Effect of the ROCK inhibitor fasudil on the brain proteomic profile in the tau transgenic mouse model of Alzheimer's disease.

Introduction: The goal of this study is to explore the pharmacological potential of the amyloid-reducing vasodilator fasudil, a selective Ras homolog (Rho)-associated kinases (ROCK) inhibitor, in the P301S tau transgenic mouse model (Line PS19) of neurodegenerative tauopathy and Alzheimer's disease (AD). Methods: We used LC-MS/MS, ELISA and bioinformatic approaches to investigate the effect of treatment with fasudil on the brain proteomic profile in PS19 tau transgenic mice. We also explored the efficacy of fasudil in reducing tau phosphorylation, and the potential beneficial and/or toxic effects of its administration in mice. Results: Proteomic profiling of mice brains exposed to fasudil revealed the activation of the mitochondrial tricarboxylic acid (TCA) cycle and blood-brain barrier (BBB) gap junction metabolic pathways. We also observed a significant negative correlation between the brain levels of phosphorylated tau (pTau) at residue 396 and both fasudil and its metabolite hydroxyfasudil. Conclusions: Our results provide evidence on the activation of proteins and pathways related to mitochondria and BBB functions by fasudil treatment and support its further development and therapeutic potential for AD.

Symptomatic remission and recovery in major psychosis: Is there a role for BDNF? A secondary analysis of the LABSP cohort data.

Remission, relapse prevention, and clinical recovery are crucial areas of interest in schizophrenia (SCZ) research. Although SCZ is a chronic disorder with poor overall outcomes, years of research demonstrated that recovery is possible. There are considerable data linking brain-derived neurotrophic factor (BDNF) to SCZ, however, evidence on the role of BDNF in remission in SCZ is scarce. This secondary analysis of the Longitudinal Assessment of BDNF in Sardinian patients (LABSP) data aimed to investigate the relationship between serum BDNF levels and symptomatic remission, simultaneous clinical and functional remission, and recovery in patients with SCZ. A total of 105 patients with SCZ or schizoaffective disorder were recruited for a longitudinal assessment of BDNF levels over 24 months. Longitudinal data were analyzed using mixed-effects linear regression models. The study found significant associations between use of long acting injectables (χ2 = 7.075, df = 1, p = 0.008), baseline serum BDNF levels (U = 701, z = -2.543, p = 0.011), and "childhood" (U = 475, z = -2.124, p = 0.034) and "general" (U = 55, z = -2.014, p = 0.044) subscales of the Premorbid Adjustment Scale (PAS) with patients maintaining remission and recovery. The diagnosis of SCZ was significantly associated with lower BDNF levels for patients with simultaneous clinical and functional remission (Z = 2.035, p = 0.0419) and recovery (Z = 2.009, p = 0.0445) compared to those without. There were no significant associations between remission in the entire sample and longitudinal serum BDNF levels or genetic variants within the BDNF gene. These findings provide further insight into the complex relationship between BDNF and SCZ.

Reduction of Phosphorylated Tau in Alzheimer's Disease Induced Pluripotent Stem Cell-Derived Neuro-Spheroids by Rho-Associated Coiled-Coil Kinase Inhibitor Fasudil.

BACKGROUND: Alzheimer's disease (AD) is the most predominant form of dementia. Rho-associated coiled coil kinase (ROCK) inhibitor, fasudil, is one of the candidate drugs against the AD progression. OBJECTIVE: We aimed to investigate possible changes of AD associated markers in three-dimensional neuro-spheroids (3D neuro-spheroids) generated from induced pluripotent stem cells derived from AD patients or healthy control subjects (HC) and to determine the impact of pharmacological intervention with the ROCK inhibitor fasudil. METHODS: We treated 3D neuro-spheroids with fasudil and tested the possible effect on AD markers by ELISA, transcriptomic and proteomic analyses. RESULTS: Transcriptomic analysis revealed a reduction in the expression of AKT serine/threonine-protein kinase 1 (AKT1) in AD neuro-spheroids, compared to HC. This decrease was reverted in the presence of fasudil. Proteomic analysis showed up- and down-regulation of proteins related to AKT pathway in fasudil-treated neuro-spheroids. We found an evident increase of phosphorylated tau at four different residues (pTau181, 202, 231, and 396) in AD compared to HC-derived neuro-spheroids. This was accompanied by a decrease of secreted clusterin (clu) and an increase of intracellular clu levels in AD patient-derived neuro-spheroids. Increases of phosphorylated tau in AD patient-derived neuro-spheroids were suppressed in the presence of fasudil. CONCLUSIONS: Fasudil modulates clu protein levels and enhances AKT1 that results in the suppression of AD associated tau phosphorylation.

Angiotensin-converting enzyme inhibitors and statins therapies-induced changes in omics profiles in humans and transgenic tau mice.

BACKGROUND: Hypertension and hyperlipidemia are considered risk factors for Alzheimer's disease (AD) and other related dementias. Clinically approved medications typically prescribed to manage these conditions have shown an association with reduced risk of developing AD and could be explored as potential repurposed therapeutics. OBJECTIVE: We aimed to explore the effects of the pharmacological treatment with angiotensin-converting enzyme inhibitors (ACEI) and statins (STAT) on AD-related neuropathology and the potential benefits of their concurrent use. METHODS: We investigated the effect of ACEI, STAT or combination of both by exploring the transcriptomic, proteomic and tau pathology profiles after treatment in both human patients and in P301S transgenic mice (PS19) modeling tauopathies and AD. We performed bioinformatic analysis of enriched pathways after treatment. RESULTS: Proteomics and transcriptomics analysis revealed proteins and genes whose expression is significantly changed in subjects receiving treatment with ACEI, STAT or combined drugs. In mice, treatment with the ACEI lisinopril significantly decreased brain levels of total tau (Tau) and phosphorylated tau (pTau)-181, while the STAT atorvastatin significantly reduced the levels of pTau-396. The combined therapy with lisinopril and atorvastatin significantly decreased Tau. Moreover, brain levels of lisinopril were negatively correlated with Tau. Among the others, CD200, ADAM22, BCAN and NCAM1 were significantly affected by treatments in both human subjects and transgenic mice. CONCLUSIONS: Our findings provide significant information that may guide future investigation of the potential use of ACEI, STAT, or the combination of the two drug classes as repurposed therapies or preventive strategies for AD and other neurodegenerative diseases.

Food restriction and hyperactivity induce changes in corticolimbic brain dopamine and serotonin levels in female rats.

Compelling data support altered dopamine (DA) and serotonin (5-HT) signaling in anorexia nervosa (AN). However, their exact role in the etiopathogenesis of AN has yet to be elucidated. Here, we evaluated the corticolimbic brain levels of DA and 5-HT in the induction and recovery phases of the activity-based anorexia (ABA) model of AN. We exposed female rats to the ABA paradigm and measured the levels of DA, 5-HT, the metabolites 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxyindoleacetic acid (5-HIAA), and the dopaminergic type 2 (D2) receptors density in feeding- and reward-implicated brain regions (i.e., cerebral cortex, Cx; prefrontal cortex, PFC; caudate putamen, CPu; nucleus accumbens, NAcc; amygdala, Amy; hypothalamus, Hyp; hippocampus, Hipp). DA levels were significantly increased in the Cx, PFC and NAcc, while 5-HT was significantly enhanced in the NAcc and Hipp of ABA rats. Following recovery, DA was still elevated in the NAcc, while 5-HT was increased in the Hyp of recovered ABA rats. DA and 5-HT turnover were impaired at both ABA induction and recovery. D2 receptors density was increased in the NAcc shell. These results provide further proof of the impairment of the dopaminergic and serotoninergic systems in the brain of ABA rats and support the knowledge of the involvement of these two important neurotransmitter systems in the development and progression of AN. Thus, providing new insights on the corticolimbic regions involved in the monoamine dysregulations in the ABA model of AN.

Converging Evidence Points to BDNF as Biomarker of Depressive Symptoms in Schizophrenia-Spectrum Disorders.

Brain-derived neurotrophic factor (BDNF) is a key modulator of neuroplasticity and has an important role in determining the susceptibility to severe psychiatric disorder with a significant neurodevelopmental component such as major psychoses. Indeed, a potential association between BDNF serum levels and schizophrenia (SCZ) and schizoaffective disorder (SAD) has been tested in diverse studies and a considerable amount of them found reduced BDNF levels in these disorders. Here, we aimed at testing the association of BDNF serum levels with several demographic, clinical, and psychometric measures in 105 patients with SCZ and SAD, assessing the moderating effect of genetic variants within the BDNF gene. We also verified whether peripheral BDNF levels differed between patients with SCZ and SAD. Our findings revealed that BDNF serum levels are significantly lower in patients affected by SCZ and SAD presenting more severe depressive symptomatology. This finding awaits replication in future independent studies and points to BDNF as a possible prognostic indicator in major psychoses.

Exploring the association between brain-derived neurotrophic factor levels and longitudinal psychopathological and cognitive changes in Sardinian psychotic patients.

BACKGROUND AND HYPOTHESIS: Schizophrenia spectrum disorders are among the most debilitating mental disorders and has complex pathophysiological underpinnings. There is growing evidence that brain-derived neurotrophic factor (BDNF) can play a role in its pathogenesis. The present study investigated the longitudinal variation of serum BDNF levels in a 24-month observational prospective cohort study of Sardinian psychotic patients and its relationship with psychopathological and cognitive changes. Furthermore, we examined whether genetic variation within the BDNF gene could moderate these relationships. STUDY DESIGN: Every 6 months, 105 patients were assessed for their BDNF serum levels, as well as for a series of psychopathological, cognitive, and social measures. We performed a targeted analysis of four tag single nucleotide polymorphisms within the BDNF gene that were selected and analyzed using polymerase chain reaction. Longitudinal data were analyzed using mixed-effects linear regression models. STUDY RESULTS: We observed a declining longitudinal trajectory of BDNF levels in psychotic patients in general, and in relation to the severity of depressive and negative symptoms. BDNF serum levels also declined in patients scoring lower in cognitive measures such as attention and speed of information processing and verbal fluency. The rs7934165 polymorphism moderated the significant association between verbal fluency and BDNF levels. CONCLUSIONS: These findings in patients from real-world settings suggest a plausible role of peripheral BDNF levels as a marker of illness burden in schizophrenia spectrum disorders.

Altered brain levels of arachidonic acid-derived inflammatory eicosanoids in a rodent model of anorexia nervosa.

Increasing evidence underline the role of inflammation in the behavioral, emotional and cognitive dysregulations displayed in anorexia nervosa (AN). Among the inflammatory mediators acting at both peripheral and central levels, growing attention receives a class of lipids derived from arachidonic acid (AA), called eicosanoids (eiCs), which exert a complex, multifaceted role in a wide range of neuroinflammatory processes, peripheral inflammation, and generally in immune system function. To date, little is known about their possible involvement in the neurobiological underpinnings of AN. The present study evaluated whether the activity-based model of AN (ABA) may alter AA-metabolic pathways by changing the levels of AA-derived eiCs in specific brain areas implicated in the development of the typical anorexic-like phenotype, i.e. in prefrontal cortex, cerebral cortex, nucleus accumbens, caudate putamen, amygdala, hippocampus, hypothalamus and cerebellum. Our results point to brain region-specific alterations of the cyclooxygenase (COX), lipoxygenase (LOX) and cytochrome P450 epoxygenase (CYP) metabolic pathways rendering altered levels of AA-derived eiCs (i.e. prostaglandins, thromboxanes and hydroxyeicosatetraenoic acids) in response to induction of and recovery from the ABA condition. These changes, supported by altered messenger RNA (mRNA) levels of genes coding for enzymes involved in eiCs-related methabolic pathways (i.e., PLA2, COX-2, 5-LOX and 15-LOX), underlie a widespread brain dysregulation of pro- and anti-inflammatory eiC-mediated processes in the ABA model of AN. These data suggest the importance of eiCs signaling within corticolimbic areas in regulating key neurobehavioral functions and highlight eiCs as biomarker candidates for monitoring the onset and development of AN, and/or as possible targets for pharmacological management.

Impaired brain endocannabinoid tone in the activity-based model of anorexia nervosa.

OBJECTIVE: Despite the growing knowledge on the functional relationship between an altered endocannabinoid (eCB) system and development of anorexia nervosa (AN), to date no studies have investigated the central eCB tone in the activity-based anorexia (ABA) model that reproduces key aspects of human AN. METHOD: We measured levels of two major eCBs, anandamide (AEA) and 2-arachidonoylglycerol (2-AG), those of two eCB-related lipids, oleoylethanolamide (OEA) and palmitoylethanolamide (PEA), and the cannabinoid type-1 receptor (CB1R) density in the brain of female ABA rats, focusing on areas involved in homeostatic and rewarding-related regulation of feeding behavior (i.e., prefrontal cortex, nucleus accumbens, caudato putamen, amygdala, hippocampus and hypothalamus). Analysis was carried out also at the end of recovery from the ABA condition. RESULTS: At the end of the ABA induction phase, 2-AG was significantly decreased in ABA rats in different brain areas but not in the caudato putamen. No changes were detected in AEA levels in any region, whereas the levels of OEA and PEA were decreased exclusively in the hippocampus and hypothalamus. Furthermore, CB1R density was decreased in the dentate gyrus of hippocampus and in the lateral hypothalamus. After recovery, both 2-AG levels and CB1R density were partially normalized in some areas. In contrast, AEA levels became markedly reduced in all the analyzed areas. DISCUSSION: These data demonstrate an altered brain eCB tone in ABA rats, further supporting the involvement of an impaired eCB system in AN pathophysiology that may contribute to the maintenance of some symptomatic aspects of the disease.

Animal Models of Eating Disorders.

Eating disorders (EDs) include a range of chronic and disabling pathologies characterized by persistent maladaptive eating habits and/or behaviors aimed at controlling body shape and size, with important consequences on physical health. Different animal models of EDs have been developed to investigate pharmacological, environmental, and genetic determinants that contribute to the development and maintenance of these disorders as well as for the identification of potential therapeutic targets. In this chapter, we will provide an overview of the most useful animal models of EDs, focusing mainly on those used to study anorexia nervosa and binge eating disorder.

Cannabinoid CB1 /CB2 receptor agonists attenuate hyperactivity and body weight loss in a rat model of activity-based anorexia.

BACKGROUND AND PURPOSE: Anorexia nervosa (AN) is a serious psychiatric condition characterized by excessive body weight loss and disturbed perceptions of body shape and size, often associated with excessive physical activity. There is currently no effective drug-related therapy of this disease and this leads to high relapse rate. Clinical data suggest that a promising therapy to treat and reduce reoccurrence of AN may be based on the use of drugs that target the endocannabinoid (EC) system, which appears dysregulated in AN patients. EXPERIMENTAL APPROACH: The activity-based anorexia (ABA) rodent model mimics the severe body weight loss and increased physical activity, as well as the neuroendocrine disturbances (i.e. hypoleptinaemia and hypercortisolaemia) in AN. This study investigated whether cannabinoid agonists can effectively modify anorexic-like behaviours and neuroendocrine changes in rats subjected to a repeated ABA regime that mimics the human condition in which patients repeatedly undergo a recovery and illness cycle. KEY RESULTS: Our data show that subchronic treatment with both the natural CB1 /CB2 receptor agonist Δ9 -tetrahydrocannabinol and the synthetic CB1 /CB2 receptor agonist CP-55,940 significantly reduced body weight loss and running wheel activity in ABA rats. These behavioural effects were accompanied by an increase in leptin signalling and a decrease in plasma levels of corticosterone. CONCLUSION AND IMPLICATIONS: Taken together, our results further demonstrate the involvement of the EC system in AN pathophysiology and that strategies which modulate EC signalling are useful to treat this disorder, specifically in patients where physical hyperactivity plays a central role in its progression and maintenance.

Longitudinal assessment of brain-derived neurotrophic factor in Sardinian psychotic patients (LABSP): a protocol for a prospective observational study.

INTRODUCTION: Brain-derived neurotrophic factor (BDNF) plays a crucial role in neurodevelopment, synaptic plasticity and neuronal function and survival. Serum and plasma BDNF levels are moderately, but consistently, decreased in patients with schizophrenia (SCZ) compared with healthy controls. There is a lack of knowledge, however, on the temporal manifestation of this decline. Clinical, illness course and treatment factors might influence the variation of BDNF serum levels in patients with psychosis. In this context, we propose a longitudinal study of a cohort of SCZ and schizophrenic and schizoaffective disorder (SAD) Sardinian patients with the aim of disentangling the relationship between peripheral BDNF serum levels and changes of psychopathology, cognition and drug treatments. METHODS AND ANALYSIS: Longitudinal assessment of BDNF in Sardinian psychotic patients (LABSP) is a 24-month observational prospective cohort study. Patients with SAD will be recruited at the Psychiatry Research Unit of the Department of Medical Science and Public Health, University of Cagliari and University of Cagliari Health Agency, Cagliari, Italy. We will collect BDNF serum levels as well as sociodemographic, psychopathological and neurocognitive measures. Structured, semistructured and self-rating assessment tools, such as the Positive and Negative Syndrome Scale for psychopathological measures and the Brief Assessment of Cognition in Schizophrenia for cognitive function, will be used. ETHICS AND DISSEMINATION: This study protocol was approved by the University of Cagliari Health Agency Ethics Committee (NP2016/5491). The study will be conducted in accordance with the principles of good clinical practice, in the Declaration of Helsinki in compliance with the regulations. Participation will be voluntary and written informed consent will be obtained for each participant upon entry into the study. We plan to disseminate the results of our study through conference presentations and publication in international peer-reviewed journals. Access to raw data will be available in anonymised form upon request to the corresponding author.

The ketamine-like compound methoxetamine substitutes for ketamine in the self-administration paradigm and enhances mesolimbic dopaminergic transmission.

RATIONALE: Recently, an increasing number of emergency cases due to a novel ketamine-like drug, methoxetamine (MXE), were reported in several countries. However, very little is known about the neuropsychopharmacological and reinforcing profile of this compound. OBJECTIVES: Our study aims to investigate the effects of MXE on self-administration (SA) behaviour in comparison to ketamine and on dopaminergic transmission. METHODS: A SA substitution study was performed in male rats trained to intravenously (IV) self-administer ketamine. At responding stability, rats were exposed to sequential phases of MXE substitution at different dosages (starting from 0.5 and then decreasing to 0.25 and 0.125 mg/kg). Standard electrophysiological techniques were used to record changes in firing activities of ventral tegmental area (VTA) dopamine neurons projecting to the nucleus accumbens (NAc) shell after acute injection of cumulative doses of MXE (0.031-0.5 mg/kg IV). Finally, in vivo microdialysis was performed in freely moving rats to evaluate the effect of acute MXE administration (0.125, 0.25 and 0.5 mg/kg IV) on dopamine release in the NAc shell. RESULTS: MXE 0.125 and 0.25 mg/kg, but not 0.5 mg/kg, substituted for ketamine SA. MXE also induced a dose-dependent stimulation of firing rate (p < 0.0001) and burst firing (p < 0.05) of NAc-projecting VTA dopamine neurons. Consistently, MXE significantly (p < 0.05) increased dopamine extracellular levels in the NAc shell at 0.5 and 0.25 mg/kg with different time onsets, i.e. at 40 and 100 min, respectively. CONCLUSIONS: This study, while confirming the reinforcing effects of MXE, highlights an electrophysiological and neurochemical profile predictive of its addictive properties.

Behavioural and neurochemical assessment of salvinorin A abuse potential in the rat.

RATIONALE: Salvinorin A is a recreational drug derived from Salvia divinorum, a sage species long used as an entheogen. While salvinorin A has potent hallucinogenic properties, its abuse potential has not been assessed consistently in controlled behavioural and neurochemical studies in rodents. OBJECTIVE: This study aimed to assess salvinorin A abuse potential by measuring its capacity to establish and maintain self-administration behaviour and to modify dopamine (DA) levels in the nucleus accumbens (NAcc) of rats. RESULTS: Male Lister Hooded (LH) and Sprague-Dawley (SD) rats were allowed to self-administer salvinorin A (0.5 or 1.0 µg/kg/infusion) intravenously 2 h/day for 20 days under a continuous schedule of reinforcement and lever pressing as operandum. LH rats discriminated between the active and inactive levers but did not reach the acquisition criterion for stable self-administration (≥12 active responses vs ≤5 inactive responses for at least 5 consecutive days). SD rats discriminated between the two levers at the lower dose only but, like LH rats, never acquired stable self-administration behaviour. Systemic salvinorin A increased extracellular DA in the NAcc shell of both LH (at ≥40 µg/kg) and SD rats (at ≥5 µg/kg), but injection into the ventral tegmental area (VTA) induced no significant change in NAcc DA concentration in LH rats and only brief elevations in SD rats. CONCLUSIONS: Salvinorin A differs from other commonly abused compounds since although it affects accumbal dopamine transmission, yet it is unable, at least at the tested doses, to sustain stable intravenous self-administration behaviour.