Behavioural and Neurodevelopmental Disruptions in Zebrafish Exposed to

Abstract

Background: The rapid expansion of nanotechnology has led to the increased production and application of aluminum oxide nanoparticles (Al₂O₃ NPs) across biomedical, industrial, and environmental sectors. Despite their widespread use, concerns regarding their potential neurotoxic and developmental effects remain insufficiently addressed. Given their nanoscale size and enhanced reactivity, Al₂O₃ nanoparticles can interact with biological systems and potentially disrupt normal physiological processes. Therefore, evaluating their toxicity using relevant biological models is essential.

Objectives: This study aimed to investigate the physicochemical properties, developmental toxicity, and neurobehavioral effects of chemically synthesized Al₂O₃ nanoparticles (<20 nm) using zebrafish (Danio rerio) as a model organism.

Methods: Al₂O₃ nanoparticles were synthesized via chemical precipitation and characterized using UV–Visible spectroscopy, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), and inductively coupled plasma–atomic emission spectroscopy (ICP–AES). Developmental toxicity was assessed using the Fish Embryo Toxicity (FET) assay in accordance with OECD guidelines. Embryos were exposed to graded nanoparticle concentrations and monitored up to 120 hours post-fertilization for survival, hatching rate, heart rate, and morphological abnormalities. Neurobehavioral assessment was conducted in larvae at 5 days post-fertilization using automated video tracking, while adult zebrafish were subjected to chronic exposure (7–21 days) followed by behavioural analysis using locomotor tracking and spatial distribution (KDE) analysis.

Results: Characterization confirmed the successful synthesis of nanoscale, crystalline Al₂O₃ nanoparticles with defined morphology and elemental composition. Developmental toxicity studies revealed a clear dose-dependent effect, with concentrations ≥6 µg/mL inducing delayed hatching, reduced viability, bradycardia, and morphological abnormalities such as pericardial edema, yolk sac edema, and body curvature. Higher concentrations resulted in severe deformities and increased mortality. Behavioural analysis of larvae demonstrated reduced swimming speed, decreased exploratory behaviour, and increased inactivity, indicating impaired neuromotor function. In adult zebrafish, chronic exposure led to reduced locomotion, increased thigmotaxis, and anxiety-like behaviour. A non-monotonic dose-response pattern was observed, characterized by mild anxiety at lower concentrations, transient hyperactivity at intermediate levels, and progressive hypoactivity at higher concentrations.

Conclusion: Al₂O₃ nanoparticles induce significant developmental and neurobehavioral toxicity in zebrafish, affecting multiple biological endpoints. The findings highlight potential ecological and human health risks associated with nanoparticle exposure and emphasize the need for further mechanistic studies using molecular and multi-omics approaches. Additionally, the results support the development of safer-by-design nanomaterials to mitigate adverse biological effects