Plant functional trait response to environmental drivers across European temperate forest understorey communities.

Functional traits respond to environmental drivers, hence evaluating trait-environment relationships across spatial environmental gradients can help to understand how multiple drivers influence plant communities. Global-change drivers such as changes in atmospheric nitrogen deposition occur worldwide, but affect community trait distributions at the local scale, where resources (e.g. light availability) and conditions (e.g. soil pH) also influence plant communities. We investigate how multiple environmental drivers affect community trait responses related to resource acquisition (plant height, specific leaf area (SLA), woodiness, and mycorrhizal status) and regeneration (seed mass, lateral spread) of European temperate deciduous forest understoreys. We sampled understorey communities and derived trait responses across spatial gradients of global-change drivers (temperature, precipitation, nitrogen deposition, and past land use), while integrating in-situ plot measurements on resources and conditions (soil type, Olsen phosphorus (P), Ellenberg soil moisture, light, litter mass, and litter quality). Among the global-change drivers, mean annual temperature strongly influenced traits related to resource acquisition. Higher temperatures were associated with taller understoreys producing leaves with lower SLA, and a higher proportional cover of woody and obligate mycorrhizal (OM) species. Communities in plots with higher Ellenberg soil moisture content had smaller seeds and lower proportional cover of woody and OM species. Finally, plots with thicker litter layers hosted taller understoreys with larger seeds and a higher proportional cover of OM species. Our findings suggest potential community shifts in temperate forest understoreys with global warming, and highlight the importance of local resources and conditions as well as global-change drivers for community trait variation.

[Botulinum Toxin in Functional Hypersalivation--All about Dosage?]. / Botulinumtoxin bei funktioneller Hypersalivation--alles eine Frage der Dosis?

BACKGROUND: The use of botulinum toxin injection in the salivary gland, is taking an increasing significance in the treatment of functional hypersalivation today. With due regard to the off -label use and the prospect of success, dosage levels are not yet standardized. MATERIAL AND METHODS: In a retrospective study, 54 patients resp. 117 treatments were analysed over a period of 5 years according to their dosage levels of botulinum toxin, outcome and side effects. RESULTS: In 90% of the cases, a reduction of saliva after botulinum toxin injections was reported, although a significant number of patients wished for an even greater effect. Compared to the first botulinum toxin injection, we therefore used a higher dosage plan in the following treatment in order to achieve better clinical results. Besides not enough saliva reduction, the main side effects were swallowing problems and thick or sticky saliva in patients with a tracheal cannula. With the exception of insufficient saliva reduction, the other described side effects were irrespective to the dosage level. CONCLUSIONS: Botulinum toxin injection as a treatment of hypersalivation is an effective method with only minor side effects, even in increased dosage levels. Nevertheless, certain modifications according to each individual treatment are required. Possible side effects such as swallowing problems or non-responding situations should always be part of informed consent, especially as the latter is even possible for higher dosage levels.

Baclofen enhances extinction of opiate conditioned place preference.

Conditioned opiate reward (COR) is rapidly acquired and slowly extinguished. The slow rate of extinction of the salience of drug-related cues contributes to drug craving and relapse. The gamma-aminobutyric acid receptor type B (GABA(B)) agonist, baclofen, attenuates the unconditioned rewarding actions of several drugs of abuse and was investigated for effects on the extinction of COR. C57BL/6 mice were utilized in an unbiased conditioned place preference (CPP) protocol using morphine (10mg/kg, s.c.) and saline. CPP was measured by increases in time spent in the morphine-associated (CS+) vs. the saline-associated (CS-) chamber in a 15-min test after four morphine and four saline alternated conditioning sessions. CPP and locomotor sensitization were produced to the CS+ chamber. Subsequently, sixteen daily extinction sessions were conducted with either vehicle or baclofen (1 or 2.5mg/kg, s.c.) treatment given either before or after the sessions. This design was used to create the baclofen drug state before or after the activation of the CPP memory trace in the extinction protocol. After morphine CPP development, its extinction was significantly facilitated in a dose-dependent manner by post-session, but not by pre-session, baclofen treatments. No significant sedative effects of baclofen were detected during any extinction training or testing phase. Baclofen treatment facilitated the extinction of COR and reduced conditioned sensitization during extinction when given after, but not before, the activation of the CPP memory trace. Baclofen appears to have disrupted reconsolidation of conditioned reward memory during extinction training and might similarly facilitate extinction learning in human opiate addiction.

Accurate detection of uniparental disomy and microdeletions by SNP array analysis in myelodysplastic syndromes with normal cytogenetics.

Progress in the management of patients with myelodysplastic syndromes (MDS) has been hampered by the inability to detect cytogenetic abnormalities in 40-60% of cases. We prospectively analyzed matched pairs of bone marrow and buccal cell (normal) DNA samples from 51 MDS patients by single nucleotide polymorphism (SNP) arrays, and identified somatically acquired clonal genomic abnormalities in 21 patients (41%). Among the 33 patients with normal bone marrow cell karyotypes, 5 (15%) had clonal, somatically acquired aberrations by SNP array analysis, including 4 with segmental uniparental disomies (UPD) and 1 with three separate microdeletions. Each abnormality was detected more readily in CD34+ cells than in unselected bone marrow cells. Paired analysis of bone marrow and buccal cell DNA from each patient was necessary to distinguish true clonal genomic abnormalities from inherited copy number variations and regions with apparent loss of heterozygosity. UPDs affecting chromosome 7q were identified in two patients who had a rapidly deteriorating clinical course despite a low-risk International Prognostic Scoring System score. Further studies of larger numbers of patients will be needed to determine whether 7q UPD detected by SNP array analysis will identify higher risk MDS patients at diagnosis, analogous to those with 7q cytogenetic abnormalities.

Temporal ontogeny of circuit activation prior to the onset of seizure susceptibility in EL/Suz mice.

The EL/Suz (EL) mouse is a model of multifactorial temporal lobe epilepsy in which seizures begin around 90 days of age, but can be hastened through increased exposure to human handling. In order to better understand seizure etiology in this mouse strain relative to seizure-resistant control mice, the present study examined region-specific neuronal activation in response to non-seizure-inducing handling implemented before the onset of seizure susceptibility. Immediate-early gene (cFos) expression emerged in EL mice by postnatal day (PND) 21 in the primary motor cortex, progressed to the locus ceruleus and prefrontal cortex by PND 35, and appeared in the hippocampus and amygdala by PND 70, as mice neared the age of onset for seizure susceptibility. Thus, mirroring the pattern observed during the course of a seizure, specific brain regions were differentially recruited to a neural network for seizure predisposition before the onset of seizure susceptibility. This developmental pattern of early and transient neural activation represents an important window for the study of causal mechanisms of seizure susceptibility following exposure to environmental triggers.

Mouse feeding behavior: ethology, regulatory mechanisms and utility for mutant phenotyping.

Ingestive behaviors, feeding and drinking, constitute unconditioned, obligatory functions that are tightly regulated in the rodent according to demands of the external and internal milieu. Dependent measures of food intake have been used extensively in rats to infer the identity and function of neurochemical pathways, which mediate energy balance. A recent interest in application of appetitive measures in mice can be attributed jointly to the discovery of novel markers of energy balance in genetically obese mice as well as systematic targeting of known feeding regulatory pathways in bioengineered mutant mice. Accordingly, this review will attempt to provide the reader interested in behavioral phenotyping of knockout or transgenic mice with information regarding the ethology of mouse eating behavior, known mechanisms of appetitive regulation and examples of successes and pitfalls encountered when studying food intake in mutant mice.

Selective stimulatory actions of corticotropin-releasing factor ligands on correlates of energy balance.

Acute administration of corticotropin-releasing factor (CRF) results in anorexic and sympathomimetic effects that suggest efficacy in chronic models of energy balance. The present studies employed a broad spectrum energy balance indices in lean and genetically obese Zucker rats in order to fully characterize the pharmacological efficacy of CRF and a CRF binding protein (CRF-BP) ligand inhibitor, CRF(6-33), which is thought to liberate CRF from CRF-BP. Acute administration of CRF(6-33) significantly increased CRF(2) receptor density by 10% within the ventromedial hypothalamic (VMH) nucleus of Zucker lean rats and decreased density by 10% in Zucker obese rats. A single infusion of CRF(6-33) increased nonshivering thermogenesis by 25-30% as measured by proton conductance in brown adipose tissue of both lean and obese rats. Chronic CRF(6-33) infusion suppressed body weight gain and elevated core temperature irrespective of genotype while increasing motor activity in obese rats without altering heart rate or blood pressure. Taken together, these results document strain-dependent, long-term effects of a CRF-BP ligand inhibitor on a select subset of physiological and behavioral measures of increased energy expenditure.

The role of corticotropin-releasing factor in drug addiction.

The goal of this article is to summarize available data examining the physiological significance of brain corticotropin-releasing factor (CRF) systems in mediating the behavioral and physiological effects of several classes of abused drugs, including opioid and psychostimulant drugs, alcohol and sedative hypnotics, nicotine, and cannabinoids. An initial discussion of CRF neurobiology is followed by consideration of the role of CRF in drug-induced activation of the hypothalamic-pituitary-adrenocortical (HPA) axis, the behavioral effects of drugs (e.g., locomotor activity, anxiogenic-like responses), drug self-administration, drug withdrawal, and relapse to drug-seeking. Subsequently, neurochemical changes in brain CRF in response to acute and chronic drug exposure are examined. A major conclusion derived from the data reviewed is that extrahypothalamic brain CRF systems are critically involved in behavioral and physiological manifestations of drug withdrawal and in relapse to drug-taking behavior induced by environmental stressors. On the other hand, it appears that hypothalamic CRF, via its action on the HPA axis, is involved in the reinforcing effects of cocaine and alcohol, and the locomotor activating effects of psychostimulant drugs. These preclinical data may provide a rationale for the development of CRF-based pharmacotherapies for the treatment of compulsive drug use in humans.

The role of CRH in behavioral responses to stress.

Corticotropin-releasing hormone (CRH) and urocortin in the central nervous system affect behavior and can enhance behavioral responses to stressors. The action of CRH-related peptides is mediated through multiple receptors that differ markedly in their pharmacological profiles and anatomical distribution. Comparative pharmacology of CRH receptor agonists suggests that CRH, urocortin, sauvagine and urotensin consistently mimic, and CRH receptor antagonists consistently lessen, functional consequences of stressor exposure. Recently, important advances have been made in understanding the CRH system and its role in behavioral responses to stress by the development of specific CRH receptor antagonists, application of antisense oligonucleotides and development of transgenic mice lacking peptides and functional receptors. This review summarizes recent findings with respect to components of the CRH system and their role in stress-induced behavioral responses.

Inhibition of anti-IgM-induced translocation of protein kinase C beta I inhibits ERK2 activation and increases apoptosis.

Expression of the COOH-terminal residues 179-330 of the LSP1 protein in the LSP1(+) B-cell line W10 increases anti-IgM- or ionomycin-induced apoptosis, suggesting that expression of this LSP1 truncate (B-LSP1) interferes with a Ca(2+)-dependent step in anti-IgM signaling. Here we show that inhibition of Ca(2+)-dependent conventional protein kinase C (cPKC) isoforms with Gö6976 increases anti-IgM-induced apoptosis of W10 cells and that expression of B-LSP1 inhibits translocation of PKCbetaI but not of PKCbetaII or PKCalpha to the plasma membrane. The increased anti-IgM-induced apoptosis is partially reversed by overexpression of PKCbetaI. This shows that the B-LSP1-mediated inhibition of PKCbetaI leads to increased anti-IgM-induced apoptosis. Expression of constitutively active PKCbetaI protein in W10 cells activates the mitogen-activated protein kinase ERK2, whereas expression of B-LSP1 inhibits anti-IgM-induced activation of ERK2, suggesting that anti-IgM-activated PKCbetaI is involved in the activation of ERK2 and that inhibition of ERK2 activation contributes to the increased anti-IgM-induced apoptosis. Pull-down assays show that LSP1 interacts with PKCbetaI but not with PKCbetaII or PKCalpha in W10 cell lysates, while in vitro LSP1 and B-LSP1 bind directly to PKCbetaI. Thus, B-LSP1 is a unique reagent that binds PKCbetaI and inhibits anti-IgM-induced PKCbetaI translocation, leading to inhibition of ERK2 activation and increased apoptosis.

Corticotropin-releasing factor (CRF) or CRF binding-protein ligand inhibitor administration suppresses food intake in mice and elevates body temperature in rats.

Corticotropin-releasing factor (CRF) receptor agonist and CRF binding-protein (CRF-BP) ligand inhibitor peptides both activate CRF systems but exert very distinct functional profiles in animal models of arousal, energy balance and emotionality. The present studies were designed to extend the dissimilar efficacy profiles of central administration of a CRF agonist, r/h CRF(1-41), versus a CRF-BP ligand inhibitor, r/h CRF(6-33), into mouse and rat models of energy balance in order to further explore in vivo efficacy of these ligands in two separate animal species. In CD-1 mice, food intake was significantly attenuated 3 h after acute administration of CRF(1-41) (0.007-0.2 nmol), but not CRF(6-33). In obese Ob/Ob mice, both CRF(1-41) (0.007-0.2 nmol) and CRF(6-33) (0.02-2.3 nmol) significantly attenuated basal feeding over 3 h following acute peptide administration. In rats, CRF(1-41) (1 nmol) and CRF(6-33) (1.5-7.7 nmol) infusion significantly increased rectal temperature. In studies employing a telemetry apparatus, core temperature was also increased by CRF(1-41) (1 nmol) and CRF(6-33) (1.5 nmol), whereas only CRF(1-41) increased locomotor activity and heart rate. These results suggest that CRF receptor agonist administration is capable of producing a global profile of negative energy balance by reducing food intake in mice and increasing energy expenditure in rats. In contrast, CRF-BP ligand inhibitor administration appears to suppress food intake in a mouse strain selective manner and to elevate rectal and core temperature in rats without accompanying cardiovascular activation.

Dissociation of arousal-like from anxiogenic-like actions of brain corticotropin-releasing factor receptor ligands in rats.

Behavioral actions of centrally administered corticotropin-releasing factor (CRF) are likely subserved by multiple brain targets and functional effector systems. The present studies compared effects of two CRF ligands, a full, post-synaptic CRF receptor agonist (rat/human CRF(1-41)) and a CRF binding protein ligand inhibitor (rat/human CRF(6-33)) in a behavioral testing battery sensitive to arousal, fear-like and aversive processes in Wistar rats. The profile of global efficacy for the centrally administered CRF receptor agonist was characterized by low dose (0.5-1.0 microg) arousal-like effects in locomotor and conditioned ambulation contexts and by high dose (5-25 microg) conditioned immobility, taste aversion and place aversion. In contrast, a profile of limited efficacy for the centrally administered CRF binding protein ligand inhibitor included only dose dependent motor activating and facilitation of fear conditioning effects without any of the anxiogenic-like or aversive properties of CRF agonist administration. These results suggest that arousal-like activation is a fundamental, physiologically relevant consequence of brain CRF system stimulation whereas aversive and anxiety-like effects reflect pharmacological actions of a CRF receptor agonist.

Therapeutic potential of CRF receptor antagonists: a gut-brain perspective.

Activation of the corticotropin-releasing factor (CRF) family of neuropeptide receptors in the brain and periphery appears to mediate stress-related changes in a variety of physiological and functional domains. Comparative pharmacology of CRF receptor agonists suggests that CRF, urocortin, sauvagine and urotensin consistently mimic, and conversely peptide CRF receptor antagonists lessen, the functional consequences of stressor exposure. Together with the development of novel non-peptide CRF receptor antagonists, a growing number of CRF receptor selective ligands are available to elucidate the neurobiology and physiological role of CRF systems. The present review considers available preclinical evidence as well as results from one Phase II clinical trial which address the hypothesis that CRF receptor antagonists may represent a new option for pharmacotherapy of stress-related disorders.

Steroid structure and pharmacological properties determine the anti-amnesic effects of pregnenolone sulphate in the passive avoidance task in rats.

Pregnenolone sulphate (PREGS) has generated interest as one of the most potent memory-enhancing neurosteroids to be examined in rodent learning studies, with particular importance in the ageing process. The mechanism by which this endogenous steroid enhances memory formation is hypothesized to involve actions on glutamatergic and GABAergic systems. This hypothesis stems from findings that PREGS is a potent positive modulator of N-methyl-d-aspartate receptors (NMDARs) and a negative modulator of gamma-aminobutyric acid(A) receptors (GABA(A)Rs). Moreover, PREGS is able to reverse the amnesic-like effects of NMDAR and GABA(A)R ligands. To investigate this hypothesis, the present study in rats examined the memory-altering abilities of structural analogs of PREGS, which differ in their modulation of NMDAR and/or GABA(A)R function. The analogs tested were: 11-ketopregnenolone sulphate (an agent that is inactive at GABA(A)Rs and NMDARs), epipregnanolone ([3beta-hydroxy-5beta-pregnan-20-one] sulphate, an inhibitor of both GABA(A)Rs and NMDARs), and a newly synthesized (-) PREGS enantiomer (which is identical to PREGS in effects on GABA(A)Rs and NMDARs). The memory-enhancing effects of PREGS and its analogs were tested in the passive avoidance task using the model of scopolamine-induced amnesia. Both PREGS and its (-) enantiomer blocked the effects of scopolamine. The results show that, unlike PREGS, 11-ketopregnenolone sulphate and epipregnanolone sulphate failed to block the effect of scopolamine, suggesting that altering the modulation of NMDA receptors diminishes the memory-enhancing effects of PREGS. Moreover, enantioselectivity was demonstrated by the ability of natural PREGS to be an order of magnitude more effective than its synthetic enantiomer in reversing scopolamine-induced amnesia. These results identify a novel neuropharmacological site for the modulation of memory processes by neuroactive steroids.

Comparison of central administration of corticotropin-releasing hormone and urocortin on food intake, conditioned taste aversion, and c-Fos expression.

Corticotropin-releasing hormone (CRH) is a potent regulator of the hypothalamic-pituitary-adrenal axis, and reduces food intake when administered into the third cerebral ventricle (i3vt). However, CRH also promotes conditioned taste aversion (CTA) learning which indicates that its anorectic effects are accompanied by aversive consequences that would reduce food intake independently of energy regulation. Urocortin (Ucn) is a closely related mammalian peptide that binds to both identified CRH receptor subtypes and also reduces food intake when administered i3vt. The present experiments compared the aversive consequences of i3vt administration of CRH and Ucn at doses that produced comparable decrements in food intake. Experiment 1 found that 1.0 microg Ucn and 2.0 microg CRH produced similar reductions in food intake. Experiment 2 demonstrated that, at these doses, CRH but not Ucn promoted robust and reliable CTA learning. A third experiment showed comparable increased c-Fos-like immunoreactivity after Ucn and CRH in forebrain and hindbrain structures associated with food intake. It is concluded that Ucn, at doses that reduce food intake to levels like that observed after administration of CRH, do not produce similarly aversive consequences.

Selective impairment of corticotropin-releasing factor1 (CRF1) receptor-mediated function using CRF coupled to saporin.

CRF is the main component in the brain neuropeptide effector system responsible for the behavioral, endocrine, and physiological activation that accompanies stress activation. Reduced CRF system activation plays a role in the etiology of a variety of psychiatric and metabolic disease states. We have developed a novel protein conjugate that joins native rat/human CRF to a ribosome-inactivating protein, saporin (CRF-SAP), for the purpose of targeted inactivation of CRF receptor-expressing cells. Cytotoxicity measurements revealed that CRF-SAP (1-100 nM) produced concentration-dependent and progressive cell death over time in CRF1 receptor-transfected L cells, but at similar concentrations had no effect on CRF2alpha receptor-transfected cells. The CRF-SAP-induced toxicity in CRF1-transfected cells was prevented by coincubation with the competitive CRF1/CRF2 receptor peptide antagonist, [D-Phe12]CRF-(12-41), or the selective nonpeptide CRF1 receptor antagonist, NBI 27914. Finally, in cultured rat pituitary cells that express native CRF1 receptors, CRF-SAP suppressed CRF-induced (1 nM) ACTH release. GnRH (1-10 nM) stimulated LH release was also assessed in the same pituitary cultures. Although there was a slight decrease in LH release from these cultures, this decrease was observed with CRF-SAP or SAP alone, suggesting that the response was nonspecific. Taken together, these results suggest the utility of CRF-SAP as a specific and subtype-selective tool for long term impairment of CRF1 receptor-expressing cells.

Influence of promoter DNA topology on sequence-specific DNA binding and transactivation by tumor suppressor p53.

Transcriptional activation by the tumor suppressor p53 is regulated at multiple levels, including posttranslational modifications of the p53 protein, interaction of p53 with various regulatory proteins, or at the level of sequence-specific DNA binding to the response elements in p53's target genes. We here propose as an additional regulatory mechanism that the DNA topology of p53-responsive promoters may determine the interaction of p53 with its target genes. We demonstrate that sequence-specific DNA binding (SSDB) and transcriptional activation by p53 of the mdm2 promoter is inhibited when this promoter is present in supercoiled DNA, where it forms a non-B-DNA structure which spans the p53-responsive elements. Relaxation of the supercoiled DNA in vitro resulted in conversion of the non-B-DNA to a B-DNA conformation within the mdm2 promoter, and correlated with an enhanced SSDB of p53 and an elevated expression of a reporter gene. In contrast, sequence specific DNA binding and transcriptional activation of the p21 promoter were not inhibited by DNA supercoiling. We propose that conformational alterations within p53-responsive sites, which either promote or prohibit sequence specific DNA binding of p53, are an important feature in orchestrating the activation of different p53 responsive promoters.

A role for corticotropin releasing factor and urocortin in behavioral responses to stressors.

Corticotropin-releasing factor (CRF) and CRF-related neuropeptides have an important role in the central nervous system to mediate behavioral responses to stressors. CRF receptor antagonists are very effective in reversing stress-induced suppression and activation in behavior. An additional CRF-like neuropeptide, urocortin, has been identified in the brain and has a high affinity for the CRF-2 receptor in addition to the CRF-1 receptor. Urocortin has many of the effects of CRF but also is significantly more potent than CRF in decreasing feeding in both meal-deprived and free-feeding rats. In mouse genetic models, mice over-expressing CRF show anxiogenic-like responses compared to wild-type mice, and mice lacking the CRF-1 receptor showed an anxiolytic-like behavioral profile compared to wild-type mice. Results to date have led to the hypothesis that CRF-1 receptors may mediate CRF-like neuropeptide effects on behavioral responses to stressors, but CRF-2 receptors may mediate the suppression of feeding produced by CRF-like neuropeptides. Brain sites for the behavioral effects of CRF include the locus coeruleus (LC), paraventricular nucleus (PVN) of the hypothalamus, the bed nucleus of the stria terminalis (BNST), and the central nucleus of the amygdala. CRF may also be activated during acute withdrawal from all major drugs of abuse, and recent data suggest that CRF may contribute to the dependence and vulnerability to relapse associated with chronic administration of drugs of abuse. These data suggest that CRF systems in the brain have a unique role in mediating behavioral responses to diverse stressors. These systems may be particularly important in situations were an organism must mobilize not only the pituitary adrenal system, but also the central nervous system in response to environmental challenge. Clearly, dysfunction in such a fundamental brain-activating system may be the key to a variety of pathophysiological conditions involving abnormal responses to stressors such as anxiety disorders, affective disorders, and anorexia nervosa.