Sub-seafloor sediments are populated by large numbers of microbial cells but not much is known about their metabolic activities, growth rates and carbon assimilation pathways. Here we introduce a new method enabling the sensitive detection of microbial lipid production and the distinction of auto- and heterotrophic carbon assimilation. Application of this approach to anoxic sediments from a Swedish fjord allowed to compare the activity of different functional groups, the growth and turnover times of the bacterial and archaeal communities. The assay involves dual stable isotope probing (SIP) with deuterated water (D(2) O) and (13) C(DIC)(dissolved inorganic carbon). Culture experiments confirmed that the D content in newly synthesized lipids is in equilibrium with the D content in labelled water, independent of whether the culture grew hetero- or autotrophically. The ratio of (13) C(DIC) to D(2) O incorporation enables distinction between these two carbon pathways in studies of microbial cultures and in environmental communities. Furthermore, D(2) O-SIP is sufficiently sensitive to detect the formation of few hundred cells per day in a gram of sediment. In anoxic sediments from a Swedish fjord, we found that > 99% of newly formed lipids were attributed to predominantly heterotrophic bacteria. The production rate of bacterial lipids was highest in the top 5 cm and decreased 60-fold below this depth while the production rate of archaeal lipids was rather low throughout the top meter of seabed. The contrasting patterns in the rates of archaeal and bacterial lipid formation indicate that the factors controlling the presence of these two lipid groups must differ fundamentally.

OBJECTIVE: The pathophysiology of depression and the effects of antidepressant treatment are hypothesized to be related to hippocampal structural changes. This study aims to investigate the effect of electroconvulsive seizures on behavior and hippocampal structure in a rat model of depression. METHODS: Flinders Sensitive Line and Flinders Resistant Line rats were treated daily for ten days with either electroconvulsive seizures or sham treatment. The behavior was evaluated using the forced swim test. Design-based stereological methods were used to quantify the hippocampal volume and the numbers of neurons and glial cells in specific hippocampal subregions. RESULTS: The basal level of hippocampal volume and neuron number differed significantly between the two rat strains, and a trend toward the FSL strain having more glial cells was found. The structural differences found between the sham treated animals were counteracted by ECS treatment, which also normalized the behavior. ECS treatment increased the number of glial cells in hilus significantly in the FRL rats, and with the same tendency for the FSL rats. CONCLUSION: Our results indicate that along with hippocampal neurogenesis, also gliogenesis may be involved in the pathophysiology of depression and in the effect of antidepressant treatment. The underlying mechanisms remain unknown and further investigations are required to clarify whether the structural changes are necessary to induce a therapeutic effect of antidepressant treatment or if they rather represent an epiphenomenon. © 2012 Wiley Periodicals, Inc.

There are conflicting results from behavioural studies regarding whether the activation or inhibition of the cGMP-nitric oxide (NO) pathway induces anxiolytic-like behaviour. Sildenafil, an inhibitor of cGMP-selective phosphodiesterase-5, increases anxiety acutely, but previous evidence suggests that its chronic administration may be anxiolytic, and could involve a cholinergic interaction. We used the Flinders Sensitive Line (FSL) rat, a genetic model of depression that presents with increased anxiety- and depression-like behaviour, to investigate the action of chronic treatment with the PDE5 inhibitors sildenafil or tadalafil, with/without atropine on social interaction behaviour, a correlate for anxiety. Fluoxetine was used as positive control, with validation performed using Flinders Resistant Line (FRL) rats. In order to relate behavioural changes to brain penetration, we determined the concentration of sildenafil in cortex and hippocampus of rats following the schedule above using LC-MS/MS. FSL rats displayed significantly reduced social interactive behaviour than FRL rats, while sildenafil, tadalafil, and fluoxetine significantly reversed these deficits. Atropine did not exert effects on social interactive behaviour, nor did it modulate the effects of sildenafil or tadalafil. Sildenafil was present in cortex and hippocampus regions in lower nanomolar concentrations after chronic treatment, in agreement with the binding to PDE5 required for pharmacological effects. This study emphasizes the complicated regulation of anxiety by the cGMP-NO system, and provides supporting evidence for an anxiolytic action after the chronic activation of this pathway. As far as we know this is also the first report to formally demonstrate that sildenafil effectively crosses the blood-brain barrier to elicit central effects.

Stress related diseases such as depression and anxiety have a high degree of co morbidity, and represent one of the greatest therapeutic challenges for the twenty-first century. The present chapter will summarize existing rodent models for research in psychiatry, mimicking depression- and anxiety-related diseases. In particular we will highlight the use of selective breeding of rodents for extremes in stress-related behavior. We will summarize major behavioral, neuroendocrine and neuronal parameters, and pharmacological interventions, assessed in great detail in two rat model systems: The Flinders Sensitive and Flinders Resistant Line rats (FSL/FRL model), and rats selectively bred for high (HAB) or low (LAB) anxiety related behavior (HAB/LAB model). Selectively bred rodents also provide an excellent tool in order to study gene and environment interactions. Although it is generally accepted that genes and environmental factors determine the etiology of mental disorders, precise information is limited: How rigid is the genetic disposition? How do genetic, prenatal and postnatal influences interact to shape adult disease? Does the genetic predisposition determine the vulnerability to prenatal and postnatal or adult stressors? In combination with modern neurobiological methods, these models are important to elucidate the etiology and pathophysiology of anxiety and affective disorders, and to assist in the development of new treatment paradigms.

Recent genetic evidence has implicated the bromodomain containing 1 gene (BRD1) with brain development and susceptibility to Schizophrenia and Bipolar Disorder. The BRD1 protein, which is essential for acetylation of histone H3K14, is a putative regulator of transcription during brain development and in the mature CNS. However, several issues remain to be clarified for example regarding the regulation of the BRD1 gene upon environmental interventions. Chronic restraint stress (CRS) in rats represents an environmental method for induction of morphological and functional changes in the hippocampus and the prefrontal cortex. In order to investigate whether the expression of the rat Brd1 gene may be regulated during such conditions, Brd1 mRNA and protein levels in hippocampus and prefrontal cortex extracts from rats subjected to either 1/2 or 6h of CRS per day for 21days were measured. We found a significant 2-fold up-regulation of long exon 7 splice variants of the Brd1 gene (Brd1-L) in hippocampus in both groups of CRS rats compared to controls. Concomitantly, we found a similar up-regulation of the BRD1 protein. In prefrontal cortex, we found no significant differences in Brd1 mRNA or protein levels. As selective histone deacetylase (HDAC) inhibitors not only preserve stress-induced hyperacetylation of histone H3K14 but also have hippocampal-dependent antidepressant-like activity, we propose that BRD1 by its intrinsic acetylation activity towards histone H3K14 is a player in the regulatory processes underlying adaptation to stress in the mature CNS.

Depression is a heterogeneous disorder displaying a range of symptoms including feelings of despair and social withdrawal. Social isolation may complicate the progression of depression and have effects on both behaviour and physiology. The aim of this study was to investigate the effects of social isolation on behavioural and metabolic parameters in a genetic rat model of depression, the Flinders Sensitive and Resistant Line (FSL/FRL) rats. Rats were housed either individually (social isolation) or pair-housed for 5weeks, and subjected to behavioural testing and metabolic evaluation. We found that social isolation erased the characteristic difference in depressive-like behaviour, measured as immobility in the forced swim test, between the FSL and FRL rats. Social isolation affected both strains equally in impairing object recognition memory, while leading to an increased explorative behaviour in the elevated plus maze test. Surprisingly, single-housed FRL rats showed an increased food intake compared to pair-housed FRL rats, whereas no difference in food intake or body weight was evident in FSL rats. Our results indicate that social isolation for 5weeks causes behavioural alterations, independent of strain. As the changes in appetite were only observed in the FRL rats, this may suggest that this strain responds to the stress of isolation by a change in feeding behaviour.

Methylene blue (MB) has been shown to act at multiple cellular and molecular targets and as a result possesses diverse medical applications. Among these is a high potency reversible inhibition of monoamine oxidase A (MAO-A) that may, at least in part, underlie its adverse effects but also its psycho- and neuromodulatory actions. MB is metabolized to yield N-demethylated products of which azure B, the monodemethyl species, is the major metabolite. Similar to MB, azure B also displays a variety of biological activities and may therefore contribute to the pharmacological profile of MB. Based on these observations, the present study examines the interactions of azure B with recombinant human MAO-A and -B. The results show that azure B is a potent MAO-A inhibitor (IC(50)=11nM), approximately 6-fold more potent than is MB (IC(50)=70nM) under identical conditions. Measurements of the time-dependency of inhibition suggest that the interaction of azure B with MAO-A is reversible. Azure B also reversibly inhibits the MAO-B isozyme with an IC(50) value of 968nM. These results suggest that azure B may be a hitherto under recognized contributor to the pharmacology and toxicology of MB by blocking central and peripheral MAO-A activity and as such needs to be considered during its use in humans and animals.

Lu AA21004 (1-[2-(2,4-dimethylphenyl-sulfanyl)-phenyl]-piperazine) is a human serotonin (h5-HT)(3A) receptor antagonist (K(i)=3.7nM), h5-HT(7) receptor antagonist (K(i)=19nM), h5-HT(1B) receptor partial agonist (K(i)=33nM), h5-HT(1A) receptor agonist (K(i)=15nM) and a h5-HT transporter (hSERT) inhibitor (K(i)=1.6nM)(Bang-Andersen et al., 2011). Here we confirm that Lu AA21004 is a partial h5-HT(1B) receptor agonist (EC(50)=460nM, IA=22%) using a whole cell cAMP-based assay and demonstrate that Lu AA21004 is a rat (r)5-HT(7) receptor antagonist (K(i)=200nM and IC(50)=2080nM). In vivo, Lu AA21004 occupies the r5-HT(1B) receptor and rSERT (ED(50)=3.2mg/kg and 0.4mg/kg, respectively) after subcutaneous (sc) administration and is a 5-HT(3) receptor antagonist in the Bezold-Jarisch reflex assay (ED(50)=0.11mg/kg sc). In rat microdialysis experiments, Lu AA21004 (2.5-10.0mg/kg sc) increased extracellular 5-HT, dopamine and noradrenaline in the medial prefrontal cortex and ventral hippocampus. Lu AA21004, 5mg/kg/day for 3 days (minipumps sc), corresponding to 41% rSERT occupancy, significantly increased extracellular 5-HT in the ventral hippocampus. Furthermore, the 5-HT(3) receptor antagonist, ondansetron potentiated the increase in extracellular levels of 5-HT induced by citalopram. Lu AA21004 has antidepressant- and anxiolytic-like effects in the rat forced swim (Flinders Sensitive Line) and social interaction and conditioned fear tests (minimal effective doses: 7.8, 2.0 and 3.9mg/kg). In conclusion, Lu AA21004 mediates its pharmacological effects via two pharmacological modalities: SERT inhibition and 5-HT receptor modulation. In vivo, this results in enhanced release of several neurotransmitters and antidepressant- and anxiolytic-like profiles at doses where targets in addition to the SERT are occupied. The multimodal activity profile of Lu AA21004 is distinct from current antidepressants.

Electroconvulsive treatment (ECT) is considered the most effective treatment of depression. Recent studies demonstrate that electroconvulsive seizures (ECS), the animal variant of ECT, robustly stimulate hippocampal cell proliferation. However, the mechanisms underlying the cellular and molecular responses to ECS are not yet fully understood. A leading hypothesis of depression suggests that neurotrophic factors/growth factors play a critical role. Particularly the vascular endothelial growth factor (VEGF) is considered important, as it has been demonstrated that hippocampal VEGF expression is induced by ECS and that signaling through the VEGF receptor, Flk-1 (VEGFR2), is required for cell proliferation. VEGF expression is believed to be regulated by two distinct mTOR (mammalian Target of Rapamycin)-containing multiprotein complexes, mTORC1 and mTORC2, respectively. The present study was undertaken to investigate the effect of ECS on the expression of VEGF, cognate receptors, mTORC1, and mTORC2 in the frontal cortex and the hippocampus. Using male Sprague-Dawley rats, the messenger RNA (mRNA) levels were measured by quantitative real time polymerase chain reaction (real-time qPCR) in 3 groups: Sham, acute (after one ECS), and repeated (ECS every day for 10 days). VEGF, VEGFR2, and components from mTORC1 were affected by repeated ECS, indicating that mediation of VEGF via mTORC1 is important for the effect of ECS. Synapse, 2011. © 2011 Wiley-Liss, Inc.

Microbial degradation of substrates to terminal products is commonly understood as a unidirectional process. In individual enzymatic reactions, however, reversibility (reverse reaction and product back flux) is common. Hence, it is possible that entire pathways of microbial degradation are associated with back flux from the accumulating product pool through intracellular intermediates into the substrate pool. We investigated carbon and sulfur back flux during the anaerobic oxidation of methane (AOM) with sulfate, one of the least exergonic microbial catabolic processes known. The involved enzymes must operate not far from the thermodynamic equilibrium. Such an energetic situation is likely to favor product back flux. Indeed, cultures of highly enriched archaeal-bacterial consortia, performing net AOM with unlabeled methane and sulfate, converted label from (14)C-bicarbonate and (35)S-sulfide to (14)C-methane and (35)S-sulfate, respectively. Back fluxes reached 5% and 13%, respectively, of the net AOM rate. The existence of catabolic back fluxes in the reverse direction of net reactions has implications for biogeochemical isotope studies. In environments where biochemical processes are close to thermodynamic equilibrium, measured fluxes of labeled substrates to products are not equal to microbial net rates. Detection of a reaction in situ by labeling may not even indicate a net reaction occurring in the direction of label conversion but may reflect the reverse component of a so far unrecognized net reaction. Furthermore, the natural isotopic composition of the substrate and product pool will be determined by both the forward and back flux. This finding may have to be considered in the interpretation of stable isotope records.