•Greater PA was significantly associated with larger P300s across inhibitory and target detection in older adults.•Accelerometer-based MVPA provide unique insight into both P300s over and above self-reported MVPA in older adults.•Accelerometer-based findings may be influenced by the scoring method, defining standards for scoring measures is crucial. Older adults who experience cognitive decline are more likely to have a reduced quality of life. Identifying lifestyle factors that may influence cognitive processing and in turn improve quality of life during older adulthood is an important area of interest. Cognitive function, as measured by the P300 event-related potential (ERP), has been noted to be modified by physical activity; however, no study to date has examined relationships between this neurophysiological measure and physical activity and sedentary time in older adults. Furthermore, there is a gap in understanding as to whether physical activity and sedentary time assessed using self-reported and accelerometer-based methods similarly relate to the P300. This study aimed to assess the P300 during a Go/No-Go task in relation to self-reported and accelerometer-based physical activity and sedentary time in a community sample of 75 older adults. Results indicated that participants engaging in more moderate-to-vigorous physical activity had larger P300 amplitudes across self-reported and accelerometer-based measurements; however, no relationships between sedentary time and P300 amplitude were observed. Notably, accelerometer-based moderate-to-vigorous physical activity explained P300 amplitudes over and above self-reported moderate-to-vigorous physical activity—an effect that remained significant even after accounting for age. Although these results highlight the importance of moderate-to-vigorous physical activity in relation to cognitive function, as measured via the P300 in older adults, a secondary analysis indicated that engaging in lifestyle activity may have similar effects on the P300 as moderate-to-vigorous physical activity. In sum, the present study highlights the role of habitual engagement in physical activity as a possible means for supporting cognitive function during the aging process.