Monika Herian, MSc - 2022

DOCTORAL DISSERTATION

Pharmacological properties of the hallucinogenic compound 25I-NBOMe

Własności farmakologiczne związku halucynogennego 25I-NBOMe

Monika Herian

Department of Pharmacology, Maj Institute of Pharmacology, Polish Academy of Sciences in Krakow

Nowadays, one of the important problems of the world is the recreational use of new psychoactive substances (NPS), also known as ‘legal highs’. The uncontrolled increase in the number of NPS’ poses an extremely high risk of permanent damage to the health of users. Phenylethylamine derivatives, including synthetic tryptamines, belong to the group of hallucinogens. The popularity of these substances has remained constant for years and new synthesized hallucinogenic compounds contribute to serious poisoning, often fatal. In recent years, phenylethylamine derivatives so-called NBOMe compounds (N-(2-methoxybenzyl)phenylethylamine) have gained popularity. Initially, these compounds were considered as a legal alternative to lysergic acid diethylamide (LSD). The peripheral effects resulting from their intake are: hallucinations, aggression, delusional psychosis, convulsions, hyperthermia, numbness in parts of the body, cognitive impairment, rhabdomyolysis and vasoconstriction. NBOMe compounds have been found to be potent at low microgram doses and many deaths have been reported following their overdose. Despite the popularity of NBOMe’s on the legal highs market, knowledge about their pharmacological properties is limited. One of the most popular representative of this series is 4-iodo-2,5-dimethoxy-N-(2- methoxybenzyl) phenylethylamine, in short 25I-NBOMe. In order to understand the 25I-NBOMe mechanism of action, the presented doctoral dissertation aimed to characterize the effect of the 25I-NBOMe compound on brain neurotransmission, animal behavior and neurotoxicity. The studies were carried out in a single (0.3, 1, 3 and 10 mg/kg) and multiple (7 days) administration regimen with a dose of 0.3 mg/kg on male Wistar Han rats. The effect of 25I-NBOMe on glutamate, serotonin, dopamine and acetylcholine release was analyzed through brain microdialysis in freely moving rats. Subsequently, the ability of 25I-NBOMe to induce hallucinogenic activity was measured by episodes of head and body shaking (WDS) counts, its effect on motor and exploratory activity as well as on cognitive functions were examined in the open field test (OF) and in the new object recognition test (NOR), respectively. Moreover, anxiety behavior under the influence of 25I-NBOMe was investigated in the light/dark box (LDB) test. The neurotoxic effect of 25I-NBOMe was assessed by measuring the oxidative DNA damage with the comet assay, the effect on the nerve cells number was determined by immunohistochemistry and the presence of an apoptotic signal was confirmed by the TUNEL assay. Moreover, the amount of 25I-NBOMe in the rat brain structures was also studied using mass spectrometry. Single administration of 25I-NBOMe increased the release of glutamate in the frontal cortex (FCx) and induced hallucinogenic activity of 25I-NBOMe which is mainly related to the activation of cortical serotonin 5-HT2A receptors. The dose-effect dependence in the glutamate release was non-linear and ‘U’-shaped with the intermediate dose of 3 mg/kg having the least effect. A similar effect was obtained in inducing hallucinogenic activity. In contrast, the opposite effect (inverted ‘U’-shaped) was observed in the influence of 25I-NBOMe on monoamine release. These relationships may be related to the modulatory role of the serotonin 5-HT2C and 5-HT1A receptors stimulated by 25I-NBOMe. After repeated administration of 25I-NBOMe at a dose of 0.3 mg/kg the effect on glutamate and monoamine release in the FCx was reduced suggesting the development of tolerance due to 5-HT2A receptor desensitization. The weakened pyramidal cells stimulation after repeated administrations of 25I-NBOMe could result in decreased responses of ascending dopamine and serotonin pathways to FCx from the ventral tegmental area (VTA) and raphe nuclei (NR), respectively, which may be important in the release of other neurotransmitters in the striatum (STR) and nucleus accumbens septi (NAS). Inhibition of motor activity in the OF test and cognitive impairment in the NOR test after multiple administrations of 25I-NBOMe may be due to changes in 5-HT2A receptor signaling and attenuation of glutamate release in the FCx. The prolonged time spent in the dark zone after single and repeated administration of 25I-NBOMe in the LDB test suggest an anxiogenic effect of the compound as a result of decreased serotonin release in the FCx. On the other hand, the increase in vertical and stereotypic activity after multiple administration may be the result of increased dopamine release in the STR and NAS. Both single and multiple injections of 25I-NBOMe induced DNA damage in the rat FCx and hippocampus (HP), possibly caused by oxidative stress. These impairments may have contributed to a reduction in the number of glial cells in the FCx regions, however, they did not induce cell death by apoptosis. In the study of the 25I-NBOMe distribution in brain tissue the drug was found to be accumulated after multiple doses, mainly in the FCx and HP, which may be an additional factor having an influence on the DNA damage and cells survival. In conclusion, the results of this doctoral dissertation, composed of a series of the four original articles, indicate that 25I-NBOMe significantly affects the brain neurotransmission and animals behavior through the cortical serotonin 5-HT2A and 5-HT2C receptors. In addition, 25I-NBOMe exhibits neurotoxic potential as evidenced by nuclear DNA damage, changes in glial cell numbers and significant compound accumulation after multiple administrations.