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Clinical studies suggest a link between depression and dysfunctional tryptophan (TRP) metabolism. Even though depression is twice as prevalent in women as men, the impact of the estrous cycle on TRP metabolism is not well-understood. Here we investigated 13 kynurenine and serotonin metabolites in female Flinders Sensitive Line (FSL) rats, a genetic rat model of depression. FSL rats and controls (Flinders Resistant Line rats), 12-20weeks old, were subject to the forced swim test (FST), a commonly used measure of depression-like behavior. Open field was used to evaluate locomotor ability and agoraphobia. Subsequently, plasma and hemispheres were collected and analyzed for their content of TRP metabolites using liquid chromatography-tandem mass spectrometry. Vaginal saline lavages were obtained daily for ⩾2 cycles. To estimate the effects of sex and FST we included plasma from unhandled, naïve male FSL and FRL rats. Female FSL rats showed a depression-like phenotype with increased immobility in the FST, not confounded by anxiety. In the brain, 3-hydroxykynurenine was increased whereas anthranilate and 5-hydroxytryptophan were decreased. In plasma, anthranilate and quinolinate levels were lower in FSL rats compared to the control line, independent of sex and FST. The estrous cycle neither impacted behavior nor TRP metabolite levels in the FSL rat. In conclusion, the female FSL rat is an interesting preclinical model of depression with altered TRP metabolism, independent of the estrous cycle. The status of the pathway in brain was not reflected in the plasma, which may indicate that an inherent local, cerebral regulation of TRP metabolism occurs., (Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.)

Rationale: The mechanisms mediating ketamine's antidepressant effect have only been partly resolved. Recent preclinical reports implicate serotonin (5-hydroxytryptamine; 5-HT) in the antidepressant-like action of ketamine. Vortioxetine is a multimodal-acting antidepressant that is hypothesized to exert its therapeutic activity through 5-HT reuptake inhibition and modulation of several 5-HT receptors., Objectives: The objective of this study was to evaluate the therapeutic-like profiles of S-ketamine, vortioxetine, and the serotonin reuptake inhibitor fluoxetine in response to manipulation of 5-HT tone., Method: Flinders Sensitive Line (FSL) rats, a genetic model of depression, were depleted of 5-HT by repeated administration of 4-chloro-DL-phenylalanine methyl ester HCl (pCPA). Using pCPA-pretreated and control FSL rats, we investigated the acute and sustained effects of S-ketamine (15 mg/kg), fluoxetine (10 mg/kg), or vortioxetine (10 mg/kg) on recognition memory and depression-like behavior in the object recognition task (ORT) and forced swim test (FST), respectively., Results: The behavioral phenotype of FSL rats was unaffected by 5-HT depletion. Vortioxetine, but not fluoxetine or S-ketamine, acutely ameliorated the memory deficits of FSL rats in the ORT irrespective of 5-HT tone. No sustained effects were observed in the ORT. In the FST, all three drugs demonstrated acute antidepressant-like activity but only S-ketamine had sustained effects. Unlike vortioxetine, the antidepressant-like responses of fluoxetine and S-ketamine were abolished by 5-HT depletion., Conclusions: These observations suggest that the acute and sustained antidepressant-like effects of S-ketamine depend on endogenous stimulation of 5-HT receptors. In contrast, the acute therapeutic-like effects of vortioxetine on memory and depression-like behavior may be mediated by direct activity at 5-HT receptors.

Inflammatory reactions are involved in a diversity of diseases, including major depressive disorder. Cytokines act as intercellular signaling molecules and mediators of inflammation between the periphery and the brain. Within the brain, evidence from animal studies of acute inflammation has shown that elevated cytokine levels are linked to behavioral responses of sickness and depression-like behavior. Although chronic inflammation is more translational to human depression than acute studies, little is known on central cytokine expression and associated behavioral responses following chronic immune challenges. The present study assessed behavioral changes and a selection of cytokines in the brain and in the blood in rats randomized to receive a single or 8week administration with either lipopolysaccharide (LPS, 600μg/kg, i.p.) or saline. Acute and long-term LPS treatments caused similar sickness and depression-like behavior. Chronic LPS administration did not have an effect on blood cytokine levels, indicating endotoxin tolerance, whereas increased fasting blood glucose was observed, indicating insulin resistance, a metabolic consequence of chronic inflammation. While a single LPS injection produced a generalized cytokine response in the brain, long-term LPS administration produced a specific central cytokine response with increased interleukin (IL)-1β and interferon (IFN)-γ. These cytokines can explain the behavioral changes observed, and could indicate microglia activation, although future studies are needed to uncover this assumption. Taken together, although the behavioral outcome was similar between acute and chronic LPS administration, the central cytokine response was distinct. As the long-term LPS paradigm also posed a metabolic demand, this setting may reflect a more translational insight into inflammatory reactions in human depression, and could prove useful for assessing cytokine down-stream effects and experimental antidepressant drug products., (Copyright © 2015. Published by Elsevier B.V.)

Obesity-related inflammation may have a causal role in the development of diabetes and insulin resistance, and studies using animal models of chronic experimental endotoxemia have shown the link. However, many studies use only males, and much less is known about the role of obesity-related inflammation in females. Therefore, we addressed how experimentally induced chronic inflammation affects body mass, energy intake, and glucose metabolism in female rats. Adult female Sprague Dawley rats were instrumented with slow release pellets that delivered a constant daily dose of 53 or 207 μg of lipopolysaccharide (LPS) per rat for 60 days. Control rats were instrumented with vehicle pellets. Due to inflammatory nature of high-fat diet (HFD) half of the rats received HFD (60% of calories from lard), while the other half remained on control diet to detect possible interactions between two modes of induced inflammation. Our results showed that chronic LPS administration increased female rat body mass and calorie intake in a dose-dependent manner, and that HFD further exacerbated these effects. Despite these effects, no effects of LPS and HFD were evident on female rat glucose metabolism. Only LPS elevated expression of inflammatory markers in the hypothalamus. To conclude, female rats respond to experimentally induced chronic inflammation by increasing body mass, but do not develop glucose intolerance in the given period of time., (© 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.)

Immunotherapy with the cytokine interferon-alpha (IFN-α) can induce symptoms of depression, and it is likely that the tryptophan-kynurenine pathway may be involved in this regard. In this study we investigated the effects of IFN-α on depression-like behaviour and central metabolites of the tryptophan-kynurenine pathway in rats. Secondly, we explored the modulating effects of an antidepressant (imipramine) and anti-inflammatory drug (celecoxib) on IFN-α-induced behavioural and pathophysiological changes in the brain. The following treatment groups were used: Control (saline), IFN-α (6×10(4)IU/kg s.c.), IFN-α+imipramine or IFN-α+celecoxib. Drugs were administered daily for 1 week. IFN-α treatment induced depression-like behaviour by increasing immobility in the forced swim test (FST), and decreased tryptophan levels in the brain. There was a trend for an increased kynurenine/tryptophan ratio, indicative of indoleamine 2,3-dioxygenase (IDO) activation, and increased quinolinic acid in the hippocampus. Imipramine decreased immobility in the FST, but did not reverse the IFN-α-induced changes in the tryptophan-kynurenine pathway. There was a trend for celecoxib to decrease immobility and to reverse the IFN-α-induced increase in the kynurenine/tryptophan ratio. Thus, our study provides further evidence for IFN-α-induced depression-like behaviour through central changes of the tryptophan-kynurenine pathway., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor A (VEGF) have been suggested to play a role in the pathophysiology of depression. The neurotrophic model of depression hypothesises that the serum level of e.g. BDNF is decreased during depression and increased in response to treatment. The aim of the present study was to investigate BDNF and VEGF as potential predictors of response to antidepressant treatment., Methods: We investigated the longitudinal associations between depression scores and serum levels of these neurotrophic factors during antidepressant treatment in 90 individuals with depression of at least moderate severity. Serum levels were measured at baseline and after 8 and 12 weeks of treatment with nortriptyline or escitalopram., Results: No baseline or longitudinal correlations between depression scores and serum levels of BDNF and VEGF were found, and the baseline serum levels did not predict the MADRS depression score after 12 weeks of treatment or the improvement in depression scores. Interestingly, we observed a significant baseline and longitudinal correlation between serum levels of BDNF and VEGF. The two classes of antidepressant treatment did not affect the results differently., Limitations: Information on potential factors influencing the serum levels is missing., Conclusion: Our results do not support the neurotrophic model of depression, since a significant decrease in serum BDNF and VEGF levels after 12 weeks of antidepressant treatment was observed. Our study encourages future studies with large sample sizes, more observations and a longer follow-up period., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Electroconvulsive therapy (ECT) remains the treatment of choice for patients with severe or drug-resistant depressive disorders, yet the mechanism behind its efficacy remains poorly characterized. In the present study, we used electroconvulsive seizures (ECS), an animal model of ECT, to identify proteins possibly involved in the preventive effect of ECS on stress-induced neuronal atrophy in the hippocampus. Rats were stressed daily using the 21-day 6h daily restraint stress paradigm and subjected to sham seizures, a single ECS on the last day of the restraint period or daily repeated seizures for 10 consecutive days during the end of the restraint period. Consistent with previous findings, dendritic atrophy was observed in the CA3c hippocampal region of chronically stressed rats. In addition, we confirmed our recent findings of increased spine density in the CA1 region following chronic restraint stress. The morphological alterations in the CA3c area were prevented by treatment with ECS. On the molecular level, we showed that the synaptic proteins Homer1 and Spinophilin are targeted by ECS. Repeated ECS blocked stress-induced up-regulation of Spinophilin protein levels and further increased the stress-induced up-regulation of Homer1. Given the roles of Spinophilin in the regulation of AMPA receptors and Homer1 in the regulation of metabotropic glutamate receptors (mGluRs), our data imply the existence of a mechanism where ECS regulate cell excitability by modulating AMPA receptor function and mGluR related calcium homeostasis. These molecular changes could potentially contribute to the mechanism induced by ECS which prevents the stress-induced morphological changes in the CA3c region., (Copyright © 2015 Elsevier B.V. and ECNP. All rights reserved.)

Objective: Gut microbiota (GM) has previously been associated with alterations in rodent behaviour, and since the GM is affected by the diet, the composition of the diet may be an important factor contributing to behavioural changes. Interestingly, a magnesium restricted diet has been shown to induce anxiety and depressive-like behaviour in humans and rodents, and it could be suggested that magnesium deficiency may mediate the effects through an altered GM., Methods: The present study therefore fed C57BL/6 mice with a standard diet or a magnesium deficient diet (MgD) for 6 weeks, followed by behavioural testing in the forced swim test (FST) to evaluate depressive-like behaviour. An intraperitoneal glucose tolerance test (GTT) was performed 2 day after the FST to assess metabolic alterations. Neuroinflammatory markers were analysed from hippocampus. GM composition was analysed and correlated to the behaviour and hippocampal markers., Results: It was found that mice exposed to MgD for 6 weeks were more immobile than control mice in the FST, suggesting an increased depressive-like behaviour. No significant difference was detected in the GTT. GM composition correlated positively with the behaviour of undisturbed C57BL/6 mice, feeding MgD diet altered the microbial composition. The altered GM correlated positively to the hippocampal interleukin-6., Conclusion: In conclusion, we hypothesise that imbalances of the microbiota-gut-brain axis induced by consuming a MgD diet, contributes to the development of depressive-like behaviour.

In the present study the central and peripheral regulation of VEGF, its cognate receptors, and regulators were examined after acute and chronic restraint stress in rats. After chronic restraint stress (6 h per day for 21 days) the protein levels of VEGF (175 ± 24%) and its receptor VEGFR-2 (169 ± 17%) increased significantly in the prefrontal cortex (A and B). mRNA levels of VEGFR-2 (132 ± 11%) were also significantly increased (D). In the hippocampus no significant changes were observed at the mRNA or protein levels. In serum there was a tendency towards increased VEGF protein expression after both acute and chronic restraint stress (C)., (© 2015 Wiley Periodicals, Inc.)

Recent studies suggest that the angiogenic cytokine vascular endothelial growth factor (VEGF) is involved in the pathogenesis of depression. However, only a few studies have investigated serum VEGF levels in individuals with depression, or the possible association between genetic variants within the VEGF gene and depression. The purpose of the present study was to investigate differences between serum VEGF levels in individuals with depression vs. control individuals, and associations between genetic markers located within VEGF and depression. In addition, determinants of the serum VEGF levels were identified. One-hundred and fifty-five depressed subjects and 280 controls were included in the study. All individuals returned a questionnaire and participated in a semi-structured diagnostic interview. Eleven single nucleotide polymorphisms were successfully analysed. VEGF levels were measured in serum by immunoassay and independent determinants of the serum VEGF level were assessed by generalized linear models.The main findings were that depression, severity of depression, previous depressive episodes, age and body mass index (BMI) were associated with higher serum VEGF levels. The genetic marker rs10434 was significantly associated with depression after correction for multiple testing, but not with the serum VEGF level. Our final model included depression and BMI as predictors of serum VEGF levels. Our study suggests a role for circulating serum VEGF in depression. Furthermore, our data also demonstrate that other factors than a diagnosis of depression influence the serum VEGF level. The importance of these factors should be emphasized when studies are compared.

Objectives: Major depressive disorder (MDD) is accompanied with an imbalance in the immune system and cardiovascular impairments, such as atherosclerosis. Several mechanisms have been pointed out to underlie this rather unexpected association, and among them the activity of myeloperoxidase (MPO). The aim of our study was to find compounds that inhibit both MPO and serotonin transporter (SERT) for treating MDD associated with cardiovascular diseases., Methods: SERT inhibition was assessed with measuring of [(3) H]-serotonin uptake using HEK-293 MSR cells. MPO inhibition was determined by taurine chloramine test on 3-(aminoalkyl)-5-fluoroindole derivatives and on clinically relevant antidepressants. All kinetic measurements were performed using a temperature-controlled stopped-flow apparatus (model SX-18 MV). Promising lead compounds were docked onto SERT 3D structure modelled using the LeuT structure complexed to tryptophan (PDB code 3F3A). Their toxicological profile was also assessed., Key Findings: 3-(aminoalkyl)-5-fluoroindole derivative with 5 carbons on the side chain and paroxetine showed the best activity on both MPO and SERT at the nanomolar range. Paroxetine was found to be the first irreversible MPO inhibitor at nanomolar concentrations., Conclusions: Our results put forward the first hybrid molecule (compound 25) and drug (paroxetine) that can be especially used in MDD associated with inflammatory syndrome., (© 2014 Royal Pharmaceutical Society.)

Background: The hippocampal volume is reduced in patients with major depression. Exercise leads to an increased hippocampal volume in schizophrenia and in healthy old adults. The effect of exercise on hippocampal volume is potentially mediated by brain derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF), and insulin like growth factor 1 (IGF-1). The aim of this trial was to assess the effect of an aerobic exercise intervention on hippocampal volume and serum BDNF, VEGF, and IGF-1 in patients with major depression., Methods: Patients were randomized to an aerobic exercise intervention (n=41) or a control condition (n=38). Both interventions consisted of three supervised sessions per week during a three months period., Results: Post-intervention the increase in maximal oxygen uptake was 3.90 ml/kg/min (SD 5.1) in the aerobic exercise group and 0.95 ml/kg/min (SD 6.2) in the control group (p=0.03). The hippocampal volume, BDNF, VEGF, or IGF-1 did not differ between the two groups. Post-hoc we found a positive association between change in hippocampal volume and verbal memory (Rho=0.27; p=0.05) and change in hippocampal volume and depressive symptoms (Rho=0.30; p=0.03)., Limitations: Participation was low in both groups corresponding to an average participation of one session per week., Conclusion: Despite a significant increase in maximal oxygen uptake, a pragmatic exercise intervention did not increase hippocampal volume or resting levels of neurotrophines in out-patients with mild to moderate major depression. Trial identifier: ClinicalTrials.gov (NCT00695552)., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The purpose of this study was to assess the association between IL-6 and CRP with depressive items and cognitive function. We included 112 outpatients with major depression from an exercise trial and 57 healthy controls. IL-6, high sensitive CRP (hsCRP), and cognitive function were assessed in all subjects. After baseline assessment, patients were randomised to either a 3months exercise intervention or an exercise control group. Post-intervention IL-6, hsCRP, depressive symptoms, and cognitive function were reassessed in the patient group. IL-6 and hsCRP were significantly increased in depressed patients compared to healthy controls (p=0.02 and 0.04). These differences were no longer significant after adjustment for lifestyle associated variables. We found no association between immune markers and specific depressive symptoms at baseline or as change over time. Regarding the cognitive tests, IL-6 was positively associated with Serial sevens (p=0.008) and hsCRP was inversely associated with Trail making A (p=0.02) and design fluency (p=0.001) at baseline. At 3months follow-up IL-6 and hsCRP levels did not significantly change from baseline and did not differ between the two patient groups. Depression scores was lower compared to baseline but did not differ between groups. Combining the two groups, a decrease in IL-6 was associated to decreased verbal fluency (p=0.02), and a decrease in hsCRP was associated with improvement in Trail making A (p=0.005). In conclusion, the level of IL-6 and hsCRP was increased in depressed outpatients but was not associated to specific depressive symptoms. In terms of cognitive function, we found that higher hsCRP levels were associated to lower psychomotor speed both at baseline and at follow-up., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The beneficial effect of exercise on hippocampal plasticity is possibly mediated by increased angiogenesis and neurogenesis. In angiogenesis, insulin-like growth factor-1 (IGF-1), vascular endothelial growth factor (VEGF), and hypoxia-inducible factor 1, alpha subunit (HIF1α) are important factors, while the induction of neurogenesis requires signaling through the VEGF receptor, Flk-1 (VEGFR-2). VEGF expression is believed to be regulated by two distinct mTOR (mammalian target of Rapamycin)-containing multiprotein complexes mTORC1 and mTORC2, respectively. This study was initiated to investigate the effect of exercise on the expression of VEGF, cognate receptors, HIF1α, mTORC1, and mTORC2 in hippocampus and frontal cortex. To this end, we measured messenger RNA (mRNA) levels in rat brain using quantitative real-time polymerase chain reaction (real-time qPCR) after forced treadmill exercise for 1 day, 2 weeks, and 8 weeks. Rats were euthanized either immediately (0 h) or 24 h after last exercise session. Here, we show that exercise affected mRNA levels of VEGF, VEGFR2, and the coreceptor neuropilin 2 (NRP2) when the rats were euthanized immediately, whereas at 24 h only the expression of mTOR was regulated after a single bout of exercise. In conclusion, the effect of treadmill exercise on the VEGF system is acute rather than chronic and there is a transient activation of mTOR. More studies are needed to understand whether this could be beneficial in the treatment of neuropsychiatric disorders., (Copyright © 2013 Wiley Periodicals, Inc.)

In the search for new drug targets, that may help point the way to develop fast-acting treatments for mood disorders, we have explored molecular pathways regulated by ketamine, an NMDA receptor antagonist, which has consistently shown antidepressant response within a few hours of administration. Using Sprague-Dawley rats we investigated the effects of ketamine on the presynaptic release machinery responsible for neurotransmitter release at 1, 2 and 4 h as well as 7 days after administration of a single subanesthetic dose of ketamine (15 mg/kg). A large reduction in the accumulation of SNARE complexes was observed in hippocampal synaptic membranes after 1, 2 and 4 h of ketamine administration. In parallel, we found a selective reduction in the expression of the synaptic vesicle protein synaptotagmin I and an increase in the levels of synapsin I in hippocampal synaptosomes suggesting a mechanism by which ketamine reduces SNARE complex formation, in part, by regulating the number of synaptic vesicles in the nerve terminals. Moreover, ketamine reduced Thr(286)-phosphorylated αCaMKII and its interaction with syntaxin 1A, which identifies CaMKII as a potential target for second messenger-mediated actions of ketamine. In addition, despite previous reports of ketamine-induced inhibition of GSK-3, we were unable to detect regulation of its activity after ketamine administration. Our findings demonstrate that ketamine rapidly induces changes in the hippocampal presynaptic machinery similar to those that are obtained only with chronic treatments with traditional antidepressants. This suggests that reduction of neurotransmitter release in the hippocampus has possible relevance for the rapid antidepressant effect of ketamine., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Due to its production of potent antimicrobial oxidants including hypochlorous acid, human myeloperoxidase (MPO) plays a critical role in innate immunity and inflammatory diseases. Thus MPO is an attractive target in drug design. (Aminoalkyl)fluoroindole derivatives were detected to be very potent MPO inhibitors; however, they also promote inhibition of the serotonin reuptake transporter (SERT) at the same concentration range. Via structure-based drug design, a new series of MPO inhibitors derived from 3-alkylindole were synthesized and their effects were assessed on MPO-mediated taurine chlorination and low-density lipoprotein oxidation as well as on inhibition of SERT. The fluoroindole compound with three carbons in the side chain and one amide group exhibited a selectivity index of 35 (Ki/IC50) with high inhibition of MPO activity (IC50 = 18 nM), whereas its effect on SERT was in the micromolar range. Structure-function relationships, mechanism of action, and safety of the molecule are discussed.

Background: Cytoglobin (Cygb) was discovered a decade ago as the fourth vertebrate heme-globin. The function of Cygb is still unknown, but accumulating evidence from in vitro studies point to a putative role in scavenging of reactive oxygen species and nitric oxide metabolism and in vivo studies have shown Cygb to be up regulated by hypoxic stress. This study addresses three main questions related to Cygb expression in the hippocampus: 1) Is the rat hippocampus a valid neuroanatomical model for the human hippocampus; 2) What is the degree of co-expression of Cygb and neuronal nitric oxide synthase (nNOS) in the rat hippocampus; 3) The effect of chronic restraint stress (CRS) on Cygb and nNOS expression., Methods: Immunohistochemistry was used to compare Cygb expression in the human and rat hippocampi as well as Cygb and nNOS co-expression in the rat hippocampus. Transcription and translation of Cygb and nNOS were investigated using quantitative real-time polymerase chain reaction (real-time qPCR) and Western blotting on hippocampi from Flinders (FSL/FRL) rats exposed to CRS., Principal Findings: Cygb expression pattern in the human and rat hippocampus was found to be similar. A high degree of Cygb and nNOS co-expression was observed in the rat hippocampus. The protein levels of nNOS and Cygb were significantly up-regulated in FSL animals in the dorsal hippocampus. In the ventral hippocampus Cygb protein levels were significantly up-regulated in the FSL compared to the FRL, following CRS., Significance: The rodent hippocampus can be used to probe questions related to Cygb protein localization in human hippocampus. The high degree of Cygb and nNOS co-expression gives support for Cygb involvement in nitric oxide metabolism. CRS induced Cygb and nNOS expression indicating that Cygb expression is stress responsive. Cygb and nNOS may be important in physiological response to stress.

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., (Copyright © 2012 Elsevier B.V. and ECNP. All rights reserved.)

Brain-derived neurotrophic factor (BDNF) has been suggested as a candidate gene for depression and numerous studies have investigated the possible association between genetic variants within BDNF and depression. Clinical studies have investigated the serum BDNF levels in individuals with depression. However, few studies have combined genetic association studies with serum BDNF measurements. The purpose of the present study was therefore to perform an investigation of BDNF using 162 individuals with depression and 289 healthy individuals. All individuals returned a completed questionnaire and participated in a semi-structured diagnostic interview. The major contribution of the present study is the integration of clinical assessment of cases and control individuals, simultaneous analyses of several genetic variants, serum BDNF measurements, and information on socio-demographic variables, lifestyle, and health indicators in a statistical model. In the present study the serum BDNF levels were increased in the depressive subjects compared to control individuals. Additionally, six SNPs were successfully analyzed, but did not associate with depression. Multiple linear regression models were applied and age, depression, gender, the Val66Met polymorphism, and the interaction between Val66Met and gender were identified as significant determinants of the serum BDNF level. In conclusion, our data demonstrate that other factors than a diagnosis of depression influence the serum BDNF level and the importance of these factors should be emphasized comparing different studies., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Depressive disorders represent a significant health concern as they are associated with high social and physical dysfunction and increased risk for suicide. Electroconvulsive therapy (ECT) is the most effective treatment for patients with drug-resistant severe depressive disorders. However, the underlying biological mechanisms of ECT are not well characterized. In particular, the regulation of transcription factors upon ECT has only just started to be unveiled. The schizophrenia and bipolar disorder associated bromodomain containing 1 (BRD1) gene is important for the acetylation of histone H3K14 and holds a key role in normal embryonic development and survival. In this study, we have measured Brd1 mRNA in the hippocampus and the frontal cortex of male Sprague-Dawley rats upon acute and repeated electroconvulsive seizures (ECS) over a period of 10 days. We found an increase in the general expression of Brd1 mRNA in the hippocampus after repeated ECS compared to sham (F = 8.108, P = 0.003). Furthermore, we provide evidence suggesting a decrease in the expression of the Brd1 mRNA variant comprising an extended version of exon 7 (Brd1-L) in the frontal cortex after repeated ECS compared to sham (F = 6.225, P = 0.023). These findings indicate that regulation of the Brd1 gene is part of the biological response to ECS and that splice variants are induced differentially in different brain regions., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

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., (Copyright © 2012 Elsevier B.V. All rights reserved.)

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. This 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 three 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., (Copyright © 2011 Wiley Periodicals, Inc.)

Neuropeptide S (NPS) and its receptor (NPSR) have been implicated in the mediation of anxiolytic-like behaviour in rodents. However, little knowledge is available regarding the NPS system in depression-related behaviours, and whether NPS also exerts anxiolytic effects in an animal model of psychopathology. Therefore, the aim of this work was to characterize the effects of NPS on depression- and anxiety-related parameters, using male and female rats in a well-validated animal model of depression: the Flinders Sensitive Line (FSL), their controls, the Flinders Resistant Line (FRL), and Sprague-Dawley (SD) rats. We found that FSL showed greater immobility in the forced swim test (FST) than FRL, confirming their phenotype. However, NPS did not affect depression-related behaviour in any rat line. No significant differences in baseline anxiety levels between the FSL and FRL strains were observed, but FSL and FRL rats displayed less anxiety-like behaviour compared to SD rats. NPS decreased anxiety-like behaviour on the elevated plus-maze in all strains. The expression of the NPSR in the amygdala, periventricular hypothalamic nucleus, and hippocampus was equal in all male strains, although a trend towards reduced expression within the amygdala was observed in FSL rats compared to SD rats. In conclusion, NPS had a marked anxiolytic effect in FSL, FRL and SD rats, but did not modify the depression-related behaviour in any strain, in spite of the significant differences in innate level between the strains. These findings suggest that NPS specifically modifies anxiety behaviour but cannot overcome/reverse a genetically mediated depression phenotype.

Although it is well established that cyclic adenosine monophosphate (cAMP) signalling via cAMP-dependent protein kinase (PKA)within neurons plays an important role in depression and antidepressant treatment, the importance of several newly discovered targets that function independently from PKA, such as exchange protein activated by cAMP (Epac), remains unexplored in this regard. In this study we used a cAMP analogue that inhibits PKA but not Epac (Rp-8-Br-cAMP), to explore the modifying actions of these two targets on immobility in the forced swim test (FST) and cerebellar cAMP response element binding protein (CREB) phosphorylation in rats. In addition, we assessed central cAMP and cGMP levels and investigated the involvement of cGMP-dependent protein kinase (PKG) on any observed effects by using a selective PKG inhibitor (Rp-8-Br-PET-cGMPS).Interestingly, Rp-8-Br-cAMPS strongly reduced immobility in the FST and induced an increase in the phosphorylation of CREB in the cerebellum, effects that were unaltered by the co-administration of Rp-8-Br-PET-cGMPS. Furthermore, Rp-8-Br-cAMPS increased the accumulation of cAMP and cGMP in the hippocampus, frontal cortex and cerebellum of these rats. Together, these results suggest that in addition to activating PKA, elevated cAMP may also stimulate other targets that mediate antidepressant activity. According to the pharmacodynamic profile of Rp-8-Br-cAMPS and taking into consideration what has recently been discovered regarding the cAMP signalling system, a likely candidate is the guanine nucleotide exchange factor, Epac., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Prolonged stress has been associated with altered synaptic plasticity but little is known about the molecular components and mechanisms involved in the stress response. In this study, we examined the effect of chronic restraint stress (CRS) on the expression of genes associated with synaptic vesicle exocytosis in rat prefrontal cortex and hippocampus. Rats were stressed daily using a 21day restraint stress paradigm, with durations of half an hour or 6h. RNA and protein were extracted from the same tissue sample and used for real-time quantitative polymerase chain reaction (real-time qPCR) and immunoblotting, respectively. Focusing on the SNARE complex, we investigated the expression of the SNARE core components syntaxin 1A, SNAP-25, and VAMP2 at both transcriptional and protein levels. In addition, the expression of 10 SNARE regulatory proteins was investigated at the transcriptional level. Overall, the prefrontal cortex was more sensitive to CRS compared to the hippocampus. In prefrontal cortex, CRS induced increased mRNA levels of VAMP2, VAMP1, syntaxin 1A, snapin, synaptotagmins I and III, and synapsins I and II, whereas SNAP-25 was down-regulated after CRS. Immunoblotting demonstrated equivalent changes in protein levels of VAMP2, syntaxin 1A, and SNAP-25. In hippocampus, we found increased mRNA levels of VAMP2 and SNAP-29 and a decrease in VAMP1 levels. Immunoblotting revealed decreased VAMP2 protein levels despite increased mRNA levels. Changes in the expression of synaptic proteins may accompany or contribute to the morphological, functional, and behavioral changes observed in experimental models of stress and may have relevance to the pathophysiology of stress-related disorders., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Depression and pathological anxiety disorders are among the most prevalent neurological diseases in the world and can be precipitated and exacerbated by stress. Prenatal stress alters both behavioral and endocrine responses to stressful stimuli in later life. We have previously observed increased basal acoustic startle response (ASR) in Wistar rats exposed to stress or dexamethasone (DEX) in utero when tested during the light phase of the circadian rhythm, and decreased prepulse inhibition (PPI) in similar animals tested during the dark phase of the cycle. We speculated that this observation of increased basal startle might be influenced by diurnal phase. In the present study, adult female Sprague Dawley rats, stressed prenatally with DEX (200 μg/kg, gestational days 14-21) and postnatally by blood sampling under restraint, were tested for the ASR during both circadian phases (light and dark). Basal startle was increased in animals tested both during the light and the dark phases of the cycle. We hereby replicated our earlier findings in a new strain and laboratory, thus strengthening the validity of our model regarding prenatal stress effects on ASR in female offspring. Our results indicate that observation of increased basal ASR is not solely dependent on diurnal phase. We found no difference in hippocampal glucocorticoid and mineral corticoid receptor expression between groups., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Objective: To identify and describe the qualitative variations in how physical activity is perceived and understood by individuals with current or previous venous leg ulcer., Design: A qualitative study using semi-structured interviews., Method: Twenty-two individuals aged 60-85 years were interviewed. The interviews were recorded, transcribed verbatim and analysed by three researchers using a phenomenographic research approach. A set of categories of descriptions and their internal relationships were constructed based on the essential features of the variation in patients' perceptions of physical activity., Results: Four categories of descriptions were identified: (i) 'self-management', (ii) 'instructions and support', (iii) 'fear of injury' and (iv) 'a wish to stay normal'. The categories could be interpreted by a two-dimensional construct: (1) perception of venous leg ulcer as a chronic or acute condition and (2) engagement or avoidance behaviour toward physical activity. Chronicity and behaviour combined together formed a 2 × 2 square housing the four qualitatively different categories. Irrespective of category, the participants reported that information given by caregivers regarding leg ulcer and physical activity was insufficient or contradictory. Written information or exercise programmes were not obtained regularly and not at all in primary care., Conclusion: A dichotomous view emerged from participants' experiences of physical activity based on (1) perception of venous leg ulcer as a chronic or acute condition and (2) engagement or avoidance behaviour toward physical activity.

Background: Electromyographic (EMG) spectral parameters and endurance tests for back muscles have long been used to estimate low-back muscle fatigue., Purpose: The present purpose was to evaluate the test-retest reliability of a fatigue-testing protocol and to investigate associations between activity limitations and fatigue in patients with lumbar disc herniation., Methods: To evaluate the reliability, endurance time, EMG spectral parameters and ratings of lumbar muscle fatigue and back pain were measured three times in 15 patients with lumbar disc herniation. The patients performed a modified Sørensen test with concurrent recordings of electromyography from the lumbar muscles at four recording sites. Ratings of their perceived fatigue and pain on a Borg scale were collected simultaneously. Recovery over five minutes was recorded. Questionnaires about activity, participation, self-efficacy and health were also used., Results: Endurance time had an ICC of 0.85 indicating good reliability. Initial, end and median frequency recovery had intraclass correlation coefficients (ICCs) over 0.6 that is acceptable but with large standard errors of measurement. The median frequency slopes for the four recording sites had ICCs of 0.19-0.48. Agreement coefficients for the Borg ratings ranged between 0.78 and 0.97. Moderate correlation was found between activity limitations and endurance time., Conclusions: Endurance time measurement and Borg ratings may be recommended for evaluating intervention both with groups of patients, i.e. for research purposes, and for individuals. The median frequency parameters may be recommended for groups only. The study shows the importance of establishing the reliability of the patient category if the results are to be used to evaluate intervention., (Copyright © 2010 John Wiley & Sons, Ltd.)

There is accumulating evidence that brain-derived neurotrophic factor (BDNF) plays a critical role in the pathophysiology of depression. Decreased serum levels have been reported in major depression, and a correlation between BDNF reduction and the severity of the disease was found. Moreover, in post-mortem hippocampal tissue, increased levels of BDNF immunoreactivity have been reported in subjects treated with antidepressants compared to untreated subjects. These findings indicate parallel changes in brain and serum BDNF levels during depression. BDNF has been measured in selected brain areas in several animal models. In investigations between Flinders Sensitive Line (FSL) and Flinders Resistant Line (FRL) rats, a genetic rat model of depression, no differences were found in BDNF levels in the frontal cortex and hippocampus, areas believed to be core brain regions in depression. However, to our knowledge brain and serum BDNF levels have never been reported in parallel for any psychiatric disease model. Therefore, we examined the levels of BDNF in whole blood, serum, cerebrospinal fluid (CSF), hippocampus, and frontal cortex in male FSL and FRL rats. BDNF levels in serum and whole blood of FSL rats were significantly increased compared to FRL rats. In contrast, in the hippocampus the BDNF level was significantly decreased in FSL compared to FRL rats while no differences were found in the frontal cortex and CSF. The differential regulation of the BDNF levels in hippocampus, serum, and whole blood in FSL/FRL rats adds to the hypothesis that neurotrophic factors are related to the pathophysiology of depression.

Low birth weight (LBW) is associated with type 2 diabetes and depression, which may be related to prenatal stress and insulin resistance as a result of chronic hypothalamic-pituitary-adrenal (HPA) axis hyperactivity. We examined whether treatment with a selective serotonin reuptake inhibitor [escitalopram (ESC)] could downregulate HPA axis activity and restore insulin sensitivity in LBW rats. After 4-5 wk of treatment, ESC-exposed LBW (SSRI-LBW) and saline-treated control and LBW rats (Cx and LBW) underwent an oral glucose tolerance test or a hyperinsulinemic euglycemic clamp to assess whole body insulin sensitivity. Hepatic phosphoenolpyruvate carboxykinase (PEPCK) mRNA expression and red skeletal muscle PKB Ser(473) phosphorylation were used to assess tissue-specific insulin sensitivity. mRNA expression of the hypothalamic mineralocorticoid receptor was fivefold upregulated in LBW (P < 0.05 vs. Cx), accompanied by increased corticosterone release during restraint stress and total 24-h urinary excretion (P < 0.05 vs. Cx), whole body insulin resistance (P < 0.001 vs. Cx), and impaired insulin suppression of hepatic PEPCK mRNA expression (P < 0.05 vs. Cx). Additionally, there was a tendency for reduced red muscle PKB Ser(473) phosphorylation. The ESC treatment normalized corticosterone secretion (P < 0.05 vs. LBW), whole body insulin sensitivity (P < 0.01) as well as postprandial suppression of hepatic mRNA PEPCK expression (P < 0.05), and red muscle PKB Ser(473) phosphorylation (P < 0.01 vs. LBW). We conclude that these data suggest that the insulin resistance and chronic HPA axis hyperactivity in LBW rats can be reversed by treatment with an ESC, which downregulates HPA axis activity, lowers glucocorticoid exposure, and restores insulin sensitivity in LBW rats.

Stress engenders the precipitation and progression of affective disorders, while stress-related release of excitatory mediators is implicated in the degenerative pathology observed especially in the hippocampus of patients with severe depression. Nitric oxide (NO) release following stress-evoked N-methyl-d-aspartate (NMDA) receptor activation modulates neurotransmission, cellular memory and neuronal toxicity. We have investigated the Flinders rat (FSL/FRL), a genetic animal model of depression, regarding the response of the hippocampal nitrergic system following exposure to an escapable stress/inescapable stress (ES-IS) paradigm. Hippocampal tissue from naive FSL/FRL rats and those exposed to ES-IS were studied with respect to constitutive nitric oxide synthase (cNOS) activity and neuronal nitric oxide synthase (nNOS) protein levels, as well as transcript expression of upstream regulatory proteins in the NMDA-NO signalling pathway, including NMDAR1, nNOS, CAPON, PIN and PSD95. Within stress-naive animals, no differences in hippocampal cNOS activity and nNOS expression or PIN were evident in FSL and FRL rats, although transcripts for NMDAR1 and CAPON were increased in FSL rats. Within the group of ES-IS animals, we found an increase in total hippocampal cNOS activity, nNOS protein levels and mRNA expression in FSL vs. FRL rats, together with an increase in PSD95 transcripts, and a reduction in PIN. In conclusion, ES-IS enhanced hippocampal cNOS activity in FSL rats, but not FRL rats, confirming the NMDA-NO cascade as an important vulnerability factor in the depressive phenotype of the FSL rat.

Compared to the classical monoamine hypotheses focus on neuroplasticity is a major new approach in studies of depression and antidepressants. Recent studies have demonstrated that vascular endothelial growth factor (VEGF) is regulated by antidepressant treatment in rodents. However, in depressive patients no significant changes were found in the serum VEGF levels compared to control subjects. To our knowledge, brain and serum VEGF levels have never been reported in parallel for any psychiatric disease model. That prompted us to examine the levels of VEGF in serum, hippocampus, frontal cortex, corpus striatum, and hypothalamus in male Flinders Sensitive Line (FSL) and Flinders Resistant Line (FRL), a genetic rat model of depression. The VEGF levels were identical in the FSL and the FRL rats in serum, corpus striatum, and hypothalamus. In hippocampus and frontal cortex, the VEGF levels were significantly decreased in the FSL rats compared to the FRL rats. The results may add to the hypothesis that altered expression of growth factors/neurotrophic factors are related to the pathophysiology of depression., (Copyright 2010 Elsevier Ireland Ltd. All rights reserved.)

Quantification of endogenous brain-derived neurotrophic factor (BDNF) can be performed with an enzyme-linked immunosorbent assay (ELISA). Although BDNF has been determined in blood and brain preparations in numerous studies with ELISA kits, the methodological issues regarding measurements of BDNF concentrations in the blood and particularly in the brain have only been superficially investigated. We aimed at validating and optimizing the BDNF ELISA kit with respect to measurements in rat blood and brain samples. We found that the pre-analytical conditions were critical for plasma samples, but not serum or whole blood samples. The intra- and inter-assay variation and the accuracy and yield of the BDNF ELISA kit in rat serum and brain tissue were conducted with the optimal dilutions of frontal cortex and hippocampus extract. The optimal dilutions of frontal cortex and hippocampus extracts were determined to be 20 and 120 times, and we established that the intra-assay coefficient of variation (CV%) was 8 in hippocampus and 2 in frontal cortex and serum. The inter-assay variation was also low with a CV% of 11 or less in hippocampus, frontal cortex, and serum. Finally, we found that the accuracy and yield of the BDNF measurements were high in serum and low in hippocampus and frontal cortex. We conclude that the BDNF ELISA kit determines serum BDNF accurately and with high reproducibility. Furthermore it can be used for measurement of BDNF in rat brain preparations when particular precautions are taken and in particular with care regarding the dilution of the brain tissue samples., (Copyright (c) 2009 Elsevier B.V. All rights reserved.)

Background and Purpose: Previous studies have shown that patients with chronic venous insufficiency are deconditioned and physically inactive. The present study aimed to examine the occurrence of fear-avoidance beliefs in patients with chronic venous insufficiency, and to investigate the role of fear-avoidance beliefs and pain severity in predicting the low level of physical activity in these patients., Method: Data were collected by a postal questionnaire sent to 146 patients with chronic venous insufficiency and current or previous venous leg ulcer. Complete data were collected from 98 patients aged 60-85 years - 63% women - giving a response rate of 67%. Fear-avoidance beliefs were assessed by the Fear-Avoidance Beliefs Questionnaire, physical activity subscale. Pain and physical activity were assessed by the Six-point Verbal Rating Scale of Pain Assessment and the Physical Activity Questionnaire, respectively., Results: Fear-avoidance beliefs were present in 81 (83%) of the patients with chronic venous insufficiency (range 0-24, median 12). Forty patients (41%) had strong fear-avoidance beliefs. One-third of the patients with healed ulcers had strong fear-avoidance beliefs. Patients with low physical activity had significantly stronger fear-avoidance beliefs and more severe pain than patients with high physical activity. Multiple logistic regression showed that the odds ratio (OR) for low physical activity were about three times higher for patients with strong fear-avoidance beliefs (OR 3.1, 95% confidence interval 1.1-8.3; p = 0.027) than for patients with weak fear-avoidance beliefs., Conclusions: Fear-avoidance beliefs were present in most patients with chronic venous insufficiency and were associated with low physical activity. Clinical implications ought to include a better recognition of fear-avoidance beliefs, early information about the negative consequences of such beliefs, and the importance of physical activity to counteract poor mobility., (Copyright (c) 2009 John Wiley & Sons, Ltd.)

Electroconvulsive therapy (ECT) remains the treatment of choice for patients with severe or drug-resistant depressive disorders, yet the mechanism behind its efficacy and the effect on neurotransmission is essentially unknown. As synaptic vesicle proteins (SVPs) are required for vesicle fusion and neurotransmitter release, we have examined the effect of single and repeated electroconvulsive seizures (ECS), an animal model of ECT, on the expression of 14 SVPs in the rat frontal cortex and the hippocampus using quantitative real-time polymerase chain reaction (real-time qPCR). Only in the frontal cortex, the mRNA level of synapsin II was significantly upregulated after repeated ECS. In contrast, the mRNA levels of 6 of the 14 SVPs were significantly regulated in the hippocampus after ECS. We found that SNAP29 was upregulated and synaptotagmin III was downregulated after one single ECS in the hippocampus. Furthermore, SNAP29, synapsin I, synapsin III, VAMP2, and VAMP5 were significantly upregulated, whereas synaptotagmin III was significantly downregulated after repeated ECS in the hippocampus. We suggest that these genes are highly important in the long-term therapeutic effect of ECS, and thus it can be hypothesized that the SVPs are involved in the pathophysiology of depression., (Published 2008 Wiley-Liss, Inc.)

Urinary incontinence (UI) is a common female dysfunction, affecting women in all ages. Experienced physiotherapists and experts on low back pain (LBP) have since long observed and discussed the empirical association between LBP and UI. The aim of this study was to describe the occurrence of UI in women with LBP and to compare this group with a reference group of comparable age, language, culture and parity. The authors of this study had previously collected all original data from the reference group. A validated questionnaire concerning UI was answered by 200 consecutive women with LBP visiting specialised physiotherapy clinics in the area of Stockholm. Inclusion criteria were LBP, female, not pregnant, Swedish speaking, age between 17 and 45 years. Seventy-eight percent of the women with LBP reported UI. In comparison with the reference group, the prevalence of UI and signs of dysfunctional pelvic floor muscles (PFM) were greatly increased (p < 0.001) in the LBP group. Logistic regression analysis showed that the condition LBP and PFM dysfunction i.e. inability to interrupt the urine flow (p < 0.001) were risk factors for UI irrespective of parity. Physiotherapists treating patients with LBP should be aware of possible incontinence problems in this patient group.

Quantitative real-time polymerase chain reaction (qPCR) has become a widely used tool in the search for disease genes. When examining gene expression with qPCR in psychiatric diseases, endogenous reference gene(s) must be used for normalization. Traditionally, genes such as beta-actin (ActB), Gapd, and 18s rRNA, assumed to be stably expressed, have been used for normalization. However, it has become clear that expression of these genes is influenced by different experimental paradigms. Since stable gene expression of houskeeping genes (HKGs), therefore, cannot be expected, alternative strategies are warranted. With the overall aim to inspect the gene expression of three target genes, NMDAR1, SORT, and CREB, in rat hippocampus, we tested a panel of eight HKGs, 18s rRNA, ActB, CycA, Gapd, Hmbs, Hprt1, Rpl13A, and Ywhaz in order to select the most stably expressed gene, using the NormFinder and geNorm software applets. Additionally, we have applied four different normalization approaches for normalization of the three target genes. We found using the NormFinder software that Ywhaz is the most stably expressed gene among the eight tested HKGs. However, the results of the analysis of the target genes are highly dependent on the choice of normalization approach. Moreover, the number of HKGs, used for selection of the most stable HKG, also influences on the result of the gene expression analysis of target genes. These results underline the importance of choosing a proper normalization strategy when analyzing gene expression with qPCR. The strategy should be unbiased and carried out in every specific experimental setting.

Numerous attempts have been made to develop suitable radiolabeled tracers for positron emission tomography or single photon emission computed tomography imaging of the serotonin transporter (SERT), but most often, negative outcomes are reported. The aim of this study is to define characteristics of a good SERT radioligand and to investigate species differences. We examined seven different selective serotonin reuptake inhibitors (SSRIs) and that except for one all have been previously tested as emission tomography ligands. The outcome of the ligands as emission tomography tracers was compared in relation with receptor density (Bmax) and/or ligand affinity (Kd) in rat and monkey cerebrum and cerebellum (reference region) membranes. [3H]-(S)-Citalopram and [3H]-(+)-McN5652 display statistically significantly lower affinity, whereas [3H]paroxetine displays statistically significantly higher affinity for SERT in monkey cortex when compared with the rat cerebrum. The affinity of [3H]MADAM, [123I]ADAM, and [11C]DASB for SERT obtained with rat cerebrum and monkey cortex are similar. In monkey cortex, Kd and Bmax could not be determined with [3H]fluoxetine. Of the seven SSRIs, [3H]-(S)-citalopram, [3H]MADAM, and [11C]DASB displayed significant specific binding to SERT in monkey cerebellum, with Bmax cortex:cerebellum ratios being 17, 3, and 4, respectively. In rat brain tissue the ratios were 12, 6, and 3, respectively. In conclusion, it can be estimated that imaging of the human SERT in a high-density region requires radioligands with Kd values between 0.03 and a maximum of 0.3 nM (at 37 degrees C). The differential specific cerebellar binding raises the question of the suitability of cerebellum as a reference region for nonspecific binding., ((c) 2007 Wiley-Liss, Inc.)

Background and Purpose: Measurements of walking speeds are commonly used as an objective measure of functional performance in patients with knee osteoarthritis (OA) and are easily performed in a clinical setting. However, the choice in which the walking speed evaluation should be performed is controversial. The aim of this study was to identify the most discriminating walking speed after surgical interventions in patients with knee osteoarthritis, and to compare the responsiveness of the different gait speeds., Method: A prospective clinical one-year follow-up study involving 54 patients with knee osteoarthritis (63 (+/-5) years of age) who were operated with either a unicompartmental knee arthroplasty or a high tibial osteotomy was undertaken. Thirty-nine patients had unilateral knee OA and 15 patients had bilateral knee OA or other symptoms from the lower extremities that could influence walking. The patients were examined at a gait laboratory before surgery, and one year after surgery. The patients were instructed to walk at slow, normal and fast walking speed., Results: All patients (n=54) walked.faster one year after the surgical intervention compared to before surgery (p = 0.001) at slow (+15%), normal (+8%) and fast (+7%) walking speed. This increase was similar for the three walking speeds (p = 0.171). Patients with unilateral knee OA (n=39) reached an average change of +0.12 m/s, which was considered clinically important, while patients with bilateral knee OA (n=15) did not increase their walking speed > 0.12 m/s. Effect size was moderate for slow walking speed and small for normal and fast walking speeds, respectively., Conclusions: The different walking speeds were equally good in detecting changes one year after surgical interventions. In this study, responsiveness favoured slow walking speed, however, the advantages of normal walking speed are discussed.

Background and Purpose: Fear-avoidance beliefs are important determinants for disability in patients with non-specific low-back pain (LBP). The association with self-reported level of physical activity is less known. The aim of the present study was to describe the level of physical activity in patients with chronic non-specific LBP and its relation to fear-avoidance beliefs and pain catastrophizing., Method: A cross-sectional study on 64 patients with chronic non-specific LBP in primary healthcare. The variables measured and the questionnaires used were: level of physical activity (six-graded scale); activity limitations (Roland Morris Disability Questionnare (RDQ)); fear-avoidance beliefs (Tampa Scale of Kinesiophobia (TSK) 13-item and sub-scales 'activity avoidance' and 'somatic focus'); and pain catastrophizing (Pain Catastrophizing Scale (PCS)). The level of physical activity was dichotomised into low and high physical activity. Individual median scores on the TSKandPCSscales were used to group the patients into different levels offear-avoidance beliefs and pain catastrophizing. Univariate logistic regressions were used to calculate odds ratios for having low physical activity., Results: Patients with low physical activity had significantly higher scores in fear-avoidance beliefs and pain catastrophizing (p < 0.05). Odds ratios for low level of physical activity were between 4 and 8 (p < 0.05) for patients with high fear-avoidance beliefs or medium/high pain catastrophizing., Conclusions: This study indicates that it seems important for physiotherapists in primary care to measure levels of fear-avoidance beliefs or pain catastrophizing. In particular, the two subscales of the TSK could be of real value for clinicians when making treatment decisions concerning physical exercise therapy for patients with chronic LBP.

To study the 5-HT(2A) receptors in the living human brain, using positron emission tomography (PET), two selective radiotracers are currently in use: [(18)F]altanserin and [(11)C]MDL 100907. It is, however, currently unknown to what extent data obtained with either tracer are directly comparable. The aim of this study was to compare binding characteristics of these two radiotracers in rat brain with respect to affinity (K(d)), receptor binding density (B(max)), binding potential (BP), and nonspecific binding. Further, binding kinetics, sensitivity towards competition with the endogenous transmitter serotonin, and the competitive/noncompetitive interaction between the two radioligands were evaluated. In addition, the selectivity of [(18)F]altanserin for the 5-HT(2A) receptor was assessed. The K(d) value of [(18)F]altanserin and [(3)H]MDL 100907 was in the order of 0.3 nM. B(max) in frontal cortex was 523 and 527 fmol/mg protein, respectively. The binding of [(18)F]altanserin was not influenced by blocking either the 5-HT(2B/2C) or the alpha(1)-adrenergic receptors. At 37 degrees C the association t(1/2) was 2.8 and 2.7 min and the dissociation t(1/2) was 11 and 13.5 min for [(18)F]altanserin and [(3)H]MDL 100907, respectively. Both radioligands were displaced by 5-HT, only at high concentrations; the K(i) value of 5-HT ranging between 650 and 3,300 nM. This indicates that binding of both radioligands in PET studies is not directly influenced by changes in endogenous 5-HT.Overall, the binding of [(18)F]altanserin and [(3)H]MDL 100907 to the 5-HT(2A) receptor was very comparable, showing selective high affinity binding in the subnanomolar range., (Copyright 2005 Wiley-Liss, Inc.)

The serotonin transporter (SERT) belongs to a family of sodium chloride-dependent transporters responsible for uptake of amino acids and biogenic amines from extracellular spaces. SERT represents the main pharmacological target in the treatment of several clinical conditions, including depression and anxiety. Serotonin-selective reuptake inhibitors and tricyclic antidepressants are the most predominantly prescribed drugs in the treatment of depression. In addition to antidepressants also psychostimulants, like cocaine and amphetamines, are important SERT antagonists. In the present study, we report the cloning and characterization of chicken SERT. Although the uptake kinetic was very similar to human SERT, the pharmacological profiles differed considerably for the two species. We find that chicken SERT is capable of discriminating between different serotonin-selective reuptake inhibitors; thus, the potency of S-citalopram and paroxetine is reduced more than 40-fold. A cross-species chimera strategy was undertaken and followed by species-scanning mutagenesis. Differences in pharmacological profiles were tracked to amino acid residues 169, 172, and 586 in human SERT. Structure-activity studies on structurally related compounds indicated that species divergences in drug sensitivity between human and chicken SERT were arising from differences in coordination or recognition of an important aminomethyl pharmacophoric substructure, which is shared by all high affinity antidepressants. Consequently, we suggest that Ala(169) and Ile(172) of human SERT are important residues in sensing the N-methylation state of SERT antagonists.

This prospective study aimed to examine whether patients with lumbar disc herniation, before and after surgery, showed impaired recovery from fatigue measured with EMG and subjective ratings as compared to healthy subjects. Forty-three patients due for, and after, lumbar disc surgery and 60 healthy subjects were fatigued using a modified Sørensen's test. Recovery of back extensor muscles was assessed using electromyography and the Borg scale ratings at 1, 2, 3, and 5 minutes of recovery, and was further analyzed using an exponential time-dependence model. The patients completed four questionnaires: the Oswestry, the Roland-Morris, a self-efficacy scale and the SF-36. The exponential time-dependence model was successful for 14 of the 43 patients and 57 of the 60 healthy subjects. The patients for whom the model succeeded had significantly better scores on the questionnaires than those for whom the model was unsuccessful. Specific items related to standing and lifting were identified. The patients' recovery was impaired compared to the healthy subjects, as determined by use of the exponential time-dependence model. The combination of fatigue and recovery measures was reflected in the questionnaire scores and is therefore important for evaluating patients with lumbar disc herniation.

Evaluations of new emission tomography ligands are usually carried out in animals. In order to keep the animals in a restricted position during the scan session, anaesthesia is almost inevitable. In ex vivo rat studies we investigated the interference of ketamine/xylazine, zoletile mixture, isoflurane and halothane with the serotonin re-uptake site, the serotonin(2A) receptor and the dopamine re-uptake site by use of [(3)H]-(S)-citalopram, [(18)F]altanserin and [(125)I]PE2I, respectively. Ketamine/xylazine decreased the target-to-background ratio (mean +/- SD) of [(3)H]-( S)-citalopram from 1.5+/-0.19 to 0.81+/-0.19 (P<0.05), whereas isoflurane and halothane increased the ratio from 1.5+/-0.19 to 1.9+/-0.24 and 2.1+/-0.13 (P<0.05), respectively. Only with the zoletile mixture did the ratio remain unaltered. None of the tested anaesthetics affected the target-to-background ratio of [(18)F]altanserin. The [(125)I]PE2I target-to-background ratio decreased with both ketamine/xylazine (from 12.4+/-0.81 to 10.1+/-1.4, P<0.05) and isoflurane (from 12.4+/-0.81 to 9.5+/-1.1, P<0.05) treated rats, whereas treatment with zoletile mixture and halothane left the ratio unaltered. It is concluded that prior to performance of neuroreceptor radioligand studies, the possible interaction between radioligands and anaesthetics should be carefully evaluated.

In this study we investigated whether the PET signal of labeled (S)-citalopram could be improved by predosing with the unlabeled distomer (R)-citalopram. Ten minutes before intravenous injection of 1.5 MBq [(3)H]-(S)-citalopram, rats were given i.v. 0.9% saline, 16 or 24 microg (R)-citalopram. Sixty minutes after injection of [(3)H]-(S)-citalopram, the brains were dissected into eight regions. The binding obtained in each brain region was compared to cerebellum as a reference region. Predosing with saline, 16 and 24 microg (R)-citalopram yielded thalamus/cerebellum ratios of 1.6 +/- 0.12, 1.6 +/- 0.12, and 1.1 +/- 0.26 (means +/- SD), respectively. It is concluded that the nonspecific binding of radiolabeled (S)-citalopram cannot be reduced by predosing with the 150 times less active enantiomer (R)-citalopram, possibly due to the pool of nonspecific sites being too large for blocking., (Copyright 2002 Wiley-Liss, Inc.)

The recovery of the median frequency of the power spectrum of the electromyogram (EMG) after fatigue has been studied to obtain reference data for healthy subjects (n = 55). In a seated position, the subjects performed a 45 s isometric back muscle contraction at 80% of maximal voluntary contraction, followed by 5 s contractions after 1, 2, 3, and 5 min in the recovery phase. In an additional reliability study (n = 11), this was performed six times. Surface EMG was recorded on four sites, namely, bilaterally from the lumbar muscles at the level of the first (L1) and fifth (L5) vertebrae. By non-linear regression, an exponential time dependence model was used to analyse the recovery of median frequency, giving recovery half-time as a resulting measure. Agreement with exponential time dependence was very good (coefficient of determination r(2) = 0.98) in the analysis of mean data (n = 55), with recovery half-times in the range 32-39 s on the four recording sites. Analysis of individual recordings, for which r(2) values in general were lower, revealed further details. Median values of the half-times in general agreed well with the half-times obtained from the analysis of mean data. Recovery half-time and median frequency slope during contraction were not correlated; this is in agreement with what may be expected from an exponential time dependence. Non-significant negative slopes occurred on apparently randomly selected sites, possibly indicating varying muscle coordination in the seated test method. The reliability of the half-time was not sufficient to allow for follow-up measurements on individuals, due to the fluctuations of the recovery data in recordings from individuals.

In order to study the influence of interelectrode distance and force level on the electromyographic (EMG) spectral parameters and on their reliability, bipolar surface EMG measurements were performed on the lumbar muscles of 15 subjects. Two test contractions (45 s) at 40% of maximal voluntary contraction (MVC) were performed, one with 2 cm interelectrode distance and the other with 4 cm, followed by two contractions at 80% MVC with the same change in interelectrode distance. Increasing the interelectrode distance from 2 to 4 cm caused a significant mean decrease (about 8%) in the initial median frequency. It is shown that this shift is of an order of magnitude that may be expected from the bipolar electrode filter factor, and we further conclude that the observed individual variations in the shift are likely to be connected to fluctuations in the shape of the power spectrum and to variations in conduction velocity. No significant change was found for the median frequency slope when changing the interelectrode distance. Increasing the force (from 40 to 80% MVC) also caused a significant mean decrease (about 10%) in the initial median frequency. The median frequency slope became significantly more negative by more than 200%. We conclude, however, that torque fluctuations during the fatigue contractions should have had only minor influence on the standard error of measurement of the initial median frequency and of the median frequency slope.

In the intensive care of sick infants, the global oxygen reserve capacity is estimated by co-oximetry (co-ox) of blood sampled from central venous catheters. Introduction of a noninvasive alternative is desirable. Near infrared spectroscopy (NIRS) offers a technique for noninvasive bedside monitoring of tissue oxygen economy. We studied the relation between peripheral venous oxyhemoglobin saturation (SvO(2)) estimated by venous occlusion and NIRS, and the central SvO(2) measured by co-ox of central venous blood. We report the high reproducibility of NIRS with a test-retest variation of only 2.51 +/- 1.41%. After bias adjusting of NIRS SvO(2) values, a nice correlation (r = 0.96, p > 0.05) between NIRS measurements of peripheral SvO(2) and co-ox of central venous blood was found. The study indicates that NIRS is practical for monitoring relative changes in central venous saturation. This might be useful in the future clinical care of newborns., (Copyright 2002 S. Karger AG, Basel)