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ObjectivePreclinical models of seizures and epilepsy in rodents contributed substantially to the discovery of currently available antiseizure medications. These were also broadly used for investigation of processes of epileptogenesis. Nevertheless, rodent models pose some limitations, thus, new models using alternative species are in high demand. The aim of this study was to describe a new model of seizures/epilepsy induced by the cholinomimetic agent, pilocarpine (PILO), in larval zebrafish.MethodsLocal field potential (LFP) recordings were conducted to analyze electroencephalographic discharges and correlate it with larval behavior. Hematoxylin and eosin (H&E) staining, as well as TUNEL staining were performed to analyze morphology and apoptosis, respectively. Real-time quantitative polymerase chain reaction (qRT-PCR) was undertaken for gene expression analysis.ResultsAcute exposure to PILO, in a concentration-dependent manner, induces electroencephalographic discharges in larval zebrafish, which behaviorally manifest as decreased locomotion and moving time, but enhanced movement velocity. The PILO-induced seizure-like activity is behaviorally distinct from this induced by the application of chemoconvulsant pentylenetetrazole (PTZ). Zebrafish larvae previously exposed to PILO (2 h), after a washing out period, exhibit spontaneous, unprovoked discharges and apoptotic changes in their brains.SignificanceHere, we comprehensively investigated a new model of PILO-induced seizures/epilepsy in larval zebrafish. We propose that this model may be used to study epileptogenesis and for antiseizure drug screening purposes.

IntroductionBlood transfusion is an intervention widely used in therapeutics (e.g., in the perioperative period), thus, knowledge about factors associated with blood donation is important. The aim of this study was to investigate the impact of sociodemographic factors, personal values and life satisfaction on blood donation in Poland.MethodsSecondary analysis of data. A multiple logistic regression was carried out to assess the relationship between sociodemographic factors, life satisfaction (Satisfaction with Life Scale), personal values (Personal Values List) and blood donation.ResultsOf the 770 respondents aged 18–65 years, 262 respondents (34%) donated blood at least once in their lives. Respondents who believed that blood donation is safe (OR = 1.71; Cl95%: 1.20 to 2.43), were male (OR = 1.47; Cl95%: 1.26 to 1.72), married (OR = 1.31; Cl95%: 1.11 to 1.54) and those with high school education (OR = 0.81; Cl95%: 0.66 to 0.99) were more often blood donors.ConclusionPerceived blood donation safety and sociodemographic factors such as gender, marital status, and education level may influence blood donation. Health education is necessary to increase knowledge and shape positive attitudes toward blood donation among the society.

Presented text is an account of Krzysztof Koszlaga, a teacher with over 50 years of experience and one of the leading activists of the NSZZ “Solidarity” Education and Upbringing, with a scholarly commentary. The narration presents the functioning of education in the People’s Republic of Poland in the years 1969–1989, as well as the impact of the socio-political changes occurring during this period on the work of teachers and their relationships with students. The text is based on an in-depth interview with a witness to history, accompanied by an introduction containing biographical data of the author and a description of the research project within the framework of which the interview was conducted, including the research methods.

Recesja wywołana pandemią COVID-19 i lockdownem była głównym czynnikiem wpływającym na sytuację finansową jednostek samorządu terytorialnego w 2020 r. Epidemia postawiła przed jednostkami samorządu terytorialnego szereg wyzwań i spowodowała konieczność szybkiego dostosowania ich funkcjonowania do nowej rzeczywistości. Celem artykułu jest odpowiedź na pytanie w jaki sposób pandemia wpłynęła na jednostki samorządu terytorialnego (JST). Przedstawiono problemy wynikające z wprowadzanego i zmienianego w wyniku pandemii ustawodawstwa dotyczącego funkcjonowania JST, a także wpływ pandemii na ich sytuację finansową. Do oceny zmian sytuacji finansowej JST zastosowano metodę analizy porównawczej. Za podstawę analiz przyjęto wielkości dochodowe, wydatkowe oraz wynikowe za lata 2018-2020 oraz ich porównanie z planami. Głównym źródłem danych są wielkości zbiorcze dostępne na stronie www.mf.gov.pl, na które składają się zestawienia zbiorcze z wykonania budżetów JST po czterech kwartałach za lata 2018, 2019 i 2020 uzupełnione w niektórych kwestiach odwołaniem do sprawozdań z wykonania budżetów Rb27s (dochody) i Rb28s (wydatki).

New pharmacological approaches that target orexin receptors (OXRs) are being developed to treat sleep disorders such as insomnia and narcolepsy, with fewer side effects than existing treatments. Orexins are neuropeptides that exert excitatory effects on postsynaptic neurons via the OXRs, and are important in regulating sleep/wake states. To date, there are three FDA-approved dual orexin receptor antagonists for the treatment of insomnia, and several small molecule oral OX2R (OXR type 2) agonists are in the pipeline for addressing the orexin deficiency in narcolepsy. To find new hypnotics and psychostimulants, rodents have been the model of choice, but they are costly and have substantially different sleep patterns to humans. As an alternative model, zebrafish larvae that like humans are diurnal and show peak daytime activity and rest at night offer several potential advantages including the ability for high throughput screening. To pharmacologically validate the use of a zebrafish model in the discovery of new compounds, we aimed in this study to evaluate the functionality of a set of known small molecule OX2R agonists and antagonists on human and zebrafish OXRs and to probe their effects on the behavior of zebrafish larvae. To this end, we developed an in vitro IP-One Homogeneous Time Resolved Fluorescence (HTRF) immunoassay, and in vivo locomotor assays that record the locomotor activity of zebrafish larvae under physiological light conditions as well as under dark-light triggers. We demonstrate that the functional IP-One test is a good predictor of biological activity in vivo. Moreover, the behavioral data show that a high-throughput assay that records the locomotor activity of zebrafish throughout the evening, night and morning is able to distinguish between OXR agonists and antagonists active on the zebrafish OXR. Conversely, a locomotor assay with alternating 30 min dark-light transitions throughout the day is not able to distinguish between the two sets of compounds, indicating the importance of circadian rhythm to their pharmacological activity. Overall, the results show that a functional IP-one test in combination with a behavioral assay using zebrafish is well-suited as a discovery platform to find novel compounds that target OXRs for the treatment of sleep disorders.

Tuberous sclerosis complex (TSC) is a multisystem genetic disorder caused by pathogenic variants in TSC1 and TSC2 genes. TSC patients present with seizures and brain abnormalities such as tubers and subependymal giant cells astrocytoma (SEGA). Despite common molecular and clinical features, the severity of the disease varies greatly, even intrafamilially. The second hit hypothesis suggests that an additional, inactivating mutation in the remaining functional allele causes a more severe phenotype and therefore explains the phenotypic variability. Recently, second hit mutations have been detected frequently in mTORopathies. To investigate the pathophysiological effects of second hit mutations, several mouse models have been developed. Here, we opted for a double mutant zebrafish model that carries a LOF mutation both in the tsc2 and the depdc5 gene. To the best of our knowledge, this is the first time a second-hit model has been studied in zebrafish. Significantly, the DEP domain-containing protein 5 (DEPDC5) gene has an important role in the regulation of mTORC1, and the combination of a germline TSC2 and somatic DEPDC5 mutation has been described in a TSC patient with intractable epilepsy. Our depdc5−/−x tsc2−/− double mutant zebrafish line displayed greatly increased levels of mammalian target of rapamycin (mTORC1) activity, augmented seizure susceptibility, and early lethality which could be rescued by rapamycin. Histological analysis of the brain revealed ventricular dilatation in the tsc2 and double homozygotes. RNA-sequencing showed a linear relation between the number of differentially expressed genes (DEGs) and the degree of mTORC1 hyperactivity. Enrichment analysis of their transcriptomes revealed that many genes associated with neurological developmental processes were downregulated and mitochondrial genes were upregulated. In particular, the transcriptome of human SEGA lesions overlapped strongly with the double homozygous zebrafish larvae. The data highlight the clinical relevance of the depdc5−/− x tsc2−/− double mutant zebrafish larvae that showed a more severe phenotype compared to the single mutants. Finally, analysis of gene-drug interactions identified interesting pharmacological targets for SEGA, underscoring the value of our small zebrafish vertebrate model for future drug discovery efforts.

Objective: Preclinical models of seizures and epilepsy in rodents contributed substantially to the discovery of currently available antiseizure medications. These were also broadly used for investigation of processes of epileptogenesis. Nevertheless, rodent models pose some limitations, thus, new models using alternative species are in high demand. The aim of this study was to describe a new model of seizures/epilepsy induced by the cholinomimetic agent, pilocarpine (PILO), in larval zebrafish. Methods: Local field potential (LFP) recordings were conducted to analyze electroencephalographic discharges and correlate it with larval behavior. Hematoxylin and eosin (H&E) staining, as well as TUNEL staining were performed to analyze morphology and apoptosis, respectively. Real-time quantitative polymerase chain reaction (qRT-PCR) was undertaken for gene expression analysis. Results: Acute exposure to PILO, in a concentration-dependent manner, induces electroencephalographic discharges in larval zebrafish, which behaviorally manifest as decreased locomotion and moving time, but enhanced movement velocity. The PILO-induced seizure-like activity is behaviorally distinct from this induced by the application of chemoconvulsant pentylenetetrazole (PTZ). Zebrafish larvae previously exposed to PILO (2 h), after a washing out period, exhibit spontaneous, unprovoked discharges and apoptotic changes in their brains. Significance: Here, we comprehensively investigated a new model of PILOinduced seizures/epilepsy in larval zebrafish. We propose that this model may be used to study epileptogenesis and for antiseizure drug screening purposes. [ABSTRACT FROM AUTHOR]

Synaptic vesicle glycoprotein 2A (SV2A) regulates action potential-dependent neurotransmitter release and is commonly known as the primary binding site of an approved anti-epileptic drug, levetiracetam. Although several rodent knockout models have demonstrated the importance of SV2A for functional neurotransmission, its precise physiological function and role in epilepsy pathophysiology remains to be elucidated. Here, we present a novel sv2a knockout model in zebrafish, a vertebrate with complementary advantages to rodents. We demonstrated that 6 days post fertilization homozygous sv2a–/– mutant zebrafish larvae, but not sv2a+/– and sv2a+/+ larvae, displayed locomotor hyperactivity and spontaneous epileptiform discharges, however, no major brain malformations could be observed. A partial rescue of this epileptiform brain activity could be observed after treatment with two commonly used anti-epileptic drugs, valproic acid and, surprisingly, levetiracetam. This observation indicated that additional targets, besides Sv2a, maybe are involved in the protective effects of levetiracetam against epileptic seizures. Furthermore, a transcriptome analysis provided insights into the neuropathological processes underlying the observed epileptic phenotype. While gene expression profiling revealed only one differentially expressed gene (DEG) between wildtype and sv2a+/– larvae, there were 4386 and 3535 DEGs between wildtype and sv2a–/–, and sv2a+/– and sv2a–/– larvae, respectively. Pathway and gene ontology (GO) enrichment analysis between wildtype and sv2a–/– larvae revealed several pathways and GO terms enriched amongst up- and down-regulated genes, including MAPK signaling, synaptic vesicle cycle, and extracellular matrix organization, all known to be involved in epileptogenesis and epilepsy. Importantly, we used the Connectivity map database to identify compounds with opposing gene signatures compared to the one observed in sv2a–/– larvae, to finally rescue the epileptic phenotype. Two out of three selected compounds rescued electrographic discharges in sv2a–/– larvae, while negative controls did not. Taken together, our results demonstrate that sv2a deficiency leads to increased seizure vulnerability and provide valuable insight into the functional importance of sv2a in the brain in general. Furthermore, we provided evidence that the concept of connectivity mapping represents an attractive and powerful approach in the discovery of novel compounds against epilepsy.

Epilepsy is a common disorder of the brain characterized by spontaneous recurrent seizures, which develop gradually during a process called epileptogenesis. The mechanistic processes underlying the changes of brain tissue and networks toward increased seizure susceptibility are not fully understood. In rodents, injection of kainic acid (KA) ultimately leads to the development of spontaneous epileptic seizures, reflecting similar neuropathological characteristics as seen in patients with temporal lobe epilepsy (TLE). Although this model has significantly contributed to increased knowledge of epileptogenesis, it is technically demanding, costly to operate and hence not suitable for high-throughput screening of anti-epileptic drugs (AEDs). Zebrafish, a vertebrate with complementary advantages to rodents, is an established animal model for epilepsy research. Here, we generated a novel KA-induced epilepsy model in zebrafish larvae that we functionally and pharmacologically validated. KA was administered by pericardial injection at an early zebrafish larval stage. The epileptic phenotype induced was examined by quantification of seizure-like behavior using automated video recording, and of epileptiform brain activity measured via local field potential (LFP) recordings. We also assessed GFP-labeled GABAergic and RFP-labeled glutamatergic neurons in double transgenic KA-injected zebrafish larvae, and examined the GABA and glutamate levels in the larval heads by liquid chromatography with tandem mass spectrometry detection (LC-MS/MS). Finally, KA-injected larvae were exposed to five commonly used AEDs by immersion for pharmacological characterization of the model. Shortly after injection, KA induced a massive damage and inflammation in the zebrafish brain and seizure-like locomotor behavior. An abnormal reorganization of brain circuits was observed, a decrease in both GABAergic and glutamatergic neuronal population and their associated neurotransmitters. Importantly, these changes were accompanied by spontaneous and continuous epileptiform brain discharges starting after a short latency period, as seen in KA rodent models and reminiscent of human pathology. Three out of five AEDs tested rescued LFP abnormalities but did not affect the seizure-like behavior. Taken together, for the first time we describe a chemically-induced larval zebrafish epilepsy model offering unique insights into studying epileptogenic processes in vivo and suitable for high-throughput AED screening purposes and rapid genetic investigations.

tRNAs are linked with their cognate amino acid by aminoacyl tRNA synthetases (ARS). Here, the authors report a developmental encephalopathy associated with biallelic VARS variants (valyl-tRNA synthetase) that lead to loss of function, as determined by several in vitro assays and a vars knockout zebrafish model.

Introduction. Warthin tumors are the second most common benign tumor of the salivary gland, located mainly in the parotid glands, sometimes bilaterally. The main risk factor is nicotine addiction. The aim of the study was to present our own experience in the diagnosis and treatment of salivary gland neoplasm, and to analyze the risk factors for the development of Warthin tumors. Materials and methods. The study group consisted of 55 patients operated on with Warthin tumors (between 2009 and 2023). 55 control individuals with no Warthin tumors were recruited. The patients underwent a retrospective analysis of risk factors for head and neck cancer. Results. Warthin tumor patients reported salivary gland diseases, such as urolithiasis, inflammation, dry mouth, nicotine addiction, and chronic diseases, such as hypercholesterolemia. In 83% of cases of fine-needle aspiration biopsy (FNAB) of Warthin tumors, results were confirmed by postoperative histopathological diagnosis. The therapy included extracapsular tumor removal, partial parotidectomy with preservation of the facial nerve, and removal of the submandibular gland. Postoperative complications were a cutaneous fistula and paresis of the marginal branch of the facial nerve. Conclusions. The study confirmed that nicotine addiction (smoking duration and number of cigarettes smoked per day) was the main risk factor for developing Warthin tumors. An increase in body-mass index (BMI), hypercholesterolemia, salivary gland diseases, and dry mouth symptoms manifested Warthin tumors. FNAB, ultrasonography (USG) and computer tomography (CT) or magnetic resonance imaging (MRI) with contrast were essential in the diagnostics and planning therapeutic strategy. The main treatment used in the clinic was extracapsular tumor removal. [ABSTRACT FROM AUTHOR]

Tuberous sclerosis complex (TSC) is a rare, genetic disease caused by loss-of-function mutations in either TSC1 or TSC2. Patients with TSC are neurologically characterized by the presence of abnormal brain structure, intractable epilepsy and TSC-associated neuropsychiatric disorders. Given the lack of effective long-term treatments for TSC, there is a need to gain greater insight into TSC-related pathophysiology and to identify and develop new treatments.In this work we show that homozygous tsc2−/− mutant zebrafish larvae, but not tsc2+/− and WT larvae, display enlarged brains, reduced locomotor behavior and epileptiform discharges at 7dpf. In addition, we pharmacologically validated the TSC model by demonstrating the dramatic rescue effect of pericardially injected rapamycin, a well-known mTOR inhibitor, on selected behavioral read-outs and at the molecular level.By means of trancriptome profiling we also acquired more insight into the neuropathology of TSC, and as a result were able to highlight possible new treatment targets. The gene expression profiles of WT and tsc2+/− larvae revealed 117 differentially expressed genes (DEGs), while between WT and tsc2−/− larvae and tsc2+/− and tsc2−/− larvae there were 1414 and 1079 DEGs, respectively. Pathway enrichment analysis from the WT and tsc2−/− DEGs, identified 14 enriched pathways from the up-regulated genes and 6 enriched pathways from the down-regulated genes. Moreover, genes related to inflammation and immune response were up-regulated in the heads of tsc2−/− larvae, in line with the findings in human brain tissue where inflammatory and immune responses appear to be major hallmarks of TSC.Taken together, our phenotypic, transcriptomic and pharmacological analysis identified the tsc2−/− zebrafish as a preclinical model that mirrors well aspects of the human condition and delineated relevant TSC-related biological pathways. The model may be of value for future TSC-related drug discovery and development programs.

Abstract Epilepsy is a chronic brain disorder characterized by recurrent seizures due to abnormal, excessive and synchronous neuronal activities in the brain. It affects approximately 65 million people worldwide, one third of which are still estimated to suffer from refractory seizures. Glutamic acid decarboxylase (GAD) that converts glutamate into GABA is a key enzyme in the dynamic regulation of neural network excitability. Importantly, clinical evidence shows that lowered GAD activity is associated with several forms of epilepsy which are often treatment resistant. In the present study, we synthetized and explored the possibility of using ethyl ketopentenoate (EKP), a lipid-permeable GAD-inhibitor, to induce refractory seizures in zebrafish larvae. Our results demonstrate that EKP evoked robust convulsive locomotor activities, excessive epileptiform discharges and upregulated c-fos expression in zebrafish. Moreover, transgenic animals in which neuronal cells express apoaequorin, a Ca2+-sensitive bioluminescent photoprotein, displayed large luminescence signals indicating strong EKP-induced neuronal activation. Molecular docking data indicated that this proconvulsant activity resulted from the direct inhibition of both gad67 and gad65. Limited protective efficacy of tested anti-seizure drugs (ASDs) demonstrated a high level of treatment resistance of EKP-induced seizures. We conclude that the EKP zebrafish model can serve as a high-throughput platform for novel ASDs discovery.

The article examines the process of longing for Odessa-Mama on the basis of Michale Boganim’s Odessa…Odessa!, Liev Schreiber’s Everything is illuminated and Sergey Ursulyak’s Liquidation. In this paper particular attention is devoted to the terms Odessan myth and Odessan text that are key elements of analysed movies and contribute to the nostalgic portrayal of Odessa.

Pleomorphic adenomas (PAs) and Warthin tumors (WTs) are the most common benign tumors that occur in the salivary gland. PA has a tendency towards malignant transformation. Thus, searching for new methods to diagnose salivary gland tumors and treatment is important. The members of the class O forehead box transcription factor (FOXO3) and mitogen-activated protein kinase 1 (MAPK1) genes participate in the cellular processes, including in cell proliferation. The aim of this study was to analyze these genes' expression in the salivary gland tissues and in salivary gland tumors. The study group consisted of 50 patients treated for salivary gland tumors. For genetic tests, fresh samples of tissue collected during the surgery were used. The expression levels of the FOXO3 and MAPK1 genes were statistically significantly lower in PA tissue than in normal salivary gland tissue and WT tissue. This research revealed that the FOXO3 and MAPK1 genes are present in benign salivary gland tumors and also indicated a role of these genes in the development of benign salivary gland tumors. The cause of the development of pleomorphic adenomas may be apoptotic disorder and the activation of the inflammatory process. The examined genes may have potential to be new therapeutic targets for the treatment of pleomorphic adenomas. [ABSTRACT FROM AUTHOR]

Mutations in aggregation prone regions of recombinant proteins often improve their solubility, although they might cause negative effects on their structure and function. Here, the authors identify proteins hot spots that can be exploited to optimize solubility without compromising stability.

In the beginning of the last decade of XX century regional policy in Poland constituted a small part of all the actions under the public policy. In the second decade of XXI century, regional policy become the main policy realizing the development policy in Poland. During this twenty years the system of regional policy has changed a lot. The changes were mainly caused by transformation of polish economy from the central planning economy to market economy as well as the accession to European Union. The research concentrates on changes of the system of regional policy. The article starts with presentation of research and conceptual framework. It continues with the short description of the historical changes of regional policy in Poland. Later the evolution of the system is described. The changes are presented in three main categories: the changes in approach, strategic planning and institutional and administrative system. The author also presents the advantages and disadvantages of the changes as well as the practical problems and costs of regional policy system modification.