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Exploring the neural mechanisms of finasteride: A proteomic analysis in the nucleus accumbens
... Epigenetic modification seems to be another candidate mechanism for depressive symptoms in prior 5-ARI users. One study found that an increase in four proteins (Syntaxin-18, Cytochrome P450 2B3, Collapsin response mediator protein 2, and phosphoribosyl pyrophosphate synthetase-1), and decreased GABA transaminase levels, in the NAc area following FIN treatment, may be linked to its anti-dopaminergic properties and the alterations of neuroactive steroids in NAc [63]. ...
... With regard to epigenetic modification, FIN may lead to the alterations of proteins involved in the regulation of steroid and pyrimidine metabolism, as well as GABAergic neurotransmission in NAc [63]. Future studies need to rigorously explore these aspects to verify these initial findings and increase information. ...
... Changes in neuroactive steroids following 5-ARI use can lead to dysfunction of the dopaminergic system, reduction of hippocampal neurogenesis, an increase in neuroinflammation, alterations of the HPA axis, and epigenetic modification. Moreover, the alterations of the neuroactive steroids, especially AP [67,96], are also linked to the alteration of central nervous system receptor functions including dopaminergic receptors [43,47], GABA-A receptors [63,67], estrogen receptors [67], and androgen receptors [67]. Unfortunately, the complete specific effects of 5-ARIs have not yet been fully characterized [67]. ...
Recent literature connects 5-alpha reductase inhibitors (5-ARIs) with neuropsychiatric adverse effects. Several clinical studies have indicated that former 5-ARIs users had a higher incidence of depressive symptoms and neuropsychiatric side effects than non-users. However, the underlying mechanisms involved in the depression in former 5-ARIs patients, a condition known as “post finasteride syndrome (PFS)”, are not thoroughly understood. This review aims to summarize and discuss the association between 5-ARIs and depression as well as possible mechanisms. We used PubMed search terms including “depression”, “depressive symptoms”, “MDD”, “anxiety”, or “suicidal idea”, and “5-alpha reductase inhibitors”, “finasteride”, “dutasteride”, “5-ARIs”. All relevant articles from in vivo and clinical studies from 2002 to 2021 were carefully reviewed. Any contradictory findings were included and debated. The potential mechanisms that link 5-ARIs and depression include alteration in neuroactive steroids, dopaminergic dysfunction, reduced hippocampal neurogenesis, increased neuroinflammation, alteration of the HPA axis, and epigenetic modifications. From this review, we hope to provide information for future studies based on animal experiments, and potential therapeutic strategies for depressive patients with PFS.
... For example, DHEA decreases the activity of monoamine oxidase, an enzyme necessary for the degradation of monoamines, in the NAc in male rats in vivo and in vitro (162). Pharmacological inhibition of 5αR suggests that DHT influences neurochemistry, particularly DA signaling, in the mesocorticolimbic system (163)(164)(165). Overall, these data suggest neurosteroids regulate DA turnover and DA signaling in the mesocorticolimbic system, which is important for regulating executive functions. ...
... The effects of apomorphine on PPI were attenuated by microinjecting (i.c.v.) the Cyp17a1 inhibitor abiraterone. Along with studies using systemic finasteride [5αR inhibitor (163,164,222,223)], these results suggest that local androgen synthesis regulates DA signaling in the mesocorticolimbic system and DA-dependent behaviors. While these studies are informative, there still remains an important gap in our understanding of how neural androgen production specifically influences executive functioning. ...
Multiple lines of evidence indicate that androgens, such as testosterone, modulate the mesocorticolimbic system and executive function. This review integrates neuroanatomical, molecular biological, neurochemical, and behavioral studies to highlight how endogenous and exogenous androgens alter behaviors, such as behavioral flexibility, decision making, and risk taking. First, we briefly review the neuroanatomy of the mesocorticolimbic system, which mediates executive function, with a focus on the ventral tegmental area (VTA), nucleus accumbens (NAc), medial prefrontal cortex (mPFC), and orbitofrontal cortex (OFC). Second, we present evidence that androgen receptors (AR) and other steroid receptors are expressed in the mesocorticolimbic system. Using sensitive immunohistochemistry and quantitative polymerase chain reaction (qPCR) techniques, ARs are detected in the VTA, NAc, mPFC, and OFC. Third, we describe recent evidence for local androgens (“neuroandrogens”) in the mesocorticolimbic system. Steroidogenic enzymes are expressed in mesocorticolimbic regions. Furthermore, following long-term gonadectomy, testosterone is nondetectable in the blood but detectable in the mesocorticolimbic system, using liquid chromatography tandem mass spectrometry. However, the physiological relevance of neuroandrogens remains unknown. Fourth, we review how anabolic-androgenic steroids (AAS) influence the mesocorticolimbic system. Fifth, we describe how androgens modulate the neurochemistry and structure of the mesocorticolimbic system, particularly with regard to dopaminergic signaling. Finally, we discuss evidence that androgens influence executive functions, including the effects of androgen deprivation therapy and AAS. Taken together, the evidence indicates that androgens are critical modulators of executive function. Similar to dopamine signaling, there might be optimal levels of androgen signaling within the mesocorticolimbic system for executive functioning. Future studies should examine the regulation and functions of neurosteroids in the mesocorticolimbic system, as well as the potential deleterious and enduring effects of AAS use.
... Other factors, in addition to neuroactive steroids, have been proposed to explain the pathogenesis of this syndrome as, for instance, alterations of dopaminergic signaling [126]. Indeed, as demonstrated in animal models, finasteride treatment was able to impair the signaling of dopamine (i.e., that is involved in the regulation of sex drive, as described above) [127,128]. ...
... However, whether this impairment is due to peripheral steroidogenesis and/or neurosteroidogenesis is still unrevealed. In addition, while serotonin signaling has been never considered, and alteration of dopaminergic pathways has been proposed [126]. However, whether this impairment still occurs after the discontinuation of finasteride (i.e., in the PFS condition) is still unclear. ...
Sexual dysfunction is a clinical condition due to different causes including the iatrogenic origin. For instance, it is well known that sexual dysfunction may occur in patients treated with antidepressants like selective serotonin reuptake inhibitors (SSRI). A similar side effect has been also reported during treatment with finasteride, an inhibitor of the enzyme 5alpha-reductase, for androgenetic alopecia. Interestingly, sexual dysfunction persists in both cases after drug discontinuation. These conditions have been named post-SSRI sexual dysfunction (PSSD) and post-finasteride syndrome (PFS). In particular, feeling of a lack of connection between the brain and penis, loss of libido and sex drive, difficulty in achieving an erection and genital paresthesia have been reported by patients of both conditions. It is interesting to note that the incidence of these diseases is probably so far underestimated and their etiopathogenesis is not sufficiently explored. To this aim, the present review will report the state of art of these two different pathologies and discuss, on the basis of the role exerted by three different neuromodulators such as dopamine, serotonin and neuroactive steroids, whether the persistent sexual dysfunction observed could be determined by common mechanisms.
... In other models, finasteride induces changes in steroid levels in specific brain regions (45,46), but because of the small size of the zebrafish brain, we used whole brains and could not achieve the same level of regional specificity. Nonetheless, we identified interesting trends, including an accumulation of several 5αR precursors and a reduction in 5αR products after treatment with finasteride. ...
Opioid use disorder (OUD) has become a leading cause of death in the US, yet current therapeutic strategies remain highly inadequate. To identify novel potential treatments for OUD, we screened a targeted selection of over 100 drugs using a recently developed opioid self-administration assay in zebrafish. This paradigm showed that finasteride, a steroidogenesis inhibitor approved for the treatment of benign prostatic hyperplasia and androgenetic alopecia, reduced self-administration of multiple opioids without affecting locomotion or feeding behavior. These findings were confirmed in rats; furthermore, finasteride reduced the physical signs associated with opioid withdrawal. In rat models of neuropathic pain, finasteride did not alter the antinociceptive effect of opioids and reduced withdrawal-induced hyperalgesia. Steroidomic analyses of the brains of fish treated with finasteride revealed a significant increase in dehydroepiandrosterone sulfate (DHEAS). Treatment with precursors of DHEAS reduced opioid self-administration in zebrafish in a fashion akin to the effects of finasteride. These results highlight the importance of steroidogenic pathways as a rich source of therapeutic targets for OUD and point to the potential of finasteride as a new treatment option for this disorder.
... Protein identification was performed according to previous studies [22,23]. Briefly, after different steps of dehydration, reduction and alkylation, the excided single spots were digested with a solution of 0.01 µg/µL of porcine trypsin (Promega, Madison, WI, USA) at 37 • C o.n., and peptides were concentrated using C18 ZipTip (Millipore, Bedford, MA, USA). ...
Antimicrobial resistance (AMR) represents one of the most critical challenges that humanity will face in the following years. In this context, a “One Health” approach with an integrated multidisciplinary effort involving humans, animals and their surrounding environment is needed to tackle the spread of AMR. One of the most common ways for bacteria to live is to adhere to surfaces and form biofilms. Staphylococcus aureus (S. aureus) can form biofilm on most surfaces and in a wide heterogeneity of environmental conditions. The biofilm guarantees the survival of the S. aureus in harsh environmental conditions and represents an issue for the food industry and animal production. The identification and characterization of biofilm-related proteins may provide interesting insights into biofilm formation mechanisms in S. aureus. In this regard, the aims of this study were: (i) to use proteomics to compare proteomes of S. aureus growing in planktonic and biofilm forms in order to investigate the common features of biofilm formation properties of different strains; (ii) to identify specific biofilm mechanisms that may be involved in AMR. The proteomic analysis showed 14 differentially expressed proteins among biofilm and planktonic forms of S. aureus. Moreover, three proteins, such as alcohol dehydrogenase, ATP-dependent 6-phosphofructokinase, and fructose-bisphosphate aldolase, were only differentially expressed in strains classified as high biofilm producers. Differentially regulated catabolites metabolisms and the switch to lower oxygen-related metabolisms were related to the sessile conformation analyzed.
... To investigate the landscape of changes induced by the treatment of opioid conditioned animals with finasteride, we used an untargeted approach to quantify steroids in the brain. In other models, finasteride induces changes in steroid levels in specific brain regions 58,59 , but because of the small size of the zebrafish brain, we used whole brains and could not achieve the same level of regional specificity. Nonetheless, we identified interesting trends, including an accumulation of several 5αR precursors and a reduction in 5αR products after treatment with finasteride. ...
Opioid use disorder (OUD) has become a leading cause of death in the US, yet current therapeutic strategies remain highly inadequate. To identify novel potential treatments for OUD, we screened a targeted selection of over 100 drugs, using a recently developed opioid self-administration assay in zebrafish. This paradigm showed that finasteride, a steroidogenesis inhibitor approved for the treatment of benign prostatic hyperplasia and androgenetic alopecia, reduced self-administration of multiple opioids without affecting locomotion or feeding behavior. These findings were confirmed in rats; furthermore, finasteride did not interfere with the antinociceptive effect of opioids in rat models of neuropathic pain. Steroidomic analyses of the brains of fish treated with finasteride revealed a significant increase in dehydroepiandrosterone sulfate (DHEAS). Treatment with precursors of DHEAS reduced opioid self-administration in zebrafish, in a fashion akin to the effects of finasteride. Our results highlight the importance of steroidogenic pathways as a rich source of therapeutic targets for OUD and point to the potential of finasteride as a new option for this disorder.
... поскольку считается, что нейростероиды обладают анксиолитическим, антидепрессивным действием, а также улучшают когнитивные функции и играют роль в нейропротекции, теоретически можно предположить отрицательное действие применения 5-АРИ на психические функции. Кроме того, было выявлено снижение допаминергической передачи сигналов, а ведь допамин является одним из основных нейротрансмиттеров, определяющих сексуальное поведение [7]. Нужно отметить, что данные изменения имели место у крайне малого числа пациентов, принимающих 5-АРИ, что может свидетельствовать о роли эпигенетических механизмов, которые не изучены при пФс. ...
5-α reductase inhibitors are effective and commonly used medications for the treatment of benign prostatic hyperplasia. However, there are some questions about the safety of such medications. A post-finasteride syndrome is a controversial syndrome associated with a constellation of sexual, physical, and psychological symptoms that develop during or after finasteride exposure and persist after discontinuation. Finasteride and dutasteride have the same mechanism of action and we can suppose that the term «post-finasteride syndrome» may apply to the whole group of 5-alfa reductase inhibitors. Nowadays, reasons and development mechanisms of this syndrome are unclear. The results of studies on sexual, physical, and psychological disorders have shown mixed results. Some experts attribute post-finasteride syndrome to a group of «mysterious diseases» based on psychosocial factors. Also, we need new studies with better design because reasons and development mechanisms of this syndrome are unclear.
Aleksandr A. Galushkin and Dmitriу G. Likhikh are employees GlaxoSmithKline Trading, Michael I. Kogan declares no conflict of interest.
... Spots of interest were cut out from gels, trypsinized as reported by Giusti et al. (21). Protein identification was performed according to Soggiu et al. (22). Briefly after the digestion protocol, peptides were Receiver-operating characteristic (ROC) curves were plotted using standard techniques to assess sensitivity and specificity of serum PSAP and QSOX1 to differentiate MPM patients from asbestos-exposed healthy subjects calculated with their 95% confidence intervals (95% CI). ...
Background:
Malignant pleural mesothelioma (MPM) a rare neoplasm linked to asbestos exposure is characterized by a poor prognosis. Soluble mesothelin is currently considered the most specific diagnostic biomarker. The aim of the study was to identify novel biomarkers by proteomic analysis of two MPM cell lines secretome.
Materials and methods:
The protein patterns of MPM cells secretome were examined and compared to a non-malignant mesothelial cell line using two-dimensional gel electrophoresis coupled to mass spectrometry. Serum levels of candidate biomarkers were determined in MPM patients and control subjects.
Results:
Two up-regulated proteins involved in cancer biology, prosaposin and quiescin Q6 sulfhydryl oxidase 1, were considered candidate biomarkers. Serum levels of both proteins were significantly higher in MPM patients than control subjects. Combining the data of each receiver-operating characteristic analysis predicted a good diagnostic accuracy.
Conclusion:
A panel of the putative biomarkers represents a promising tool for MPM diagnosis.
... In addition to the reduction in HPA axis hormones, other mechanisms may be involved in the effects of FIN. For example, the effects of this drug on the reduction of responses to salient stimuli may reflect the marked antidopaminergic effects of FIN in the prefrontal cortex and nucleus accumbens, which have been documented by our previous studies [26,[61][62][63]; indeed, dopaminergic signaling in these brain areas is pivotal to allowing for behavioral arousal and salience appraisal [64,65]. Future studies are needed to understand the involvement of the HPA axis, dopamine pathways, or other neural mechanisms in the behavioral effects of FIN. ...
Finasteride (FIN) is the prototypical inhibitor of steroid 5α-reductase (5αR), the enzyme that catalyzes the rate-limiting step of the conversion of progesterone and testosterone into their main neuroactive metabolites. FIN is clinically approved for the treatment of benign prostatic hyperplasia and male baldness; while often well-tolerated, FIN has also been shown to cause or exacerbate psychological problems in vulnerable subjects. Evidence on the psychological effects of FIN, however, remains controversial, in view of inconsistent clinical reports. Here, we tested the effects of FIN in a battery of tests aimed at capturing complementary aspects of mood regulation and stress reactivity in rats. FIN reduced exploratory, incentive, prosocial, and risk-taking behavior; furthermore, it decreased stress coping, as revealed by increased immobility in the forced-swim test (FST). This last effect was also observed in female and orchiectomized male rats, suggesting that the mechanism of action of FIN does not primarily reflect changes in gonadal steroids. The effects of FIN on FST responses were associated with a dramatic decrease in corticotropin release hormone (CRH) mRNA and adrenocorticotropic hormone (ACTH) levels. These results suggest that FIN impairs stress reactivity and reduces behavioral activation and impulsive behavior by altering the function of the hypothalamus–pituitary–adrenal (HPA) axis.
... In addition, finasteride-induced epigenetic changes manifested in upregulation of AR suggest that such epigenetic modifications may manifest itself in the symptoms of PFS [23,42,52,144]. Finally, finasteride-induced impairment of dopaminergic signaling [145] may contribute to this poorly understood epigenetic response [146,147]. Thus, finasteride inhibition of 5α-Rs results in reduced biosynthesis and metabolism of several key neurosteroids. ...
Purpose of Review
Post-finasteride syndrome (PFS) is a disorder characterized by a set of clinical symptoms experienced during use or after drug discontinuation. This cluster of symptoms encompasses overall sexual dysfunction (SD), erectile dysfunction (ED), loss of libido, depression, suicidal ideation, anxiety, panic attacks, insomnia, and cognitive dysfunction. To date, there is lack of comprehensive understanding of the biochemical and pathophysiological mechanisms responsible for the adverse effects of finasteride. More importantly, there is lack of knowledge and effective clinical tools for treatments of this condition, resulting in outright dismissal of complaints by individuals afflicted with this syndrome. Psychological symptoms and cognitive dysfunction of PFS are far more serious and difficult to treat than sexual dysfunction symptoms and may lead young men to contemplate, attempt, or even commit suicide. Therefore, an urgent need exists to fill the knowledge gap in physiology, pathophysiology, and clinical management of patients with PFS.
Recent Findings
Finasteride treatment impairs biosynthesis and function of neurosteroids, which are critical regulators of central (CNS) as well as peripheral nervous system functions and modulate a host of neurotransmitter receptors, such as gamma amino butyric acid receptors. Thus, finasteride-induced neuroendocrine disruption of biosynthesis of critical signaling molecules results in pathophysiological states, which contribute to inhibition of biochemical pathways responsible for a host of physiological functions, ranging from sexual activity, mood, and cognition. In addition, finasteride-induced epigenetic changes in gene expression, including upregulation of androgen receptors (AR), increased histone acetylation, and methylation results in undesirable biological outcomes such as impairment of dopaminergic signaling and modulation of other neurotransmitter receptors, may be the underlying mechanism causing persistent or permanent adverse effects, manifested in anxiety, depression, and suicidal ideation.
Summary
The medical community has an obligation not to turn a blind eye on this rare yet debilitating condition in young men. Patients with this condition should not be stereotyped or stigmatized by untrained and unprepared clinicians, due to lack of awareness and knowledge pertaining to this new and rare syndrome. Greater awareness and education is needed among the medical and scientific communities in order to develop better approaches for managing men with PFS. It is paramount that steps are taken to develop better understanding of the underlying mechanisms contributing to the onset and progression of PFS and to promote educational and training programs to increase awareness and improve management of this condition.
... These analyses may prove essential to help clarify the male predominance of TS, as well as potential mechanisms of comorbidity with other neuropsychiatric problems, including ADHD and OCD. Fourth, our analyses were only limited to the PFC of adult mice; however, it is likely that the effects of other neurosteroids may differ with age; furthermore, other regions, such as the nucleus accumbens, may be involved in the effects of finasteride 47 . Given the limitations in the size of this region, however, further improvements in our ability to detect neurosteroid levels will be needed to address this issue. ...
Tourette syndrome (TS) is a neuropsychiatric disorder characterized by multiple tics and sensorimotor abnormalities, the severity of which is typically increased by stress. The neurobiological underpinnings of this exacerbation, however, remain elusive. We recently reported that spatial confinement (SC), a moderate environmental stressor, increases tic-like responses and elicits TS-like sensorimotor gating deficits in the D1CT-7 mouse, one of the best-validated models of TS. Here, we hypothesized that these adverse effects may be mediated by neurosteroids, given their well-documented role in stress-response orchestration. Indeed, SC increased the levels of progesterone, as well as its derivatives 5α-dihydroprogesterone and allopregnanolone, in the prefrontal cortex (PFC) of D1CT-7 mice. Among these steroids, however, only allopregnanolone (5–15 mg/kg, IP) dose-dependently exacerbated TS-like manifestations in D1CT-7, but not wild-type littermates; these effects were countered by the benchmark anti-tic therapy haloperidol (0.3 mg/kg, IP). Furthermore, the phenotypic effects of spatial confinement in D1CT-7 mice were suppressed by finasteride (25–50 mg/kg, IP), an inhibitor of the main rate-limiting enzyme in allopregnanolone synthesis. These findings collectively suggest that stress may exacerbate TS symptoms by promoting allopregnanolone synthesis in the PFC, and corroborate previous clinical results pointing to finasteride as a novel therapeutic avenue to curb symptom fluctuations in TS.
... This could explain the andrological and psychiatric features observed in PFS patients. However, even if the present observations add another piece of information to what has been so far proposed by others, such as an alteration of dopaminergic signalling in the nucleus accumbens (i.e., a brain region that is critical for normal libido and mood regulation) [63], lateralization process of the brain [64] or pre-existing familial mental health condition [65] a clear demonstration of the pathogenic mechanism underlying the PFS is not yet understood. ...
Recent reports show that, in patients treated with finasteride for male pattern hair loss, persistent side effects including sexual side effects, depression, anxiety and cognitive complaints may occur. We here explored the psychiatric and andrological features of patients affected by post-finasteride syndrome (PFS) and verified whether the cerebrospinal fluid (CSF) and plasma levels of neuroactive steroids (i.e., important regulators of nervous function) are modified. We found that eight out of sixteen PFS male patients considered suffered from a DSM-IV major depressive disorder (MDD). In addition, all PFS patients showed erectile dysfunction (ED); in particular, ten patients showed a severe and six a mild-moderate ED. We also reported abnormal somatosensory evoked potentials of the pudendal nerve in PFS patients with severe ED, the first objective evidence of a neuropathy involving peripheral neurogenic control of erection. Testicular volume by ultrasonography was normal in PFS patients. Data obtained on neuroactive steroid levels also indicate interesting features. Indeed, decreased levels of pregnenolone, progesterone and its metabolite (i.e., dihydroprogesterone), dihydrotestosterone and 17beta-estradiol and increased levels of dehydroepiandrosterone, testosterone and 5alpha-androstane-3alpha,17beta-diol were observed in CSF of PFS patients. Neuroactive steroid levels were also altered in plasma of PFS patients, however these changes did not reflect exactly what occurs in CSF. Finally, finasteride did not only affect, as expected, the levels of 5alpha-reduced metabolites of progesterone and testosterone, but also the further metabolites and precursors suggesting that this drug has broad consequence on neuroactive steroid levels of PFS patients.
Affective disorders such as depression and anxiety are among the most prevalent psychiatric illnesses and causes of disability worldwide. The recent FDA-approval of a novel antidepressant treatment, ZULRESSO® (Brexanolone), a synthetic neurosteroid has fueled interest into the role of neurosteroids in the pathophysiology of depression as well as the mechanisms mediating the antidepressant effects of these compounds. The majority of studies examining the impact of neurosteroids on affective states have relied on the administration of exogenous neurosteroids; however, neurosteroids can also be synthesized endogenously from cholesterol or steroid hormone precursors. Despite the well-established influence of exogenous neurosteroids on affective states, we still lack an understanding of the role of endogenous neurosteroids in modulating affective tone. This review aims to summarize the current literature supporting the influence of neurosteroids on affective states in clinical and preclinical studies, as well as recent evidence suggesting that endogenous neurosteroids may set a baseline affective tone.
The enzyme steroid 5α-reductase 2 (5αR2) catalyzes the conversion of testosterone into the potent androgen 5α-dihydrotestosterone. Previous investigations showed that 5αR2 is expressed in key brain areas for emotional and socio-affective reactivity, yet the role of this enzyme in behavioral regulation remains mostly unknown. Here, we profiled the behavioral characteristics of 5αR2 heterozygous (HZ) and knockout (KO) mice, as compared with their wild-type (WT) littermates. While male 5αR2 KO mice displayed no overt alterations in motoric, sensory, information-processing and anxiety-related behaviors, they exhibited deficits in neurobehavioral correlates of dominance (including aggression against intruders, mating, and tube dominance) as well as novelty-seeking and risk-taking responses. Furthermore, male 5αR2 KO mice exhibited reduced D2-like dopamine receptor binding in the shell of the nucleus accumbens – a well-recognized molecular signature of social dominance. Collectively, these results suggest that 5αR2 is involved in the establishment of social dominance and its behavioral manifestations. Further studies are warranted to understand how the metabolic actions of 5αR2 on steroid profile may be implicated in social ranking, impulse control, and the modulation of dopamine receptor expression in the nucleus accumbens.
PANTHER (Protein Analysis THrough Evolutionary Relationships, http://pantherdb.org) is a widely used online resource for comprehensive protein evolutionary and functional classification, and includes tools
for large-scale biological data analysis. Recent development has been focused in three main areas: genome coverage, functional
information (‘annotation’) coverage and accuracy, and improved genomic data analysis tools. The latest version of PANTHER,
10.0, includes almost 5000 new protein families (for a total of over 12 000 families), each with a reference phylogenetic
tree including protein-coding genes from 104 fully sequenced genomes spanning all kingdoms of life. Phylogenetic trees now
include inference of horizontal transfer events in addition to speciation and gene duplication events. Functional annotations
are regularly updated using the models generated by the Gene Ontology Phylogenetic Annotation Project. For the data analysis
tools, PANTHER has expanded the number of different ‘functional annotation sets’ available for functional enrichment testing,
allowing analyses to access all Gene Ontology annotations—updated monthly from the Gene Ontology database—in addition to the annotations that have been inferred
through evolutionary relationships. The Prowler (data browser) has been updated to enable users to more efficiently browse
the entire database, and to create custom gene lists using the multiple axes of classification in PANTHER.
Neurosteroids exert diverse modulatory actions on dopamine neurotransmission and signaling. We previously documented that the enzyme 5α-reductase, which catalyzes the main rate-limiting step in neurosteroid synthesis, is required for the behavioral responses of Sprague-Dawley rats to non-selective dopaminergic agonists, such as the D1-D2 receptor agonist apomorphine. Specifically, systemic and intra-accumbal administrations of the 5α-reductase inhibitor finasteride countered apomorphine-induced deficits of sensorimotor gating, as measured by the prepulse inhibition (PPI) of the startle reflex; the classes of dopamine receptors involved in these effects, however, remain unknown. Prior rodent studies have revealed that the contributions of dopamine receptors to PPI regulation vary depending on the genetic background; thus, we analyzed the effect of finasteride on the PPI deficits induced by selective dopamine receptor agonists in Long-Evans (a strain exhibiting PPI deficits in response to both D1 and D2 receptor agonists) and Sprague-Dawley rats (which display PPI reductions following treatment with D2, and D3, but not D1 receptor agonists). In Long-Evans rats, finasteride opposed the PPI deficits induced by activation of D1, but not D2 receptors; conversely, in Sprague-Dawley rats, finasteride prevented the reductions in %PPI and accumbal dopamine extracellular levels caused by selective stimulation of D3, but not D2 receptors; however, the effects on %PPI were not confirmed by analyses on absolute PPI values. Our findings suggest that 5α-reductase modulates the effects of D1, but not D2 receptor agonists on sensorimotor gating. These data may help elucidate the role of neurosteroids in neuropsychiatric disorders featuring PPI deficits, including schizophrenia and Tourette syndrome.
N-methyl-D-aspartate receptors (NMDARs) mediate synaptic plasticity, and their dysfunction is implicated in multiple brain disorders. NMDARs can be allosterically modulated by numerous compounds, including endogenous neurosteroid pregnanolone sulfate. Here, we identify the molecular basis of the use-dependent and voltage-independent inhibitory effect of neurosteroids on NMDAR responses. The site of action is located at the extracellular vestibule of the receptor’s ion channel pore and is accessible after receptor activation. Mutations in the extracellular vestibule in the SYTANLAAF motif disrupt the inhibitory effect of negatively charged steroids. In contrast, positively charged steroids inhibit mutated NMDAR responses in a voltage-dependent manner. These results, in combination with molecular modeling, characterize structure details of the open configuration of the NMDAR channel. Our results provide a unique opportunity for the development of new therapeutic neurosteroid-based ligands to treat diseases associated with dysfunction of the glutamate system.
The brain has high energy requirements to maintain neuronal activity. Consequently impaired mitochondrial function will lead to disease. Normal aging is associated with several alterations in neurosteroid production and secretion. Decreases in neurosteroid levels might contribute to brain aging and loss of important nervous functions, such as memory. Up to now, extensive studies only focused on estradiol as a promising neurosteroid compound that is able to ameliorate cellular bioenergetics, while the effects of other steroids on brain mitochondria are poorly understood or not investigated at all. Thus, we aimed to characterize the bioenergetic modulating profile of a panel of seven structurally diverse neurosteroids (progesterone, estradiol, estrone, testosterone, 3α-androstanediol, DHEA and allopregnanolone), known to be involved in brain function regulation. Of note, most of the steroids tested were able to improve bioenergetic activity in neuronal cells by increasing ATP levels, mitochondrial membrane potential and basal mitochondrial respiration. In parallel, they modulated redox homeostasis by increasing antioxidant activity, probably as a compensatory mechanism to a slight enhancement of ROS which might result from the rise in oxygen consumption. Thereby, neurosteroids appeared to act via their corresponding receptors and exhibited specific bioenergetic profiles. Taken together, our results indicate that the ability to boost mitochondria is not unique to estradiol, but seems to be a rather common mechanism of different steroids in the brain. Thus, neurosteroids may act upon neuronal bioenergetics in a delicate balance and an age-related steroid disturbance might be involved in mitochondrial dysfunction underlying neurodegenerative disorders.
α-Synuclein is thought to regulate neurotransmitter release through multiple interactions with presynaptic proteins, cytoskeletal elements, ion channels, and synaptic vesicles membrane. α-Synuclein is abundant in the presynaptic compartment, and its release from neurons and glia has been described as responsible for spreading of α-synuclein-derived pathology. α-Synuclein-dependent dysregulation of neurotransmitter release might occur via its action on surface-exposed calcium channels. Here, we provide electrophysiological and biochemical evidence to show that α-synuclein, applied to rat neurons in culture or striatal slices, selectively activates Cav2.2 channels, and said activation correlates with increased neurotransmitter release. Furthermore, in vivo perfusion of α-synuclein into the striatum also leads to acute dopamine release. We further demonstrate that α-synuclein reduces the amount of plasma membrane cholesterol and alters the partitioning of Cav2.2 channels, which move from raft to cholesterol-poor areas of the plasma membrane. We provide evidence for a novel mechanism through which α-synuclein acts from the extracellular milieu to modulate neurotransmitter release and propose a unifying hypothesis for the mechanism of α-synuclein action on multiple targets: the reorganization of plasma membrane microdomains.
Collapsin response mediator protein 2 (CRMP2) is traditionally viewed as an axonal growth protein involved in axon/dendrite specification. Here, we describe novel functions of CRMP2. A 15-amino acid peptide from CRMP2, fused to the tat cell penetrating motif of the HIV-1 protein, tat-CBD3, but not CBD3 sans tat, attenuated N-methyl-D-aspartate receptor (NMDAR) activity and protected neurons against glutamate-induced Ca(2+) dysregulation, suggesting the key contribution of CRMP2 in these processes. In addition, tat-CBD3, but not CBD3 sans tat or tat-scramble peptide, inhibited increases in cytosolic Ca(2+) mediated by the plasmalemmal Na(+)/Ca(2+) exchanger (NCX) operating in the reverse mode. Co-immunoprecipitation experiments revealed an interaction between CRMP2 and NMDAR as well as NCX3 but not NCX1. Tat-CBD3 disrupted CRMP2-NMDAR interaction without change in NMDAR localization. In contrast, tat-CBD3 augmented the CRMP2-NCX3 co-immunoprecipitation, indicating increased interaction or stabilization of a complex between these proteins. Immunostaining with an anti-NCX3 antibody revealed that tat-CBD3 induced NCX3 internalization, suggesting that both reverse and forward modes of NCX might be affected. Indeed, the forward mode of NCX, evaluated in experiments with ionomycin-induced Ca(2+) influx into neurons, was strongly suppressed by tat-CBD3. Knockdown of CRMP2 with short interfering RNA (siRNA) prevented NCX3 internalization in response to tat-CBD3 exposure. Moreover CRMP2 downregulation strongly attenuated tat-CBD3-induced inhibition of reverse NCX. Overall, our results demonstrate that CRMP2 interacts with NCX and NMDAR and that tat-CBD3 protects against glutamate-induced Ca(2+) dysregulation most likely via suppression of both NMDAR and NCX activities. Our results further clarify the mechanism of action of tat-CBD3 and identify a novel regulatory checkpoint for NMDAR and NCX function based on CRMP2 interaction with these proteins.
Bupropion is indicated to promote smoking cessation. Animal studies suggest that the pharmacologic activity of bupropion can be mediated by its major metabolite, hydroxybupropion. We measured plasma bupropion and its metabolite levels in a double-blind, placebo controlled, randomized smoking-cessation trial. Among the treatment-adherent individuals, higher hydroxybupropion concentrations (per μg/ml) resulted in better smoking-cessation outcomes (week 3, 7, and 26 odds ratio (OR) = 2.82, 2.96, and 2.37, respectively, P = 0.005-0.040); this was not observed with bupropion levels (OR = 1.00-1.03, P = 0.59-0.90). Genetic variation in CYP2B6, the enzyme that metabolizes bupropion to hydroxybupropion, was identified as a significant source of variability in hydroxybupropion formation. Our data indicate that hydroxybupropion contributes to the pharmacologic effects of bupropion for smoking cessation, and that variability in response to bupropion treatment is related to variability in CYP2B6-mediated hydroxybupropion formation. These findings suggest that dosing of bupropion to achieve a hydroxybupropion level of 0.7 μg/ml or increasing bupropion dose for CYP2B6 slow metabolizers could improve bupropion's cessation outcomes.Clinical Pharmacology & Therapeutics (2012); advance online publication 14 November 2012. doi:10.1038/clpt.2012.186.
Studies addressing behavioral functions of dopamine (DA) in the nucleus accumbens septi (NAS) are reviewed. A role of NAS DA in reward has long been suggested. However, some investigators have questioned the role of NAS DA in rewarding effects because of its role in aversive contexts. As findings supporting the role of NAS DA in mediating aversively motivated behaviors accumulate, it is necessary to accommodate such data for understanding the role of NAS DA in behavior. The aim of the present paper is to provide a unifying interpretation that can account for the functions of NAS DA in a variety of behavioral contexts: (1) its role in appetitive behavioral arousal, (2) its role as a facilitator as well as an inducer of reward processes, and (3) its presently undefined role in aversive contexts. The present analysis suggests that NAS DA plays an important role in sensorimotor integrations that facilitate flexible approach responses. Flexible approach responses are contrasted with fixed instrumental approach responses (habits), which may involve the nigro-striatal DA system more than the meso-accumbens DA system. Functional properties of NAS DA transmission are considered in two stages: unconditioned behavioral invigoration effects and incentive learning effects. (1) When organisms are presented with salient stimuli (e.g., novel stimuli and incentive stimuli), NAS DA is released and invigorates flexible approach responses (invigoration effects). (2) When proximal exteroceptive receptors are stimulated by unconditioned stimuli, NAS DA is released and enables stimulus representations to acquire incentive properties within specific environmental context. It is important to make a distinction that NAS DA is a critical component for the conditional formation of incentive representations but not the retrieval of incentive stimuli or behavioral expressions based on over-learned incentive responses (i.e., habits). Nor is NAS DA essential for the cognitive perception of environmental stimuli. Therefore, even without normal NAS DA transmission, the habit response system still allows animals to perform instrumental responses given that the tasks take place in fixed environment. Such a role of NAS DA as an incentive-property constructor is not limited to appetitive contexts but also aversive contexts. This dual action of NAS DA in invigoration and incentive learning may explain the rewarding effects of NAS DA as well as other effects of NAS DA in a variety of contexts including avoidance and unconditioned/conditioned increases in open-field locomotor activity. Particularly, the present hypothesis offers the following interpretation for the finding that both conditioned and unconditioned aversive stimuli stimulate DA release in the NAS: NAS DA invigorates approach responses toward `safety'. Moreover, NAS DA modulates incentive properties of the environment so that organisms emit approach responses toward `safety' (i.e., avoidance responses) when animals later encounter similar environmental contexts. There may be no obligatory relationship between NAS DA release and positive subjective effects, even though these systems probably interact with other brain systems which can mediate such effects. The present conceptual framework may be valuable in understanding the dynamic interplay of NAS DA neurochemistry and behavior, both normal and pathophysiological.
Drug resistance in food-borne bacterial pathogens is an almost inevitable consequence of the use of antimicrobial drugs, used either therapeutically or to avoid infections in food-producing animals. In the past decades, the spread and inappropriate use of antibiotics have caused a considerable increase of antibiotics to which bacteria have developed resistance and, moreover, bacteria are becoming resistant to more than one antibiotic simultaneously. Understanding mechanisms at the molecular level is extremely important to control multi-resistant strains and to develop new therapeutic strategies. In the present study, comparative proteomics was applied to characterize membrane and cytosolic proteome in order to investigate the regulation of protein expression in multi-resistance E. coli isolated from young never vaccinated water buffalo. Results highlighted differentially expressed proteins under multi drug resistance conditions giving new insights about mechanisms involved in resistance, as quorum sensing mechanisms, and suggesting possible novel bacterial targets to develop alternative antibiotic drugs.
The use of N-type voltage-gated calcium channel (CaV2.2) blockers to treat pain is limited by many physiological side effects. Here we report that inflammatory and neuropathic hypersensitivity can be suppressed by inhibiting the binding of collapsin response mediator protein 2 (CRMP-2) to CaV2.2 and thereby reducing channel function. A peptide of CRMP-2 fused to the HIV transactivator of transcription (TAT) protein (TAT-CBD3) decreased neuropeptide release from sensory neurons and excitatory synaptic transmission in dorsal horn neurons, reduced meningeal blood flow, reduced nocifensive behavior induced by formalin injection or corneal capsaicin application and reversed neuropathic hypersensitivity produced by an antiretroviral drug. TAT-CBD3 was mildly anxiolytic without affecting memory retrieval, sensorimotor function or depression. At doses tenfold higher than that required to reduce hypersensitivity in vivo, TAT-CBD3 caused a transient episode of tail kinking and body contortion. By preventing CRMP-2-mediated enhancement of CaV2.2 function, TAT-CBD3 alleviated inflammatory and neuropathic hypersensitivity, an approach that may prove useful in managing chronic pain.
The nucleus accumbens (NAc) is involved in many reward-related behaviors. The NAc has two major components, the core and the shell. These two areas have different inputs and outputs, suggesting that they contribute differentially to goal-directed behaviors. Using a discriminative stimulus (DS) task in rats and inactivating the NAc by blocking excitatory inputs with glutamate antagonists, we dissociated core and shell contributions to task performance. NAc core but not shell inactivation decreased responding to a reward-predictive cue. In contrast, inactivation of either subregion induced a general behavioral disinhibition. This reveals that the NAc actively suppresses actions inappropriate to the DS task. Importantly, selective inactivation of the shell but not core significantly increased responding to the nonrewarded cue. To determine whether the different contributions of the NAc core and shell depend on the information encoded in their constituent neurons, we performed electrophysiological recording in rats performing the DS task. Although there was no firing pattern unique to either core or shell, the reward-predictive cue elicited more frequent and larger magnitude responses in the NAc core than in the shell. Conversely, more NAc shell neurons selectively responded to the nonrewarded stimulus. These quantitative differences might account for the different behavioral patterns that require either core or shell. Neurons with similar firing patterns could also have different effects on behavior due to their distinct projection targets.
Some hippocampally-influenced affective and/or cognitive processes decline with aging. The role of androgens in this process is of interest. Testosterone (T) is aromatized to estrogen, and reduced to dihydrotestosterone (DHT), which is converted to 5alpha-androstane, 3alpha, 17alpha-diol (3alpha-diol). To determine the extent to which some age-related decline in hippocampally-influenced behaviors may be due to androgens, we examined the effects of variation in androgen levels due to age, gonadectomy, and androgen replacement on cognitive (inhibitory avoidance, Morris water maze) and affective (defensive freezing, forced swim) behavior among young (4 months), middle-aged (13 months), and aged (24 months) male rats. Plasma and hippocampal levels of androgens were determined. In experiment 1, comparisons were made between 4-, 13-, and 24-month-old rats that were intact or gonadectomized (GDX) and administered a T-filled or empty silastic capsule. There was age-related decline in performance of the inhibitory avoidance, water maze, defensive freezing, and forced swim tasks, and hippocampal 3alpha-diol levels. Chronic, long-term (1-4 weeks) T-replacement reversed the effects of GDX in 4- and 13-month-old, but not 24-month-old, rats in the inhibitory avoidance task. Experiments 2 and 3 assessed whether acute subcutaneous T or 3alpha-diol, respectively, could reverse age-associated decline in performance. 3alpha-diol, but not T, compared to vehicle, improved performance in the inhibitory avoidance, water maze, forced swim, and defensive freezing tasks, irrespective of age. Thus, age is associated with a decrease in 3alpha-diol production and 3alpha-diol administration reinstates cognitive and affective performance of aged male rats.
Streptococcus thermophilus is a major component of dairy starter cultures used for the manufacture of yoghurt and cheese. In this study, the CO(2) metabolism of S. thermophilus DSM 20617(T), grown in either a N(2) atmosphere or an enriched CO(2) atmosphere, was analysed using both genetic and proteomic approaches. Growth experiments performed in a chemically defined medium revealed that CO(2) depletion resulted in bacterial arginine, aspartate and uracil auxotrophy. Moreover, CO(2) depletion governed a significant change in cell morphology, and a high reduction in biomass production. A comparative proteomic analysis revealed that cells of S. thermophilus showed a different degree of energy status depending on the CO(2) availability. In agreement with proteomic data, cells grown under N(2) showed a significantly higher milk acidification rate compared with those grown in an enriched CO(2) atmosphere. Experiments carried out on S. thermophilus wild-type and its derivative mutant, which was inactivated in the phosphoenolpyruvate carboxylase and carbamoyl-phosphate synthase activities responsible for fixing CO(2) to organic molecules, suggested that the anaplerotic reactions governed by these enzymes have a central role in bacterial metabolism. Our results reveal the capnophilic nature of this micro-organism, underlining the essential role of CO(2) in S. thermophilus physiology, and suggesting potential applications in dairy fermentation processes.
Several lines of evidence suggest that the major inhibitory neuro-transmitter, gamma-aminobutyric acid (GABA) is involved, both directly and indirectly, in the pathogenesis of certain neurological and psychiatric disorders. The main enzyme responsible for GABA catabolism is gamma-aminobutyrate aminotransferase (GABA-T). Inhibition of this enzyme produces a considerable elevation of brain GABA concentrations, and such elevation has been correlated with many pharmacological effects. There seems to be that, as is discussed below, GABA-T activity in the brain and/or blood platelets is related to some neuro-psychiatric disorders such as alcoholism, epilepsy and Alzheimer's disease. GABA-T has been identified in the blood platelets with similar characteristics to those of brain GABA-T. In this way, studies on GABA-T activity in neuro-psychiatric disorders could be performed to understand, diagnosis and treat GABA-related disorders of the central nervous system (CNS).
In vivo intracellular recordings of spontaneous activity of neostriatal spiny cells revealed two-state behavior, i.e., characteristic shifts of membrane potential between two preferred levels. The more polarized level, called the Down state, varied among neurons from -61 to -94 mV. The more depolarized level, called the Up state, varied among neurons form -71 to -40 mV. For any one neuron, the membrane potential in the Up and Down states was constant over the period of observation (from 15 min to 4 hr), and the cells spent little time in transition between states. The level of membrane potential noise was higher in the Up state than in the Down state. Spontaneous membrane potential fluctuations were not abolished by experimental alteration of the membrane potential, but the time spent in each state was altered when intracellular current was used to vary the baseline membrane potential. Neither the sodium nor the calcium action potential that could be evoked by depolarization of spiny neurons was required for the occurrence of spontaneous shifts of membrane potential. Blockade of these action potentials using intracellular injection of QX314 and D890, respectively, altered neither the incidence of the membrane potential shifts nor the preferred membrane potential in either state. In contrast, antagonism of voltage-dependent potassium channels with intracellular cesium altered membrane potential shifts. In the presence of QX314 and D890, intracellular injection of cesium caused little or no change in the Down state and a large depolarizing shift in the Up state (to about -20 mV). Under these circumstances, the neuron responded to current in a nearly linear manner, and membrane conductance was found to be increased in the Up state, attributable to a membrane conductance with the same reversal potential as that of the synaptic potential evoked by cortical stimulation. These results indicate that the event underlying the Up state is a maintained barrage of synaptic excitation, but that the membrane potential achieved during the Up state in neostriatal spiny neurons is determined by dendritic potassium channels that clamp the membrane potential at a level determined by their voltage sensitivity. Neostriatal spiny neurons ordinarily receive enormously powerful excitation, which would drive the cells to saturation, and probably destroy them, if it were not for these potassium currents.
We measured the timing of spontaneous membrane potential fluctuations and action potentials of medial and lateral agranular corticostriatal and striatal neurons with the use of in vivo intracellular recordings in urethan-anesthetized rats. All neurons showed spontaneous subthreshold membrane potential shifts from 7 to 32 mV in amplitude, fluctuating between a hyperpolarized down state and depolarized up state. Action potentials arose only during the up state. The membrane potential state transitions showed a weak periodicity with a peak frequency near 1 Hz. The peak of the frequency spectra was broad in all neurons, indicating that the membrane potential fluctuations were not dominated by a single periodic function. At frequencies >1 Hz, the log of magnitude decreased linearly with the log of frequency in all neurons. No serial dependence was found for up and down state durations, or for the time between successive up or down state transitions, showing that the up and down state transitions are not due to superimposition of noisy inputs onto a single frequency. Monte Carlo simulations of stochastic synaptic inputs to a uniform finite cylinder showed that the Fourier spectra obtained for corticostriatal and striatal neurons are inconsistent with a Poisson-like synaptic input, demonstrating that the up state is not due to an increase in the strength of an unpatterned synaptic input. Frequency components arising from state transitions were separated from those arising from the smaller membrane potential fluctuations within each state. A larger proportion of the total signal was represented by the fluctuations within states, especially in the up state, than was predicted by the simulations. The individual state spectra did not correspond to those of random synaptic inputs, but reproduced the spectra of the up and down state transitions. This suggests that the process causing the state transitions and the process responsible for synaptic input may be the same. A high-frequency periodic component in the up states was found in the majority of the corticostriatal cells in the sample. The average size of the component was not different between neurons injected with QX-314 and control neurons. The high-frequency component was not seen in any of our sample of striatal cells. Corticostriatal and striatal neurons' coefficients of variation of interspike intervals ranged from 1.0 to 1.9. When interspike intervals including a down state were subtracted from the calculation, the coefficient of variation ranged from 0.4 to 1.1, indicating that a substantial proportion of spike interval variance was due to the subthreshold membrane potential fluctuations.
A novel subunit of the gamma-aminobutyrate, type A (GABAA) receptor family has been identified in human and rat tissues. The subunit displays 30-40% amino acid identity with known family members and represents a distinct subunit class (termed pi). Transcripts of the pi subunit were detected in several human tissues and were particularly abundant in the uterus. The pi subunit protein can assemble with known GABAA receptor subunits and confer unique ligand binding properties to the recombinant receptors in which it combines. Most notably, the presence of the pi subunit alters the sensitivity of recombinant receptors to the endogenous steroid, pregnanolone. Identification of the pi subunit indicates a new target for pharmacological manipulation of GABAA receptors that are located outside of the central nervous system.
Background: Delays between actions and their outcomes severely hinder reinforcement learning systems, but little is known of the neural mechanism by which animals overcome this problem and bridge such delays. The nucleus accumbens core ( AcbC), part of the ventral striatum, is required for normal preference for a large, delayed reward over a small, immediate reward ( self-controlled choice) in rats, but the reason for this is unclear. We investigated the role of the AcbC in learning a free-operant instrumental response using delayed reinforcement, performance of a previously-learned response for delayed reinforcement, and assessment of the relative magnitudes of two different rewards. Results: Groups of rats with excitotoxic or sham lesions of the AcbC acquired an instrumental response with different delays ( 0, 10, or 20 s) between the lever-press response and reinforcer delivery. A second ( inactive) lever was also present, but responding on it was never reinforced. As expected, the delays retarded learning in normal rats. AcbC lesions did not hinder learning in the absence of delays, but AcbC-lesioned rats were impaired in learning when there was a delay, relative to sham-operated controls. All groups eventually acquired the response and discriminated the active lever from the inactive lever to some degree. Rats were subsequently trained to discriminate reinforcers of different magnitudes. AcbC-lesioned rats were more sensitive to differences in reinforcer magnitude than sham-operated controls, suggesting that the deficit in self-controlled choice previously observed in such rats was a consequence of reduced preference for delayed rewards relative to immediate rewards, not of reduced preference for large rewards relative to small rewards. AcbC lesions also impaired the performance of a previously-learned instrumental response in a delay-dependent fashion. Conclusions: These results demonstrate that the AcbC contributes to instrumental learning and performance by bridging delays between subjects' actions and the ensuing outcomes that reinforce behaviour.
Steroids are important physiological orchestrators of endocrine as well as peripheral and central nervous system functions. One of the key processes for regulation of these molecules lies in their enzymatic processing by a family of 5α-reductase (5α-Rs) isozymes. By catalyzing a key rate-limiting step in steroidogenesis, this family of enzymes exerts a crucial role not only in the physiological control but also in pathological events. Indeed, both 5α-R inhibition and supplementation of 5α-reduced metabolites are currently used or have been proposed as therapeutic strategies for a wide array of pathological conditions. In particular, the potent 5α-R inhibitors finasteride and dutasteride are used in the treatments of benign prostatic hyperplasia (BPH), as well as in male pattern hair loss (MPHL) known as androgenetic alopecia (AGA). Recent preclinical and clinical findings indicate that 5α-R inhibitors evoke not only beneficial, but also adverse effects. Future studies should investigate the biochemical and physiological mechanisms that underlie the persistence of the adverse sexual side effects to determine why a subset of patients is afflicted with such persistence or irreversible adverse effects. Also a better focus of clinical research is urgently needed to better define those subjects who are likely to be adversely affected by such agents. Furthermore, research on the non-sexual adverse effects such as diabetes, psychosis, depression, and cognitive function are needed to better understand the broad spectrum of the effects these drugs may elicit during their use in treatment of AGA or BPH. In this review, we will summarize the state of art on this topic, overview the key unresolved questions that have emerged on the pharmacological targeting of these enzymes and their products, and highlight the need for further studies to ascertain the severity and duration of the adverse effects of 5α-R inhibitors, as well as their biological underpinnings.
The 5α-reductase inhibitor finasteride blocks the conversion of testosterone to dihydrotestosterone (DHT), the androgen responsible for male pattern hair loss (androgenetic alopecia) in genetically predisposed men.
Results of phase III clinical studies in 1879 men have shown that oral finasteride 1 mg/day promotes hair growth and prevents further hair loss in a significant proportion of men with male pattern hair loss. Evidence suggests that the improvement in hair count reported after 1 year is maintained during 2 years’ treatment. In men with vertex hair loss, global photographs showed improvement in hair growth in 48% of finasteride recipients at 1 year and in 66% at 2 years compared with 7% of placebo recipients at each time point. Furthermore, hair counts in these men showed that 83% of finasteride versus 28% of placebo recipients had no further hair loss compared with baseline after 2 years.
The clinical efficacy of oral finasteride has not yet been compared with that of topical minoxidil, the only other drug used clinically in patients with male pattern hair loss.
Therapeutic dosages of finasteride are generally well tolerated. In phase III studies, 7.7% of patients receiving finasteride 1 mg/day compared with 7.0% of those receiving placebo reported treatment-related adverse events. The overall incidence of sexual function disorders, comprising decreased libido, ejaculation disorder and erectile dysfunction, was significantly greater in finasteride than placebo recipients (3.8 vs 2.1%). All sexual adverse events were reversed on discontinuation of therapy and many resolved in patients who continued therapy. No other drug-related events were reported with an incidence ≥1% in patients receiving finasteride. Most events were of mild to moderate severity. Oral finasteride is contraindicated in pregnant women because of the risk of hypospadias in male fetuses.
Conclusions:Oral finasteride promotes scalp hair growth and prevents further hair loss in a significant proportion of men with male pattern hair loss. With its generally good tolerability profile, finasteride is a new approach to the management of this condition, for which treatment options are few. Its role relative to topical minoxidil has yet to be determined.
Pharmacodynamic Properties
Finasteride specifically inhibits the type II 5α-reductase enzyme which converts testosterone to dihydrotestosterone (DHT), the androgen responsible for the development of male pattern hair loss in genetically predisposed men. Oral finasteride 1 mg/day significantly reduced serum DHT levels by a median 68.4% in men with male pattern hair loss treated for 1 year. A corresponding 9.1% (median) increase in testosterone levels from baseline was reported, but these levels remained within the normal physiological range.
Oral finasteride 0.2 to 5 mg/day for 4 to 6 weeks reduced DHT levels by up to 65% in the scalp, the desired site of action of finasteride in men with male pattern hair loss.
Finasteride did not affect serum luteinising hormone or follicle-stimulating hormone responses to gonadotropin-releasing hormone stimulation in healthy volunteers, and therefore does not appear to influence the hypothalamic-pituitary-testicular axis. In addition, the drug did not alter serum prolactin, sex hormone-binding globulin, aldosterone or cortisol levels in healthy volunteers. Slight increases in serum estradiol levels were seen but, like testosterone levels, these remained within the normal physiological range and the ratio of testosterone to estradiol was unaltered.
Finasteride 1 mg/day does not appear to significantly affect ejaculate volume or other measures of testosterone-mediated semen production such as sperm motility, morphology and number. In addition, this dosage of finasteride had no clinically significant effects on prostate volume in healthy men aged ≤41 years, but caused a slight reduction in serum prostate-specific antigen levels. Evidence suggests that finasteride has no adverse effects on lipid or bone metabolism.
Pharmacokinetic Properties
Peak plasma concentrations of finasteride (9.2 µg/L) were reached 1 to 2 hours after drug administration in healthy volunteers who received a 1 mg/day dosage of finasteride for 17 days. Modest accumulation of finasteride in plasma was reported with repeated administration, but trough concentrations appeared to reach steady state within 3 days.
The oral bioavailability of finasteride (80%) is not affected by the presence of food.
Finasteride undergoes wide tissue distribution (volume of distribution = 76L), with ≈90% of circulating finasteride being protein bound. The drug has been detected in nanogram quantities in seminal fluid but these low levels have no clinical significance.
After oral administration, finasteride is extensively metabolised in the liver to compounds which are then eliminated in the bile (56.8%) and the urine (39.1%). Virtually no unchanged drug is recovered after an oral dose of finasteride. The mean terminal elimination half-life of finasteride 1 mg/day after repeated administration is 4.8 hours.
The elimination of finasteride is slower in elderly (≥70 years) than in younger (45 to 60 years) volunteers, but no dosage adjustment is warranted in the former age group, nor in patients with renal impairment. Reduced renal excretion of finasteride is compensated for by an increase in faecal elimination. There are currently no data on the pharmacokinetic properties of finasteride in patients with hepatic impairment.
Although finasteride is principally metabolised by cytochrome P450 3A4 enzymes within the liver, no clinically significant interactions have been reported between finasteride and digoxin, propranolol, aminophylline, warfarin, glibenclamide (glyburide) or antipyrine.
Therapeutic Efficacy
Oral finasteride 1 mg/day has shown efficacy in men with male pattern hair loss. The clinical use of this dosage of finasteride has been assessed in 3 phase III studies involving 1879 men with vertex or frontal hair loss who were treated for up to 2 years. Finasteride produced statistically significant increases in scalp hair growth from baseline within several months of starting treatment, a finding documented by hair count, patient self-assessment, investigator assessment and pre- and post-treatment clinical assessment based on standardised photos. Importantly, finasteride prevented the further hair loss seen in placebo recipients. In men with vertex hair loss, the mean improvement in hair count reported after 1 year with finasteride was maintained during a further 12 months’ treatment. Global photographs of the vertex area showed improvement in hair growth in 48% of finasteride recipients at 1 year (versus 7% with placebo) and in 66% at 2 years (versus 7% with placebo). Furthermore, vertex hair counts showed that 83% of finasteride versus 28% of placebo recipients had no further hair loss compared with baseline after 2 years. Hair growth was enhanced in patients who switched from placebo to finasteride after 12 months but declined progressively in those switched from finasteride to placebo.
Tolerability
Available data from 1879 patients with male pattern hair loss who received either finasteride 1 mg/day or placebo for 1 year in phase III studies show that finasteride is generally well tolerated. Overall, 7.7% of finasteride and 7.0% of placebo recipients reported mild to moderate treatment-related adverse events (1.4 and 1.6% withdrew). The only events reported more frequently in finasteride than placebo recipients were sexual disorders (3.8 vs 2.1%; p = 0.041), which comprised decreased libido (1.8 vs 1.3%), ejaculation disorders (1.2 vs 0.7%) and erectile dysfunction (1.3 vs 0.7%). These resolved in many men who reported them but remained on therapy and in all men who discontinued therapy because of these adverse events. No other drug-related events were reported with an incidence ≥1% in finasteride recipients.
The incidence of drug-related laboratory events was similar in finasteride and placebo groups (2.6 vs 2.4%). Finasteride had no significant effects on non-scalp body hair.
Dosage and Administration
Finasteride is indicated for the treatment of men with male pattern hair loss. The recommended dosage of finasteride in male pattern hair loss is 1 mg/day, taken with or without food. Daily treatment for 3 months or more is necessary before results are seen, and continued treatment is essential to sustain benefit. Furthermore, the effects of the drug are reversed within 12 months after treatment cessation.
There are no current data to support the use of finasteride in women with androgenetic alopecia. Moreover, pregnant women should not be directly exposed to finasteride by using or handling crushed tablets because of the risk of hypospadias developing in a male fetus.
Mycobacterium avium subsp. paratuberculosis (MAP) is the cause of a chronic enteritis of ruminants (bovine paratuberculosis-Johne disease) that is associated with enormous worldwide economic losses for the animal production industries. Diagnosis is based on observation of clinical signs, on the detection of antibodies in milk or serum or on evaluation of bacterial culture from feces. The limit of these methods is that they are not able to detect the disease in the subclinical stage and are applicable only when the disease is already in an advanced status. For this reason the main purpose of this study is to use the MAP proteome to detect novel immunoreactive proteins that may be helpful for paratuberculosis diagnoses. 2D electrophoresis and 2D immunoblotting of MAP proteins were performed using sera of control cattle and paratuberculosis infected cattle in order to highlight the specific immunoreactive proteins. Among the assigned identifiers to immunoreactive spots it was found that most of them correspond to surface-located proteins while three of them have never been described before as antigens. The identification of these proteins improves scientific knowledge that could be useful for paratuberculosis diagnoses. The sequence of the identified protein can be used for the synthesis of immunoreactive peptides that could be screened for their immunoreaction against bovine sera infected with MAP. All MS data have been deposited in the ProteomeXchange Consortium with identifier PXD001159 and DOI 10.6019/PXD001159.This article is protected by copyright. All rights reserved
Background: Androgenetic alopecia (male pattern hair loss) is caused by androgen-dependent miniaturization of scalp hair follicles, with scalp dihydrotestosterone (DHT) implicated as a contributing cause. Finasteride, an inhibitor of type II 5α-reductase, decreases serum and scalp DHT by inhibiting conversion of testosterone to DHT. Objective: Our purpose was to determine whether finasteride treatment leads to clinical improvement in men with male pattern hair loss. Methods: In two 1-year trials, 1553 men (18 to 41 years of age) with male pattern hair loss received oral finasteride 1 mg/d or placebo, and 1215 men continued in blinded extension studies for a second year. Efficacy was evaluated by scalp hair counts, patient and investigator assessments, and review of photographs by an expert panel. Results: Finasteride treatment improved scalp hair by all evaluation techniques at 1 and 2 years (P < .001 vs placebo, all comparisons). Clinically significant increases in hair count (baseline = 876 hairs), measured in a 1-inch diameter circular area (5.1 cm2 ) of balding vertex scalp, were observed with finasteride treatment (107 and 138 hairs vs placebo at 1 and 2 years, respectively; P < .001). Treatment with placebo resulted in progressive hair loss. Patients’ self-assessment demonstrated that finasteride treatment slowed hair loss, increased hair growth, and improved appearance of hair. These improvements were corroborated by investigator assessments and assessments of photographs. Adverse effects were minimal. Conclusion: In men with male pattern hair loss, finasteride 1 mg/d slowed the progression of hair loss and increased hair growth in clinical trials over 2 years. (J Am Acad Dermatol 1998;39:578-89.)
mRNA encoding the recently cloned g-aminobuytyric acidA receptor (GABAR) p subunit is expressed in the hippocampus and in several non-neuronal tissues including the uterus and ovaries. Whereas native GABARs are pentamers composed primarily of abg, abd ,o rabe subunits, it has not been demon- strated clearly that the p subunit incorporates into functional GABARs to form abp receptors and, if so, with what properties. We provide electrophysiological evidence that the p subunit can coassemble with either a5b 3o ra5b3g3 subunits to pro- duce recombinant GABARs with distinct pharmacological and biophysical properties. Compared with a5b3 receptors, GABARs produced by coexpression of a5b3p subunits had a lower GABA EC50 value, were enhanced to a lesser extent by loreclezole, had different IC50 values for pregnenolone sulfate and lanthanum, and were insensitive to benzodiazepines. In- corporation of both p and g3 subunits into an a5b3g3p isoform was suggested by reduced enhancement by diazepam and a high zinc IC50 value. Current-voltage relations for the a5b3p subunit combination outwardly rectified more than currents from a5b3g3 but less than a5b3 combination GABARs. Single- channel a5b3 GABAR currents had a main conductance state of 15.2 picoSeimens (pS). Coexpression of the p subunit with a5b3 subtypes increased the conductance level to 23.8 pS, similar to the conductance level of a5b3g3 GABARs (26.9 pS). We conclude that the p subunit coassembles with a, b, and g subunits to form functional abp or abgp GABARs and, thus, could have a significant impact on the function of native GA- BARs expressed in the brain or non-neuronal tissue.
In the last few decades a negative association between the level of milk production and fertility has been observed. Currently, the most utilized method of measuring male fertility employed by the livestock industry is related to the Non-Return Rate (NRR). Through differential proteome analysis, this study evaluated changes in the expression of the protein profile of spermatozoa collected from 16 bulls with different levels of field fertility expressed as an estimated relative conception rate (ERCR). The main aim is to identify putative protein markers to be used as putative indices of fertility. Two dimensional electrophoresis coupled with mass spectrometry analysis was used for protein separation and identification. To improve differential proteome analysis among experimental groups, a part of shotgun MS analysis was also performed. Three protein spots showed a differential expression pattern among all ERCR classes. Alpha enolase was significantly down-regulated in the ERCR- group, while two other proteins, isocitrate dehydrogenase and triosephosphate isomerase, were up-regulated in ERCR- in comparison to ERCR+. Alpha-enolase and isocitrate dehydrogenase subunit alpha (IDH-alpha) have been described in the literature for having a potential role in bull fertility. The possibility of determining protein biomarkers for fertility is more useful and less expensive than ERCR for acquiring rapid estimation of fertility because it does not require the use of field insemination trials. Shotgun MS analysis conducted on the same samples revealed 7 proteins down-regulated in the ERCR- group and 1 protein up-regulated. Among these proteins, calmodulin, ATP synthase mitochondrial subunits alpha and delta, malate dehydrogenase and sperm equatorial segment protein 1 were shown to be linked with sperm fertility.
Finasteride, a commonly prescribed medication for male pattern hair loss, has recently been associated with persistent sexual side effects. In addition, depression has recently been added to the product labeling of Propecia (finasteride 1 mg). Finasteride reduces the levels of several neuroactive steroids linked to sexual function and depression. This study assesses depressive symptoms and suicidal thoughts in former users of finasteride who developed persistent sexual side effects despite the discontinuation of finasteride.
In 2010-2011, former users of finasteride (n = 61) with persistent sexual side effects for ≥ 3 months were administered standardized interviews that gathered demographic information, medical and psychiatric histories, and information on medication use, sexual function, and alcohol consumption. All former users were otherwise healthy men with no baseline sexual dysfunction, chronic medical conditions, current or past psychiatric conditions, or use of oral prescription medications before or during finasteride use. A control group of men (n = 29), recruited from the community, had male pattern hair loss but had never used finasteride and denied any history of psychiatric conditions or use of psychiatric medications. The primary outcomes were the prevalence of depressive symptoms and the prevalence of suicidal thoughts as determined by the Beck Depression Inventory II (BDI-II); all subjects self-administered this questionnaire at the time of the interview or up to 10 months later.
Rates of depressive symptoms (BDI-II score ≥ 14) were significantly higher in the former finasteride users (75%; 46/61) as compared to the controls (10%; 3/29) (P < .0001). Moderate or severe depressive symptoms (BDI-II score ≥ 20) were present in 64% (39/61) of the finasteride group and 0% of the controls. Suicidal thoughts were present in 44% (27/61) of the former finasteride users and in 3% (1/29) of the controls (P < .0001).
Clinicians and potential users of finasteride should be aware of the potential risk of depressive symptoms and suicidal thoughts. The preliminary findings of this study warrant further research with controlled studies.
NMDA receptors are glutamatergic ionotropic receptors involved in excitatory neurotransmission, synaptic plasticity and excitotoxic cell death. Many allosteric modulators can influence the activity of these receptors positively or negatively, with behavioural consequences. 20-Oxo-5β-pregnan-3α-yl sulphate (pregnanolone sulphate; PA-6) is an endogenous neurosteroid that inhibits NMDA receptors and is neuroprotective. We tested the hypothesis that the interaction of PA-6 with the plasma membrane is critical for its inhibitory effect at NMDA receptors.
Electrophysiological recordings and live microscopy were performed on heterologous HEK293 cells expressing GluN1/GluN2B receptors and cultured rat hippocampal neurons.
Our experiments showed that the kinetics of the steroid inhibition were slow and not typical of drug-receptor interaction in an aqueous solution. In addition, the recovery from steroid inhibition was accelerated by β- and γ-cyclodextrin. Values of IC(50) assessed for novel synthetic C3 analogues of PA-6 differed by more than 30-fold and were positively correlated with the lipophilicity of the PA-6 analogues. Finally, the onset of inhibition induced by C3 analogues of PA-6 ranged from use-dependent to use-independent. The onset and offset of cell staining by fluorescent analogues of PA-6 were slower than those of steroid-induced inhibition of current responses mediated by NMDA receptors.
We conclude that steroid accumulation in the plasma membrane is the route by which it accesses a binding site on the NMDA receptor. Thus, our results provide a possible structural framework for pharmacologically targeting the transmembrane domains of the receptor.
Cogent evidence highlights a key role of neurosteroids and androgens in schizophrenia. We recently reported that inhibition of steroid 5α-reductase (5αR), the rate-limiting enzyme in neurosteroid synthesis and androgen metabolism, elicits antipsychotic-like effects in humans and animal models, without inducing extrapyramidal side effects. To elucidate the anatomical substrates mediating these effects, we investigated the contribution of peripheral and neural structures to the behavioral effects of the 5αR inhibitor finasteride (FIN) on the prepulse inhibition (PPI) of the acoustic startle reflex (ASR), a rat paradigm that dependably simulates the sensorimotor gating impairments observed in schizophrenia and other neuropsychiatric disorders. The potential effect of drug-induced ASR modifications on PPI was excluded by measuring this index both as percent (%PPI) and absolute values (ΔPPI). In both orchidectomized and sham-operated rats, FIN prevented the %PPI deficits induced by the dopamine (DA) receptor agonists apomorphine (APO, 0.25mg/kg, SC) and d-amphetamine (AMPH, 2.5mg/kg, SC), although the latter effect was not corroborated by ΔPPI analysis. Conversely, APO-induced PPI deficits were countered by FIN infusions in the brain ventricles (10μg/1μl) and in the nucleus accumbens (NAc) shell and core (0.5μg/0.5μl/side). No significant PPI-ameliorating effect was observed following FIN injections in other brain regions, including dorsal caudate, basolateral amygdala, ventral hippocampus and medial prefrontal cortex, although a statistical trend was observed for the latter region. The efflux of DA in NAc was increased by systemic, but not intracerebral FIN administration. Taken together, these findings suggest that the role of 5αR in gating regulation is based on post-synaptic mechanisms in the NAc, and is not directly related to alterations in DA efflux in this region.
Collapsin response mediator protein-2 (DPYSL2 or CRMP2) is a multifunctional adaptor protein within the central nervous system. In the developing brain or cell cultures, CRMP2 performs structural and regulatory functions related to cytoskeletal dynamics, vesicle trafficking and synaptic physiology whereas CRMP2 functions in adult brain are still being elucidated. CRMP2 has been associated with several neuropathologic or psychiatric conditions including Alzheimer's disease (AD) and schizophrenia, either at the level of genetic polymorphisms; protein expression; post-translational modifications; or protein/protein interactions. In AD, CRMP2 is phosphorylated by glycogen synthase kinase-3β (GSK3β) and cyclin dependent protein kinase-5 (CDK5), the same kinases that act on tau protein in generating neurofibrillary tangles (NFTs). Phosphorylated CRMP2 collects in NFTs in association with the synaptic structure-regulating SRA1/WAVE1 (specifically Rac1-associated protein-1/WASP family verprolin-homologous protein-1) complex. This phenomenon could plausibly contribute to deficits in neural and synaptic structure that have been well documented in AD. This review discusses the essential biology of CRMP2 in the context of nascent data implicating CRMP2 perturbations as either a correlate of, or plausible contributor to, diverse neuropathologies. A discussion is made of recent findings that the atypical antidepressant tianeptine increases CRMP2 expression, whereas other, neuroactive small molecules including the epilepsy drug lacosamide and the natural brain metabolite lanthionine ketimine appear to bind CRMP2 directly with concomitant affects on neural structure. These findings constitute proofs-of-concept that pharmacological manipulation of CRMP2 is possible and hence, may offer new opportunities for therapy development against certain neurological diseases.
5α-reductase inhibitors (5α-RIs), finasteride and dutasteride, have been approved for treatment of lower urinary tract symptoms, due to benign prostatic hyperplasia, with marked clinical efficacy. Finasteride is also approved for treatment of hair loss (androgenetic alopecia). Although the adverse side effects of these agents are thought to be minimal, the magnitude of adverse effects on sexual function, gynecomastia, depression, and quality of life remains ill-defined.
The goal of this review is to discuss 5α-RIs therapy, the potential persistent side effects, and the possible mechanisms responsible for these undesirable effects.
We examined data reported in various clinical studies from the available literature concerning the side effects of finasteride and dutasteride.
Data reported in the literature were reviewed and discussed. Results. Prolonged adverse effects on sexual function such as erectile dysfunction and diminished libido are reported by a subset of men, raising the possibility of a causal relationship.
We suggest discussion with patients on the potential sexual side effects of 5α-RIs before commencing therapy. Alternative therapies may be considered in the discussion, especially when treating androgenetic alopecia.
This study examined the allosteric modulation of the NMDA receptor by nanomolar concentrations of neurosteroids in rats treated long term with morphine. The neurosteroids dehydroepiandrosterone sulfate (DHEAS), pregnenolone sulfate (PS) and pregnanolone sulfate (3α5βS) are important mediators in the central nervous system. They induce rapid responses by non-classical steroidal mechanisms, e.g. via interaction with the N-methyl-D-aspartate (NMDA) receptor, and are known to modify the binding of ifenprodil to the NMDA receptor subunit NR2B. The NMDA receptor is involved in several processes, including memory, learning, synaptic plasticity and neuronal development. Morphine, a μ-opioid receptor agonist, has an important role in the clinical treatment of pain. The main drawback of morphine treatment is the associated development of dependence and tolerance. The mechanisms behind these phenomena are still to be elucidated, but several reports suggest the involvement of the NMDA receptor. The results of the present study indicate that the allosteric modulation induced by the neurosteroids DHEAS, PS and 3α5βS was similar in all tested brain regions. This suggests that the NR2B receptor subunit behaves independently of its site of expression. Moreover, the NR2B subunit was up-regulated in the frontal cortex but not in the hippocampus or hypothalamus. It is concluded that morphine does not affect the neurosteroid modulatory effect on ifenprodil binding in the rat hippocampus or hypothalamus but does significantly affect both the expression of the NR2B subunit and the 3α5βS modulatory effect on ifenprodil binding in the frontal cortex. It is suggested that the observed effect of long term morphine on the properties of NR2B in the frontal cortex may be associated with the mechanism underlying the development of opiate dependence.
As neuroactive steroids modulate several ionotropic receptors, we assessed whether the ATP-gated currents elicited by P2X(4) receptors are modulated by these compounds. We transfected HEK293 cells or injected Xenopus laevis oocytes with the cDNA coding for rat P2X(4) receptor. Application of 0.1-10 microM alfaxolone potentiated within 60-s the 1 microM ATP-evoked currents with a maximal potentiation of 1.8 and 2.6-fold in HEK293 or oocytes cells respectively. Allopregnalolone or 3alpha, 21-dihydroxy-5alpha-pregnan-20-one (THDOC) also potentiated the ATP-gated currents but with a maximal effect only averaging 1.25 and 1.35-fold respectively. In contrast, 0.3-10 microM pregnanolone, but not its sulfated derivative, inhibited the ATP-gated currents; the maximal inhibition reached 40% in both cell types. THDOC, but not other neurosteroids increased significantly the tau(off) of the ATP-evoked currents, revealing another mode of neurosteroid modulation. Sexual steroids such as 17beta-estradiol or progesterone were inactive revealing explicit structural requirements. Alfaxolone or THDOC at concentrations 30- to 100-fold larger than required to modulate the receptor, gated the P2X(4) receptor eliciting ATP-like currents that were reduced with suramin or brilliant blue G, but potentiated the P2X(4) receptor more than 10-fold by 10 microM zinc. In conclusion, neurosteroids rapidly modulate via non-genomic mechanisms and with nanomolar potencies, the P2X4 receptor interacting likely at distinct modulator sites.
The isolation-rearing (IR) paradigm, consisting of the social deprivation for 6-9 weeks after weaning, induces a spectrum of aberrant behaviors in adult rats. Some of these alterations such as sensorimotor gating deficits are reminiscent of the dysfunctions observed in schizophrenia patients. Although gating impairments in IR rats have been linked to impairments in the cortico-mesolimbic system, the specific molecular mechanisms underlying this relation are unclear. To elucidate the neurochemical modifications underlying the gating disturbances exhibited by IR rats, we compared their pre-pulse inhibition (PPI) of the acoustic startle reflex with that of socially reared (SR) controls, and correlated this index to the results of proteomic analyses in prefrontal cortex and nucleus accumbens from both groups. As expected, IR rats exhibited significantly lower startle amplitude and PPI than their SR counterparts. Following behavioral testing, IR and SR rats were killed and protein expression profiles of their brain regions were examined using two-dimensional electrophoresis based proteomics. Image analysis in the Coomassie blue-stained gel revealed that three protein spots were differentially expressed in the nucleus accumbens of IR and SR rats. Mass spectrometry (matrix-assisted laser desorption ionization-time of flight and MS/MS) identified these spots as heat shock protein 60 (HSP60), alpha-synuclein (alpha-syn), and 14-3-3 protein zeta/delta. While accumbal levels of HSP60 was decreased in IR rats, alpha-syn and 14-3-3 proteins were significantly increased in IR in comparison with SR controls. Notably, these two last alterations were significantly correlated with different loudness intensity-specific PPI deficits in IR rats. In view of the role of these proteins in synaptic trafficking and dopaminergic regulation, these findings might provide a neurochemical foundation for the gating alterations and psychotic-like behaviors in IR rats.
Estrogens have cell-specific effects on a variety of physiological endpoints including regulation of mitochondrial biogenesis and activity. Estrogens regulate gene transcription by the classical genomic mechanism of binding to estrogen receptors alpha and beta (ERalpha and ERbeta) as well as the more recently described nongenomic pathways involving plasma membrane-associated ERs that activate intracellular protein kinase-mediated phosphorylation signaling cascades. Here I will review the rapid and longer-term effects of estrogen on mitochondrial function. The identification of ERalpha and ERbeta within mitochondria of various cells and tissues is discussed with a model of estrogen regulation of the transcription of nuclear respiratory factor-1 (NRF-1, NRF1). NRF-1 subsequently promotes transcription of mitochondrial transcription factor Tfam (mtDNA maintenance factor, also called mtTFA) and then Tfam targets mtDNA-encoded genes. The nuclear effects of estrogens on gene expression directly controlling mitochondrial biogenesis, oxygen consumption, mtDNA transcription, and apoptosis are reviewed. Overall, we are just beginning to evaluate the many direct and indirect effects of estrogens on mitochondrial activities.
1. Intracellular recordings were made from neurons in slices cut from the rat nucleus accumbens septi. Membrane currents were measured with a single-electrode voltage-clamp amplifier in the potential range -50 to -140 mV. 2. In control conditions (2.5 mM potassium), the resting membrane potential of the neurons was -83.4 +/- 1.1 (SE) mV (n = 157). Steady state membrane conductance was voltage dependent, being 34.8 +/- 1.7 nS (n = 25) at -100 mV and 8.0 +/- 0.7 nS (n = 25) at -60 mV. 3. Barium (1 microM) markedly reduced the inward rectification and caused a small inward current (40.6 +/- 8.7 pA, n = 8) at the resting potential. These effects became larger with higher barium concentrations, and, in 100 microM barium, the current-voltage relation was straight. 4. The block of the inward current by barium (at -130 mV) occurred with an exponential time course; the time constant was approximately 1 s at 1 microM barium and less than 90 ms with 100 microM. Strontium had effects similar to those of barium, but 1000-fold higher concentrations were required. Cesium chloride (2 mM) and rubidium chloride (2 mM) also blocked the inward rectification; their action reached steady state within 50 ms. 5. It is concluded that the nucleus accumbens neurons have a potassium conductance with many features of a typical inward rectifier and that this contributes to the potassium conductance at the resting potential.
4-Aminobutyric acid (GABA), a major inhibitory neurotransmitter of mammalian central nervous system, is found in a wide range of organisms, from prokaryotes to vertebrates. GABA is widely distributed in nonneural tissue including peripheral nervous and endocrine systems. GABA acts on GABAA and GABAB receptors. GABAA receptors are ligand-gated chloride channels modulated by a variety of drugs. GABAB receptors are essentially presynaptic, usually coupled to potassium or calcium channels, and they function via a GTP binding protein. In neural and nonneural tissues, GABA is metabolized by three enzymes--glutamic acid decarboxylase (GAD), which produces GABA from glutamic acid, and the catabolic enzymes GABA-transaminase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH). Production of succinic acid by SSADH allows entry of the GABA carbon skeleton into the tricarboxylic acid cycle. Alternate sources of GABA include putrescine, spermine, spermidine and ornithine, which produce GABA via deamination and decarboxylation reactions, while L-glutamine is an additional source of glutamic acid via deamination. GAD from mammalian brain occurs in two molecular forms, GAD65 and GAD67 (referring to subunit relative molecular weight (Mr) in kilodaltons). These different forms of GAD are the product of different genes, differing in nucleotide sequence, immunoreactivity and subcellular localization. The presence and characteristics of GAD have been investigated in a wide variety of nonneural tissues including liver, kidney, pancreas, testis, ova, oviduct, adrenal, sympathetic ganglia, gastrointestinal tract and circulating erythrocytes. In some tissues, one form (GAD65 or GAD67) predominates. GABA-T has been located in most of the same tissues, primarily through histochemical and/or immunochemical methods; GABA-T is also present in a variety of circulating cells, including platelets and lymphocytes. SSADH, the final enzyme GABA catabolism, has been detected in some of the tissues in which GAD and GABA-T have been identified, although the presence of this enzyme has not been in mammalian pancreas, ova, oviduct, testis or sympathetic ganglia.
Sex hormones are known to exert direct and indirect effects on cardiovascular function, but their effects on cardiac repolarization have not been elucidated. The repolarization phase of the cardiac action potential or QT interval of the ECG is regulated largely by potassium channels such as the delayed rectifier currents HK2 and IsK.
The effects of ovariectomy (OVX) and estradiol (E2) or dihydrotestosterone (DHT) treatment were evaluated on HK2, HERG, and IsK mRNA levels, QT duration, and quinidine-induced changes in QT interval in isolated rabbit hearts. HK2 and 0.7-kilobase IsK mRNA were downregulated in cardiac ventricular tissue from OVX rabbits treated with either E2 or DHT. The QT interval was prolonged in E2- and DHT-treated animals (OVX + vehicle, 223 +/- 6 ms; OVX + DHT, 236 +/- 10 ms; and OVX + DHT, 245 +/- 6 ms; P < .05).
The association between hormone-induced changes in baseline QT interval and the mRNA level for these channels suggests that sex hormones may play a critical role in regulating cardiac repolarization. However, the changes in baseline QT and potassium channel mRNA after hormone treatment were not concordant with the changes in QT interval after the infusion of quinidine, after which E2-treated animals responded similarly to controls (18.4 +/- 4.6% and 19.3 +/- 4.6% increase in QT interval, respectively) and DHT-treated animals exhibited less QT prolongation (11.4 +/- 3.8% increase; P < .03).
The rat CYP2B gene subfamily includes CYP2B1, CYP2B2 and CYP2B3. Translation of an alternatively spliced hepatic CYP2B2 mRNA would generate a CYP2B2 variant, CYP2B2v, having eight additional amino acid residues inserted between CYP2B2 positions 274 and 275. The presence of CYP2B3 and CYP2B2v in rat liver has yet to be demonstrated. cDNA expression vectors were obtained for CYP2B1, CYP2B2, CYP2B3 and CYP2B2v. All four proteins react with an anti-CYP2B1 antibody and can be resolved by SDS-PAGE. A CYP2B3-specific polyclonal antibody raised against an undecapeptide (SPVDPNTIDMT) from near the C-terminus of CYP2B3 detected a constitutive protein on immunoblots of rat liver microsomes, thus demonstrating that the CYP2B3 mRNA is translated in the liver. Similarly, a CYP2B2v-specific polyclonal antibody was raised against a peptide containing the eight additional amino acid residues (VSPAWMRE) predicted to be present in the CYP2B2v protein. It detected a phenobarbital- and Aroclor 1254-inducible protein in rat liver microsomes. Microsomes of Ad293 cells expressing cDNAs for CYP2B2 and CYP2B2v were used to metabolize 7,12-dimethylbenz[a]anthracene (DMBA), and the metabolites produced were compared with those generated by microsomes of cells expressing CYP2B1 cDNA. CYP2B2v had activity similar to that of CYP2B2 for DMBA metabolism. Both CYP2B2 forms preferentially catalyzed 12-hydroxylation, whereas CYP2B1 preferred 7-hydroxylation and exhibited turnover that was strongly suppressed as previously reported. These results demonstrate the existence in rat liver of two new CYP2B proteins: CYP2B3, the major constitutive CYP2B form, and CYP2B2v, which represents a rare case of non-aberrant alternative splicing among xenobiotic-metabolizing P450s.
Previous studies in this laboratory have determined the lack of specificity of several antibody and substrate probes of CYP2B6. The goals of the current study were to examine the expression of CYP2B6 in a bank of human liver microsome (HLM) samples using a new specific monoclonal antibody (MAb 49-10-20) and to further characterize the substrate specificity of CYP2B6. A 100-fold variability in expression of immunodetectable CYP2B6 was demonstrated in a bank of 19 HLM samples (0.7 pmol/mg protein to 71. 1 pmol/mg protein) using MAb 49-10-20. CYP2B6 levels were found to significantly (P < .0001) correlate with S-mephenytoin N-demethylation to nirvanol (r2 = 0.89), 7-hydroxy-4-trifluoromethylcoumarin formation (r2 = 0.81) and several markers of CYP3A levels and activity. The relationships between nirvanol formation and CYP3A levels or activity were found to depend on two HLM samples. Km (apparent) values were generated for benzyloxyresorufin O-deethylation (1.3 microM), benzphetamine N-demethylation (93.4 microM), 3-cyano 7-ethoxycoumarin O-deethylation (71.3 microM), midazolam 1'-hydroxylation (46.1 microM) and 4-chloromethyl-7-ethoxycoumarin O-deethylation (33.7 microM) using expressed CYP2B6. Testosterone 16beta-hydroxylation by expressed CYP2B6 resulted in atypical kinetics characteristic of substrate activation. The data best fit the Hill equation with a Km (apparent) of 50.5 microM and an n of 1.3 (n = number of sites bound by activator). In conclusion, the highly specific MAb 49-10-20 was used to provide further confirmation that S-mephenytoin N-demethylation to nirvanol is a CYP2B6 selective probe. Finally, some, but not all substrates of CYP2B6 demonstrate autoactivation.
Androgenetic alopecia (male pattern hair loss) is caused by androgen-dependent miniaturization of scalp hair follicles, with scalp dihydrotestosterone (DHT) implicated as a contributing cause. Finasteride, an inhibitor of type II 5alpha-reductase, decreases serum and scalp DHT by inhibiting conversion of testosterone to DHT.
Our purpose was to determine whether finasteride treatment leads to clinical improvement in men with male pattern hair loss.
In two 1-year trials, 1553 men (18 to 41 years of age) with male pattern hair loss received oral finasteride 1 mg/d or placebo, and 1215 men continued in blinded extension studies for a second year. Efficacy was evaluated by scalp hair counts, patient and investigator assessments, and review of photographs by an expert panel.
Finasteride treatment improved scalp hair by all evaluation techniques at 1 and 2 years (P < .001 vs placebo, all comparisons). Clinically significant increases in hair count (baseline = 876 hairs), measured in a 1-inch diameter circular area (5.1 cm2) of balding vertex scalp, were observed with finasteride treatment (107 and 138 hairs vs placebo at 1 and 2 years, respectively; P < .001). Treatment with placebo resulted in progressive hair loss. Patients' self-assessment demonstrated that finasteride treatment slowed hair loss, increased hair growth, and improved appearance of hair. These improvements were corroborated by investigator assessments and assessments of photographs. Adverse effects were minimal.
In men with male pattern hair loss, finasteride 1 mg/d slowed the progression of hair loss and increased hair growth in clinical trials over 2 years.
Several experimental approaches have been applied to examine the significance of the available pool of 5-phosphoribosyl-1-pyrophosphate (PRPP) for purine and pyrimidine nucleotide metabolism in the rat heart. In a series of studies including some pentoses and pentitols, in particular ribose, it was shown that these sugars were all capable of elevating the cardiac PRPP pool and stimulating the rate of adenine nucleotide biosynthesis. In several pathophysiological situations that were characterized by a decrease in ATP content, the increase in adenine nucleotide biosynthesis elicited by ribose was of such magnitude that the ATP level was replenished partially or completely in a considerably shorter period of time than that without any intervention. In two experimental models, in cardiac hypertrophy induced by aortic constriction with additional isoproterenol administration and in the noninfarcted rat heart after permanent coronary artery ligation, there was also an improvement in global heart function under the influence of ribose. The myocardial cell damage induced by isoproterenol was prevented by ribose. Combination of ribose with adenine or inosine led to an even quicker ATP normalization in the isoproterenol-stimulated rat heart than with either intervention alone. Ribose had no functional effects on the cardiovascular system, whereas adenine, inosine, and orotic acid were demonstrated to have different hemodynamic influences. Adenine and inosine had negative chronotropic and inotropic effects in the intact rat, whereas orotic acid had a positive influence both on the left and right rat heart. On the basis of these experimental studies, a new therapeutic strategy is suggested in which elevation of the available PRPP plays a key role. Once this has been elevated by ribose, additional substrates, such as adenine, inosine, and orotic acids, should be included. This would exploit the full potential of a therapeutic approach that corrects a natural metabolic deficiency of the heart that is the low capacity of the oxidative pentose phosphate pathway in which PRPP is generated.
Two voltage-gated potassium channels, hKv1.1 and hKv2.1, were stably expressed in Chinese hamster ovary cells. The effects of neurosteroids, pregnenolone sulfate (PS), 5alpha-pregnan-3alpha-ol-20-one sulfate (5alpha3alphaP), and 5beta-pregnan-3alpha-ol-20-one sulfate (5beta3alphaP), on hKv1.1 and hKv2.1 were studied using the patch-clamp method. PS, 5alpha3alphaP, and 5beta3alphaP shifted current/voltage (I/V) relationship of both channels to the left by about 10 mV. The neurosteroids also increased the current amplitude voltage dependently. The activation time constants (taua) for hKv1.1 and hKv2.1 were voltage dependent and decreased with membrane depolarization. At -20 mV, PS reduced taua for hKv1.1 by 8 ms and 15 ms at 10 and 100 microM respectively. PS also significantly decreased the taua of hKv2.1 by 30 ms (10 microM) and 38 ms (100 microM) at +10 mV. However, only at 100 microM did 5alpha3alphaP or 5beta3alphaP significantly alter the taua of hKv1.1. Neither 5alpha3alphaP nor 5beta3alphaP affected the taua of hKv2.1. Deactivation time constants (taud) were also voltage dependent and, in contrast to taua, taud was prolonged with membrane depolarization. At 100 microM, PS, 5alpha3alphaP, and 5beta3alphaP significantly increased the taud for hKv1.1 but did not affect the taud for hKv2.1.
Unlabelled:
The 5alpha-reductase inhibitor finasteride blocks the conversion of testosterone to dihydrotestosterone (DHT), the androgen responsible for male pattern hair loss (androgenetic alopecia) in genetically predisposed men. Results of phase III clinical studies in 1879 men have shown that oral finasteride 1 mg/day promotes hair growth and prevents further hair loss in a significant proportion of men with male pattern hair loss. Evidence suggests that the improvement in hair count reported after 1 year is maintained during 2 years' treatment. In men with vertex hair loss, global photographs showed improvement in hair growth in 48% of finasteride recipients at 1 year and in 66% at 2 years compared with 7% of placebo recipients at each time point. Furthermore, hair counts in these men showed that 83% of finasteride versus 28% of placebo recipients had no further hair loss compared with baseline after 2 years. The clinical efficacy of oral finasteride has not yet been compared with that of topical minoxidil, the only other drug used clinically in patients with male pattern hair loss. Therapeutic dosages of finasteride are generally well tolerated. In phase III studies, 7.7% of patients receiving finasteride 1 mg/day compared with 7.0% of those receiving placebo reported treatment-related adverse events. The overall incidence of sexual function disorders, comprising decreased libido, ejaculation disorder and erectile dysfunction, was significantly greater in finasteride than placebo recipients (3.8 vs 2.1%). All sexual adverse events were reversed on discontinuation of therapy and many resolved in patients who continued therapy. No other drug-related events were reported with an incidence > or =1% in patients receiving finasteride. Most events were of mild to moderate severity. Oral finasteride is contraindicated in pregnant women because of the risk of hypospadias in male fetuses.
Conclusions:
Oral finasteride promotes scalp hair growth and prevents further hair loss in a significant proportion of men with male pattern hair loss. With its generally good tolerability profile, finasteride is a new approach to the management of this condition, for which treatment options are few. Its role relative to topical minoxidil has yet to be determined.
Four mammalian Kv3 genes have been identified, each of which generates, by alternative splicing, multiple protein products differing in their C-terminal sequence. Products of the Kv3.1 and Kv3.2 genes express similar delayed-rectifier type currents in heterologous expression systems, while Kv3.3 and Kv3.4 proteins express A-type currents. All Kv3 currents activate relatively fast at voltages more positive than -10 mV, and deactivate very fast. The distribution of Kv3 mRNAs in the rodent CNS was studied by in situ hybridization, and the localization of Kv3.1 and Kv3.2 proteins has been studied by immunohistochemistry. Most Kv3.2 mRNAs (approximately 90%) are present in thalamic-relay neurons throughout the dorsal thalamus. The protein is expressed mainly in the axons and terminals of these neurons. Kv3.2 channels are thought to be important for thalamocortical signal transmission. Kv3.1 and Kv3.2 proteins are coexpressed in some neuronal populations such as in fast-spiking interneurons of the cortex and hippocampus, and neurons in the globus pallidus. Coprecipitation studies suggest that in these cells the two types of protein form heteromeric channels. Kv3 proteins appear to mediate, in native neurons, similar currents to those seen in heterologous expression systems. The activation voltage and fast deactivation rates are believed to allow these channels to help repolarize action potentials fast without affecting the threshold for action potential generation. The fast deactivating current generates a quickly recovering after hyperpolarization, thus maximizing the rate of recovery of Na+ channel inactivation without contributing to an increase in the duration of the refractory period. These properties are believed to contribute to the ability of neurons to fire at high frequencies and to help regulate the fidelity of synaptic transmission. Experimental evidence has now become available showing that Kv3.1-Kv3.2 channels play critical roles in the generation of fast-spiking properties in cortical GABAergic interneurons.