Filament Extrusion‐Based Conductive TPU Composite Scaffolds Enable Superior Neuronal Growth and Synaptic Maturation In Vitro

ABSTRACT Fused filament fabrication (FFF) three‐dimensional (3D) printing technologies offer new opportunities for fabricating customizable, low‐cost platforms for tissue engineering applications. Here, we developed and characterized 3D‐printed scaffolds using conductive thermoplastic polyurethane (cTPU) filaments and evaluated their mechanical, electrical, and biological performance in vitro. Dynamic mechanical analysis (DMA) across a range of temperatures and frequencies revealed that both TPU and cTPU exhibit temperature‐ and rate‐dependent elastic moduli, with cTPU showing enhanced mechanical stiffness due to the incorporation of conductive fillers. Electrical testing confirmed that cTPU exhibited a stable conductivity (∼1–2 mS/cm) resembling physiological conditions. Surface characterization showed that cTPU was significantly more hydrophilic and exhibited higher nanoscale roughness, both of which are favorable for cell‐material interactions. Mouse embryonic fibroblasts (MEFs) cultured on both scaffolds showed high viability (>85%) and significant proliferation. Notably, immunofluorescence analysis of cultured hippocampal neurons revealed significantly higher density of neuronal networks represented by higher microtubule‐associated protein 2 (MAP‐2)‐positive cell density, greater MAP‐2 area coverage, larger average MAP‐2 cell area, and enhanced postsynaptic density protein 95 (PSD‐95) expression on cTPU scaffolds. Together, these results demonstrate that FFF 3D‐printed cTPU platforms can support long‐term neuronal growth and synaptic maturation, offering promising applications in neural tissue modeling and bioelectronic interfaces. Practical Application: Characterizing soft viscoelastic materials whose properties strongly depend on temperature and strain rate is challenging and typically requires extensive testing across multiple conditions. Using a single‐specimen Dynamic Mechanical Analysis‐based mechanical testing method and a viscoelastic–elastic transformation that converts frequency‐domain viscoelastic measurements into elastic constants over a broad range of test conditions, validated by tensile tests, we efficiently generated reliable modulus data across a range of conditions, enhancing testing throughput without sacrificing accuracy. As a case study, we demonstrate the successful fabrication and comprehensive characterization of FDM 3D‐printed conductive TPU (cTPU) scaffolds for potential applications in neural tissue modeling and bioelectronic interfaces, with the results positioning cTPU composites as cost‐effective, tunable, cytocompatible, and electrically active platforms capable of supporting neuronal growth and function.