Polystyrene microplastics disrupt skeletal development in marine medaka Oryzias melastigma

Polystyrene microplastics (PS MPs) are prevalent marine pollutants with documented toxicity in aquatic organisms. However, their impacts on vertebrate skeletal development, particularly in fish, remain poorly understood. This study investigated the effects of chronic (28-day) exposure to PS MPs (7.3 μm; 0, 0.2, 2, 20 mg L^-1) on skeletal development in marine medaka (Oryzias melastigma). Fluorescence imaging and histology confirmed PS MP bioaccumulation, primarily in the intestine and gills. Behavioral analysis revealed concentration-dependent alterations, including initial hyperactivity and subsequent hypoactivity at low doses. Micro-computed tomography and histopathology demonstrated significant vertebral abnormalities, reduced bone volume/surface, and impaired mineralization at higher concentrations, while a low dose induced enhanced bone formation. Scanning electron microscopy with energy-dispersive X-ray spectroscopy and Fourier transform infrared spectroscopy revealed disrupted mineralization, decreased Ca/P ratios, and altered collagen/mineral matrix composition. Gene expression analysis indicated dysregulation of osteogenic (Runx2, Sp7, Bmp4, AcvR1b), osteoclastic (Ctsk), and morphogenic (Shh) pathways. These multi-scale findings demonstrate that PS MPs impair skeletal development through disrupted mineralization, matrix abnormalities, and inflammation. The observed non-monotonic dose responses highlight complex chronic toxicity mechanisms, posing significant ecological risks to marine fish populations.