We have analyzed the immune response induced by a 9mer synthetic peptide derived from the male histocompatibility antigen H-Y and containing Db-binding motifs in C57BL/6 mice. In this study we report that a single, subcutaneous injection of the peptide emulsified in IFA gave rise to the development of male-specific CD8+ T cells which displayed H-Y-specific proliferative response in vitro and showed a Tc1-type pattern of cytokine production (i.e. they secreted IFN-gamma and IL-2, but not IL-4 and IL-10). Development of a strong cytotoxic activity required in vitro stimulation with specific peptide and IL-2: under these culture conditions, we were able to generate potent CD8+ CTLs that lysed both male cells and peptide-pulsed female cells. Continuous administration of soluble peptide, delivered over a 7-day period by a mini-osmotic pump implanted subcutaneously, inhibited proliferative and cytotoxic responses and IFN-gamma production in lymph node cells from C57BL/6 mice subsequently primed with peptide in adjuvant. This decreased responses were associated with a strong increase in the secretion of IL-4 by antigen-specific CD8+ T lymphocytes. Subcutaneous administration of the H-Y-peptide in adjuvant significantly accelerates rejection of male skin graft, while continuous administration of peptide in soluble form did not modify the time course of rejection.