Nano-roughness has shown great potential in enhancing high-fidelity electrogenic cell interfaces, owing to its characteristic topography comparable to proteins and lipids, which influences a wide range of cellular mechanical responses. Gaining a comprehensive understanding of how cells respond to nano-roughness at the single-cell level is not only imperative for implanted devices but also essential for tissue regeneration and interaction with complex biomaterial surfaces. In this study, we quantify cell adhesion and biomechanics of single cells to nano-roughened surfaces by measuring neural cell adhesion and biomechanics via fluidic-based single-cell force spectroscopy (SCFS). For this, we introduce nanoscale topographical features on polyimide (PI) surfaces achieving roughness up to 25 nm without chemical modifications. Initial adhesion experiments show cell-specific response to nano-roughness for neuroblastoma cells (SH-SY5Y) compared to human astrocytes (NHA) around 15 and 20 nm surface roughness. In addition, our SCFS measurements revealed a remarkable 2.5-fold increase in adhesion forces (150-164 nN) for SH-SY5Y cells cultured on roughened PI (rPI) surfaces compared to smooth surfaces (60-107 nN). Our data also shows that cells can distinguish changes in nano-roughness as small 2 nm (close to the diameter of a single lipid) and show roughness dependence adhesion while favoring 15 nm. Notably, this enhanced adhesion is accompanied by increased cell elongation upon cell detachment without any significant differences in cell area spreading. The study provides valuable insights into the interplay between nano-topography and cellular responses and offers practical implications for designing biomaterial surfaces with enhanced cellular interactions.