Combined effects of thinning and decline on fine root dynamics in a Quercus robur L. forest adjoining the Italian Pre-Alps.

Aims: Oak decline is a complex phenomenon, characterized by symptoms of canopy transparency, bark cracks and root biomass reduction. Root health status is one of the first stress indicators, and root turnover is a key process in plant adaptation to unfavourable conditions. In this study, the combined effects of decline and thinning were evaluated on fine root dynamics in an oak forest adjoining the Italian Pre-Alps by comparison of acute declining trees with non-declining trees, both with and without thinning treatment of surrounding trees. Methods: Dynamics of volumetric root length density (RLD V ) and tip density (RTD V ), root tip density per unit length of root (RTD L ), diameter, branching index (BI) and mycorrhizal colonization were monitored by soil coring over 2 years as possible descriptors of decline. Key Results: At the beginning of the experiment, the relationship between canopy transparency and root status was weak, declining trees having slightly lower RLD V (-20 %) and RTD V (-11 %). After a 1 year lag, during which the parameters were almost unaffected, BI and RLD V , together with tip density, tip vitality and mycorrhizal colonization, became the descriptors most representative of both decline class and thinning. Thinning of declining trees increased RLD V (+12 %) and RTD V (+32 %), but reduced tip mycorrhizal colonization and vitality over time compared with non-thinned trees, whereas the opposite occurred in healthy trees, together with a marked decrease in branching. After thinning, there was an initial reduction in the structure of the ectomycorrhizal community, although recovery occurred about 10 months later, regardless of decline severity. Conclusions: Decline causes losses of fine root length, and a moderate recovery can be achieved by thinning, allowing better soil exploration by oak roots. The close correlation between root vitality and mycorrhizal colonization and their deterioration after thinning indicates that decline does not benefit from reduced root competition, excluding the hypothesis of limited water and nutrient availability as a possible cause of the syndrome in this forest.

The effects of sex and neonatal stress on pituitary adenylate cyclase-activating peptide expression.

NEW FINDINGS: What is the central question of this study? Does sex or neonatal stress affect the expression of pituitary adenylate cyclase-activating peptide or its receptors? What is the main finding and its importance? Neonatal-maternal separation stress has little long-lasting effect on the expression of pituitary adenylate cyclase-activating peptide or its receptors, but sex differences exist in these genes between males and females at baseline. Sex differences in classic stress hormones have been studied in depth, but pituitary adenylate cyclase-activating peptide (PACAP), recently identified as playing a critical role in the stress axes, has not. Here we studied whether baseline levels of PACAP differ between sexes in various stress-related tissues and whether neonatal-maternal separation stress has a sex-dependent effect on PACAP gene expression in stress pathways. Using quantitative RT-PCR, we found sex differences in PACAP and PACAP receptor gene expression in several respiratory and/or stress-related tissues, while neonatal-maternal separation stress did little to affect PACAP signalling in adult animals. We propose that sex differences in PACAP expression are likely to contribute to differences between males and females in responses to stress.

The geographical and environmental determinants of genetic diversity for four alpine conifers of the European Alps.

Climate is one of the most important drivers of local adaptation in forest tree species. Standing levels of genetic diversity and structure within and among natural populations of forest trees are determined by the interplay between climatic heterogeneity and the balance between selection and gene flow. To investigate this interplay, single nucleotide polymorphisms (SNPs) were genotyped in 24 to 37 populations from four subalpine conifers, Abies alba Mill., Larix decidua Mill., Pinus cembra L. and Pinus mugo Turra, across their natural ranges in the Italian Alps and Apennines. Patterns of population structure were apparent using a Bayesian clustering program, STRUCTURE, which identified three to five genetic groups per species. Geographical correlates with these patterns, however, were only apparent for P. cembra. Multivariate environmental variables [i.e. principal components (PCs)] were subsequently tested for association with SNPs using a Bayesian generalized linear mixed model. The majority of the SNPs, ranging from six in L. decidua to 18 in P. mugo, were associated with PC1, corresponding to winter precipitation and seasonal minimum temperature. In A. alba, four SNPs were associated with PC2, corresponding to the seasonal minimum temperature. Functional annotation of those genes with the orthologs in Arabidopsis revealed several genes involved in abiotic stress response. This study provides a detailed assessment of population structure and its association with environment and geography in four coniferous species in the Italian mountains.

The role of extracellular matrix in mouse and human corneal neovascularization.

Corneal neo-vascularization (CNV) is a highly prevalent medical condition which impairs visual acuity. The role of specific proteins in modulating CNV has been extensively reported, although no studies have described the entire human proteome in CNV corneas. In this paper, we performed a proteomic analysis of vascularized vs healthy corneal stroma, in a CNV mouse model and in CNV-affected patients, with a specific focus on extracellular matrix (ECM) proteins. We identified and quantified 2315 murine proteins, 691 human proteins and validated 5 proteins which are differentially expressed in vascularized samples and conserved in mice and humans: tenascin-C and fibronectin-1 were upregulated, while decorin, lumican and collagen-VI were downregulated in CNV samples. Interestingly, among CNV patients, those affected with Acanthamoeba keratitis showed the highest levels of fibronectin-1 and tenascin-C, suggesting a specific role of these two proteins in Acanthamoeba driven corneal CNV. On a broader picture, our findings support the hypothesis that the corneal stroma in CNV samples is disorganized and less compact. We are confident that the dissection of the human corneal proteome may shed new light on the complex pathophysiology of human CNV, and finally lead to improved treatments.

Presynaptic dopamine autoreceptors and second messengers controlling tyrosine hydroxylase activity in rat brain.

In brain areas enriched of dopaminergic nerve terminals presynaptic dopamine (DA) autoreceptors control the state of activation of tyrosine hydroxylase (TH) by regulating the extent of phosphorylation of the enzyme. Evidence is presented indicating that this autoinhibitory control may involve a decrease in the cyclic AMP-dependent activation of TH through an inhibitory coupling of presynaptic DA autoreceptors to adenylate cyclase. As indicated by the insensitivity of the DA inhibition of TH to changes in the extracellular concentrations of Ca++, to the addition of the Ca++ ionophore A 23187 and of different K+ channel blockers, a reduction of Ca++ influx and an increase in the K+ channel activity do not seem to be involved in the presynaptic regulation of TH activity by DA autoreceptors at least under basal conditions.

Corticotropin-releasing hormone stimulates adenylyl cyclase activity in the retinas of different animal species.

In the present study we investigated the presence of corticotropin-releasing hormone (CRH)-stimulated adenylyl cyclase activity in the retinas of different animal species. CRH significantly stimulated adenylyl cyclase activity in homogenates of calf, pig, rabbit and guinea pig retinas. The stimulatory effects were concentration-dependent with half-maximal responses occurring at 20-30 nM CRH. The enzyme activities increased by 37-80% at the maximal concentration of CRH (1 microM). On the other hand, adenylyl cyclase activities of chicken and pigeon retinas were poorly stimulated by CRH. In calf, pig and rabbit retinas, the CRH effect was completely antagonized by the CRH receptor antagonist alpha-helical CRH 9-41 and required the presence of GTP. The stimulatory response elicited by CRH was also found to be not additive with that produced by either vasoactive intestinal peptide or dopamine. These results provide evidence for the presence in retinas of different animal species of functional CRH receptors, an important criterion for the classification of CRH as a retinal neurotransmitter.

Evidence for dopamine autoreceptors in mesocortical dopamine neurons.

The effect of different treatments which are thought to modify dopamine (DA) synthesis by an action on DA autoreceptors was compared in the caudate nucleus and two frontal cortical areas: the medial prefrontal cortex and dorsolateral frontal cortex, having the highest and lowest DA concentration, respectively, but having equal concentrations of norepinephrine (NE); the NE to DA ratio being 3:2 and 8:1, respectively. DA synthesis was measured by the rate of DOPA accumulation after inhibition of DOPA decarboxylase. Gamma-butyrolactone (GBL) (750 mg/kg) increased DOPA accumulation by 200% in the caudate nucleus but only by 40% in the medial prefrontal cortex and was ineffective in the dorsolateral frontal cortex. Apomorphine (25-100 micrograms/kg) decreased DOPA accumulation by 7-30% in the medial prefrontal cortex and by 20-40% in the caudate nucleus in a dose-dependent manner. N-n-propylnorapomorphine (NPA) produced a similar effect within the dose range of 2.5-10 micrograms/kg. Both DA agonists were completely ineffective in the dorsolateral frontal cortical area. Haloperidol (0.5 mg/kg) increased DOPA accumulation by 80 and 220% in the medial prefrontal cortex and the caudate nucleus, respectively. It is concluded that DA autoreceptors regulate DA synthesis in the medial prefrontal cortex as in the caudate nucleus. Moreover, it was found that DOPA accumulation was approximately equal in the medial prefrontal cortex, with dense dopaminergic innervation, as in the dorsolateral area, devoid of dopaminergic terminals, suggesting that only a small fraction of cortical DA synthesis takes place in dopaminergic neurons, while the major part occurs in noradrenergic neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Suppression by progabide of ethanol withdrawal syndrome in rats.

Progabide, a specific and clinically used GABA receptor agonist, was tested for its ability to suppress ethanol withdrawal syndrome. Male rats were rendered physically dependent on ethanol by feeding for 12 days on a liquid diet in which ethanol isocalorically replaced dextrose. Progabide (100-400 mg/kg i.p.), administered 8 h after ethanol was withdrawn, produced a dose-related inhibition of both tremors and audiogenically induced seizures. A single dose of 400 mg/kg of progabide completely suppressed all ethanol withdrawal reactions. Seizures were more sensitive to the drug than tremors. The results support the view that a decrease in GABA transmission plays a role in ethanol withdrawal symptoms and suggest that progabide may be tested as a possible treatment of ethanol withdrawal syndrome in man.

Effect of spontaneous ingestion of ethanol on brain dopamine metabolism.

The effect of ethanol, either administered by gavage or voluntarily ingested, on brain dopamine (DA) metabolism was studied in alcohol-preferring and alcohol non-preferring rats. In alcohol non-preferring rats ethanol administration (2 g/kg) increased 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) and reduced DA levels in the caudate nucleus and olfactory tubercle but was ineffective in the medial prefrontal cortex. In alcohol-preferring rats ethanol effect was greater than in non-preferring animals and ethanol influenced DA metabolism also in the medial prefrontal cortex. The effect of voluntary ethanol ingestion was studied in alcohol-preferring rats trained to consume their daily fluid intake within 2 hrs. Voluntary ingestion of ethanol (3.1 +/- 0.7 g/kg in 1 hr) increased DA metabolites and reduced DA levels in the caudate nucleus, olfactory tubercle and medial prefrontal cortex. The results suggest that voluntary ethanol ingestion increases the release of DA from nigro-striatal and meso-limbic DA neurons.

Alcohol-preferring rats: genetic sensitivity to alcohol-induced stimulation of dopamine metabolism.

The effect of ethanol on brain dopamine (DA) metabolism in the caudate nucleus (CN), olfactory tubercle (OT) and medial prefrontal cortex (MPFC) was compared in two selectively bred rat lines, one ethanol preferring and the other ethanol nonpreferring. Male rats from the 16th and 17th generations of both lines that never experienced ethanol beforehand were used. No differences in the basal concentrations of DA and its metabolites, DOPAC and HVA, in the above brain regions were found between the two lines. The oral administration of 2 g/kg of ethanol to ethanolnonpreferring rats increased DOPAC and HVA and reduced DA levels in the CN and OT but was ineffective in the MPFC. On the other hand, ethanol administration to ethanol-preferring rats decreased DA content and increased DOPAC and HVA levels, not only in the CN and OT, but also in the MPFC. Moreover, the changes induced by ethanol on DA metabolism in the latter group were significantly greater than in ethanol nonpreferring rats. These results indicate that ethanol preferring rats have a genetic high sensitivity to the ethanol effect on DA metabolism, and suggest that such a trait might play a role in ethanol preference.

Ethanol prevents stress-induced increase in cortical DOPAC: reversal by RO 15-4513.

Electric foot-shock increased DOPAC and decreased DA levels by about 70 and 20% respectively in the medial prefrontal cortex in rats. Pretreatment with diazepam (5 mg/kg IP) or ethanol (1.2 g/kg orally) prevented these stress-induced changes. The protective effect of diazepam and ethanol was eliminated by RO 15-4513 (5 mg/kg IP) a partial inverse benzodiazepine agonist.

Suppression of ethanol intake in ethanol-preferring rats by 1,4-butanediol.

The oral administration of 1,4-butanediol (1,4-BD) at doses ranging from 100 to 300 mg/kg, twice daily, produced a dose-dependent reduction (40 to 85%) in the voluntary ethanol intake in rats selectively bred for high preference for ethanol. Treatment with 1,4-BD did not reduce total fluid intake. Repeated 1,4-BD administration (300 mg/kg twice daily for 7 days) suppressed ethanol intake almost completely. After suspension of 1,4-BD treatment, the inhibitory effect on ethanol intake remained significantly low for 2 days. 1,4-BD failed to inhibit aldehyde dehydrogenase to a concentration of 10 mM in rat liver homogenate.

Voluntary alcohol drinking increases brain dopamine metabolism in rats.

The effect of ethanol on dopamine (DA) metabolism in two selectively bred lines of rats, one alcohol-preferring (sP) and the other--non-preferring (sNP), was studied. Ethanol administration (2 g/kg per os) produced in the two lines of rats a decrease of DA content and an increased concentration of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the caudate nucleus, olfactory tubercle and medial prefrontal cortex in both lines, but the effect was significantly greater in sP than in sNP rats. Moreover, in sP rats, the voluntary consumption of ethanol increased DOPAC and HVA levels in the above areas. In these animals, DOPAC and HVA accumulation was associated with a small depletion in DA content, suggesting that ethanol releases DA from stores.

[Protection conferred by a hyperimmune serum and its fractions on rats infected with the Junin virus]. / Protección conferida por un suero hiperinmune y sus fracciones a ratas infectadas con virus Junín.

Suckling rats infected by ic route with 10(3)LD50 of the XJC13 strain of JV were passively immunized with homologous hyperimmune serum (HIS). Animals treated at 2 days pi with HIS showed a significant increase in survival vs. non-treated infected controls (82% vs 5%). However, at 4 days pi, transfer failed to modify survival. By means of DEAE Sephadex A25 column chromatography, the presence of neutralizing immunoglobulin closely correlated with protective antibodies, but were not restricted to the IgG-containing fraction. Employing Sephadex G200, chromatography demonstrated the absence of neutralizing and protective activity in the high molecular weight protein fraction. Results show that the success of HIS treatment depends on early administration. Besides, it was found that fractions capable of conferring protection exhibited high neutralizing antibody titers.

[Role of macrophages in the dissemination of the Junin virus in the rat]. / Papel del macrófago en la diseminación del virus Junín en la rata.

The 2-day-old rat is known to be susceptible to infection by ip route with the XJ strain of Junin virus but resists inoculation with XJC13 strain (85% vs 15% mortality). In order to determine whether peritoneal macrophages play a role in modulating the course of infection, viral replication in adherent peritoneal cells infected with either strain was studied. XJ was found to replicate 30-fold as regards XJC13 at day 3 pi. Besides, silica blockage of peritoneal macrophages was also evaluated. Following silica treatment, 60% survival was recorded for XJ-infected animals vs 15% for untreated infected controls. No significant differences were recorded for silica-treated XJC13-infected rats vs untreated infected controls. These preliminary findings indicate that, upon replication within the host's macrophages at the peritoneal inoculation site, virus spreads readily to reach the target organs. Effective silica blockage lends support to this pathway, as shown by significantly greater survival in XJ-infected rats.

Methylxanthine reversal of opioid-induced respiratory depression in the neonatal rat: mechanism and location of action.

Methylxanthines like caffeine and theophylline have long been used to treat apnea of prematurity. Despite their success in stimulating neonatal breathing, their mechanism of action remains poorly understood. Methylxanthines can act as both non-specific adenosine receptor antagonists and inhibitors of cAMP-dependent phosphodiesterases, sarcoplasmic/endoplasmic reticulum calcium ATPases or receptor-coupled anion channels, depending on the dose used. Though there is evidence for methylxanthine action at the level of the carotid body, the consensus is that methylxanthines stimulate the respiratory centers of the brainstem. Here we used the in situ neonatal rat working heart-brainstem preparation and the ex vivo neonatal rat carotid body preparation to test the hypothesis that methylxanthines act at the level of the carotid body. We conclude that although the neonatal carotid body has active adenosine receptors, the effects of methylxanthine therapy are likely mediated centrally, predominantly via inhibition of cAMP-dependent phosphodiesterase-4.