Brief stimulation of the raphe–spinal pathway in the turtle spinal cord releases serotonin (5‐HT) onto motoneurones to enhance excitability. However, intense release of 5‐HT via prolonged stimulation results in 5‐HT spillover to the motoneurone axon initial segment to activate inhibitory 5‐HT1A receptors, thus providing a potential spinal mechanism for exercise‐induced central fatigue. We examined how increased extracellular concentrations of 5‐HT affect the ability to perform brief, as well as sustained, maximal voluntary contractions (MVCs) in humans. Paroxetine was used to enhance 5‐HT concentrations by reuptake inhibition, and three studies were performed. Study 1 (n = 14) revealed that 5‐HT reuptake inhibition caused an ∼4% increase in elbow flexion MVC. However, when maximal contractions were sustained, time‐to‐task failure was reduced and self‐perceived fatigue was higher with enhanced availability of 5‐HT. Study 2 (n = 11) used twitch interpolation to reveal that 5‐HT‐based changes in motor performance had a neural basis. Enhanced 5‐HT availability increased voluntary activation for the unfatigued biceps brachii and decreased voluntary activation of the biceps brachii by 2–5% following repeated maximal elbow flexions. The final study (n = 8) investigated whether altered motoneurone excitability may contribute to 5‐HT changes in voluntary activation. F‐waves of the abductor digiti minimi (ADM) were unaffected by paroxetine for unfatigued muscle and marginally affected following a brief 2‐s MVC. However, F‐wave area and persistence were significantly decreased following a prolonged 60‐s MVC of the ADM. Overall, high serotonergic drive provides a spinal mechanism by which higher concentrations of 5‐HT may contribute to central fatigue.