Antagonism by theophylline of respiratory inhibition induced by adenosine

The effects on respiration of an analogue of adenosine, L-2-N6-(phenylisopropyl)adenosine (PIA), and of the methylxanthine, theophylline, were determined in 19 vagotomized glomectomized cats whose end-tidal PCO2 was kept constant by means of a servo-controlled ventilator. Integrated phrenic nerve activity was used to represent respiratory output. Our results show that PIA, whether given systemically or into the third cerebral ventricle, depressed respiration. Systemically administered theophylline stimulated respiration. Theophylline given intravenously, or into the third ventricle not only reversed the depressive effects of previously administered PIA but caused further increases of respiration above the control level. Prior systemic administration of theophylline blocked both respiratory and hypotensive effects of subsequently administered PIA. Effects of either agent on medullary extracellular fluid pH did not explain the results. We conclude that the adenosine analogue PIA, acts to inhibit neurons in the brain that are involved in the control of respiration and that its effects are blocked by theophylline. We suggest that adenosine acts as a tonic modulator of respiration and that theophylline stimulates breathing by competitive antagonism of adenosine at neuronal receptor sites.     

The fertile island effect varies with aridity and plant patch type across an extensive continental gradient

Plant and Soil - Perennial plants play important roles in maintaining ecosystem functions by forming fertile islands beneath their canopies. Little is known about how the fertile island effect...     

Serological evidence of California serogroup virus activity in Oregon

We wished to demonstrate evidence of the presence of California serogroup viruses in Oregon and to test for the presence of certain other arboviruses in large ungulates. Blood samples from black-tailed deer (Odocoileus hemionus columbianus), mule deer (O. hemionus hemionus), and Roosevelt elk (Cervus elaphus roosevelti) from nine counties in Oregon were tested by serum-dilution plaque reduction neutralization for antibody to California serogroup viruses, including snowshoe hare, California encephalitis, and Jamestown Canyon, as well as to Cache Valley (Bunyamwera serogroup) and Klamath, an ungrouped rhabdovirus. Of 132 samples tested, 60 (46%) were found to be seropositive at a dilution of greater than or equal to 1:10 for at least one of the five different arboviruses. Forty (30%) samples contained antibody to more than one arbovirus, and 15 samples (11%) contained antibody to all five. Of these 15, 14 were from 75 black-tailed deer sera collected in Lincoln County, Oregon. Seropositivity rates for black-tailed deer ranged from 23% to 35%, with all five arboviruses represented. Positive reactions for all five arboviruses were represented among mule deer sera at rates from 5% to 29%. Elk sera were found to be positive for four of the viruses (none for Klamath virus). Although Cache Valley and Klamath viruses have been reported from Oregon, these data represent the first evidence of a California serogroup virus in the state.     

A protein induced by NGF in PC12 cells is stored in secretory vesicles and released through the regulated pathway.

We have previously described the isolation of a cDNA clone corresponding to an mRNA rapidly induced to high levels in PC12 cells by treatment with NGF. We report here the complete amino acid sequence of the protein (named VGF8a) as deduced by nucleotide sequencing of overlapping cDNA clones. VGF8a is particularly rich in proline residues and has a conspicuous number of short stretches of basic amino acid residues which may represent potential targets for proteolytic cleavage. Antibodies directed against recombinant VGF8a-beta-galactosidase fusion proteins were used for immunofluorescent staining of the protein in PC12 cells as well as for its localization, by Western blot analysis, in subfractions of cell homogenates. We demonstrate that in PC12 cells, VGF8a protein is stored in secretory vesicles and is released in response to a variety of stimuli that are known to induce the regulated secretion of neurotransmitters.     

Heparan sulfate proteoglycans in the substratum adhesion sites of human neuroblastoma cells: modulation of affinity binding to fibronectin

Tissue culture substratum adhesion sites from EGTA-detached Platt human neuroblastoma cells were extracted with a buffer containing ocytlglucoside, NaCl, guanidine hydrochloride, and a variety of protease inhibitors, an extraction which resulted in quantitative solubilization of the 35SO4 = -radiolabeled proteoglycans and 3H-leucine-radiolabeled proteins. Of the sulfate-radiolabeled material, the vast majority was heparan sulfate proteoglycan (Kav = 0.15 on Sepharose C14B columns) and the remainder was chondroitin sulfate chains (no single chains of heparan sulfate were observed). This extract was then fractionated on DEAE-Sephadex columns under two different buffer elution conditions. Under DEAE-I conditions in low ionic strength acetate buffer, two major peaks of 35SO4 = -radiolabeled material (A,B) and a minor peak (C) could be resolved in the NaCl gradient; however, three-fourths of the material required 4 M guanidine hydrochloride to elute it from the column (peak D). Under DEAE-II conditions in acetate buffer supplemented with 8 M urea, the vast majority of the proteoglycan material could be eluted in the NaCl gradient as peak AB. Peak D material was shown to contain aggregated proteoglycan, along with nonproteoglycan protein, which high concentrations of urea or guanidine could dissociate, but not nonionic or zwitterionic detergents. Three different affinity chromatography systems were used to further characterize these components. Approximately 60% of peak A heparan sulfate proteoglycan from DEAE-I binds to the hydrophobic matrix, octyl-Sepharose, while 80% of the proteoglycan in DEAE-I peak D binds to this hydrophobic column. A sizable fraction of peak A proteoglycan fails to bind to plasma fibronectin but does bind to platelet factor-4 affinity columns. In contrast, peak AB proteoglycan from DEAE-II columns yields a much higher proportion of molecules which do bind to fibronectin. To examine the basis for these differences in affinity binding, nonproteoglycan protein from these adhesion sites was mixed with peak AB proteoglycan prior to affinity chromatography; proteoglycan binding to fibronectin decreased markedly while binding to platelet factor-4 was unaffected. This modulating activity involves the binding of nonproteoglycan protein in adhesion site extracts to both fibronectin on the column, as well as to heparan sulfate proteoglycan itself, and it could not be mimicked by a number of known proteins in adhesion site extracts or several other proteins. These results demonstrate selectivity and specificity in this modulation and indicate that a previously unidentified protein(s) is responsible.(ABSTRACT TRUNCATED AT 400 WORDS)     

Characterization of membrane-bound electron transport enzymes from castor bean glyoxysomes and endoplasmic reticulum

Membranes purified from castor bean endosperm glyoxysomes by washing with sodium carbonate exhibited integral NADH:ferricyanide and NADH:cytochrome c reductase activities. The enzyme activities could not be attributed to contamination by other endomembranes. Purified endoplasmic reticulum membranes also contained the redox activities; and marker enzyme analysis indicated minimum cross contamination between glyoxysomal and endoplasmic reticulum fractions. The glyoxysomal redox activities were optimally solubilized at detergent to protein ratios (weight to weight) of 10 (Triton X-100), 50 (3-[3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate), and 100 (octylglucoside). Detergent in excess of the solubilization optimum was stimulatory to NADH:ferricyanide reductase and inhibitory to NADH:cytochrome c reductase. Endoplasmic reticulum redox activity solubilization profiles were similar to those obtained for glyoxysomal enzymes using Triton X-100. Purification of the glyoxysomal and endoplasmic reticulum NADH:ferricyanide reductases was accomplished using dye-ligand affinity chromatography on Cibacron blue 3GA agarose. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of NADH:ferricyanide reductase preparations purified by rate-zonal density gradient centrifugation, affinity chromatography, and nondenaturing electrophoresis of detergent-solubilized glyoxysomal and endoplasmic reticulum membranes consistently displayed 32- and 33-kDa silver-stained polypeptide bands, respectively.     

Correlation between LH and estrogen receptor turnover in pituitary and hypothalamus of castrate rats following estrogen agonists and antagonists

A series of studies was undertaken to correlate the short-term dynamics of LH secretion and depletion-replenishment patterns of estrogen receptors (ER) in hypothalamic and pituitary cytosols of ovariectomized rats. Animals castrated for 2 weeks were administered various test compounds and analyzed at 1, 3, 5, 10 and 15 h post-treatment. A single injection of 10 micrograms 17 beta-estradiol (E2) to ovariectomized rats elicited a rapid depletion of ER in both pituitary and hypothalamus and a dramatic, though delayed, fall in serum LH. ER replenishment occurred in both tissues through 15 h and LH recovered in a similar manner. When cycloheximide was administered along with E2, ER replenishment was completely inhibited in both tissues; serum LH fell and failed to recover. Actinomycin D injected with E2 blocked replenishment in pituitary but not hypothalamus; serum LH recovered in parallel with the hypothalamic ER pattern. 17 alpha-E2 elicited only slight changes in ER and LH was suppressed 10-20% through 15 h. CI-628 caused a near total depletion of pituitary ER with no subsequent replenishment, whereas hypothalamic ER content was virtually unaltered; serum LH was suppressed and later recovered. Orchidectomized rats given 5 micrograms E2 demonstrated a less complete ER depletion in hypothalamus, and an earlier replenishment than that seen in pituitary or hypothalamus of similarly treated ovariectomized females. Serum LH rebounded to 157% of control levels at 15 h. The results indicate that the acute feedback suppression of LH by exposure to estrogens correlates with binding to ER and nuclear translocation. Replenishment and/or retention of cytoplasmic ER in hypothalamus appears to be required for full resumption of LH secretion, following acute suppression.