This study investigated the trajectory of physiological and psychological functioning during the second half of pregnancy and compared responsiveness to a laboratory stressor between pregnant and non-pregnant women. Monitoring of 137 pregnant women at 20, 24, 28, 32, 36, and 38 weeks of pregnancy included measures of heart period (HP), heart period variability (HPV), skin conductance (SCL), respiratory period (RP), respiratory sinus arrhythmia (RSA), and self-report of mood disturbance. HP and RSA declined during this period; SCL and mood disturbance increased. Parity was a significant moderator. HP and SCL responsiveness to the Stroop color-word task was assessed twice in pregnant participants and compared to a sample of 27 non-pregnant women. Physiologic responsiveness was reduced in pregnant women. Pregnant women perceived the Stroop to be more difficult, but performance was unaffected. Despite buffered responsivity to stressful stimuli during pregnancy, advancing gestation is associated with escalating sympathetic tone and declining parasympathetic tone.