The reticulocytic rat: a model for analysis of methotrexate polyglutamate dynamics in situ.

The metabolism of the antifol methotrexate in rat erythrocytes in order to determine the basis for its persistence in this tissue was investigated. By using molecular sieve high-performance liquid chromatography and a methotrexate radioassay, we have detected methotrexate polyglutamates through the heptaglutamate in red cells of rats given a single i.p. injection of 10 mg/kg of methotrexate. Higher concentrations of methotrexate and its polyglutamates were found in blood from animals made reticulocytic with phenylhydrazine than in normal erythrocytes, lending credence to the hypothesis that red cell methotrexate polyglutamates arise from developing erythrocytes in the marrow. This was borne out in discrete populations of red cells and reticulocytes separated by density centrifugation through Percoll. Reticulocytes also were shown to have no significant gamma-glutamylcarboxypeptidase (conjugase) activity. We present the reticulocytic rat as an in vivo model for studying polyglutamate metabolism.