Release of Purines, Noradrenaline, and GABA from Rat Hippocampal Slices by Field Stimulation

Labelled adenine, noradrenaline (NA), and gamma-aminobutyric acid (GABA) were taken up by the transversely cut hippocampal slice. [3H]NA and [14C]GABA were retained as such, [3H]- (or [14C]-) adenine mainly as adenine nucleotides. There was a spontaneous overflow of all three types of compounds ranging from 0.1 (GABA) to 0.21 (NA) %/min. The rate of [3H]NA overflow increased rapidly during electrical field stimulation. The release rate was well maintained over a 15-min period. The rate of [14C]GABA release also increased rapidly but it was not maintained over a 15-min period even if uptake and/or metabolism was inhibited by nipecotic acid (1 mM) and aminooxyacetic acid (AOAA, 0.1 mM). The bulk of the purines was released after the stimulation period. For all compounds the amounts released were frequency- and calcium-dependent. At a frequency of 3 Hz a 10 V stimulation was sufficient to cause a maximal [3H]NA release and 20 V to cause maximal [14C]GABA release, but 14C-purine release was increased further by increasing the voltage to 40 V. The evoked purine release was inhibited by a nucleoside uptake inhibitor (dipyridamole). On stimulation of [3H]NA-labelled slices the released radioactivity was composed of greater than 95% unchanged NA. The specific activities of NA in the slice and in the superfusate were practically identical. In [3H]adenine-labelled slices the released radioactivity was composed of adenosine, inosine, and hypoxanthine, but the activity in the slice of ATP, ADP, and AMP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Arachidonic Acid Release and Catecholamine Secretion from Digitonin-Treated Chromaffin Cells: Effects of Micromolar Calcium, Phorbol Ester, and Protein Alkylating Agents

The relationship between catecholamine secretion and arachidonic acid release from digitonin-treated chromaffin cells was investigated. Digitonin renders permeable the plasma membranes of bovine adrenal chromaffin cells to Ca2+, ATP, and proteins. Digitonin-treated cells undergo exocytosis of catecholamine in response to micromolar Ca2+ in the medium. The addition of micromolar Ca2+ to digitonin-treated chromaffin cells that had been prelabeled with [3H]arachidonic acid caused a marked increase in the release of [3H]arachidonic acid. The time course of [3H]arachidonic acid release paralleled catecholamine secretion. Although [3H]arachidonic acid release and exocytosis were both activated by free Ca2+ in the micromolar range, the activation of [3H]arachidonic acid release occurred at Ca2+ concentrations slightly lower than those required to activate exocytosis. Pretreatment of the chromaffin cells with N-ethylmaleimide (NEM) or p-bromophenacyl bromide (BPB) resulted in dose-dependent inhibition of 10 microM Ca2+-stimulated [3H]arachidonic acid release and exocytosis. The IC50 of NEM for both [3H]arachidonic acid release and exocytosis was 40 microM. The IC50 of BPB for both events was 25 microM. High concentrations (5-20 mM) of Mg2+ caused inhibition of catecholamine secretion without altering [3H]arachidonic acid release. A phorbol ester that activates protein kinase C, 12-O-tetradecanoylphorbol-13-acetate (TPA), caused enhancement of both [3H]arachidonic acid release and exocytosis. The findings demonstrate that [3H]arachidonic acid release is stimulated during catecholamine secretion from digitonin-treated chromaffin cells and they are consistent with a role for phospholipase A2 in exocytosis from chromaffin cells. Furthermore the data suggest that protein kinase C can modulate both arachidonic acid release and exocytosis.     

Cytological studies of triploids and their progeny from male-sterile (ms1) soybean

Summary Triploids (2n=3X=60) were obtained from genetic male-sterile (ms1 ms1) soybean [Glycine max (L.) Merr.] plants. Meiosis, pollen fertility, and chromosome number of their progeny were studied. Studies of meiosis in fertile and sterile triploids revealed no distinguishable differences in chromosome associations. Male-sterile plants formed coenocytic microspores characteristic of the ms1 mutant. Restitution of some dyad and tetrad nuclei were observed in male-sterile plants. Chromosomes of the triploids tended to occur in trivalents during diakinesis and metaphase I (MI), but multivalents, bivalents, and univalents also were observed. Average types and frequencies of chromosome associations per cell in diakinesis and MI from 542 pollen mother cells were 0.004 IX + 0.06 VI + 0.002 V + 0.005 IV + 16.99 III + 1.79 II + 5.03 I. Some secondary associations, nonhomologous pairing, and aberrant nucleolar distributions occasionally were observed. Such behavior support the hypothesis of duplicated genomes and the polyploid origin of soybean. Pollen fertility in male-fertile triploid plants (Ms1 ms1 ms1) varied from 57% to 82%, with an average of about 71%. Chromosome numbers of progenies obtained from these fertile triploids varied from 2n=40 to 2n=71, and exhibited a near-random distribution, with the majority (about 60%) being between 56 and 65. Progenies of the fertile triploids gave segregation ratios for the ms1 allele, which confirmed the Ms1 ms1 ms1 genotype.Joint contribution: Agricultural Research Service, U.S. Department of Agriculture, and Journal Paper No. J-11672 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA 50011, USA, Project 2471     

Presynaptic modulation by noradrenaline and an opioid of the substance P-induced release of [3H]acetylcholine from the myenteric plexus

Substance P (7.5-750 nM) applied in superfusion dose-dependently released 3H from isolated strips of myenteric plexus-longitudinal muscle of the guinea-pig ileum loaded with [3H]choline. Separation of the [3H]acetylcholine and [3H]choline components of the released radioactivity revealed that in response to substance P (SP) administration only the release of [3H]acetylcholine increased above resting level. A slowly developing tachyphylaxis to the effect of SP was observed. Evidence has been obtained that the slow tachyphylaxis developed to the acetylcholine-releasing effect of SP was not due to the exhaustion of releasable acetylcholine pool. Release of acetylcholine by 150 nM SP was completely prevented by tetrodotoxin or in a Ca2+-free medium and greatly reduced in the presence of noradrenaline or the opioid receptor agonist (D-Met2,Pro5)-enkephalinamide. The effect of noradrenaline and the opioid peptide was apparently prevented by yohimbine and naloxone, respectively.     

Radioimmunoassay for fibroblast growth factor (FGF): release by the bovine anterior pituitary in vitro

A radioimmunoassay (RIA) was developed to measure fibroblast growth factor (FGF) using antiserum generated against a synthetic replicate of [Tyr¹⁰]FGF(1–10). The antisera, previously shown to be capable of inhibiting the biological action of FGF on bovine aortic arch endothelial cells in vitro [1], are highly specific for the amino-terminus of FGF. In the RIA, the antisera recognize the decapeptide antigen [Tyr¹⁰]FGF(1–10) and the intact mitogen on an equimolar basis and show less than 0.01% cross-reactivity with N-acetyl-[Tyr¹⁰]FGF(1–10).

Bovine adenohypophysial cells maintained in primary monolayer culture release and ir-FGF which is indistinguishable from the intact mitogen in as much as it is retained on heparin-Sepharose affinity columns and shows a dose-dependent and parallel displacement in RIA. The release of ir-FGF by the bovine adenohypophysis can be increased with forskolin (10⁻⁵ M) or KCl (50 mM). Preincubation of pituitary cells with 17β-estradiol has no measurable effects on basal ir-FGF, but increases the release after KCl treatment 2–3-fold. These results show that ir-FGF can be released by the bovine adenohypophysis in vitro and lend credence to the hypothesis that FGF plays a physiological role in the homeostatic mechanisms regulating mesoderm-derived cell growth.     

Positioning of protein-coding genes on the soybean chloroplast genome

Seven major plastid protein encoding genes were positioned on the soybean chloroplast DNA by heterologous hybridization. These include the genes for the alpha, beta and epsilon subunits of the CF₁ component of ATP synthase (atpA, atpB and atpE respectively), for subunit III of the CF₀ component of ATP synthase (atpH), for the cytochrome f (cytF), for the ‘32 Kd’ thylakoid protein (psbA), and for the large subunit of ribulose-1,5-bisphosphate carboxylase-oxygenase (rbcL), all of which map in the large single copy region. The atpB, atpE and rbcL genes are located in the region adjacent to one of the segments of the inverted repeat. The genetic organization of the soybean chloroplast DNA is compared to that of other plastid genomes.     

Peptostreptococcus barnesae sp. nov., a Gram-positive,anaerobic, obligately purine utilizing coccus from chicken feces

Anaerobic, Gram-positive cocci were obtained from chicken feces by direct isolation, which grew on the purines uric acid, xanthine, 6,8-dihydroxypurine, guanine, and hypoxanthine. Adenine and glycine were fermented, but not as readily. Acetate, formate, ammonia, and CO₂ were products. The isolated strains were nutritionally non-fastidious, however, they required selenite, molybdate, and tungstate as micronutrients. The cells were spherical and 0.5–0.9 m in diameter. The addition of bile salts enhanced the growth rate in most cases. The organisms proved to be quite resistant to lysis. The guanosine-plus-cytosine (G+C) content of their deoxyribonucleic acid was 33.6 to 34.8 mol%. The peptidoglycan was of the same structure (Gly-Lys-d-Asp) as reported for the anaerobic cocci of Hare group IX. However, the latter strains could only utilize glycine, not purines. Therefore, it is proposed to form a new species, Peptostreptococcus barnesae sp. nov.This paper is dedicated to Prof. Dr. Norbert Pfennig on the occasion of his 60th birthday     

一种快速、灵敏的血小板释放功能检测方法及其在抗血小板药物研究中的应用

本文报道了一种快速、灵敏的血小板释放功能检测方法:利用荧光素-荧光素酶在有ATP、Mg~(2+)、O_2存在时产生的生物发光素测定血小板ATP的释放量,以反映血小板的释放功能;研究了ADP、AA、胶原、凝血酶等四种诱导剂对血小板释放功能的作用,发现ADP的诱导释放能力较其他三者为弱;观察在不同剂量ADP和AA的诱导下,血小板聚集强度和释放能力之间的关系,研究了血小板数等因素对ATP释放功能测定的影响。应用该方法研究了Aspirin及活血化淤药物川芎嗪,毛冬青甲素对血小板释放功能的影响,发现Aspirin对AA诱导的释放反应有强烈的抑制作用。在以ADP诱导的释放反应中,川芎嗪的抑制作用较毛冬青甲素更为强烈。     

Effects of ultraviolet-B radiation on plants during mild water stress. IV. The insensitivity of soybean internal water relations to ultraviolet-B radiation

The combined effects of ultraviolet-B (UV-B, 280–320 nm) radiation and water stress were investigated on the water relations of greenhouse grown soybean [ Glycine max (L.) Merr. cv. Essex]. On a weighted (Caldwell 1971), total daily dose basis, plants received either 0 or 3 000 effective J m² UV-B_BE supplied by filtered FS-40 sunlamps. The latter dose simulated the solar UV-B radiation anticipated at College Park, Maryland, U.S.A. (39°N latitude) in the event that the global stratospheric ozone column is reduced by 25%. Plants were either well-watered or preconditioned by drought stress cycles. Diurnal measurements of water potential and stomatal conductance were made on the youngest fully expanded leaf. Various internal water relations parameters were determined for detached leaves. Plants were monitored before, during and after water stress. There were no significant differences in leaf water potential or stomatal conductance between treatments before plants were preconditioned to water stress. However, drought stress resulted in significantly lower midday and afternoon leaf water potentials and lower leaf conductances as compared to well-watered plants. UV-B radiation had no additional effect on leaf water potential; however, UV did result in lower leaf conductances in plants preconditioned to water stress. Turgid weight:dry weight ratio, elastic modulus, bound water and relative water content were unaffected by UV-B radiation. Osmotic potentials at full and zero turgor were significantly lower in the drought stressed treatments as compared to well-watered plants.     

Effects of ultraviolet-B radiation on plants during mild water stress. III. Effects on photosynthetic recovery and growth in soybean

Soybean { Glycine max (L.) Merr. ev. Essex} was grown from seed in a greenhouse under ultraviolet-B (UV-B, 280–320 nm) radiation supplied by filtered FS-40 sunlamps. On a weighted, total daily dose basis these plants received either 0 (control) or 2875 effective J m⁻² day⁻¹ UV-B_BE. When weighted with the generalized plant action spectrum (Caldwell 1971), this simulated the solar ultraviolet-B irradiance expected to occur at College Park, Maryland, USA (39°N) in the event the global stratospheric ozone column is reduced by 23%. The effects of ultraviolet radiation on the photosynthetic recovery from water stress were measured with an infrared gas analyzer. These effects were examined in plants which were either well-watered or previously preconditioned to water stress, during two distinct phenological stages of development. During the early stages of soybean growth, enhanced levels of UV-B reduced net photosynthesis by 25%, and water stress also reduced photosynthesis to nearly the same extent (by 20%). The combination of these two stresses resulted in smaller biomass than that produced by plants exposed to either stress independently. Photosynthesis in older, larger plants was much more sensitive to water stress and was reduced by as much as 50–60% in non-preconditioned plants. Although non-irradiated, non-preconditioned (control) plants recovered to only within 60% of their prestressed value, preconditioned plants recovered to within 70–80% during the 3 day recovery period. Both water stress and UV-B radiation affected non-stomatal conductance, while stomatal conductance was primarily affected by water stress.