The anemia associated with chronic renal failure is one of the best target diseases for erythropoietin (Epo) gene transfer. We previously reported a short-term (1 month) study of continuous rat Epo delivery by muscle-targeted gene transfer of plasmid DNA expressing rat Epo (pCAGGS-Epo) using in vivo electroporation in normal rats. Here, we performed a long-term pharmacokinetic study of continuous Epo delivery by this method in normal rats and uremic five-sixths nephrectomized rats. In normal rats, Epo gene expression and sufficient erythropoiesis occurred with Epo gene transfer in a dose-dependent manner, and persisted for at least 11 weeks. Repeated administration of the plasmid DNA effectively produced erythropoiesis. Similar erythropoiesis was observed in the uremic rats, and persisted for more than 15 weeks. Both normal and uremic rats showed a significant decrease in platelet count. Moreover, the uremic rats showed Epo-induced hypertension, which is the major side-effect of recombinant human Epo. These results demonstrate that muscle-targeted pCAGGS-Epo transfer by in vivo electroporation is a useful procedure for the long-term continuous delivery of Epo in both normal and uremic rats.