Anabolic Androgenic Steroids : Neurobiological Effects of Nandrolone, Testosterone, Trenbolone, and Stanozolol

The use of anabolic androgenic steroids (AAS) for recreational purposes is a health concern, as long-term AAS-use in supraphysiological doses is associated with severe physical and psychological adverse effects. Several behavioral and cognitive problems are reported after long-term AAS-use, and alterations in brain morphology as well as neurotransmitter systems have been reported. The AAS-induced negative impact on the brain may depend on the type of AAS used, but the rationale behind the adverse effects observed is still not clear. The aim of the present thesis was to investigate the neurobiological impact of supraphysiological doses of structurally diverse AAS; testosterone, nandrolone, stanozolol, and trenbolone, as well as of the prodrugs nandrolone decanoate, testosterone undecanoate, testosterone decanoate, and trenbolone decanoate. Wistar rats were used to study the influence on behavior, effects on the brain, and additional somatic effects. Furthermore, in vitro models of immature and mature primary rat cortical cell cultures were used to examine the potential toxic properties of the AAS administered. In the in vitro studies, nandrolone and trenbolone were identified as the most toxic of the AAS investigated, due to their adverse impact on mitochondrial function, membrane integrity, apoptosis, and neurite outgrowth. In vivo, the AAS demonstrated diverse somatic outcomes affecting body weight development, and organ weights to different degrees. In addition, nandrolone decanoate caused a reduced general activity, an effect possibly induced by increased stress vulnerability and alterations in the oxytocinergic system. Furthermore, nandrolone decanoate induced impaired memory in the novel object recognition test. Overall, nandrolone decanoate was identified as the most harmful steroid investigated due to its prominent impact on body weight development, affecting multiple organs, and being the only AAS causing impaired cognitive function.  In conclusion, the structurally different AAS exerted diverse effects on cell viability, neurite development as well as regarding physical impairments and impact on behavior, suggesting that harmful physiological, neurological, and psychological outcomes may be expected after AAS-use. These findings highlight that the severity and type of adverse effects depend on the type of AAS used, which is valuable information to consider in order to provide good healthcare and treatment options to AAS-users.  

Anabolic Androgenic Steroids : Neurobiological Effects of Nandrolone, Testosterone, Trenbolone, and Stanozolol

The use of anabolic androgenic steroids (AAS) for recreational purposes is a health concern, as long-term AAS-use in supraphysiological doses is associated with severe physical and psychological adverse effects. Several behavioral and cognitive problems are reported after long-term AAS-use, and alterations in brain morphology as well as neurotransmitter systems have been reported. The AAS-induced negative impact on the brain may depend on the type of AAS used, but the rationale behind the adverse effects observed is still not clear. The aim of the present thesis was to investigate the neurobiological impact of supraphysiological doses of structurally diverse AAS; testosterone, nandrolone, stanozolol, and trenbolone, as well as of the prodrugs nandrolone decanoate, testosterone undecanoate, testosterone decanoate, and trenbolone decanoate. Wistar rats were used to study the influence on behavior, effects on the brain, and additional somatic effects. Furthermore, in vitro models of immature and mature primary rat cortical cell cultures were used to examine the potential toxic properties of the AAS administered. In the in vitro studies, nandrolone and trenbolone were identified as the most toxic of the AAS investigated, due to their adverse impact on mitochondrial function, membrane integrity, apoptosis, and neurite outgrowth. In vivo, the AAS demonstrated diverse somatic outcomes affecting body weight development, and organ weights to different degrees. In addition, nandrolone decanoate caused a reduced general activity, an effect possibly induced by increased stress vulnerability and alterations in the oxytocinergic system. Furthermore, nandrolone decanoate induced impaired memory in the novel object recognition test. Overall, nandrolone decanoate was identified as the most harmful steroid investigated due to its prominent impact on body weight development, affecting multiple organs, and being the only AAS causing impaired cognitive function.  In conclusion, the structurally different AAS exerted diverse effects on cell viability, neurite development as well as regarding physical impairments and impact on behavior, suggesting that harmful physiological, neurological, and psychological outcomes may be expected after AAS-use. These findings highlight that the severity and type of adverse effects depend on the type of AAS used, which is valuable information to consider in order to provide good healthcare and treatment options to AAS-users.  

Nandrolone decanoate and testosterone undecanoate differently affect stress hormones, neurotransmitter systems, and general activity in the male rat.

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

Toxic Impact of Anabolic Androgenic Steroids in Primary Rat Cortical Cell Cultures

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.Även finansierat av Kjell och Märta Beijers stiftelse</p

The Protective and Restorative Effects of Growth Hormone and Insulin-Like Growth Factor-1 on Methadone-Induced Toxicity In Vitro

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.Även finansierat av Kjell och Märta Beijer stiftelsen.</p

Anabolic androgenic steroids exert a selective remodeling of the plasma lipidome that mirrors the decrease of the de novo lipogenesis in the liver

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)

Acute caffeine differently affects risk-taking and the expression of BDNF and of adenosine and opioid receptors in rats with high or low anxiety-like behavior

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

Structurally different anabolic androgenic steroids reduce neurite outgrowth and neuronal viability in primary rat cortical cell cultures

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

Hydrogen Peroxide Induced Toxicity Is Reversed by the Macrocyclic IRAP-Inhibitor HA08 in Primary Hippocampal Cell Cultures

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