094 | 2-octyl-2h-isothiazol-3-one and its role in zebrafish larvae neurogenesis and neuronal function

Vanessa D’Altilia¹, Ludovica Sulcanese¹, Valentina Iezzi², Silvana Zugaro², Annamaria Iannetta¹, Tommaso Silvestrini^1|3, Elisabetta Benedetti², Monia Perugini¹. | ¹Department of Bioscience and Agro-Food and Environmental Technology, University of Teramo, Teramo, Italy; ²Department of Life, Health and Environmental Sciences, University of L’Aquila, L’Aquila, Italy; ³University of Camerino, International School of Advanced Studies, Camerino, Italy.

The zebrafish (Danio rerio) larva is a well-established model for investigating developmental toxicity. Accordingly, this study employs this model to evaluate the neurotoxicity of 2-octyl-2H-isothiazol-3-one (OIT). Embryos were treated with 0.1 and 0.2 mg/L OIT during their early development and imaging, molecular and behaviour analysis were performed after the exposure. First morphometric analysis with Alcian blue-stained larvae demonstrated that OIT impairs craniofacial development, leading to a reduction in head and brain size. Moreover, the reduced and less widespread distribution of neurons throughout the brain was  investigated with the evaluation of fluorescence intensity in Tg(NeuroD:GFP) larvae with both OIT 0.1 and 0.2 mg/L treatments. At 96 hours post-fertilization (hpf) it has also been explored by RT-PCR the mRNA expression of important genes related with neuronal growth, scaffold and function such as Bdnf, Shank3a and Cyp19a. Gene expression analysis revealed that OIT disrupts neurodevelopmental gene regulation in a dose-dependent manner, impairing neurotrophic support, synaptic scaffolding, and oestrogen-mediated pathways. Then, the protein analysis made by Western blot technique of cytokines TNF-α and IL-1β indicate that OIT induces a neuroinflammatory phenotype. Furthermore, the light/dark behaviour test revealed that OIT alters sensorimotor behaviour in a dose-dependent manner. At 0.1 mg/L, OIT enhanced light-driven locomotion, whereas at 0.2 mg/L it caused generalized hypoactivity. Both doses impaired dark-phase activity, consistent with neurodevelopmental disruption of circuits controlling movement and sensory processing, such as the dopaminergic system. Besides, the protein dosage of important catecholamines highlighted dysregulation in its levels,in particular a decrease of dopamine. Overall, it turns out that OIT, an industrial biocide, has a neurotoxic role interfering with normal craniofacial and neural development, promotes a pro-inflammatory state in the brain, and leads to locomotor and sensory dysregulation. Overall, these findings indicate that OIT may represent a potential risk factor for impaired neurogenesis and neuronal function, possibly contributing to neurodegenerative pathologies such as Parkinson-like diseases.