Introduction
The abuse of anabolic androgenic steroids (AASs) is a source of public concern because of their adverse effects. Supratherapeutic doses of AASs are known to be hepatotoxic and regulate ...the lipoproteins in plasma by modifying the metabolism of lipids in the liver, which is associated with metabolic diseases. However, the effect of AASs on the profile of lipids in plasma is unknown.
Objectives
To describe the changes in the plasma lipidome exerted by AASs and to discuss these changes in the light of previous research about AASs and de novo lipogenesis in the liver.
Methods
We treated male
Wistar
rats with supratherapeutic doses of nandrolone decanoate and testosterone undecanoate. Subsequently, we isolated the blood plasma and performed lipidomics analysis by liquid chromatography-high resolution mass spectrometry.
Results
Lipid profiling revealed a decrease of sphingolipids and glycerolipids with palmitic, palmitoleic, stearic, and oleic acids. In addition, lipid profiling revealed an increase in free fatty acids and glycerophospholipids with odd-numbered chain fatty acids and/or arachidonic acid.
Conclusion
The lipid profile presented herein reports the imprint of AASs on the plasma lipidome, which mirrors the downregulation of de novo lipogenesis in the liver. In a broader perspective, this profile will help to understand the influence of androgens on the lipid metabolism in future studies of diseases with dysregulated lipogenesis (e.g. type 2 diabetes, fatty liver disease, and hepatocellular carcinoma).
Evidence to date suggests that opioids such as methadone may be associated with cognitive impairment. Growth hormone (GH) and insulin-like growth factor-1 (IGF-1) are suggested to be neuroprotective ...and procognitive in the brain and may therefore counteract these effects. This study aims to explore the protective and restorative effects of GH and IGF-1 in methadone-treated cell cultures. Primary cortical cell cultures were harvested from rat fetuses and grown for seven days in vitro. To examine the protective effects, methadone was co-treated with or without GH or IGF-1 for three consecutive days. To examine the restorative effects, methadone was added for the first 24 h, washed, and later treated with GH or IGF-1 for 48 h. At the end of each experiment, mitochondrial function and membrane integrity were evaluated. The results revealed that GH had protective effects in the membrane integrity assay and that both GH and IGF-1 effectively recovered mitochondrial function and membrane integrity in cells pretreated with methadone. The overall conclusion of the present study is that GH, but not IGF-1, protects primary cortical cells against methadone-induced toxicity, and that both GH and IGF-1 have a restorative effect on cells pretreated with methadone.
Angiotensin IV (Ang IV), a metabolite of Angiotensin II, is a bioactive hexapeptide that inhibits the insulin-regulated aminopeptidase (IRAP). This transmembrane zinc metallopeptidase with many ...biological functions has in recent years emerged as a new pharmacological target. IRAP is expressed in a variety of tissues and can be found in high density in the hippocampus and neocortex, brain regions associated with cognition. Ang IV is known to improve memory tasks in experimental animals. One of the most potent IRAP inhibitors known today is the macrocyclic compound HA08 that is significantly more stable than the endogenous Ang IV. HA08 combines structural elements from Ang IV and the physiological substrates oxytocin and vasopressin, and binds to the catalytic site of IRAP. In the present study we evaluate whether HA08 can restore cell viability in rat primary cells submitted to hydrogen peroxide damage. After damaging the cells with hydrogen peroxide and subsequently treating them with HA08, the conceivable restoring effects of the IRAP inhibitor were assessed. The cellular viability was determined by measuring mitochondrial activity and lactate dehydrogenase (LDH) release. The mitochondrial activity was significantly higher in primary hippocampal cells, whereas the amount of LDH was unaffected. We conclude that the cell viability can be restored in this cell type by blocking IRAP with the potent macrocyclic inhibitor HA08, although the mechanism by which HA08 exerts its effects remains unclear.
Anxiety disorders affect up to one third of the population. Caffeine, an adenosine receptor antagonist, is thought to have a dose-dependent effect on anxiety. We recently showed that a high dose of ...caffeine (50 mg/kg) differentially affected anxiety-like behavior in rats with high or low baseline anxiety-like behavior, replicating findings using relatively high doses in human patient samples. It is not known if low doses of caffeine have similar effects. The elevated plus maze (EPM) was used to categorize male Wistar rats (13 weeks of age) into groups of high or low anxiety-like behavior. Behavior was evaluated using the multivariate concentric square field (MCSF) test and the EPM after a low 10 mg/kg dose of caffeine. Multivariate data analysis demonstrated that caffeine decreased the differences between the high and low anxiety group, whereas the separation remained for the high and low control groups. For the caffeine treated rats, univariate statistics showed an increase in parameters regarding activity in the EPM and duration in the slope of the MCSF. Regarding risk-taking, shelter-seeking, and exploratory behavior, caffeine did not affect the groups differently. In conclusion, these results demonstrate increased activity in the caffeine-treated rats, together with a potentially anxiolytic effect and increased impulsivity that did not differ between the baseline anxiety groups. In contrast to high caffeine doses, a low dose does not generally affect rats with high anxiety at baseline differently than rats with low anxiety-like behavior. Further studies are warranted to fully elucidate the effects of caffeine in anxiety.
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•Rats were categorized as having a high or low anxiety-like behavior using EPM.•Low dose of caffeine seems to diminish the difference in baseline anxiety.•Low dose caffeine increased open arm time in rats with low baseline anxiety.•Caffeine alters the behavioral profile in male rats.
Although the number of studies that have examined the impact of opioids on cell viability is very limited, it has clearly shown that opioids commonly used in the clinic can both decrease neurogenesis ...and induce cell death. These negative effects induced by opioids are worrying and there is a need for further in-depth investigations addressing the impact of opioids on cell function and cell viability. A useful in vitro approach for studying the effects of opioids on cellular function and viability is using primary cortical cell cultures obtained from embryonic day 17 (E17) rat embryos. These cell cultures contain both neurons and glial cells that provide a more physiologically relevant culture condition when compared to the use of various commercially available cell lines. The primary cortical cells can be cultivated in 96-well plates, treated with various concentrations of opioids, and cell viability functions such as mitochondrial function and membrane integrity can easily be assessed using specific colorimetric assays.
Anabolic androgenic steroids (AAS) are frequently used to improve physical appearance and strength. AAS are known to affect muscle growth, but many AAS-users also experience psychiatric and ...behavioral changes after long-term use. The AAS-induced effects on the brain seem to depend on the type of steroid used, but the rationale behind the observed effect is still not clear. The present study investigated and compared the impact of nandrolone decanoate and testosterone undecanoate on body weight gain, levels of stress hormones, brain gene expression, and behavioral profiles in the male rat. The behavioral profile was determined using the multivariate concentric squared field test (MCSF-test). Blood plasma and brains were collected for further analysis using ELISA and qPCR. Nandrolone decanoate caused a reduction in body weight gain in comparison with both testosterone undecanoate and control. Rats receiving nandrolone decanoate also demonstrated decreased general activity in the MCSF. In addition, nandrolone decanoate reduced the plasma levels of ACTH in comparison with the control and increased the levels of corticosterone in comparison with testosterone undecanoate. The qPCR analysis revealed brain region-dependent changes in mRNA expression, where the hypothalamus was identified as the region most affected by the AAS. Alterations in neurotransmitter systems and stress hormones may contribute to the changes in behavior detected in the MCSF. In conclusion, both AAS affect the male rat, although, nandrolone decanoate has more pronounced impact on the physiological and the behavioral parameters measured.
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•Anabolic androgenic steroids caused physiological and psychological effects•Structurally diverse anabolic androgenic steroids differently affected the male rat•Neurotransmitter systems in the hypothalamus were particularly affected•The multivariate concentric square field™ was used to determine behavioral profiles•Nandrolone affected physical health and behavior more compared to testosterone
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•This study was performed using live cell imaging for four hours.•Methadone and fentanyl induced significant changes to mitochondrial morphology.•Morphine had no effect on ...mitochondrial morphology.
The important role of mitochondria in maintaining normal brain cell function has been demonstrated in several neurodegenerative diseases where mitochondrial dysfunction is a prominent feature. Accumulating evidence indicates that opioids may induce neuronal cell death and inhibit neurogenesis, two factors that are dependent on normal mitochondrial function. The aim of the present study was to examine the effects of morphine, methadone, and fentanyl on MitoTracker-stained mitochondria. Cells from the neuroblastoma/glioma hybrid cell-line NG108−15 were seeded on 96-well cell culture plates and treated with MitoTracker for 30 min prior to opioid treatment. Morphine, methadone, and fentanyl were added at various concentrations and images of mitochondria were acquired every 30 min for four hours using a high-content imaging device. The parameters total mitochondrial area, mitochondrial network, as well as the number and mean area of mitochondrial objects were analyzed using automated image analysis. Methadone and fentanyl, but not morphine, decreased the mitochondrial network, the number of mitochondrial objects, and increased the mean area of mitochondrial objects. Both methadone and fentanyl altered mitochondrial morphology with no effects seen from morphine treatment. These data suggest that methadone and fentanyl impact mitochondrial morphology negatively, which may be associated with neuronal cell death.
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•Structurally diverse anabolic steroids cause cellular toxicity.•Testosterone, nandrolone, and trenbolone induce apoptosis.•Stanozolol causes cell death, probably through ...necrosis.•The toxic steroid effects are primarily mediated by the androgen receptor.
The use of anabolic androgenic steroids (AASs) among non-athletes is a public health-problem, as abusers underestimate the negative effects associated with these drugs. The present study investigated the toxic effects of testosterone, nandrolone, stanozolol, and trenbolone, and aimed to understand how AAS abuse affects the brain. Mixed cortical cultures from embryonic rats were grown in vitro for 7 days and thereafter treated with increasing concentrations of AASs for 24 h (single-dose) or 3 days (repeated exposure). Cells were co-treated with the androgen-receptor (AR) antagonist flutamide, to determine whether the potential adverse effects observed were mediated by the AR. Cellular toxicity was determined by measuring mitochondrial activity, lactate dehydrogenase (LDH) release, and caspase-3/7 activity. Nandrolone, unlike the other AASs studied, indicated an effect on mitochondrial activity after 24 h. Furthermore, single-dose exposure with testosterone, nandrolone and trenbolone increased LDH release, while no effect was detected with stanozolol. However, all of the four steroids negatively affected mitochondrial function and resulted in LDH release after repeated exposure. Testosterone, nandrolone, and trenbolone caused their toxic effects by induction of apoptosis, unlike stanozolol that seemed to induce necrosis. Flutamide almost completely prevented AAS-induced toxicity by maintaining mitochondrial function, cellular integrity, and inhibition of apoptosis. Overall, we found that supra-physiological concentrations of AASs induce cell death in mixed primary cortical cultures, but to different extents, and possibly through various mechanisms. The data presented herein suggest that the molecular interactions of the AASs with the AR are primarily responsible for the toxic outcomes observed.
Anxiety disorders are common psychiatric conditions with a partially elucidated neurobiology. Caffeine, an unspecific adenosine receptor antagonist, is a common psychostimulant with anxiogenic ...effects in sensitive individuals. High doses of caffeine produce anxiety-like behavior in rats but it is not known if this is specific for rats with high baseline anxiety-like behavior. Thus, the aim of this study was to investigate general behavior, risk-taking, and anxiety-like behavior, as well as mRNA expression (adenosine A2A and A1, dopamine D2, and, μ, κ, δ opioid, receptors, BDNF, c-fos, IGF-1) in amygdala, caudate putamen, frontal cortex, hippocampus, hypothalamus, after an acute dose of caffeine. Untreated rats were screened using the elevated plus maze (EPM), giving each rat a score on anxiety-like behavior based on their time spent in the open arms, and categorized into a high or low anxiety-like behavior group accordingly. Three weeks after categorization, the rats were treated with 50 mg/kg caffeine and their behavior profile was studied in the multivariate concentric square field (MCSF) test, and one week later in the EPM. qPCR was performed on selected genes and corticosterone plasma levels were measured using ELISA. The results demonstrated that the high anxiety-like behavior rats treated with caffeine spent less time in risk areas of the MCSF and resituated towards the sheltered areas, a behavior accompanied by lower mRNA expression of adenosine A2A receptors in caudate putamen and increased BDNF expression in hippocampus. These results support the hypothesis that caffeine affects individuals differently depending on their baseline anxiety-like behavior, possibly involving adenosine receptors. This highlights the importance of adenosine receptors as a possible drug target for anxiety disorders, although further research is needed to fully elucidate the neurobiological mechanisms of caffeine on anxiety disorders.
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•Rats with high or low anxiety-like behavior respond differently to caffeine.•Caffeine reduced risk-taking behavior in rats with a high anxiety-like behavior.•Striatal A2A expression differs in rats with high or low anxiety-like behavior.•Caffeine increased BDNF in hippocampus in rats with a high anxiety-like behavior.
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•Structurally different anabolic steroids exhibit diverse neurotoxic properties.•Trenbolone, testosterone, and nandrolone reduce neurite outgrowth.•Trenbolone downregulates the Tubb3 ...gene expression and decreases cell viability.•Altered neurons may contribute to the mental health problems seen in AAS users.
The illicit use of anabolic androgenic steroids (AAS) among adolescents and young adults is a major concern due to the unknown and unpredictable impact of AAS on the developing brain and the consequences of this on mental health, cognitive function and behaviour. The present study aimed to investigate the effects of supra-physiological doses of four structurally different AAS (testosterone, nandrolone, stanozolol and trenbolone) on neurite development and cell viability using an in vitro model of immature primary rat cortical cell cultures. A high-throughput screening image-based approach, measuring the neurite length and number of neurons, was used for the analysis of neurite outgrowth. In addition, cell viability and expression of the Tubb3 gene (encoding the protein beta-III tubulin) were investigated. Testosterone, nandrolone, and trenbolone elicited adverse effects on neurite outgrowth as deduced from an observed reduced neurite length per neuron. Trenbolone was the only AAS that reduced the cell viability as indicated by a decreased number of neurons and declined mitochondrial function. Moreover, trenbolone downregulated the Tubb3 mRNA expression. The adverse impact on neurite development was neither inhibited nor supressed by the selective androgen receptor (AR) antagonist, flutamide, suggesting that the observed effects result from another mechanism or mechanisms of action that are operating apart from AR activation. The results demonstrate a possible AAS-induced detrimental effect on neuronal development and regenerative functions. An impact on these events, that are essential mechanisms for maintaining normal brain function, could possibly contribute to behavioural alterations seen in AAS users.
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