This study revises and enhances a guided inquiry activity designed to introduce undergraduate and high school students to the physiological concepts of exercise fatigue during a 400-meter dash. This activity has been employed since 2016 in entry-level human physiology courses at the University of Minnesota and across 30 high schools. Using simulated data, students analyze variables such as blood lactate, pH, heart rate, and oxygen consumption to identify factors contributing to fatigue in a 400-meter dash. Recognizing that limitations existed in the original dataset (e.g. blood lactate and ventilatory data that rose too high or too quickly), this work incorporates revised data derived from a treadmill-based 400-meter dash, supplemented with peer-reviewed benchmarks to better reflect physiological responses in the severe and extreme exercise intensity domains. The updated activity includes adaptations for both entry-level as well as upper-level human and exercise physiology courses. The upper-level version introduces students to exercise intensity domains. These revisions provide students with a data-driven exploration of the mechanisms underlying exercise fatigue and expand opportunities for analyzing and interpreting data. The activity aligns with Next Generation Science Standards by fostering skills in data analysis, collaborative learning, and critical thinking. Future directions include further refinement of the dataset using real-world physiological measurements, enabling an even closer alignment with contemporary exercise science research.