activity, a variety of kinetic behaviors could be observed with more or less complex gating patterns. Three conductance levels: 22, 45 and 78 pS were routinely observed in the same excised membrane patch, sometimes combining to give a larger level. These channels were significantly permeable to divalent cations and showed little or no permeability to potassium or sodium ions. The inorganic blockers of voltage-gated Ca channels, cobalt (2mm), cadmium (0.5mm) or nickel (3mm), had no apparent effect on these spontaneous unitary currents carried by barium ions. Under 10^–5 m bay K 8644 or nitrendipine, the activity was clearly increased in about half of the tested excised inside-out membrane patches. Both dihydropyridines enhanced openings of the larger conductance level, which was only very occasionally seen under control conditions. When the single-channel activity became sustained under 5×10^–6 m Bay K 8644, it was possible to calculate the mean unitary current at different membrane potentials and show that the mean current value increased with membrane potential.     

Effects of dl-2-Amino-5-Phosphonovalerate on Metabolism of Catecholamines in Synaptosomes from Rat Brain

Incubation of synaptosomes from rat brain with DL-2-amino-5-phosphonovalerate (APV) stimulated an increased release of dopamine, and this effect was strictly dependent on the extrasynaptosomal calcium level. APV increased biosynthesis of dopamine from tyrosine by 30%, whereas monoamine oxidase activity was inhibited by 30%. When synaptosomes were incubated with radioactive dopamine, APV caused a large decrease in incorporation of label into 3,4-dihydroxyphenylacetic acid but greatly increased incorporation into norepinephrine and its N-methyl derivatives. Quantification of dopamine and its metabolites in synaptosomes, using electrochemical detection, indicated that the presence of APV resulted in changes in the absolute levels of the aforementioned dopamine metabolites similar to the changes in radiolabel incorporation. Omission of Ca2+ from the extrasynaptosomal medium greatly diminished the APV-induced changes in catecholamine metabolism. The metabolic changes appear to largely result from an increased intrasynaptosomal Ca2+ level due to the APV-induced increase in calcium permeability of the plasma membrane.     

Studies on zinc- and cadmium-bound proteins in bovine kidneys by biochemical and neutron activation techniques

Bovine kidneys were found to contain about 78 ppm Zn and 0.78 ppm Cd. Approximately 45% of Zn and 60% of Cd were present in the cytosol fraction. More than 95% of these two metals were bound to macromolecules. Both Zn- and Cd-protein complexes were observed to be stable between pH 7 and 10.5. Separation and characterization of these proteins were carried out using several chromatographic and electrophoretic techniques in conjunction with instrumental neutron activation analysis (INAA). The results showed the presence of at least four Zn-binding proteins with mol wt>300,000, 260,000, 89,000, and 27,000 and at least three Cd-binding proteins of mol wt>300,000, 32,000, and 13,000.     

Sustained biological control of the cassava mealybugPhenacoccus manihoti [Hom.: Pseudococcidae] byEpidinocarsis lopezi [Hym.: Encyrtidae] in Nigeria

Following the successful introduction ofEpidinocarsis lopezi (De Santis) for biological control of the cassava mealybug (CM)Phenacoccus manihoti Mat.-Ferr. in southwestern Nigeria in 1981 and 1982, 11 groups of cassava fields were sampled every 2 weeks up to 1988 for impact assessment. After 1984, CM populations remained mostly below 10 per tip despite the presence of native hyperparasitoids, demonstrating the long-term success of biological control byE. lopezi in the region. Indigenous polyphagous coccinellids were found only during peak host densities, whereas the specificE. lopezi was common throughout the year. During some periods, percentage parasitism indicated delayed density dependence. Since 89% of all sampled cassava tips had no CM at all and the parasitisme is very mobile, parasitization rates were also calculated for individual infested tips (N=4,878). Parasitism increased slightly with host density on tips having between 1 and 10 CM of the 3^rd and 4^th instars, indicating positive density dependence. Such tips comprised 64% of all infested tips. At higher host densities, parasitism rates fell rapidly. The results are discussed in view of different theories on population regulation by biological control agents.