Bioinspired organic coatings regulate electrochemical interfacial reactivity in sensing, catalysis, and energy applications1,2 However, polydopamine (PDA) coatings remain underexplored for systematically distinguishing how ultrathin interlayers modulate outer and inner-sphere electron transfer3 as tunneling barriers and interfacial reorganization can obscure reaction-specific charge-transfer mechanisms. Here, we examine how PDA interlayers regulate electron-transfer pathways at polycrystalline gold electrodes. Ultrathin PDA films were deposited by cyclic voltammetry via dopamine electrooxidation at pH 8.5 using different cycle numbers (e.g., 10 and 50 cycles). Film formation was evidenced by the initial appearance and subsequent suppression of dopamine redox peaks, accompanied by progressively decreasing currents, indicating increasing surface coverage. Bare and PDA-coated gold electrodes were characterized using contact angle goniometry, optical microscopy, ellipsometry, and atomic force microscopy, confirming uniform and homogeneous coating formation. The influence of PDA films on electron-transfer pathways was evaluated using two model redox systems: potassium ferricyanide as an outer-sphere probe and glucose oxidation as an inner-sphere process. Cyclic voltammetry was conducted at the bulk scale using conventional method and at the nanoscale using scanning electrochemical cell microscopy (SECCM), showing that increasing PDA deposition cycles progressively limited electron transfer for both mechanisms. Notably, SECCM enabled spatially resolved mapping of local electrochemical activity, providing insights inaccessible to conventional bulk scale measurements. Overall, this study provides mechanistic insight into how PDA coating differentially regulates surface-dependent electrochemical reactivity and provide a framework for rationally designing functionalized electrode interfaces for sensing, bioelectrochemical, and energy applications.