A number of studies have assessed the effects of psychoactive drugs on stress biology, the neuroadaptations resulting from chronic drug use on stress biology, and their effects on addiction risk and relapse. This review mainly covers human research on the acute effects of different drugs of abuse (i.e., nicotine, cannabis, psychostimulants, alcohol, and opioids) on the hypothalamic-pituitary-adrenal (HPA) axis and the autonomic nervous system (ANS) responses. We review the literature on acute peripheral stress responses in naïve or light recreational users and binge/heavy or chronic drug users. We also discuss evidence of alterations in tonic levels, or tolerance, in the latter relative to the former and associated changes in the phasic stress responses. We discuss the impact of the stress system tolerance in heavy users on their response to drug- and stress-related cue responses and craving as compared to control subjects. A summary is provided of the effects of glucocorticoid responses and their adaptations on brain striatal and prefrontal cortices involved in the regulation of drug seeking and relapse risk. Finally, we summarize important considerations, including individual difference factors such as gender, co-occurring drug use, early trauma and adversity and drug use history and variation in methodologies, that may further influence the effects of these drugs on stress biology.