Impact of micro-scale topographies on neurite growth and morphology

• Right-angle turns in microtopographies enhance neurite alignment compared to acute and obtuse angles. • Shorter gratings facilitate gap-crossing and neurite network formation, while longer gratings enforce linear growth, revealing a trade-off between connectivity and alignment. • Trench patterns require longer gratings than ridge patterns for effective neurite guidance, highlighting feature-specific thresholds. • Neurons prefer continuous pathways, with even small barriers at corners disrupting growth continuity. Mimicking the complex topography of the nervous system using in vitro scaffolds is a major challenge in neural tissue engineering. Limited understanding of neuronal responses to microtopographical discontinuities, such as angular turns, gaps, and misaligned pathways, hinders scaffold development. This study examines the influence of geometric discontinuities on the growth and morphology of primary cortical neurons using polydimethylsiloxane platforms. Photolithography was employed to fabricate zigzag gratings with 45°, 90°, 135° turns, featuring arm lengths of 100–200 µm, segmented gratings with lengths of 50–200 µm, misaligned gratings, and trench versus ridge patterns. Results revealed that neurite guidance highly depends on pattern geometry and continuity. Zigzag gratings with 90° turns provided the strongest directional guidance, with arm lengths of 200 µm improved alignment at 45° turns. Shorter segmented gratings of 50 and 100 µm facilitated neurite gap-crossing and network formation, while 200 µm segments promoted linear growth and maintained alignment across offsets. Ridge patterns required 200 µm segment lengths for effective guidance. Concentric square ring patterns showed neurite growth along continuous pathways, while barriers disrupted alignment. These findings contribute to the development of scaffolds that regulate neuronal growth and connectivity, potentially supporting treatments for neurological injuries and diseases.