Estrogen and prolactin have been shown to modulate autoimmunity in the NZB/NZW F1 (B/W) mouse model of systemic lupus erythematosus (SLE). However, estrogen stimulates prolactin secretion. The goal of this study was to examine differential effects of estrogen and prolactin in the female B/W mouse model of SLE. B/W females were manipulated to create combinations of low and high concentrations of serum estrogen and prolactin. Hyperprolactinemic mice with either low or high serum estrogen levels had accelerated development of albuminuria at 24 and 32 weeks of age compared to normal and hypoprolatinemic mice. High estrogen/high prolactin mice also had a higher percentage of anti-DNA antibodies compared to mice in the low estrogen/low prolactin and the high estrogen/low prolactin groups. IgG levels were not significantly different between groups. Mean survival was shortest in the high estrogen/high prolactin group (34 1.0 weeks) and longest in the high estrogen/low prolactin group (42 1.2 weeks; P < 0.05). High levels of serum estrogen were associated with depressed in vitro lymphoproliferation and IL-2 production. This study suggests that high prolactin levels in either high or low serum estrogen states are associated with accelerated autoimmunity in the B/W mouse. This study further demonstrates that high estrogen levels do not accelerate murine SLE when the prolactin-stimulating property of estrogen is suppressed by bromocriptine. Further investigation of hormonal interactions in autoimmunity will provide a better understanding of hormonal immunoregulation and, perhaps, lead to improved clinical application of hormonal immunomodulation.