During early pregnancy, there are marked increases in cardiac output (CO) and global arterial compliance (AC), as well as decreases in systemic vascular resistance (SVR). We recently reported that administration of recombinant human relaxin to nonpregnant female rats elicits changes in systemic hemodynamics and arterial mechanical properties similar to those observed during normal pregnancy. In the present study, we directly tested whether endogenous relaxin mediates the cardiovascular adaptations of pregnancy by neutralizing circulating relaxin with monoclonal antibodies during early gestation. Relaxin neutralizing antibodies were administered daily, beginning on d 8 of rat gestation, to block the functional effects of circulating relaxin. Systemic hemodynamics and arterial properties were assessed between gestational d 11 and 15 using techniques we have previously reported. Pregnant rats administered the neutralizing antibodies failed to exhibit the gestational increases in stroke volume, CO, and global AC or decreases in SVR that were observed in control pregnant rats administered an irrelevant antibody against fluorescein or PBS. In fact, in the pregnant rats administered the relaxin neutralizing antibodies, cardiovascular parameters were not statistically different from those in virgin rats. Interestingly, small renal and first-order mesenteric arteries isolated from midterm pregnant rats administered either relaxin-neutralizing or control antibodies did not exhibit any changes in passive mechanical properties compared with virgin rats. These findings indicate that circulating relaxin mediates the transition of the systemic circulation from the virgin to the pregnant state in the gravid rat model, suggesting a potential role for aberrant relaxin regulation in abnormal pregnancies wherein these cardiovascular adaptations are inadequate or excessive.