Procognitive, Anxiolytic, and Antidepressant-like Properties of Hyperoside and Protocatechuic Acid Corresponding with the Increase in Serum Serotonin Level after Prolonged Treatment in Mice.

Two polyphenols-hyperoside (HYP) and protocatechuic acid (PCA) were reported to exert antidepressant activity in rodents after acute treatment. Our previous study also showed that this activity might have been influenced by the monoaminergic system and the upregulation of the brain-derived neurotropic factor (BDNF) level. A very long-term pharmacological therapy is required for the treatment of a patient with depression. The repetitive use of antidepressants is recognized to impact the brain structures responsible for regulating both emotional and cognitive behaviors. Thus, we investigated the antidepressant, anxiolytic, and procognitive effects of HYP and PCA in mice after acute and prolonged treatment (14 days). Both polyphenols induced an anxiogenic-like effect after acute treatment, whereas an anxiolytic effect occurred after repetitive administration. PCA and HYP showed procognitive effects when they were administered acutely and chronically, but it seems that their influence on long-term memory was stronger than on short-term memory. In addition, the preset study showed that the dose of 7.5 mg/kg of PCA and HYP was effective in counteracting the effects of co-administered scopolamine in the long-term memory impairment model induced by scopolamine. Our experiments revealed the compounds have no affinity for 5-HT1A and 5-HT2A receptors, whereas a significant increase in serum serotonin level after prolonged administration of PCA and HYP at a dose of 3.75 mg/kg was observed. Thus, it supports the involvement of the serotonergic system in the polyphenol mechanisms. These findings led us to hypothesize that the polyphenols isolated from Impatiens glandulifera can hold promise in treating mental disorders with cognitive dysfunction. Consequently, extended studies are necessary to delve into their pharmacological profile.

Research in the Field of Drug Design and Development.

The processes used by academic and industrial scientists to discover new drugs have recently experienced a true renaissance, with many new and exciting techniques being developed over the past 5-10 years alone. Drug design and discovery, and the search for new safe and well-tolerated compounds, as well as the ineffectiveness of existing therapies, and society's insufficient knowledge concerning the prophylactics and pharmacotherapy of the most common diseases today, comprise a serious challenge. This can influence not only the quality of human life, but also the health of whole societies, which became evident during the COVID-19 pandemic. In general, the process of drug development consists of three main stages: drug discovery, preclinical development using cell-based and animal models/tests, clinical trials on humans and, finally, forward moving toward the step of obtaining regulatory approval, in order to market the potential drug. In this review, we will attempt to outline the first three most important consecutive phases in drug design and development, based on the experience of three cooperating and complementary academic centers of the Visegrád group; i.e., Medical University of Lublin, Poland, Masaryk University of Brno, Czech Republic, and Comenius University Bratislava, Slovak Republic.

Inhibitors of Endocannabinoids' Enzymatic Degradation as a Potential Target of the Memory Disturbances in an Acute N-Methyl-D-Aspartate (NMDA) Receptor Hypofunction Model of Schizophrenia in Mice.

Treating schizophrenia with the available pharmacotherapy is difficult. One possible strategy is focused on the modulation of the function of the endocannabinoid system (ECS). The ECS is comprised of cannabinoid (CB) receptors, endocannabinoids and enzymes responsible for the metabolism of endocannabinoids (fatty acid hydrolase (FAAH) and monoacylglycerol lipase (MAGL)). Here, the aim of the experiments was to evaluate the impact of inhibitors of endocannabinoids' enzymatic degradation in the brain: KML-29 (MAGL inhibitor), JZL-195 (MAGL/FAAH inhibitor) and PF-3845 (FAAH inhibitor), on the memory disturbances typical for schizophrenia in an acute N-methyl-D-aspartate (NMDA) receptor hypofunction animal model of schizophrenia (i.e., injection of MK-801, an NMDA receptor antagonist). The memory-like responses were assessed in the passive avoidance (PA) test. A single administration of KML-29 or PF-3845 had a positive effect on the memory processes, but an acute administration of JZL-195 impaired cognition in mice in the PA test. Additionally, the combined administration of a PA-ineffective dose of KML-29 (5 mg/kg) or PF-3845 (3 mg/kg) attenuated the MK-801-induced cognitive impairment (0.6 mg/kg). Our results suggest that the indirect regulation of endocannabinoids' concentration in the brain through the use of selected inhibitors may positively affect memory disorders, and thus increase the effectiveness of modern pharmacotherapy of schizophrenia.

Characteristics of Seltorexant-Innovative Agent Targeting Orexin System for the Treatment of Depression and Anxiety.

Twenty-five years have passed since the discovery of the orexin system, during which time we have learned more and more about it. A number of studies have been conducted showing the role of the orexin system in insomnia, as well as its potential use in the treatment of obesity and depression. In this review, we present the role of the orexin system in the development of depressive illness and show the characteristics of seltorexant, a potential drug for the treatment of depression. This review describes the structure and synthesis of the compound as well as its pharmacodynamics and pharmacokinetics. Pre-clinical and clinical studies are also described, including side effects. There is evidence that the use of seltorexant is considered safe, with no clear or major clinically significant side effects, which makes it a promising candidate for the treatment of depression and anxiety disorders.

Recent Studies on the Development of Nicotine Abuse and Behavioral Changes Induced by Chronic Stress Depending on Gender.

Nowadays, stressful situations are an unavoidable element of everyday life. Stressors activate a number of complex mental and physiological reactions in the organism, thus affecting the state of health of an individual. Stress is the main risk factor in the development of mental disorders, such as depression and other disorders developing as a result of addiction. Studies indicate that women are twice as likely as men to develop anxiety, depression and therefore addiction, e.g., to nicotine. Even though the data presented is indicative of significant differences between the sexes in the prevalence of these disorders, the majority of preclinical animal models for investigating stress-induced disorders use predominantly male subjects. However, the recent data indicates that this type of studies has also been launched in female rodents. Therefore, conducting research on both sexes allows for a more accurate understanding and assessment of the impact of stress on stress-induced behavioral, peripheral and molecular changes in the body and brain. In this manuscript we have gathered the data from 41 years (from 1981-2022) on the influence of stress on the development of depression and nicotine addiction in both sexes.

Funnel metadynamics and behavioral studies reveal complex effect of D2AAK1 ligand on anxiety-like processes.

Anxiety is a troublesome symptom for many patients, especially those suffering from schizophrenia. Its regulation involves serotonin receptors, targeted e.g. by antipsychotics or psychedelics such as LSD. 5-HT2A receptors are known for an extremely long LSD residence time, enabling minute doses to exert a long-lasting effect. In this work, we explore the changes in anxiety-like processes induced by the previously reported antipsychotic, D2AAK1. In vivo studies revealed that the effect of D2AAK1 on the anxiety is mediated through serotonin 5-HT1A and 5-HT2A receptors, and that it is time-dependent (anxiogenic after 30 min, anxiolytic after 60 min) and dose-dependent. The funnel metadynamics simulations suggest complicated ligand-5HT2AR interactions, involving an allosteric site located under the third extracellular loop, which is a possible explanation of the time-dependency. The binding of D2AAK1 at the allosteric site results in a broader opening of the extracellular receptor entry, possibly altering the binding kinetics of orthosteric ligands.

Promising Advances in Pharmacotherapy for Patients with Spinal Cord Injury-A Review of Studies Performed In Vivo with Modern Drugs.

Spinal cord injury (SCI) is a pathological neurological condition that leads to significant motor dysfunction. It is a condition that occurs as a result of tragic accidents, violent acts, or as a consequence of chronic diseases or degenerative changes. The current treatments for patients with SCI have moderate efficacy. They improve the quality of life of patients, but they are still doomed to long-term disability. In response to the modern directions of research on possible therapeutic methods that allow for the recovery of patients with SCI, a scientific review publication is needed to summarize the recent developments in this topic. The following review is focused on the available pharmacological treatments for SCIs and the problems that patients face depending on the location of the injury. In the following review, the research team describes problems related to spasticity and neuropathic pain; possible therapeutic pathways are also described for neuroprotection and the improvement of neurotransmission within the injured spinal cord, and the review focuses on issues related to oxidative stress.

New Atypical Antipsychotics in the Treatment of Schizophrenia and Depression.

Schizophrenia and depression are heterogeneous disorders. The complex pathomechanism of the diseases imply that medication responses vary across patients. Many psychotropic drugs are available but achieving optimal therapeutic effect can be challenging. The evidence correlates well with clinical observations, suggesting that new atypical antipsychotic drugs are effective against negative and cognitive symptoms of schizophrenia, as well as against affective symptoms observed in depression. The purpose of this review presents the background and evidence for the use of the new second/third-generation antipsychotics (aripiprazole, cariprazine, lurasidone, asenapine, brexpiprazole, lumateperone, pimavanserin) in treatment of schizophrenia and depression. We have first provided a brief overview of the major neurobiological underpinnings of schizophrenia and depression. We then shortly discuss efficacy, safety and limitations of ongoing pharmacotherapy used in depression and schizophrenia. Mainly, we have focused this review on the therapeutic potential of new atypical antipsychotic drugs-currently existing-to be effective in psychotic, as well as in affective disorders.

A Comprehensive Review of Daridorexant, a Dual-Orexin Receptor Antagonist as New Approach for the Treatment of Insomnia.

Insomnia affects 4.4-4.8% of the world's population, but because the effect of hypnotic drugs is limited and may cause harmful side-effects, scientists are turning their attention to developing drugs that act on the orexin system. Daridorexant, a selective dual-orexin receptor antagonist (DORA), has exhibited promising results in both animal and human studies. Its activity was evaluated based on the physiology-based pharmacodynamic and pharmacokinetic model. The use of daridorexant is considered safe, with no clinically significant side-effects including deprivation of next-morning residual effects. In this manuscript we conducted a comprehensive review of daridorexant including pharmacodynamics, animal and human research, pharmacokinetics and safety.

The Influence of CB2-Receptor Ligands on the Memory-Related Responses in Connection with Cholinergic Pathways in Mice in the Passive Avoidance Test.

Background: Dysfunction of the cholinergic system is associated with the development of Alzheimer's disease (AD). One of the new possible strategies for the pharmacological modulation of memory-related problems typical of AD, is connected with the endocannabinoid system (ECS) and the cannabinoid (CB: CB1 and CB2) receptors. Methods: The aim of the study was to determine the influence of the selective CB2 receptor ligands: agonist (JWH 133) and antagonist (AM 630) on different stages of memory and learning in mice, in the context of their interaction with cholinergic pathways. To assess and understand the memory-related effects in mice we used the passive avoidance (PA) test. Results: We revealed that co-administration of non-effective dose of JWH 133 (0.25 mg) or AM 630 (0.25 mg/kg) with the non-effective dose of cholinergic receptor agonist - nicotine (0.05 mg/kg) enhanced cognition in the PA test in mice; however, an acute injection of JWH 133 (0.25 mg/kg) or AM 630 (0.25 mg/kg) had no influence on memory enhancement induced by the effective dose of nicotine (0.1 mg/kg). Co-administration of JWH 133 (0.25 mg) or AM 630 (0.25 mg/kg) with the effective dose of the cholinergic receptor antagonist scopolamine (1 mg/kg) attenuated the scopolamine-induced memory impairment in the PA test in mice. Conclusion: Our experiments have shown that CB2 receptors participate in the modulation of memory-related responses, especially those in which cholinergic pathways are implicated.

Cannabidiol Attenuates MK-801-Induced Cognitive Symptoms of Schizophrenia in the Passive Avoidance Test in Mice.

Schizophrenia is a chronic mental disorder that disturbs feelings and behavior. The symptoms of schizophrenia fall into three categories: positive, negative, and cognitive. Cognitive symptoms are characterized by memory loss or attentional deficits, and are especially difficult to treat. Thus, there is intense research into the development of new treatments for schizophrenia-related responses. One of the possible strategies is connected with cannabidiol (CBD), a cannabinoid compound. This research focuses on the role of CBD in different stages of memory (acquisition, consolidation, retrieval) connected with fear conditioning in the passive avoidance (PA) learning task in mice, as well as in the memory impairment typical of cognitive symptoms of schizophrenia. Memory impairment was provoked by an acute injection of the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (animal model of schizophrenia). Our results revealed that an acute injection of CBD (30 mg/kg; intraperitoneally (i.p.) improved all phases of long-term fear memory in the PA test in mice. Moreover, the acute injection of non-effective doses of CBD (1 or 5 mg/kg; i.p.) attenuated the memory impairment provoked by MK-801 (0.6 mg/kg; i.p.) in the consolidation and retrieval stages of fear memory, but not in the acquisition of memory. The present findings confirm that CBD has a positive influence on memory and learning processes in mice, and reveals that this cannabinoid compound is able to attenuate memory impairment connected with hypofunction of glutamate transmission in a murine model of schizophrenia.

Type I and type II positive allosteric modulators of α7 nicotinic acetylcholine receptors induce antidepressant-like activity in mice by a mechanism involving receptor potentiation but not neurotransmitter reuptake inhibition. Correlation with mTOR intracellular pathway activation.

The aim of this study is to determine whether type I and type II positive allosteric modulators (PAMs) of α7 nicotinic acetylcholine receptors (nAChRs) induce antidepressant-like activity in mice after acute, subchronic, and chronic treatments, and to assess whether α7-PAMs inhibit neurotransmitter transporters and activate mTOR (mammalian target of rapamycin) and/or ERK (extracellular signal-regulated protein kinases) signaling. The forced swim (FST) and tail suspension (TST) test results indicated that NS-1738 (type I PAM), PNU-120596 and PAM-2 (type II PAMs) induce antidepressant-like activity after subchronic treatment, whereas PAM-2 was also active after chronic treatment. Methyllycaconitine (α7-antagonist) inhibited the observed effects, highlighting the involvement of α7 nAChRs in this process. Drug interaction studies showed synergism between PAM-2 and bupropion (antidepressant), but not between PAM-2 and DMXBA (α7-agonist). The studied PAMs showed no high affinity (< 1 µM) for the human dopamine, serotonin, and noradrenaline transporters, suggesting that transporter inhibition is not the underlying mechanism for the observed activity. To assess whether mTOR and ERK signaling pathways are involved in the activity of α7-PAMs, the phosphorylation status of key signaling nodes was determined in prefrontal cortex and hippocampus from mice chronically treated with PAM-2. In conclusion, the antidepressant-like activity of type I and type II PAMs is mediated by a mechanism involving α7 potentiation but not α7 desensitization or neurotransmitter transporter blockade, and is correlated with activation of both mTOR and ERK signaling pathways. These results support the view that α7-PAMs might be clinically used to ameliorate depression disorders .

Effect of Berberine Isolated from Barberry Species by Centrifugal Partition Chromatography on Memory and the Expression of Parvalbumin in the Mouse Hippocampus Proper.

Neurodegenerative diseases associated with memory disturbances are important health issues occurring due to a prolonged life span. This article presents the results of a study targeting the emergence of a drug candidate with antiamnesic properties. The effect of berberine (BBR), an isoquinoline alkaloid isolated from the overground parts of Berberis sibirica Pall., on memory and expression of parvalbumin in the mouse hippocampus proper were determined. High-purity BBR was isolated by centrifugal partition chromatography from a methanolic extract from B. sibirica by using a methyl-tert-butyl ether and water (1:1 v/v) solvent system with 10 mmol/L of triethylamine and hydrochloric acid. In an in vivo study, we assessed the influence of the chronic administration of BBR on different stages of memory-related responses in mice. Our results indicated that the chronic administration of BBR in a higher dose (5 mg/kg) improves long-term memory acquisition in mice, as determined in the passive avoidance test. The hippocampal CA1-CA3 fields showed an increased number of parvalbumin-immunoreactive neurons (PV-IR) and nerve fibers as compared to the control. No significant changes in the dentate gyrus were observed between the groups. The HPLC-ESI-QTOF-MS/MS analysis of the biological material revealed the content of BBR as 363.4 ± 15.0 ng (4.11% of RSD) per brain, 15.06 ± 0.89 ng (5.91% of RSD) per hippocampus, and 54.45 ± 1.40 (4.05% of RSD) ng in 100 µL plasma. The study showed that BBR could be a factor influencing the expression of PV in hippocampal neurons. We speculate that BBR may modulate the level of Ca2+ in neurons and thus potentially act as a neuroprotective factor against neuronal damages.

N-(3-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}propyl)-1H-indazole-3-carboxamide (D2AAK3) as a potential antipsychotic: In vitro, in silico and in vivo evaluation of a multi-target ligand.

Schizophrenia is a mental illness of not adequately understood causes that is not satisfactorily enough treated by current antipsychotics. In search for novel potential antipsychotics we performed structure-based virtual screening aimed to identify new dopamine D2 receptor antagonists. We found compound D2AAK3 with affinity to dopamine D2 receptor of 115 nM. D2AAK3 possesses additional nanomolar or low micromolar affinity to D1, D3, 5-HT1A, 5-HT2A and 5-HT7 receptors, which makes it a good hit for further development as a multifunctional ligand. The compound has also some affinity to M1 and H1 receptors. We used homology modeling, molecular docking and molecular dynamics to study interactions of D2AAK3 with its molecular targets at the molecular level. In behavioral studies D2AAK3 decreases amphetamine-induced hyperactivity (when compared to the amphetamine-treated group) measured as spontaneous locomotor activity in mice. In addition, passive avoidance test demonstrated that D2AAK3 improves memory consolidation after acute treatment in mice. Elevated plus maze tests indicated that D2AAK3 induces anxiogenic activity 30 min after acute treatment, whereas this effect has no longer been observed 60 min after administration of the studied compound in mice.

Response of immature rats to a low dose of nanoparticulate silver: Alterations in behavior, cerebral vasculature-related transcriptome and permeability.

The increasing production and use of silver nanoparticles (AgNPs) as antimicrobial agents in medicinal and commercial products creates a substantial risk of exposure, especially for infants and children. Our current knowledge concerning the impact of AgNPs on developing brain is insufficient. Therefore we investigated the temporal profile of transcriptional changes in cellular components of the neurovascular unit in immature rats exposed to a low dose of AgNPs. The behavior of animals under these conditions was also monitored. Significant deposition of AgNPs in brain of exposed rats was identified and found to persist over the post-exposure time. Substantial changes were noted in the transcriptional profile of tight junction proteins such as occludin and claudin-5, and pericyte-related molecules such as angiopoietin-1. Moreover, downregulation of platelet-derived growth factor (PDGFß) and its receptor (PDGFßR) which constitute the main signaling pathway between endothelial cells and pericytes was observed. These were long-lasting effects, accompanied by overexpression of astroglial-specific GFAP mRNA and endothelial cell adhesion molecule, ICAM-1, involved in the pathomechanism of neuroinflammation. The profile of changes indicates that even low doses of AgNPs administered during the early stage of life induce dysregulation of neurovascular unit constituents which may lead to disintegration of the blood-brain barrier. This was confirmed by ultrastructural analysis that revealed enhanced permeability of cerebral microvessels resulting in perivascular edema. Changes in the behavior of exposed rats indicating pro-depressive and anti-anxiety impacts were also identified. The results show a high risk of using AgNPs in medical and consumer products dedicated for infants and children.

Identification of a Potent and Selective 5-HT1A Receptor Agonist with In Vitro and In Vivo Antinociceptive Activity.

Opioids are the gold standard drugs for the treatment of acute and chronic severe pain, although their serious side effects constitute a big limitation. In the search for new and safer drugs, 5-HT1AR agonists are emerging as potential candidates in pain relief therapy. In this work, we evaluated the affinity and activity of enantiomers of the two newly synthesized, potent 5-HT1AR agonists N-[(2,2-diphenyl-1,3-dioxolan-4-yl)methyl]-2-[2-(pyridin-4-yl)phenoxy]ethan-1-ammonium hydrogenoxalate (rac-1) and N-((2,2-diphenyl-1,3-dioxolan-4-yl)methyl)-2-(2-(1-methyl-1H-imidazol-5-yl)phenoxy)ethan-1-ammonium hydrogenoxalate (rac-2) in vitro and in vivo. The role of chirality in the interaction with 5-HT1AR was evaluated by molecular docking. The activity of the rac-1 was tested in mouse models of acute pain (hot plate) and severe tonic nociceptive stimulation (intraplantar formalin test). Rac-1 was active in the formalin test with a reduction in paw licking in both phases at 10 mg/kg, and its effect was abolished by the selective 5-HT1AR antagonist, WAY-100635. The eutomer (S)-1, but not the racemate, was active during the hot plate test at 10 and 20 mg/kg, and this effect was abolished by 30 min treatment with WAY-100635 at 30 min. Similarly to 8-OH-DPAT, (S)-1 evoked a slow outward current and depressed spontaneous glutamatergic transmission in superficial dorsal horn neurons, more effectively than rac-1. The eutomer (S)-1 showed promising developability properties, such as high selectivity over 5-HT subtypes, no interaction with the µ receptors, and low hepato- and cardiotoxicity. Therefore, (S)-1 may represent a potential candidate for the treatment of acute and chronic pain without having the adverse effects that are commonly associated with the classic opioid drugs.

The Antipsychotic D2AAK1 as a Memory Enhancer for Treatment of Mental and Neurodegenerative Diseases.

The treatment of memory impairments associated with the central nervous system diseases remains an unmet medical need with social and economic implications. Here we show, that a multi-target ligand of aminergic G protein-coupled receptors with antipsychotic activity in vivo (D2AAK1) stimulates neuron growth and survival and promotes neuron integrity. We focused on the multilevel evaluation of the D2AAK1-related effects on neurons in terms of behavioral, cellular, molecular, and biochemical features in vivo and in vitro, such as memory-related responses, locomotor activity, tissue sections analysis, metabolic activity, proliferation level, neurons morphology, and proteins level involved in intracellular signaling pathways. In silico studies indicate that activation of calcium/calmodulin-dependent protein kinase I (CaMKI) may underline some of the observed activities of the compound. Furthermore, the compound increases hippocampal neuron proliferation via the activation of neurotrophic factors and cooperating signals responsible for cell growth and proliferation. D2AAK1 improves memory and learning processes in mice after both acute and chronic administration. D2AAK1 also causes an increase in the number of hippocampal pyramidal neurons after chronic administration. Because of its neuroprotective properties and pro-cognitive activity in behavioral studies D2AAK1 has the potential for the treatment of memory disturbances in neurodegenerative and mental diseases.

Bergapten Improves Scopolamine-Induced Memory Impairment in Mice via Cholinergic and Antioxidative Mechanisms.

Bergapten is a furanocoumarin naturally occurring in the Apiaceae family and it is a well-known photosensitizing agent used in photochemotherapy. In this study, we investigated the influence of bergapten on cognitive function and mechanism underlying these effects in scopolamine-induced memory impairment in male Swiss mice. The passive avoidance test was used to evaluate the efficiency of memory acquisition and consolidation. The results demonstrated that both single and repeated administration of bergapten improved not only the acquisition but also consolidation of memory. The behavioral tests showed that bergapten prevented memory impairment induced by administration of scopolamine. Observed effects may result from the inhibition of acetylcholinesterase activity in the hippocampus and prefrontal cortex. Also, bergapten caused significant anti-oxidative effects. These new findings provide pharmacological and biochemical support for the development of the coumarin's potential in cognitive deficits.

Contribution of CB2 receptors in schizophrenia-related symptoms in various animal models: Short review.

Schizophrenia is a severe and chronic mental disease with a high prevalence and a variety of symptoms. Data from behavioural studies suggest that it is rational to investigate the endocannabinoid system (ECS) and its cannabinoid receptor (CBr) because they seem to underlie susceptibility to schizophrenia, and these findings have pointed to several lines of future research. Currently, most available studies address the role of CBr type 1 in schizophrenia-like responses. Here, we present for the first time, a review that demonstrates the pivotal role of CBr type 2 in the regulation of neurobiological processes underlying cognition, psychosis- and mood-related (anxiety, depression) behaviours, all of which may be included in schizophrenia symptoms. This review is based on available evidence from the PubMed database regarding schizophrenia-like symptoms induced via CB2r modulation in various animal models. The data presented in this manuscript indicate that CB2r could be a promising new key target in the treatment of different central nervous system (CNS) disorders, which manifest as psychosis, mood-related disturbances and/or memory impairment.

Toxicity of Carlina Oxide-A Natural Polyacetylene from the Carlina acaulis Roots-In Vitro and In Vivo Study.

There are several reports indicating that the roots of the Carlina acaulis L. used to be commonly applied as a treatment measure in skin diseases and as an antiparasitic agent, starting from antiquity to the 19th century; however, nowadays, it has lost its importance. Currently, numerous studies are being conducted assessing the possibility of reintroducing C. acaulis-derived extracts to phytotherapy. Determining the safety profile of the main constituents of the plant material is crucial for achieving this goal. Here, we aimed to determine the toxicity profile of carlina oxide, one of the most abundant components of the C. acaulis root extract. We obtained the carlina oxide by distillation of C. acaulis roots in the Deryng apparatus. The purity of the standard was evaluated using GC-MS, and the identity was confirmed by IR, Raman, and NMR spectroscopy. In vitro cytotoxicity was assessed using a panel of human cell lines of skin origin, including BJ normal fibroblasts and UACC-903, UACC-647, and C32 melanoma cells. This was accompanied by an in vivo zebrafish acute toxicity test (ZFET). In vitro studies showed a toxic effect of carlina oxide, as demonstrated by an induction of apoptosis and necrosis in both normal and melanoma cells. Decreased expression of AKT kinase and extracellular signal-regulated kinase 1/2 (ERK1/2) was noted in the UACC-647 melanoma cell line. It was also observed that carlina oxide modified the expression of programmed cell death-ligand 1 (PD-L1) in tested cell lines. Carlina oxide exhibited high in vivo toxicity, with LC50 = 10.13 µg/mL upon the 96 h of exposure in the ZFET test. Here, we demonstrate that carlina oxide displays toxic effects to cells in culture and to living organisms. The data indicate that C. acaulis-based extracts considered for therapeutic use should be completely deprived of carlina oxide.