We investigated changes in lymphocyte subsets (total, CD4+ and CD8+ T cells, as well as non-T cells) associated with human immunodeficiency virus type I (HIV-1) seroconversion in 321 homosexual or bisexual men in the Multicenter AIDS Cohort Study (MACS). These subjects had serial lymphocyte characterizations for up to 4 years before and 5 years after seroconversion. CD4+ lymphocyte levels declined rapidly in the first 18 months following seroconversion and less rapidly thereafter, while CD8 lymphocytes increased with similar kinetics. In contrast, total T (CD3+) lymphocytes declined only slightly in the first 18 months following seroconversion, and then remained stable. These results support the hypothesis of physiologic regulation of the total number of circulating T cells, such that lost CD4+ lymphocytes are replaced by newly generated CD4+ and CD8+ lymphocytes; over time, continued loss of CD4+ lymphocytes due to HIV-1 infection would result in net replacement of lost CD4+ lymphocytes with CD8+ lymphocytes. Non-T (CD3-) lymphocytes also declined after seroconversion, and this decline paralleled that of CD4+ lymphocytes. Thus, changes in both T and non-T lymphocytes after HIV-1 seroconversion may reflect the operation of homeostatic or regulatory mechanisms. Whether these mechanisms contribute to the development of immune deficiency requires further study.