NKT cells expressing the invariant Valpha14 antigen receptor constitute a novel lymphocyte subpopulation with immunoregulatory functions. Stimulation via their invariant Valpha14 receptor with anti-CD3 or a ligand, alpha-galactosylceramide (alpha-GalCer), triggers activation of Valpha14 NKT cells, resulting in a rapid cytokine production such as IFN-gamma and IL-4. Soon after their receptor activation, Valpha14 NKT cells disappeared as judged by staining with CD1d tetramer loaded with alpha-GalCer (alpha-GalCer/CD1d tetramer), which has been believed to be due to apoptotic cell death. Here we show that such a disappearance was largely attributed to down-regulation of the Valpha14 receptor. In fact, Valpha14 NKT cells were relatively resistant to apoptosis compared to the conventional T cells as evidenced by less staining with Annexin-V, a limited DNA fragmentation, and their preferential expression of anti-apoptotic genes such as NAIP and MyD118. Furthermore, they did not become tolerant, and maintained their proliferative capacity and cytokine production even after their receptor down-regulation. These as yet unrecognized facets of Valpha14 NKT cells are discussed in relation to their regulatory functions.