Short- and Long-Term Alterations of Gene Expression in Limbic Structures by Repeated Electroconvulsive-Induced Seizures

Rats were submitted to a series of 10 daily electroconvulsive shocks (ECS). A first group of animals was killed 1 day after the last seizure and a second group 30 days later. Tyrosine hydroxylase (TH) activity was measured using an in vitro assay in the nucleus caudatus, anterior cortex, amygdala, substantia nigra, ventral tegmental area, and locus ceruleus. The mRNA corresponding to this enzyme (TH-mRNA) was evaluated using a cDNA probe at the cellular level in the ventral tegmental area, substantia nigra, and locus ceruleus. Met-enkephalin (MET)-immunoreactivity and the mRNA coding for the preproenkephalin (PPE-mRNA) were assayed in striatum and the central nucleus of the amygdala. The day after the last ECS an increase of TH activity was observed in the ventral tegmental area, locus ceruleus, and substantia nigra in parallel with a similar increase in the amygdala and striatum; in the anterior cortex TH activity remained unchanged. TH-mRNA was increased in the locus ceruleus, evidencing the presence in this structure of a genomic activation. The amounts of MET and PPE-mRNA were unaffected in the striatum but increased in the amygdala. Thirty days after the last ECS we observed a decrease of TH activity in the amygdala and of TH-mRNA amount in the ventral tegmental area. In the locus ceruleus TH-mRNA remained higher in treated animals than in controls whereas TH activity returned to control levels. These results demonstrate that a series of ECS induces an initial increase of the activity of mesoamygdaloid catecholaminergic neurons followed by a sustained decrease through alterations of TH gene expression which could mediate the clinical effect of the treatment.     

Molecular cloning of a gene encoding a Chlamydia psittaci 57-kDa protein that shares antigenic determinants with ca. 60-kDa proteins present in many Gram-negative bacteria

In order to develop reagents to study the immune response of guinea pigs to infection by Chlamydia psittaci guinea pig inclusion conjunctivitis strain (GPIC), we constructed a plasmid clone bank with C. psittaci DNA. One of the recombinant clones isolated produced large amounts of a 57-kilodalton (kDa) protein that was immunoreactive with sera from GPIC infected guinea pigs. While investigating this recombinant protein, we discovered that all the Gram-negative bacteria analyzed so far have immunoreactive proteins of similar size. This protein seems to be a 'common antigen' already described in various Gram-negative bacteria.     

A new bacterial alcohol dehydrogenase active on degraded lignin and several low molecular weight aromatic compounds

A new intracellular bacterial dehydrogenase has been purified. It was active in the reversible reduction by NADH of conjugated carbonyl groups in partially degraded lignin. It was also active on various aromatic aldehydes such as vanillin, syringaldehyde and cinnamaldehyde, but had no effect on acetovanillone and lignin models carrying a conjugated ketone. It is proposed that this enzyme functions as a broadly specific lignin dehydrogenase at the level of aldehydic groups that are present in the lignin preparations.     

In Vivo and In Vitro Evidence for the Biosynthesis of Testosterone in the Telencephalon of the Female Frog

Abstract: Neurons and glial cells are capable of synthesizing various steroid hormones, but biosynthesis of testosterone in the CNS has never been reported. The aim of the present study was to demonstrate the synthesis of testosterone in the frog brain. The presence of 17β-hydroxysteroid dehydrogenase (17β-HSD)-like immunoreactivity was detected in a population of glial cells located in the telencephalon. Reversed-phase HPLC analysis of brain tissue extracts combined with radioimmunoassay detection revealed the presence of substantial amounts of testosterone and 5α-dihydrotestosterone (5α-DHT) in the telencephalon where 17β-HSD-positive cells were visualized. In male frogs, castration totally suppressed testosterone and 5α-DHT in the blood and in the rhombencephalon but did not affect the concentration of these two steroids in the telencephalon. Chemical characterization of testosterone in female frog telencephalon extracts was performed by coupling HPLC analysis with gas chromatography-mass spectrometry. Using the pulse-chase technique with [³H]pregnenolone as a precursor, the formation of a series of metabolites was observed, including dehydroepiandrosterone, androstenedione, testosterone, 5α-DHT, and estradiol. These data demonstrate the existence of an active form of 17β-HSD in the frog telencephalon, which is likely involved in testosterone biosynthesis within the brain.     

Short-term effects of urea amended with the urease inhibitor N-(n-butyl) thiophosphoric triamide on perennial ryegrass

The current study investigated the short-term physiological implications of plant nitrogen uptake of urea amended with the urease inhibitor N-(n-butyl) thiophosphoric triamide (nBTPT) under both greenhouse and field conditions. ¹⁵N labelled urea amended with 0.0, 0.01, 0.1 and 0.5% nBTPT (w/w) was surface applied at a rate equivalent to 100 kg N ha^–1 to perennial ryegrass in a greenhouse pot experiment. Root, shoot and soil fractions were destructively harvested 0.75, 1.75, 4, 7 and 10 days after fertilizer application. Urease activity was determined in each fraction together with ¹⁵N recovery and a range of chemical analyses. The effect of nBTPT amended urea on leaf tip scorch was evaluated together with the effect of the inhibitor applied on its own on plant urease activity.nBTPT-amended urea dramatically reduced shoot urease activity for the first few days after application compared to unamended urea. The higher the nBTPT concentration the longer the time required for shoot activity to return to that in the unamended treatment. At the highest inhibitor concentration of 0.5% shoot urease activity had returned to that of unamended urea by 10 days. Root urease activity was unaffected by nBTPT in the presence of urea but was affected by nBTPT in the absence of urea.Transient leaf tip scorch was observed approximately 7–15 days after nBTPT + urea application and was greatest with high concentrations of nBTPT and high urea-N application rates. New developing leaves showed no visual sign of tip necrosis.Urea hydrolysis of unamended urea was rapid with only 1.3% urea-N remaining in the soil after 1.75 days. N uptake and metabolism by ryegrass was rapid with ¹⁵N recovery from unamended urea, in the plant (shoot + root) being 33% after 1.75 days. Most of the ¹⁵N in the soil following the urea+0.5% nBTPT application was still as urea after 1.75 days, yet ¹⁵N plant recovery at this time was 25% (root+shoot). This together with other evidence, suggests that if urea hydrolysis in soil is delayed by nBTPT then urea can be taken up by ryegrass as the intact molecule, albeit at a significantly slower initial rate of uptake than NH₄ ⁺-N. Protein and water soluble carbohydrate content of the plant were not significantly affected by amending urea with nBTPT however, there was a significant effect on the composition of amino acids in the roots and shoots, suggesting a difference in metabolism.Although nBTPT-amended urea affected plant urease activity and caused some leaf-tip scorch the effects were transient and short-lived. The previously reported benefit of nBTPT in reducing NH₃ volatilization of urea would appear to far outweigh any of the observed short-term effects, as dry-matter production of ryegrass is increased.     

Biological activity and metabolic clearance of recombinant human follicle stimulating hormone produced in Sp2/0 myeloma cells

Human follicle stimulating hormone is a pituitary glycoprotein that is essential for the maintenance of ovarian follicle development and testicular spermatogenesis. Like other members of the glycoprotein hormone family, it contains a common a subunit and a hormone specific subunit. Each subunit contains two glycosylation sites. The specific structures of the oligosaccharides of human follicle stimulating hormone have been shown to influence both thein vitro andin vivo bioactivity. Since the carbohydrate structure of a protein reflects the glycosylation apparatus of the host cells in which the protein is expressed, we examined the isoform profiles,in vitro bioactivity and metabolic clearance of a preparation of purified recombinant human follicle stimulating hormone derived from a stable, transfected Sp2/0 myeloma cell line, and pituitary human follicle stimulating hormone. Isoelectric focussing and chromatofocussing studies of human follicle stimulating hormone preparations both showed a more basic isoform profile for the recombinant human follicle stimulating hormone compared to that of pituitary human follicle stimulating hormone. The recombinant human follicle stimulating hormone had a significantly higher radioreceptor activity compared to that of pituitary human follicle stimulating hormone, consistent with a greaterin vitro potency. Pharmacokinetic studies in rats indicated a similar terminal half life (124 min) to that of the pituitary human follicle stimulating hormone (119 min). Preliminary carbohydrate analysis showed recombinant human follicle stimulating hormone to contain high mannose and/or hybrid type, in addition to complex type carbohydrate chains, terminating with both2,3 and2,6 linked sialic acids. These results demonstrate that recombinant human follicle stimulating hormone made in the Sp2/0 myeloma cells is sialylated, has a more basic isoform profile, and has a greaterin vitro biological potency compared to those of the pituitary human follicle stimulating hormone.     

The elimination ofO-linked glycans from glycoproteins under non-reducing conditions

A simple procedure is described for the elimination ofO-linked glycans from bovine submaxillary mucin under non-reducing conditions, using triethylamine in aqueous hydrazine. The glycans were isolated as the hydrazones, which were converted to the reducing glycans by exchange with acetone in neutral aqueous solution. The glycan alditols obtained after reduction corresponded to those obtained by the reductive -elimination ofO-glycans.     

EVOLUTIONARY SHIFTS IN THE SPECTRAL PROPERTIES OF SPIDER SILKS

We measured the reflectance properties of unpigmented silks spun by a systematic array of primitive (Deinopoidea) and derived (Araneoidea) aerial, web-spinning spiders, as well as silks spun by Araneomorphae and Mygalomorphae spiders that do not spin aerial webs. Our data show that all of the primitive aerial web spinners produce catching silks with a spectral peak in the ultraviolet (UV), and cladistic analysis suggests that high UV reflection is the primitive character state for silk spectral properties. In contrast, all of the derived aerial web spinners produce silks that are spectrally flat or characterized by reduced reflectance in the UV. Correlated with the evolution of these catching silks is a 37-fold increase in species number and apparent habitat expansion. This suggests that the unique silk proteins spun by the araneoids have been important to their ecological and evolutionary diversity.