Exercise elicits intensity-dependent increases in epinephrine (Epi), norepinephrine (NE), cortisol (CORT), and lactate (Lac), whereas at rest the hormonal responses to mental stress include increases in Epi, NE, and CORT, but not Lac. Additionally, elevations in CORT at rest are associated with negative affect. Finally, as exercise intensity increases, perceptions of effort (RPE) increase and affect becomes more negative. The purpose of this study was to examine the responses of Epi, NE, CORT, Lac, RPE, and affect during incremental increases in exercise to maximum. Seven highly-trained male runners completed a discontinuous treadmill protocol that included 10 minutes at 60%, 10 minutes at 75%, 5 minutes at 90%, and 2 minutes at 100% of JOURNAL/jscr/04.02/00124278-200702000-00050/ENTITY_OV0312/v/2017-07-20T235329Z/r/image-pngO2max. Blood samples were collected through an intravenous catheter before, during, and following exercise during the experimental condition, and during a time-matched control condition. RPE and affect were assessed just prior to each blood draw. The Epi, NE, CORT, and Lac displayed the expected incremental increases, however affect became more negative at higher exercise intensities, and RPE increased as exercise intensity increased. Data suggest that CORT follows the same pattern of response to graded exercise as affect. Since perceptual and affective responses to exercise are associated with motivation, these documented negative affective shifts during exercise should be considered in order to develop strategies that lead to enhanced exercise adherence and performance.