Synergistic anti-depressive effect of combination treatment of Brexpiprazole and selective serotonin reuptake inhibitors on forced swimming test in mice.

AIM: Selective serotonin reuptake inhibitors (SSRIs) are used to treat major depressive disorder (MDD) and other psychiatric disorders (e.g., obsessive compulsive disorder, social anxiety disorder, and panic disorder). In MDD treatment, SSRIs do not show remission in approximately 30% of patients, indicating a need for a better treatment option. Forced swimming test (FST) is a behavioral assay to evaluate depression-like behavior and antidepressant efficacy in rodents. In the present study, we evaluated the combination effect of brexpiprazole with SSRIs on FST in mice, in order to investigate their synergistic effect. METHODS: Brexpiprazole (0.003 mg/kg) was intraperitoneally injected to mice 15 min before testing. Escitalopram (10 mg/kg), fluoxetine (75 mg/kg), paroxetine (10 mg/kg), or sertraline (15 mg/kg) were orally administered to mice 60 min before testing. Then, the mice were placed in water and immobility time was measured. Data from animals treated with escitalopram, fluoxetine, paroxetine, and sertraline were pooled as SSRI-treated group data. RESULTS: Combination treatment of brexpiprazole with SSRIs reduced immobility time in FST more than vehicle or each single treatment. A significant interaction effect was confirmed in the combination of brexpiprazole and SSRIs (p = 0.0411). CONCLUSION: Efficacy of adjunctive brexpiprazole has already been demonstrated in clinical trials in MDD patients not adequately responding to antidepressants including escitalopram, fluoxetine, paroxetine, and sertraline. The synergistic antidepressant-like effect of brexpiprazole with SSRIs found in this study supports the already known clinical findings.

Prenatal methotrexate injection increases behaviors possibly associated with depression and/or autism in rat offspring; A new animal model for mental disorder, based on folate metabolism deficit during pregnancy.

BACKGROUND: Deficiency of folate, an essential vitamin for DNA synthesis and methylation, is reported as a risk factor for mental disorders. Considering a possibility that folate metabolism deficit during pregnancy may disturb CNS development and increase mental disorders in offspring, we treated pregnant rats with methotrexate (MTX), an inhibitor of folate metabolic enzyme, and evaluated offspring behaviors. METHODS: Saline or MTX was intraperitoneally administered to female SD rats on gestational day 17. Offspring behaviors were evaluated during approximately 6-9 weeks old; prepulse inhibition (PPI), social interaction (SI), locomotor activity (LA), and forced swimming test (FST) for evaluation of schizophrenia, depression, and autism related behaviors; the elevated plus maze (EPM) and the light-dark box (LD) test for evaluation of anxiety. RESULTS: Compared to saline-treated group, MTX-treated group showed decrease of SI and increase of immobility time in FST. In addition, increases of time spent in the light box and shuttling between the light-dark boxes were observed in LD test. On the other hand, no changes were confirmed in EPM, LA, and PPI. CONCLUSION: Decrease of SI and increase of immobility time in FST may suggest association of this animal model with depression and/or autism. Increase of time spent in the light box and shuttling between the light-dark boxes may indicate changes in anxiety or cognitive level to environment, or repetitive behaviors in autism. Although further studies are warranted to characterize this animal model, at least we can say that prenatal MTX exposure, possibly causing folate metabolism deficit, affects offspring behaviors.

Brexpiprazole has a low risk of dopamine D2 receptor sensitization and inhibits rebound phenomena related to D2 and serotonin 5-HT2A receptors in rats.

BACKGROUND: Long-term antipsychotic treatment in patients with schizophrenia can induce supersensitivity psychosis and tardive dyskinesia which is thought to be caused by dopamine D2 receptor sensitization. We evaluated the effects of brexpiprazole on D2 receptor sensitivity after subchronic treatment in rats. We also evaluated whether brexpiprazole could suppress enhanced response to D2 receptors in rats subchronically dosed with another atypical antipsychotic. METHODS: The maximum D2 receptor density (Bmax ) and apomorphine (a D2 receptor agonist)-induced stereotypy were measured in rats orally dosed with vehicle, haloperidol (1 mg/kg), or brexpiprazole (4 or 30 mg/kg for Bmax , 6 or 30 mg/kg for stereotypy) for 21 days. Then, effects of oral administrations of brexpiprazole (3 mg/kg), aripiprazole (10 mg/kg), and olanzapine (3 mg/kg) against increases in apomorphine-induced hyperlocomotion and (±)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI: a 5-HT2A receptor agonist)-induced head twitches were evaluated in rats subcutaneously treated with risperidone (1.5 mg/kg/d) via minipumps for 21 days. RESULTS: Haloperidol and brexpiprazole (30 mg/kg: approximately tenfold ED50 of anti-apomorphine-induced stereotypy) but not brexpiprazole (4 or 6 mg/kg) significantly increased the Bmax and apomorphine-induced stereotypy. Brexpiprazole (3 mg/kg) and olanzapine (3 mg/kg) significantly suppressed both increases in apomorphine-induced hyperlocomotion and also DOI-induced head twitches in rats subchronically treated with risperidone, but aripiprazole (10 mg/kg) significantly suppressed only apomorphine-induced hyperlocomotion. CONCLUSION: Brexpiprazole has a low risk of D2 receptor sensitization after a repeated administration and suppresses the rebound phenomena related to D2 and 5-HT2A receptors after a repeated administration of risperidone.

A Low Mortality, High Morbidity Reduced Intensity Status Epilepticus (RISE) Model of Epilepsy and Epileptogenesis in the Rat.

Animal models of acquired epilepsies aim to provide researchers with tools for use in understanding the processes underlying the acquisition, development and establishment of the disorder. Typically, following a systemic or local insult, vulnerable brain regions undergo a process leading to the development, over time, of spontaneous recurrent seizures. Many such models make use of a period of intense seizure activity or status epilepticus, and this may be associated with high mortality and/or global damage to large areas of the brain. These undesirable elements have driven improvements in the design of chronic epilepsy models, for example the lithium-pilocarpine epileptogenesis model. Here, we present an optimised model of chronic epilepsy that reduces mortality to 1% whilst retaining features of high epileptogenicity and development of spontaneous seizures. Using local field potential recordings from hippocampus in vitro as a probe, we show that the model does not result in significant loss of neuronal network function in area CA3 and, instead, subtle alterations in network dynamics appear during a process of epileptogenesis, which eventually leads to a chronic seizure state. The model's features of very low mortality and high morbidity in the absence of global neuronal damage offer the chance to explore the processes underlying epileptogenesis in detail, in a population of animals not defined by their resistance to seizures, whilst acknowledging and being driven by the 3Rs (Replacement, Refinement and Reduction of animal use in scientific procedures) principles.

Global Trends in Alzheimer Disease Clinical Development: Increasing the Probability of Success.

PURPOSE: Alzheimer disease (AD) is a growing global health and economic issue as elderly populations increase dramatically across the world. Despite the many clinical trials conducted, currently no approved disease-modifying treatment exists. In this commentary, the present status of AD drug development and the grounds for collaborations between government, academia, and industry to accelerate the development of disease-modifying AD therapies are discussed. METHODS: Official government documents, literature, and news releases were surveyed by MEDLINE and website research. FINDINGS: Currently approved anti-AD drugs provide only short-lived symptomatic improvements, which have no effect on the underlying pathogenic mechanisms or progression of the disease. The failure to approve a disease-modifying drug for AD may be because the progression of AD in the patient populations enrolled in clinical studies was too advanced for drugs to demonstrate cognitive and functional improvements. The US Food and Drug Administration and the European Medicines Agency recently published draft guidance for industry which discusses approaches for conducting clinical studies with patients in early AD stages. For successful clinical trials in early-stage AD, however, it will be necessary to identify biomarkers highly correlated with the clinical onset and the longitudinal progress of AD. In addition, because of the high cost and length of clinical AD studies, support in the form of global initiatives and collaborations between government, industry, and academia is needed. IMPLICATIONS: In response to this situation, national guidance and international collaborations have been established. Global initiatives are focusing on 2025 as a goal to provide new treatment options, and early signs of success in biomarker and drug development are already emerging.

MK-801-induced deficits in social recognition in rats: reversal by aripiprazole, but not olanzapine, risperidone, or cannabidiol.

Deficiencies in social activities are hallmarks of numerous brain disorders. With respect to schizophrenia, social withdrawal belongs to the category of negative symptoms and is associated with deficits in the cognitive domain. Here, we used the N-methyl-D-aspartate receptor antagonist dizocilpine (MK-801) for induction of social withdrawal in rats and assessed the efficacy of several atypical antipsychotics with different pharmacological profiles as putative treatment. In addition, we reasoned that the marijuana constituent cannabidiol (CBD) may provide benefit or could be proposed as an adjunct treatment in combination with antipsychotics. Hooded Lister rats were tested in the three-chamber version for social interaction, with an initial novelty phase, followed after 3 min by a short-term recognition memory phase. No drug treatment affected sociability. However, distinct effects on social recognition were revealed. MK-801 reduced social recognition memory at all doses (>0.03 mg/kg). Predosing with aripiprazole dose-dependently (2 or 10 mg/kg) prevented the memory decline, but doses of 0.1 mg/kg risperidone or 1 mg/kg olanzapine did not. Intriguingly, CBD impaired social recognition memory (12 and 30 mg/kg) but did not rescue the MK-801-induced deficits. When CBD was combined with protective doses of aripiprazole (CBD-aripiprazole at 12 : or 5 : 2 mg/kg) the benefit of the antipsychotic was lost. At the same time, activity-related changes in behaviour were excluded as underlying reasons for these pharmacological effects. Collectively, the combined activity of aripiprazole on dopamine D2 and serotonin 5HT1A receptors appears to provide a significant advantage over risperidone and olanzapine with respect to the rescue of cognitive deficits reminiscent of schizophrenia. The differential pharmacological properties of CBD, which are seemingly beneficial in human patients, did not back-translate and rescue the MK-801-induced social memory deficit.

Brexpiprazole II: antipsychotic-like and procognitive effects of a novel serotonin-dopamine activity modulator.

Brexpiprazole (OPC-34712, 7-{4-[4-(1-benzothiophen-4-yl)piperazin-1-yl]butoxy}quinolin-2(1H)-one) is a novel serotonin-dopamine activity modulator with partial agonist activity at serotonin 1A (5-HT1A) and D2/3 receptors, combined with potent antagonist effects on 5-HT2A, α1B-, and α2C-adrenergic receptors. Brexpiprazole inhibited conditioned avoidance response (ED50 = 6.0 mg/kg), apomorphine- or d-amphetamine-induced hyperactivity (ED50 = 2.3 and 0.90, respectively), and apomorphine-induced stereotypy (ED50 = 2.9) in rats at clinically relevant D2 receptor occupancies. Brexpiprazole also potently inhibited apomorphine-induced eye blinking in monkeys. The results suggest that brexpiprazole has antipsychotic potential. Brexpiprazole induced catalepsy (ED50 = 20) well above clinically relevant D2 receptor occupancies, suggesting a low risk for extrapyramidal side effects. Subchronic treatment with phencyclidine (PCP) induced cognitive impairment in both novel object recognition (NOR) and attentional set-shifting (ID-ED) tests in rats. Brexpiprazole reversed the PCP-induced cognitive impairment in the NOR test at 1.0 and 3.0 mg/kg, and in the ID-ED test at 1.0 mg/kg. However, aripiprazole (10 mg/kg) was ineffective in both tests, despite achieving relevant D2 occupancies. In the NOR test, the 5-HT1A agonist buspirone and the 5-HT2A antagonist M100907 [(R)-(2,3-dimethoxyphenyl)[1-(4-fluorophenethyl)piperidin-4-yl]methanol] partially but significantly reversed PCP-induced impairment. Furthermore, the effect of brexpiprazole was reversed by cotreatment with the 5-HT1A antagonist WAY100635 (N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinylcyclohexanecarboxamide maleate). The results indicate that brexpiprazole has antipsychotic-like activity and robust efficacy in relevant models of cognitive impairment associated with schizophrenia. The effects of brexpiprazole in the cognitive tests are superior to those of aripiprazole. We propose that the pharmacologic profile of brexpiprazole be based on its balanced effects on 5-HT1A, D2, and 5-HT2A receptors, with possible modulating activity through additional monoamine receptors.

Brexpiprazole I: in vitro and in vivo characterization of a novel serotonin-dopamine activity modulator.

Brexpiprazole (OPC-34712, 7-{4-[4-(1-benzothiophen-4-yl)piperazin-1-yl]butoxy}quinolin-2(1H)-one) is a novel drug candidate in clinical development for psychiatric disorders with high affinity for serotonin, dopamine, and noradrenaline receptors. In particular, it bound with high affinity (Ki < 1 nM) to human serotonin 1A (h5-HT1A)-, h5-HT2A-, long form of human D2 (hD2L)-, hα1B-, and hα2C-adrenergic receptors. It displayed partial agonism at h5-HT1A and hD2 receptors in cloned receptor systems and potent antagonism of h5-HT2A receptors and hα1B/2C-adrenoceptors. Brexpiprazole also had affinity (Ki < 5 nM) for hD3-, h5-HT2B-, h5-HT7-, hα1A-, and hα1D-adrenergic receptors, moderate affinity for hH1 (Ki = 19 nM), and low affinity for hM1 receptors (Ki > 1000 nM). Brexpiprazole potently bound to rat 5-HT2A and D2 receptors in vivo, and ex vivo binding studies further confirmed high 5-HT1A receptor binding potency. Brexpiprazole inhibited DOI (2,5-dimethoxy-4-iodoamphetamine)-induced head twitches in rats, suggestive of 5-HT2A antagonism. Furthermore, in vivo D2 partial agonist activity of brexpiprazole was confirmed by its inhibitory effect on reserpine-induced DOPA accumulation in rats. In rat microdialysis studies, brexpiprazole slightly reduced extracellular dopamine in nucleus accumbens but not in prefrontal cortex, whereas moderate increases of the dopamine metabolites, homovanillic acid and DOPAC (3,4-dihydroxy-phenyl-acetic acid), in these areas also suggested in vivo D2 partial agonist activity. In particular, based on a lower intrinsic activity at D2 receptors and higher binding affinities for 5-HT1A/2A receptors than aripiprazole, brexpiprazole would have a favorable antipsychotic potential without D2 receptor agonist- and antagonist-related adverse effects. In conclusion, brexpiprazole is a serotonin-dopamine activity modulator with a unique pharmacology, which may offer novel treatment options across a broad spectrum of central nervous system disorders.

Cannabidivarin (CBDV) suppresses pentylenetetrazole (PTZ)-induced increases in epilepsy-related gene expression.

To date, anticonvulsant effects of the plant cannabinoid, cannabidivarin (CBDV), have been reported in several animal models of seizure. However, these behaviourally observed anticonvulsant effects have not been confirmed at the molecular level. To examine changes to epilepsy-related gene expression following chemical convulsant treatment and their subsequent control by phytocannabinoid administration, we behaviourally evaluated effects of CBDV (400 mg/kg, p.o.) on acute, pentylenetetrazole (PTZ: 95 mg/kg, i.p.)-induced seizures, quantified expression levels of several epilepsy-related genes (Fos, Casp 3, Ccl3, Ccl4, Npy, Arc, Penk, Camk2a, Bdnf and Egr1) by qPCR using hippocampal, neocortical and prefrontal cortical tissue samples before examining correlations between expression changes and seizure severity. PTZ treatment alone produced generalised seizures (median: 5.00) and significantly increased expression of Fos, Egr1, Arc, Ccl4 and Bdnf. Consistent with previous findings, CBDV significantly decreased PTZ-induced seizure severity (median: 3.25) and increased latency to the first sign of seizure. Furthermore, there were correlations between reductions of seizure severity and mRNA expression of Fos, Egr1, Arc, Ccl4 and Bdnf in the majority of brain regions in the CBDV+PTZ treated group. When CBDV treated animals were grouped into CBDV responders (criterion: seizure severity ≤3.25) and non-responders (criterion: seizure severity >3.25), PTZ-induced increases of Fos, Egr1, Arc, Ccl4 and Bdnf expression were suppressed in CBDV responders. These results provide the first molecular confirmation of behaviourally observed effects of the non-psychoactive, anticonvulsant cannabinoid, CBDV, upon chemically-induced seizures and serve to underscore its suitability for clinical development.

Plasma and brain pharmacokinetic profile of cannabidiol (CBD), cannabidivarine (CBDV), Δ9-tetrahydrocannabivarin (THCV) and cannabigerol (CBG) in rats and mice following oral and intraperitoneal administration and CBD action on obsessive-compulsive behaviour.

RATIONALE: Phytocannabinoids are useful therapeutics for multiple applications including treatments of constipation, malaria, rheumatism, alleviation of intraocular pressure, emesis, anxiety and some neurological and neurodegenerative disorders. Consistent with these medicinal properties, extracted cannabinoids have recently gained much interest in research, and some are currently in advanced stages of clinical testing. Other constituents of Cannabis sativa, the hemp plant, however, remain relatively unexplored in vivo. These include cannabidiol (CBD), cannabidivarine (CBDV), Δ(9)-tetrahydrocannabivarin (Δ(9)-THCV) and cannabigerol (CBG). OBJECTIVES AND METHODS: We here determined pharmacokinetic profiles of the above phytocannabinoids after acute single-dose intraperitoneal and oral administration in mice and rats. The pharmacodynamic-pharmacokinetic relationship of CBD (120 mg/kg, ip and oral) was further assessed using a marble burying test in mice. RESULTS: All phytocannabinoids readily penetrated the blood-brain barrier and solutol, despite producing moderate behavioural anomalies, led to higher brain penetration than cremophor after oral, but not intraperitoneal exposure. In mice, cremophor-based intraperitoneal administration always attained higher plasma and brain concentrations, independent of substance given. In rats, oral administration offered higher brain concentrations for CBD (120 mg/kg) and CBDV (60 mg/kg), but not for Δ(9)-THCV (30 mg/kg) and CBG (120 mg/kg), for which the intraperitoneal route was more effective. CBD inhibited obsessive-compulsive behaviour in a time-dependent manner matching its pharmacokinetic profile. CONCLUSIONS: These data provide important information on the brain and plasma exposure of new phytocannabinoids and guidance for the most efficacious administration route and time points for determination of drug effects under in vivo conditions.

Reduction of NR1 and phosphorylated Ca2+/calmodulin-dependent protein kinase II levels in Alzheimer's disease.

Ca2+ influx through the N-methyl-D-aspartate-type glutamate receptor leads to activation and postsynaptic accumulation of Ca2+/calmodulin-dependent protein kinase II. NR1 and NR2B subunits of N-methyl-D-aspartate receptor serve as high-affinity Ca2+/calmodulin-dependent protein kinase II docking sites in dendritic spines on autophosphorylation of Ca2+/calmodulin-dependent protein kinase II. By comparative Western blot analysis, we show a reduction of NR1 and phosphorylated Ca2+/calmodulin-dependent protein kinase II levels in the frontal cortex and hippocampus of Alzheimer's disease brains. We also found a significant correlation between phosphorylated Ca2+/calmodulin-dependent protein kinase II and NR1 levels. Our study extends the view that N-methyl-D-aspartate receptor deficiency underlies memory impairment in Alzheimer's disease, and that this process likely involves insufficient activation of Ca2+/calmodulin-dependent protein kinase II.

A novel rat orthologue and homologue for the Drosophila crooked neck gene in neural stem cells and their immediate descendants.

The crooked neck (crn) gene of Drosophila melanogaster encodes a scaffold protein carrying multiple tetratricopeptide repeat (TPR) motifs, and its mutation results in a reduction in the number of neuroblasts and lethality during larval stages. Here, we isolated two structurally related genes from a rat embryonic brain cDNA library. One gene is the rat orthologue of crn, which encodes 690 amino acids including 16 copies of TPR. The other gene, ATH55, encodes an 855 amino acid protein including 21 TPR motifs, which presumably represents a rat crn homologue and an orthologue of human XAB2. Both genes are highly expressed in embryonic brain but their expressions decrease during development. ATH55-like immunoreactivity is present in the ventricular zone and newly formed cortical plate, while CRN-like immunoreactivity is more abundant in a younger ventricular zone. In agreement, both proteins were found to be enriched in cultured neural stem cells and to decrease in response to cell differentiation signals. As indicated for the yeast CRN-like protein, ATH55 and CRN immunoreactivities were both recovered in the nuclear fraction and detected in the splicing complex carrying pre-mRNA. These findings suggest that both TPR-motif-containing proteins are involved in RNA processing of mammalian neural stem cells and their immediate descendants.