OBJECTIVE To investigate the effect of portal venous inoculation of donor splenocytes combined with cyclosporin A (CsA) administration on cardiac allograft survival in mice. METHODS Heterotopic cardiac transplantation between fully allogenic NIH/q and BALB/C strain mice was performed. A modified procedure of neonatal heart-in-ear transplantation, as originally described by Fulmer et al, was adopted. We prepared donor splenocytes from NIH/q or third-party C57BL/6 spleens for BALB/C recipients, which were injected preoperatively via the recipient portal vein or the systemic vein 1 week before the heart-in-ear transplantation. The recipients were subsequently treated with a short course of the immunosuppressive agent, CsA (4 mg/kg starting from 7 d before the operation till 5 d after it). RESULTS Portal venous inoculation of donor splenocytes combined with CsA significantly prolonged cardiac graft survival (n=6, P<0.05) that reached 31.00+/-3.23 d, and 2 of the 6 allografts survived for more than 35 d. Donor splenocytes injected via the systemic vein or third-party C57BL/6 mice splenocytes injected via the portal vein did not prolong graft survival (P>0.05). CsA alone or portal venous inoculation of donor-specific splenocytes alone also prolonged graft survival (P<0.05), with mean graft survival time of 18.50+/-2.59 d and 16.11+/-1.97 d respectively. CONCLUSIONS Combination of portal venous inoculation of donor-specific splenocytes and CsA can prolong murine cardiac allograft survival, which is donor antigen-specific.