Prior studies have reported cortisol secretion to be primarily influenced by negative affect, but less is known about cortisol activity during states of activation involving increased positive affect and decreased negative affect. On separate days, 30 healthy young men experienced: an activating and humorous video; a speech stressor; and a resting control period. Cortisol was measured in saliva before and after each 30-min mood induction. Positive affect (activation) was increased similarly by both the video and the speech compared to rest (p < .0001). Negative affect increased during the speech and decreased during the video (p < .001). Cortisol increased only during the speech (p < .0001). Following the video, however, cortisol was decreased significantly (p < .0001). Rest day cortisol revealed no differences across periods (p > .1). These results suggest that the hypothalamic-pituitary-adrenocortical axis is a dynamic system influenced by changes in negative affect irrespective of the experience of generalized activation.