Rationale Individuals with a family history of depression may be more likely to develop depression due to an innate vulnerability of their serotonergic system. However, even though serotonergic vulnerability may constitute a risk factor in the development of depression, it does not seem to be sufficient to cause a depressive episode. Based on previous data, it is suggested that stress may be a mediating factor. Objectives This study examined the role of serotonin (5-HT) in stress coping in individuals with or without a family history of depression. Materials and methods Nineteen healthy first-degree relatives of depressive patients (FH+) and 19 healthy controls without a family history of depression (FH-) were tested in a double-blind placebo-controlled design for affective processing under acute stress exposure, following acute tryptophan depletion (ATD) or placebo. Results Significant negative effects were found of stress on affective processing in FH- and FH+. In addition, FH- responded slower to positive words after stress only following ATD, whereas FH+ responded marginally slower under stress already after placebo and before stress following ATD. Conclusion Acute stress exposure reduces positive affective bias; supporting the role of stress as an important predecessor in the development of depression. Furthermore, FH+ may be more susceptible than FH- to the negative effects of stress as well as to the negative effects of ATD. The results support the assumption that the 5-HT system is involved in stress resilience and may be more vulnerable in first-degree relatives of depression.