Thevinols and their 3-O-demethylated relatives, orvinols, are derivatives of the Diels-Alder adduct of natural alkaloid thebaine with methyl vinyl ketone. Taken together, thevinols and orvinols constitute an important family of opioid receptor (OR) ligands playing an important role in both the OR mediated antinociception and OR antagonism. Herein, we disclose for the first time the antagonist activity of the N-allyl substituted orvinol derivative fluorinated within the pharmacophore associated with C(20) and its surrounding. This compound was prepared via a novel synthetic sequence from 18,19-dihydrothevinone bearing an allyl substituent at N(17) and three fluorine atoms at C(21). Preliminary trials reported earlier demonstrated that the compound exhibited no analgesic activity. However, in vivo experiments conducted in an acute pain model (tail-flick test in mice) demonstrated that this fluorinated compound, when administered at doses of 5-10 mg/kg (sc) 30 min before morphine, exhibited antagonistic activity at the level of naloxone (1 mg/kg, sc) for a longer duration (at least 120 min) compared to naloxone (60 min). Together with the analgesic activity that has been reported for the C(21)-trifluorinated relatives bearing methyl or cyclopropylmethyl substituent at N(17), this result highlights C(21)-fluorinated thevinols and orvinols as the family of opioid receptor ligands (structurally related to buprenorphine, diprenorphine, etc.) covering the full range of activity profiles from agonists to antagonists, which is promising for tuning of their pharmacological properties via a substitution of hydrogen atoms within the pharmacophore associated with C(20) and its surrounding for fluorine.

Housing conditions and prior life experiences are known to affect animal behavior and brain regions that regulate important neural and physiological functions, such as memory and stress responses. Stressors of low to moderate intensity can provide an enriched environment and promote cognitive functions. In contrast, exposures of excessive severity and/or duration can lead to maladaptation and distress, altering cognitive functions. Such excessive stressors can thus be used to model pathological conditions and test diverse therapeutic approaches. In this study, we addressed whether inescapable exposure of rats to aversive water environment of the “forced swim test” would interfere with their cognitive performance on the subsequent “water escape” test. The data show that a single exposure to a 15-min forced swim session interferes significantly with the ability to escape the water environment. This is manifested by a significant increase in the latency of diving under the cylinder and a reduction in the proportion of rats capable of completing the test task during the initial session. Additionally, there is a lack of development in diving skills during repeated sessions at 15-minute intervals. These negative effects can persist for at least 2 weeks. The use of the “forced swim” procedure prior to the “water escape” test may, therefore, provide a novel experimental approach to modeling stress-induced cognitive dysfunctions and evaluating the effects of pharmacological agents with potential antidepressant and procognitive properties.

Determining the estrous cycle stages in mice is essential for optimizing breeding strategies, synchronizing experimental timelines, and facilitating studies in behavior, drug testing, and genetics. It is critical for reducing the production of genetically unmodified offspring in the generation and investigation of genetically modified animal models. An accurate detection of the estrus cycle is particularly relevant in the context of the 3Rs—Replacement, Reduction, and Refinement. The estrous cycle, encompassing the reproductive phases of mice, is key to refining experimental designs and addressing ethical issues related to the use of animals in research. This study presents results from two independent laboratories on the efficacy of the Mouse Estrus Detector (MED) from ELMI Ltd. (Latvia) for the accurate determination of the estrus phase. The female mice of five strains/stocks (CD1, FVB/N, C57Bl6/J, B6D2F1, and Swiss) were used. The results showed that the MEDProTM is a low-traumatic, simple, rapid, and painless method of estrus detection that supports the principles of the 3Rs. The use of the MEDProTM for estrus detection in mice caused minimal stress, enhanced mating efficiency, facilitated an increase in the number of embryos for in vitro fertilization, and allowed the production of the desired number of foster animals.

The unique physicochemical and pharmacological properties of synthetic drugs are responsible for both the morbidity and mortality associated with their overdose. Among the consequences resulting from misuse of these potent opioids, or in the case of mass casualties involving the large-scale use of weaponised synthetic opioids, is the need for higher and repeated doses of the competitive antagonist naloxone, currently the only drug available to reverse opioid overdose. We reviewed the reversal agents available worldwide, discuss the need for stronger, faster and longer-acting opioid antagonists to reverse opioid overdose, and look at possible areas of functionalization of morphinan-based opioid ligands.

Parkinson's disease is the second most common neurodegenerative pathology. Due to the limitations of existing therapeutic approaches, novel anti-parkinsonian medicines with non-dopamine mechanisms of action are clearly needed. One of the promising pharmacological targets for anti-Parkinson drug development is phosphodiesterase (PDE) 10A. The stimulating motor effects of PDE10A inhibition were detected only under the conditions of partial dopamine depletion. The results raise the question of whether PDE10A inhibitors are able to restore locomotor activity when dopamine levels are very low. To address this issue, we (1) developed and validated the rat model of acute severe dopamine deficiency and (2) tested the action of PDE10A inhibitor MP-10 in this model. All experiments were performed in dopamine transporter knockout (DAT-KO) rats. A tyrosine hydroxylase inhibitor, α-Methyl-DL-tyrosine (αMPT), was used as an agent to cause extreme dopamine deficiency. In vivo tests included estimation of locomotor activity and catalepsy levels in the bar test. Additionally, we evaluated the tissue content of dopamine in brain samples by HPLC analysis. The acute administration of αMPT to DAT-KO rats caused severe depletion of dopamine, immobility, and catalepsy (Dopamine-Deficient DAT-KO (DDD) rats). As expected, treatment with the L-DOPA and carbidopa combination restored the motor functions of DDD rats. Strikingly, administration of MP-10 also fully reversed immobility and catalepsy in DDD rats. According to neurochemical studies, the action of MP-10, in contrast to L-DOPA + carbidopa, seems to be dopamine-independent. These observations indicate that targeting PDE10A may represent a new promising approach in the development of non-dopamine therapies for Parkinson's disease.

Thevinols and their 3-O-demethylated relatives, orvinols, are derivatives of the Diels-Alder adduct of natural alkaloid thebaine with methyl vinyl ketone. Taken together, thevinols and orvinols constitute an important family of opioid receptor (OR) ligands playing an important role in both the OR mediated antinociception and OR antagonism. Herein, we disclose for the first time the OR activity of orvinols fluorinated within the pharmocophore associated with C(20) and its surrounding along with a dependence of the activity profile on the substituent at N(17). Starting from thevinone and 18,19-dihydrothevinone, a family of C(21)-fluorinated orvinols bearing methyl, cyclopropylmethyl (CPM), and allyl substituent at N(17) was synthesized. The fluorinated compounds were evaluated for OR activity. The orvinols bearing three fluorine atoms at C(21) were found to retain the properties of OR ligands and their activity profile depends on the substituent at N(17). Pilot in vivo experiments in a model of acute pain (tail-flick test in mice) revealed that 6-O-desmethyl-21,21,21-trifluoro-20-methylorvinol at doses 1.0-10.0 mg/kg (s.c.) exhibits analgesic activity at the level of morphine for a duration of 30-180 min. Its N(17)-CPM counterpart demonstrated the partial opioid agonist properties. The N(17)-allyl substituted derivative showed no analgesic activity. In vivo evaluation of an analgesic activity indicates that 21,21,21-trifluoro-20-methylorvinols represent a novel family of OR ligands related to buprenorphine, diprenorphine, etc. These compounds are promising for the structure-activity relationship studies among the thevinol/orvinol series as well as for a search for new OR ligands with potentially valuable pharmacological profiles.

Introduction. Phosphodiesterases (PDEs) are a group of enzymes that hydrolyze the phosphodiester bond in cyclic nucleotides. PDE10A is mainly present in the medium-sized spiny neurons of the striatum. Functionally, PDE10A inhibition imitates the effect of D1-like agonists and D2-like dopamine receptor antagonists, and simultaneously modulating “direct” and “indirect” striato-thalamo-cortical brain pathway. To date, the effects of PDE10A inhibition have been characterized mainly, reproducing the inhibitory motor activity of D2-like dopamine receptor antagonists.The objective was to evaluate the stimulating motor activity of the effect of PDE10A inhibitors, as well as the possible development of tolerance to these effects when they are re-administered.Methods and materials. The hypodopaminergic state in male Wistar stock rats was modeled by administration of the VMAT2 inhibitor tetrabenazine (3 mg/kg). The effects of selective inhibitors PDE10A, MP-10 (0.3–5 mg/kg) and RO5545965 (0.1–0.9 mg/kg), on the motor activity of rats were evaluated with single and repeated administration (5 and 10 days).Results. Switching off PDE10A dose-dependently stimulated the motor activity of rats after administration of tetrabenazine. However, repeated administration of PDE10A inhibitors was accompanied by the development of tolerance to their paradoxical stimulating effect.Conclusion. The development of tolerance may limit the potential clinical use of PDE10A inhibitors to correct hypodopaminergic symptoms in patients with Parkinson’s disease. It is necessary to study the molecular mechanism of this phenomenon.

Introduction . Cognitive impairments are important components of the clinical picture of many neuropsychiatric disorders, and are in dire need of evidence-based pharmacotherapeutic approaches. The objective was to test a model of delayed cognitive impairments due to neonatal administration of NMDA receptor antagonists (7th, 9 th, and 11 th days of life). Methods and materials . Male Wistar rats were administered with phencyclidine, 10 mg/kg (Experiment 1), or (+)MK-801, 1 mg/kg (Experiment 2); then «2-choice serial reaction time task», or «reinforcement learning task based on response patterning under interval schedules of reinforcement» in the same adult rats were performed. Results . Experiment 1: rats after neonatal NMDA-blockade performed operant tasks more accurately and made fewer missed attempts as compared to control. Experiment 2: switching to another schedule of reinforcement increased the pause after reinforced responses in both groups; in the experimental group, the duration of the post-reinforcement pause was shorter. Conclusion . Neonatal NMDA receptor blockade affects inhibitory control and attention to sensory stimuli, which manifests, respectively, as increased impulsivity and hyperfocusing in limited-choice conditions.

Introduction. Changes in the expression of the dopamine transporter (DAT) have been identified in patients with a number of neuropsychiatric disorders, but their significance for pathogenesis remains unclear. DAT knockout rats are a promising model of frontostriatal dysfunctions involved in adapting learning processes to current organism's needs, motivation, and experience.The objective was to evaluate the effect of DAT disruption on the interaction of classical and instrumental conditioning processes (associative learning).Methods and materials. Rats from a local colony were used: knockouts (n=31), heterozygotes (n=32), and wild type rats (n=24). DAT knockout (Slc6a3) was detected by PCR followed by restriction analysis. The rats were kept individually with limited access to food and unlimited access to water. In the operant chambers equipped with a MED-PC interface (MED Associates, USA), 3 experiments were performed: 1) classical conditioning; 2) autoshaping of operant responding; 3) reward learning based on secondary reinforcement. Statistical analysis was performed using SigmaPlot 12.5 (Systat Software Inc., USA) and SPSS Statistics 21 (IBM, USA).Results. We revealed that DAT disruption in rats did not affect the formation of conditioned reflex connections in classical conditioning (1) but was accompanied by impairments in the autoshaping of the operant response (2) and learning based on secondary reinforcement (3).Conclusion. The observed impairments of associative learning might be associated with a decreased incentive value of stimuli in hyperdopaminergic state.

There is experimental evidence that chronic social defeat stress is accompanied by the development of an anxiety, development of a depression-like state, and downregulation of serotonergic genes in midbrain raphe nuclei of male mice. Our study was aimed at investigating the effects of chronic lithium chloride (LiCl) administration on anxiety behavior and the expression of serotonergic genes in midbrain raphe nuclei of the affected mice. A pronounced anxiety-like state in male mice was induced by chronic social defeat stress in daily agonistic interactions. After 6 days of this stress, defeated mice were chronically treated with saline or LiCl (100 mg/kg, i.p., 2 weeks) during the continuing agonistic interactions. Anxiety was assessed by behavioral tests. RT-PCR was used to determine Tph2, Htr1a, Htr5b, and Slc6a4 mRNA expression. The results revealed anxiolytic-like effects of LiCl on social communication in the partition test and anxiogenic-like effects in both elevated plus-maze and social interaction tests. Chronic LiCl treatment upregulated serotonergic genes in midbrain raphe nuclei. Thus, LiCl effects depend on the treatment mode, psycho-emotional state of the animal, and experimental context (tests). It is assumed that increased expression of serotonergic genes is accompanied by serotonergic system activation and, as a side effect, by higher anxiety.

To determine the difference in stress response between two chromosomal forms of a common vole, the “arvalis” and “obscurus”, the behavior of adult (6–12 months) lab-born vole males were evaluated in a forced swim test. During the test, they were carefully placed into glass cylinders (d=12 cm, h=20 cm) filled with water (h=15 cm, 24±1°C) and remained there for up to 6 min. Their behavioral patterns were video recorded and continuously monitored (Ethograph, ver. 2.7, RITEC, Russia) to document the duration of swimming and paddling (orientation), floating (immobility) and climbing (escape). Two series of experiments were performed, in May 2016 and June 2017, with independent groups: “arvalis” (N=9 each year) and “obscurus” (N=9 and 22 each year, respectively). The «arvalis» and «obscurus» significantly differed in their responses to the stress test. Over 35% of obscurus males (2 of 9 in 2016 and 9 of 22 in 2017) failed to complete the test, within 3 min these animals had to be rescued to prevent drowning (significant differences from “arvalis” according to Fisher's exact test). “Arvalis” voles displayed higher duration of escape attempts (F(1.14)=3.5; P=0.08 – for first; F(1.20)=15.6; P<0.001 – for second series of experiment), shorter duration of swimming and paddling (significantly in the first series – F(1.14)=14.3; P<0.005), and shorter duration of immobility (significantly in the second series – (F(1.20)=6.1; P<0.001). The results suggest that two chromosomal forms of common vole have major differences in stress response behavior. In the forced swim test, the “arvalis” voles are displaying higher endurance, associated with active escape behavior, when compared to “obscurus” animals. Further studies need to address their responses to other stress paradigms and to water environment, in general. The latter may play a major role in vole migration when they meet water barriers in their natural environment.

Trace amine-associated receptor 1 (TAAR1) is a widely recognized new perspective target for the neuropsychiatric pharmacological treatment. Despite a growing number of studies investigating TAAR1 role in the animal models of different pathologies, information of TAAR1 agonists impact on executive cognitive functions is limited. The goal of the present study was to evaluate the activity of highly selective partial TAAR1 agonist RO5263397 on various executive cognitive functions. The results of the present study demonstrated that the pretreatment with RO5263397 was able to increase attention and decrease cognitive flexibility in rats. The analysis of the RO5263397 action on impulsivity demonstrated that the TAAR1 activation failed to affect premature responding but was able to slightly modify impulsive choice. Problem solving was resistant to the pharmacological intervention.

A technique to study effect of various baffles on liquid oscillations in partially filled rigid toroidal tanks is first developed by extending the semi-analytical scaled boundary finite element method (SBFEM), which utilizes the advantages of both the boundary element (BEM) and finite element methods (FEM). As found in the BEM, only the boundary is discretized to reduce the space by one, and no fundamental solution is needed unlike the BEM. The calculated liquid domain is divided into several simple sub-domains so that the liquid velocity potential in each liquid sub-domain becomes the class C1 with continuity boundary conditions. Based on the linear potential theory and weighted residual method, the semi-analytical solutions of the liquid velocity potential corresponding to each sub-domain are obtained by means of SBFEM, where the geometry of each sub-domain is transformed into scaled boundary coordinates, including the radial and circumferential coordinates by using a scaling centre, and the finite-element approximation of the circumferential coordinate yields the analytical equation in the radial coordinate. By discretizing the flow boundaries, the integral equation governed on the boundary is formulated into a general matrix eigenvalue problem. Based on the eigenvalue problem and multimodal method, an efficient methodology is adopted to computer the sloshing masses and sloshing force. Accuracy, simple and fast numerical computations are observed by the convergence study, and excellent agreements have been achieved in the comparison of results obtained by the proposed approach with other methods. Meanwhile, several baffle configurations are considered including the horizontal bottom-mounted and surface-piercing ring baffles as well as their combination form, bottom-mounted and surface-piercing ring baffles as well as their combination, and free surface-touching baffles. The effects of baffled arrangement, the ratio b/a of elliptical cross section, liquid fill level, and baffles' length upon the sloshing frequencies, the associated sloshing mode shapes and sloshing forces are investigated in detail and some conclusions are outlined. The results show that the present method allows for the simulation of complex 3D sloshing phenomena using a relative small number of degrees of freedom while the mesh consists of two-dimensional elements only.

Genetic and physiological studies have implicated the striatum in bipolar disorder (BD). Although Glycogen synthase kinase 3 beta (GSK3β) has been suggested to play a role in the pathophysiology of BD since it is inhibited by lithium, it remains unknown how GSK3β activity might be involved. Therefore we examined the functional roles of GSK3β and one of its substrates, CRMP2, within the striatum.Using CRISPR-Cas9 system, we specifically ablated GSK3β in the striatal neurons in vivo and in vitro. Sholl analysis was performed for the structural studies of medium spiny neurons (MSNs) and amphetamine-induced hyperlocomotion was measured to investigate the effects of gene ablations on the mania-like symptom of BD.GSK3β deficiency in cultured neurons and in neurons of adult mouse brain caused opposite patterns of neurite changes. Furthermore, specific knockout of GSK3β in the MSNs of the indirect pathway significantly suppressed amphetamine-induced hyperlocomotion. We demonstrated that these phenotypes of GSK3β ablation were mediated by CRMP2, a major substrate of GSK3β.Amphetamine-induced hyperlocomotion only partially recapitulate the symptoms of BD. It requires further study to examine whether abnormality in GSK3β or CRMP2 is also involved in depression phase of BD. Additionally, we could not confirm whether the behavioral changes observed in GSK3β-ablated mice were indeed caused by the cellular structural changes observed in the striatal neurons.Our results demonstrate that GSK3β and its substrate CRMP2 critically regulate the neurite structure of MSNs and their functions specifically within the indirect pathway of the basal ganglia network play a critical role in manifesting mania-like behavior of BD. Moreover, our data also suggest lithium may exert its effect on BD through a GSK3β-independent mechanism, in addition to the GSK3β inhibition-mediated mechanism.

Rat line lacking the dopamine transporter (DAT-KO rats) was recently developed. Lack of DAT in these mutants manifests behaviorally as spontaneous hyperactivity and cognitive deficits. It has been shown that d-amphetamine and methylphenidate paradoxically calm down these animals. Also, the inhibition of dopamine synthesis in DAT-KO rats represents a straightforward approach for developing the model of severe dopamine deficiency exhibiting characteristic akinetic phenotype which can be reversed by treatment with L-DOPA that used in Parkinson’s disease. These results support the usefulness of DAT-KO rats as models of the disorders associated with dopaminergic dysfunction.

Across the world, people living with schizophrenia (PLS) are often cared for by family caregivers. This is especially true in China where community-based systems of care for PLS are underdeveloped. Thus, family caregivers of PLS may experience emotional distress and burden in the absence of community-based services. Pearlin (1990) proposed a highly influential model for understanding the stress process and its relationship to health and wellbeing. Although the model has been examined worldwide, it has yet to be studied in China for caregivers of PLS. In the present study, we use Pearlin's stress process model to examine the mediating effect of perceived caregiver burden, family functioning, and caregivers' positive feelings about caregiving on PLS global functioning and caregiver depressive symptoms. We also examine whether the model differs for parent and spouse caregivers. A total of 327 primary family caregivers of PLS in Hunan province, China, were assessed cross-sectionally using culturally-validated measures. Bootstrap methods with Hayes's PROCESS Macro were used to test multiple mediation models of global functioning scores of PLS and caregiver depressive symptoms. In this first study of the stress process model of family caregivers of PLS in China, the results generally support the model. Caregiver perceived burden, family functioning, and positive caregiver feelings about caregiving all mediate the relationship between PLS global functioning and caregiver depression, with perceived caregiver burden having the strongest indirect effect. Analyses also show that the stress process differs between parent and spouse caregivers, with perceived caregiver burden a mediator for both groups but family functioning only a mediator for parent depressive symptoms. We discuss implications of these findings for extending the stress process model cross-culturally; suggest hypotheses for future research to examine parent and spouse differences; and discuss fruitful directions for intervention.

To evaluate the diagnostic value of intravoxel incoherent motion (IVIM) diffusion-weighted (DWI) magnetic resonance imaging (MRI) and semi-quantitative dynamic contrast-enhanced MRI (DCE-MRI) to help diagnose indeterminate solitary pulmonary lesions (SPLs) and the subgroups of lung cancer (LC), and to explore the relationship between IVIM and DCE-MRI.Sixty-four consecutive patients (44 male, 20 female; age, 52.77±10.46 years) from February 2014 to September 2016 with SPLs, were involved in this prospective study. Total apparent diffusion coefficient (ADCtotal), tissue diffusivity (D), pseudo-diffusion coefficient (D*), perfusion fraction (F), maximum enhancement ratio (MER), Tmax, slope, and washout were compared between the lung cancer (LC) and benign group and among the subtypes of LC. Time–intensity curves (TICs) were drawn. Receiver operating characteristic (ROC) curves were constructed to estimate the diagnostic performance. The correlation of both tools was assessed.ADCtotal, D, and Tmax were significantly higher for benignity than for LC (p=0.005, p=0.002 and p<0.001 respectively). D* and slope were significantly higher in LC than benignity (p=0.005 and p=0.011, respectively). D and Tmax had the highest sensitivity and accuracy, respectively. A combination of D and Tmax improved the sensitivity to 90.5%, the specificity to 86.4%, and the accuracy to 89.1%. Poor correlations were found between parameters derived from IVIM and DCE-MRI. ADCtotal values of SCC and SCLC were found to be significantly lower compared with that in adenocarcinoma.Both IVIM-DWI and DCE-MRI were useful for discriminating benignity from LC. ADCtotal was helpful for distinguishing adenocarcinoma and non-adenocarcinoma. A combination of DCE-MRI and IVIM could provide a robust method to determine the microstructural characteristics of SPLs.

Lutein is a bioactive found in dark leafy vegetables that may be used as a nutraceutical agent in foodstuff and an inhibitor of key enzymes of the human body such as those involved in the cholinergic system. However, its high hydrophobicity leads to low bioavailability and must be overcome if lutein is to be added in foods. The objective of this study was to evaluate the influence of nanoencapsulated lutein in the activity of the acetylcholinesterase enzyme. The in vitro study was carried out using water in order to evaluate the impact of encapsulation on the hydrophilicity of lutein. In vitro assays showed that lutein, both free and nanoencapsulated, presented a mixed-type inhibition behavior, and encapsulated lutein was able to inhibit acetylcholinesterase activity even in an aqueous medium. Inhibition was also showed by the in silico docking results which show that lutein interacted with the pocket region of the enzyme.