Evidence for Ascending and Descending Intraspinal as Well as Primary Sensory Somatostatin Projections in the Rat Spinal Cord

Somatostatin distribution was measured quantitatively in the rat spinal cord by radioimmunoassay. Rostro-caudally, somatostatin content was about 50% higher in lumbar-sacral cord than in cervical or thoracic levels. The dorso-ventral distribution is more uneven: somatostatin is highest in the dorsal horn, where the peptide is 15 times as concentrated as it is in the ventral white matter, the region of lowest concentration. However, measurable amounts of the peptide were found in all regions studied. Dorsal root ganglionectomy decreased somatostatin levels in the dorsal cord, supporting the previously proposed role for this peptide as a primary sensory neurotransmitter or modulator; but somatostatin content also was decreased both rostral and caudal to spinal transection, indicating the presence of ascending and descending somatostatin pathways within the spinal cord. Brain levels did not change. Met-enkephalin and substance P were also measured after the above surgical manipulations. Met-enkephalin content was not altered and substance P content was lowered significantly only after ganglionectomy. Although this study confirms the primary sensory neuron as the origin of a part of spinal cord somatostatin, it further indicates the presence of ascending and descending somatostatin pathways within the rat spinal cord.     

Sulfite-induced lipid peroxidation in chloroplasts as determined by ethane production

Ethane formation, as a measure of lipid peroxidation, was studied in spinach (Spinacia oleracea L.) chloroplasts exposed to sulfite. Ethane formation required sulfite and light, and occurred with concomitant oxidation of sulfite to sulfate. In the dark, both ethane formation and sulfite oxidation were inhibited. Ethane formation was stimulated by ferric or ferrous ions and inhibited by ethylenediamine tetraacetate. The photosynthetic electron transport modulators, 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) and phenazine methosulfate, inhibited both sulfite oxidation and ethane formation. Methyl viologen greatly stimulated ethane formation, but had little effect on sulfite oxidation. Methyl viologen, in the absence of sulfite, caused only a small amount of ethane formation in comparison to that produced with sulfite alone. Sulfite oxidation and ethane formation were effectively inhibited by the radical scavengers, 1,2-dihydroxybenzene-3,5-disulfonic acid and ascorbate. Ethanol, a hydroxyl radical scavenger, inhibited ethane formation only to a small degree; formate, which converts hydroxyl radical to superoxide radical, caused a small stimulation in both sulfite oxidation and ethane formation. Superoxide dismutase inhibited ethane formation by 50% when added at a concentration equivalent to that of the endogenous activity. Singlet oxygen did not appear to play a role in ethane formation, inasmuch as the singlet oxygen scavengers, sodium azide and 1,4-diazobicyclo-[2,2,2]-octane, were not inhibitory. These data are consistent with the view that O₂ is reduced by the photosynthetic electron transport system to superoxide anion, which in turn initiates the free radical oxidation of sulfite, and the free radicals produced during sulfite oxidation were responsible for the peroxidation of membrane lipids, resulting in the formation of ethane.     

Dark metabolism of carbon monoxide in lettuce leaf discs

In the dark, leaf tissue of crisphead lettuce (Lactuca sativa L.) metabolized ¹⁴CO to ¹⁴CO₂ and acid-stable products. Tissue incubated at 2.5°C for 3.5 hours and 48 hours converted about 1% and 17%, respectively, of the applied ¹⁴CO to ¹⁴CO₂, and incorporated about 0.04% and 0.6% of the ¹⁴C in acid-stable products. Examination of soluble acid-stable products from ¹⁴CO and ¹⁴CO₂-treated leaf tissue revealed that the labeling patterns of both treatments were identical during the 3.5-hour and the 48-hour incubation periods. Malate, citrate, and aspartate together comprised 70% or more of the soluble radioactivity from both treatments. Incorporation of radioactivity from CO into soluble acid-stable products during a 3-hour incubation period at 2.5°C was inhibited 90% by adding 3% nonradioactive CO₂. These results indicate that in head lettuce in the dark ¹⁴CO is metabolized primarily to ¹⁴CO₂ which is the precursor of acid-stable products. In leaf discs at 2.5°C, the apparent K_m for CO oxidation to CO₂ was 5.3 microliters per liter and the V_max was 9.7 nanoliters per gram per hour. The mitochondrial fraction of the leaf homogenate was the most active fraction to oxidize CO to CO₂, and this activity was heat-labile and cyanide-sensitive.     

Inhibition of ethylene synthesis in tomato plants subjected to anaerobic root stress

Enhanced ethylene production and leaf epinasty are characteristic responses of tomato (Lycopersicon esculentum Mill.) to waterlogging. It has been proposed (Bradford, Yang 1980 Plant Physiol 65: 322-326) that this results from the synthesis of the immediate precursor of ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC), in the waterlogged roots, its export in the transpiration stream to the shoot, and its rapid conversion to ethylene. Inhibitors of the ethylene biosynthetic pathway are available for further testing of this ACC transport hypothesis: aminooxyacetic acid (AOA) or aminoethoxyvinylglycine (AVG) block the synthesis of ACC, whereas CO²⁺ prevents its conversion to ethylene. AOA and AVG, supplied in the nutrient solution, were found to inhibit the synthesis and export of ACC from anaerobic roots, whereas Co²⁺ had no effect, as predicted from their respective sites of action. Transport of the inhibitors to the shoot was demonstrated by their ability to block wound ethylene synthesis in excised petioles. All three inhibitors reduced petiolar ethylene production and epinasty in anaerobically stressed tomato plants. With AOA and AVG, this was due to the prevention of ACC import from the roots as well as inhibition of ACC synthesis in the petioles. With Co²⁺, conversion of both root- and petiole-synthesized ACC to ethylene was blocked. Collectively, these data support the hypothesis that the export of ACC from low O₂ roots to the shoot is an important factor in the ethylene physiology of waterlogged tomato plants.     

Stimulation of ethylene production in citrus leaf discs by mannitol

Wound ethylene formation induced in leaf tissue of citrus (Citrus sinensis [L.] Osbeck cv. “Washington Navel”) by excision was significantly stimulated by mannitol after a lag period of about 6 hours. The extent of stimulation was dependent upon the concentration of mannitol (10 to 100 millimolar). This increased ethylene production was not simply due to osmotic effect or water stress as other osmoticums tested failed to exert such an effect. The stimulatory effect of mannitol resulted from both the enhancement of 1-aminocyclopropane-1-carboxylic acid (ACC) formation and the conversion of ACC to ethylene. The effect on the latter step was particularly pronounced in aged discs. The use of labeled mannitol showed that it was taken up by the leaf discs, utilized for respiration, and metabolized to sucrose, but no radioactivity was detected in the ethylene.     

Relationship between excision repair and the cytotoxic and mutagenic effect of the ‘anti’ 7,8-diol-9,10-epoxide of benzo[a]pyrene in human cells

The cytotoxic and mutagenic effect of (±)-7β,8α-dihydroxy-9α,10α-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (anti BPDE) in normally excision diploid human cells treated just prior to onset of S was compared with that of cells allowed ～ 16 h for excision repair before onset of S and with that observed in excision-deficient serodema pigmentosum (SP12BE) cells. The cells were synchronized by release from density inhibition of cell replication. DNA synthesis began ～ 22 h after the cells were plated at lower density (i.e., 1.4 × 10⁴ cells/cm²). The frequency of thioguanine-resistant mutants induced in normal cells treated just prior to onset of S was ～ 12- to 16-fold higher than that observed in cells treated in early G₁ or treated in G₀ (confluence) and then plated at lower density. The frequency approximated that expected for XP12BE cells from extrapolation of data obtained at lower doses. The frequency of mutants measured in normal cells treated in exponential growth was also much higher than that in the cells treated in early G₁ or in G₀, No such difference could be seen in XP12BE cells treated in exponential growth or in G₀. In contrast to the mutagenicity data in the normal cells, there was no significant difference in the slope of the survival curve of normal cells treated at various times prior to S phase at low densities. However, normal cells treated even at the onset of S exhibited survival equal to XP12BE cells give a 4- to 5-fold lower dose. The data support the hypothesis that DNA synthesis is the cellular event which converts unexcised DNA lesions into mutations. However, they indicate that S is not the event primarily responsible for translating DNA damage into cell death. Accompanying studies on the rate of excision of anti BPDE adducts from the normal cells during the period priot to S support the conclusions.     

The effect of plant-hormone pretreatments on ethylene production and synthesis of 1-aminocyclopropane-1-carboxylic acid in water-stressed wheat leaves

Excised wheat (Triticum aestivum L.) leaves, when subjected to drought stress, increased ethylene production as a result of an increased synthesis of 1-aminocyclopropane-1-carboxylic acid (ACC) and an increased activity of the ethyleneforming enzyme (EFE), which catalyzes the conversion of ACC to ethylene. The rise in EFE activity was maximal within 2 h after the stress period, while rehydration to relieve water stress reduced EFE activity within 3 h to levels similar to those in nonstressed tissue. Pretreatment of the leaves with benzyladenine or indole-3-acetic acid prior to water stress caused further increase in ethylene production and in endogenous ACC level. Conversely, pretreatment of wheat leaves with abscisic acid reduced ethylene production to levels produced by nonstressed leaves; this reduction in ethylene production was accompanied by a decrease in ACC content. However, none of these hormone pretreatments significantly affected the EFE level in stressed or nonstressed leaves. These data indicate that the plant hormones participate in regulation of water-stress ethylene production primarily by modulating the level of ACC.Abbreviations ABA abscisic acid - ACC 1-aminocyclopropane-1-carboxylic acid - BA N⁶-benzyladenine - EFE ethylene-forming enzyme - IAA indole-3-acetic acid     

Release of heat pretreatment-induced dormancy in lettuce seeds by ethylene or cytokinin in relation to the production of ethylene and the synthesis of 1-aminocyclopropane-1-carboxylic acid during germination

The germination of lettuce (Lactuca sativa L.) seeds was greatly reduced when the seeds were heated at 97°C for 30 h prior to imbibition. This dormancy was effectively released when ethylene (1–100 ppm) or benzyladenine (BA) (0.005–0.05 mM) was applied during the imbibition period. Ethylene was not required during the early part of imbibition, but was essential during the period immediately prior to radicle protrusion. Treatment with 1-aminocyclopropane-1-carboxylic acid (ACC) (0.1–10 mM) stimulated germination, but was not as effective as ethylene or cytokinin treatment. During the germination of nondormant lettuce seeds, ethylene production increased rapidly and reached a peak at 24 h, which coincided with the emergence of the radicle, and then declined; the level of ACC increased as ethylene production rate increased, but remained at a high level after radicle protrusion. In heat-pretreated dormant lettuce seeds, the increases in percent germination, ethylene production, and ACC levels were all delayed and lower than those of nondormant seeds, and these increases were accelerated by treatment with ethylene or cytokinin.     

Regio- and stereo-selective metabolism of 4-methylbenz[a]anthracene by the fungus Cunninghamella elegans.

Metabolism of 4-methylbenz[a]anthracene by the fungus Cunninghamella elegans was studied. C. elegans metabolized 4-methylbenz[a]anthracene primarily at the methyl group, this being followed by further metabolism at the 8,9- and 10,11-positions to form trans-8,9-dihydro-8,9-dihydroxy-4-hydroxymethylbenz[a]anthracene and trans-10,11-dihydro-10,11-dihydroxy-4-hydroxymethylbenz[a]anthracene. There was no detectable trans-dihydrodiol formed at the methyl-substituted double bond (3,4-positions) or at the 'K' region (5,6-positions). The metabolites were isolated by reversed-phase high-pressure liquid chromatography and characterized by the application of u.v.-visible-absorption-, 1H-n.m.r.- and mass-spectral techniques. The 4-hydroxymethylbenz[a]anthracene trans-8,9- and -10,11-dihydrodiols were optically active. Comparison of the c.d. spectra of the trans-dihydrodiols formed from 4-methylbenz[a]anthracene by C. elegans with those of the corresponding benz[a]anthracene trans-dihydrodiols formed by rat liver microsomal fraction indicated that the major enantiomers of the 4-hydroxymethylbenz[a]anthracene trans-8,9-dihydrodiol and trans- 10,11-dihydrodiol formed by C. elegans have S,S absolute stereochemistries, which are opposite to those of the predominantly 8R,9R- and 10R,11R-dihydrodiols formed by the microsomal fraction. Incubation of C. elegans with 4-methylbenz[a]anthracene under 18O2 and subsequent mass-spectral analysis of the metabolites indicated that hydroxylation of the methyl group and the formation of trans-dihydrodiols are catalysed by cytochrome P-450 mono-oxygenase and epoxide hydrolase enzyme systems. The results indicate that the fungal mono-oxygenase-epoxide hydrolase enzyme systems are highly stereo- and regio-selective in the metabolism of 4-methylbenz[a]anthracene.     

甘蔗节间伸长过程赤霉素生物合成关键基因的表达及相关植物激素动态变化

以甘蔗(Saccharum officinarum)优良品种桂糖42号(GT42)为研究材料, 分别于未伸长期(9-10叶龄以前) (Ls1)、伸长初期(12-13叶龄) (Ls2)和伸长盛期(15-16叶龄) (Ls3)取甘蔗第2片真叶(自顶部起)对应的节间组织, 测定其赤霉素(GA)、生长素(IAA)、油菜素甾醇(BR)、细胞分裂素(CTK)、乙烯(ETH)和脱落酸(ABA)的含量, 并通过实时荧光定量PCR (qRT-PCR)分析赤霉素合成途径关键基因GA₂₀氧化酶基因(GA20-Oxidase1)、赤霉素受体基因(GID1)和DELLA蛋白编码基因(GAI)的差异表达。结果表明, 在甘蔗伸长期间, GA和IAA含量呈现上升趋势, CTK和ABA含量呈下降趋势, ETH含量先上升后下降, BR含量则变化不明显; GA20-Oxidase1和GID1的表达呈上升趋势, 而GAI的表达则呈下降趋势, 这与相关植物激素的变化基本一致。综上, 甘蔗节间伸长过程主要与GA和IAA相关, 其次为CTK和ABA, 而ETH受到IAA的调控影响节间伸长; 植物激素间通过相互作用调控GA20-Oxidase1、GID1和GAI的表达, 影响GA含量和GA的信号转导过程, 进而影响甘蔗节间的伸长。该研究揭示了甘蔗节间伸长过程中赤霉素生物合成途径和信号转导关键基因的差异表达及植物激素含量的动态变化规律。