Prostaglandin H Synthetase-Mediated Metabolism of Dopamine: Implication for Parkinson's Disease

Abstract: Differences in prostaglandin H synthetase (PHS) activity in the substantia nigra of age- and post-mortem interval-matched parkinsonian, Alzheimer's, and normal control brain tissue were assessed. Prostaglandin E₂ (PGE₂, an index of PHS activity) was higher in substantia nigra of parkinsonian brain tissue than Alzheimer's or control tissue. Incubation of substantia nigra slices with arachidonic acid (AA) increased PGE₂ synthesis. Dopamine stimulated PHS synthesis of PGE₂. [³H]Dopamine was activated by PHS to electrophilic intermediate(s) that covalently bound to DNA, microtubulin protein, bovine serum albumin, and sulfhydryl reagents. When AA was replaced by hydrogen peroxide, PHS/H₂O₂-supported binding proceeded at rates similar to those observed with PHS/AA. Indomethacin and aspirin inhibited AA-mediated cooxidation of dopamine but not H₂O₂-mediated metabolism. PHS-mediated metabolism of dopamine was not affected by monoamine oxidase inhibitors. Substrate requirements and effects of specific inhibitors suggest cooxidation of dopamine is mediated by the hydroperoxidase activity of PHS. ³²P-postlabeling was used to detect dopamine-DNA adducts. PHS/AA activation of dopamine in the presence of DNA resulted in the formation of five dopamine-DNA adducts, i.e., 23, 43, 114, 70, and 270 amol/µg DNA. DNA adduct formation was PHS, AA, and dopamine dependent. PHS catalyzed cooxidation of dopamine in dopaminergic neuronal degeneration is discussed.     

Compatibility of Soil Amendments with Entomopathogenic Nematodes

The impact of inorganic and organic fertilizers on the infectivity, reproduction, and population dynamics of entomopathogenic nematodes was investigated. Prolonged (10- to 20-day) laboratory exposure to high inorganic fertilizer concentrations inhibited nematode infectivity and reproduction, whereas short (1-day) exposures increased infectivity. Heterorhabditis bacteriophora was more sensitive to adverse effects than were two species of Steinernema. In field studies, organic manure resulted in increased densities of a native population of Steinernema feltiae, whereas NPK fertilizer suppressed nematode densities regardless of manure applications. Inorganic fertilizers are likely to be compatible with nematodes in tank mixes and should not reduce the effectiveness of nematodes used for short-term control as biological insecticides, but may interfere with attempts to use nematodes as inoculative agents for long-term control. Organic manure used as fertilizer may encourage nematode establishment and recycling.     

Growth,graviresponsiveness and abscisic-acid content of Zea mays seedlings treated with Fluridone

Ten-d-old seedlings of Zea mays L. cv. Tx 5855 treated with 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone (Fluridone) were analyzed for abscisic acid (ABA) content using high-performance liquid chromatography with an analysis sensitivity of 2.5 ng ABA g^-1 fresh weight (FW). Seedlings were divided into three portions: leaves, detipped roots, and root tips (terminal 1.5 mm). Control plants (water treatment only; no Fluridone) were characterized by the following amounts of ABA: leaves, 0.114±0.024 (standard deviation) g ABA g^-1 FW; detipped roots, 0.260±0.039±g ABA g^-1 FW; root tips, no ABA detected. We did not detect any ABA in tissues of Fluridone-treated plants. Primary roots of treated and untreated seedlings were strongly graviresponsive, with no significant differences between the curvatures or the growth rates of primary roots of Fluridone-treated and control seedlings. These results indicate that 1) Fluridone completely inhibits ABA synthesis, and 2) ABA is not necessary for positive gravitropism by primary roots of Zea mays.Abbreviations ABA abscisic acid - Fluridone 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone - FW fresh weight - SD standard deviation     

Graft formation in Solanum pennellii (Solanaceae)

Three phases of cohesion were observable during the development of compatible autografts in Solanum pennellii. Phase I cohesion 1) lasted 4–5 d after grafting, 2) was characterized by an average increase in tensile strength of 4 g breaking weight (BW) mm^–2 graft area (GA) d^–1, and 3) correlated positively with cellular interdigitation at the graft interface. The fresh weight of the scion increased by approximately 5% d^–1 during the first 2 d after grafting. Phase II cohesion occurred 5–15 d after grafting, during which time 1) the tensile strength of the graft union increased by 14 g BW mm^–2 GA d^–1, 2) vascular differentiation across the graft interface was completed, and 3) the fresh weight of the scion increased by 9% d^–1. Phase III cohesion occurred subsequent to 15 d after grafting, during which time 1) the tensile strength of the graft union leveled off at a value similar to that of an ungrafted internode, and 2) the fresh weight of the scion increased by 14% d^–1. These results are discussed relative to mechanisms underlying the formation of compatible grafts.     

Cloning of a Phosphate-Regulated Hemolysin Gene (Phospholipase C) from Pseudomonas aeruginosa

Phospholipase C (heat-labile hemolysin) of Pseudomonas aeruginosa is a phosphate (P_i)-regulated extracellular protein which may be a significant virulence factor of this organism. The gene for this hemolytic enzyme was cloned on a 4.1-megadalton (Mdal) fragment from a BamHI digest of P. aeruginosa PAO1 genomic DNA and was inserted into the BamHI sites of the multicopy Escherichia coli(pBR322) and P. aeruginosa(pMW79) vectors. The E. coli and P. aeruginosa recombinant plasmids were designated pGV26 and pVB81, respectively. A restriction map of the 4.1-Mdal fragment from pGV26 was constructed, using double and single digestions with BamHI and EcoRI and several different restriction enzymes. Based on information from this map, a 2.4-Mdal BamHI/BglII fragment containing the gene for phospholipase C was subcloned to pBR322. The hybrid plasmids pGV26 and pVB81 direct the synthesis of enzymatically active phospholipase C, which is also hemolytic. The plasmid-directed synthesis of phospholipase C in E. coli or P. aeruginosa is not repressible by P_i as is the chromosomally directed synthesis in P. aeruginosa. Data are presented which suggest that the synthesis of phospholipase C from pGV26 and pVB81 is directed from the tetracycline resistance gene promoter. The level of enzyme activity produced by E. coli(pGV26) is slightly higher than the levels produced by P. aeruginosa(pMW79) under repressed conditions. In contrast, the levels produced by P. aeruginosa(pVB81) are at least 600-fold higher than the levels produced by P. aeruginosa(pMW79) under repressed conditions and approximately 20-fold higher than those produced by P. aeruginosa(pMW79) under derepressed conditions. The majority (85%) of the enzyme produced by E. coli(pGV26) remained cell associated, whereas >95% of the enzyme produced by P. aeruginosa(pVB81) was extracellular. Analysis of extracellular proteins from cultures of P. aeruginosa(pMW79) and P. aeruginosa(pVB81) by high-performance liquid chromotography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the phospholipase C gene was cloned intact, and it is likely that several additional genes were cloned on the 4.1-Mdal fragment of DNA. It was also found that some of these genes encode proteins which are the same molecular weight as some previously described P_i-repressible proteins of P. aeruginosa. The existence of a P_i regulon of P. aeruginosa is proposed. It is likely that one of these genes also regulates the level of pyocyanin production by P. aeruginosa and that one or more play a role in transport or binding of P_i. The availability of the hybrid plasmids described herein will be useful in further studies on the role of this hemolysin in the virulence of P. aeruginosa and in the study of the genetics and physiology of P_i-regulated proteins.     

A comparison of some hydrolytic and gas chromatographic procedures used in methylation analysis of the carbohydrate units of glycopeptides

The polysaccharide secreted by Klebsiella aerogenes type 54 strain A3 was isolated, methylated, the ester carboxyl-reduced, and the product partially hydrolyzed. The resulting, partially O-methylated oligosaccharides were reduced and ethylated, and the mixture of products was fractionated by l.c. The l.c. fractions containing per-O-alkylated oligosaccharide-alditols were analyzed by e.i.-m.s. Pure per-O-alkylated oligosaccharide-alditols were also analyzed by ¹H-n.m.r. spectroscopy. The products obtained by base-catalyzed degradation and subsequent ethylation of the per-O-methylated polysaccharide were fractionated by l.c. The main product isolated was analyzed by e.i.-m.s., c.i.-m.s., and ¹H-n.m.r. spectroscopy. The results of these studies, in conjunction with results of analytical methods commonly used in the elucidation of polysaccharide structures, unambiguously characterized the primary glycosyl structure of the polysaccharide. Base-labile substituents, previously reported to be present in the polysaccharide, were not studied. Structure 1 revises, and complements, previously reported structures.

     

Thermoregulatory, behavioral and seizure modulatory effects of AVP in the gerbil

Recent reports suggest that arginine vasopressin (AVP) may be an endogenous antipyretic peptide and a mediator of febrile convulsions [10,12]. The spontaneously seizing Mongolian gerbil was used to investigate the thermoregulatory, behavioral and seizure modulatory effects of AVP. Injection of AVP (1.0 and 5.0 μg IV and 0.01–1.0 mg/kg SC) caused dose-related falls in body temperature. Stereotypic scratching, terminated by a body shake, was observed after AVP (1.0–5.0 μg IV). However, such behavior was not observed after subcutaneous injection of AVP. AVP did not potentiate seizure induction in the gerbils but rather reduced the seizure incidence. The data demonstrate that AVP can reduce body temperature and cause specific behaviors, but it does not appear to play a role in the pathogenesis of seizures in the seizure sensitive strain of Mongolian gerbil.     

Location of the origin of replication for the 7.5-kbChlamydia trachomatis plasmid

The hypothetical origin of replication for the 7.5-kb plasmid common toChlamydia trachomatis is believed to be in a region of the plasmid that contains four 22-bp tandem repeats preceded by an A-T-rich region. To test this hypothesis, replication of plasmid DNA in metabolically active reticulate bodies of the Lymphogranuloma venereum biovar ofC. trachomatis was examined by electron microscopy. The results presented show that the origin of replication appears to be near the tandem repeats of pCHL2. In addition, replication of the 7.5-kb plasmid is unidirectional, and the copy number during replication is 7–10. The evidence presented suggests thatC. trachomatis has a homologue to theEscherichia coli dnaA gene and that this homologue might be involved in replication of theC. trachomatis 7.5-kb plasmid.     

New mechanical aid to physiotherapy in cystic fibrosis.

The force of impact and frequency of percussion by physiotherapists and parents of children with cystic fibrosis were analysed on a special test rig and incorporated in a prototype percussor. In adult volunteers and cadavers a maximum intrathoracic pressure could be achieved by a critical frequency of mechanical percussion which was higher than that reached by physiotherapists and parents. Consequently the percussor was redesigned to operate at this optimum frequency. It was then discovered that if the percussor was pressed firmly enough against the chest, this maximum intrathoracic pressure could be indicated by quivering of the voice. In a continuing study of intrathoracic pressures obtained mechanically and manually the Salford percussor''s produced higher pressures than the physiotherapists'' and maintained them constantly, while the physiotherapists'' efforts and results varied from one to another. Hospital and domiciliary use of the percussor have shown it to help in the first stage of the physiotherapy routine for patients with cystic fibrosis. The percussor should enable adolescents and adults to treat themselves and encourage twice-daily and more effective chest treatments. It is easy to apply and its speed and efficiency should enable parents to improve the quality of their chest therapy for younger children at home. Its long-term benefits are difficult to assess because of the nature of the disease.     

Studies of the regulation of purine nucleotide catabolism.

Ehrlich ascites tumor cells containing radioactive ATP were incubated in vitro with a range of concentrations of 2-deoxyglucose in order to produce different rates of ATP catabolism. Concentrations of all radioactive products of ATP catabolism were measured, and apparent rates of adenylate deaminase and inosinate dehydrogenase and of adenylate and inosinate dephosphorylation were calculated. It was concluded that these processes were reggulated primarily by the rate of formation of substrate, and to a lesser extent in some cases, by substrate concentration. No evidence was obtained for regulation of these processes by the concentration of ATP. The deoxyglucose-induced catabolism of radioactive GTP was also studied. When ATP catabolism was induced by incubation with 2,4-dinitrophenol, time courses of accumulation of purine nucleoside monophosphates and rates of alternative pathways of their metabolism were quite different than when deoxyglucose was used.