COBALTOUS CHLORIDE-MEDIATED INDUCTION OF RAT HEPATIC TRYPTOPHAN 2,3-DIOXYGENASE - IMPLICATIONS FOR THE USE OF THE ENZYME TO PROBE THE HEPATIC FREE HEME POOL

Subcutaneous administration of CoCl2, a well recognized inhibitor of h epatic heme synthesis, to rats results in the functional stimulation o f total (holo-+apo) tryptophan 2,3 dioxygenase (TDO), a hemoprotein an d the key rate-limiting enzyme in the oxidative metabolism of tryptoph an to formylkynurenine. Because basal holo-TDO activity is not altered , TDO stimulation appears to be entirely due to CoCl2-mediated increas e of its apoprotein. This apoTDO increase was blocked by conventional inhibitors of protein synthesis (actinomycin D, cycloheximide), thereb y revealing that such CoCl2-mediated apoprotein increase truly reflect ed TDO induction. To determine whether the CoCl2-mediated TDO inductio n involved the action of its natural physiological inducers (glucocort icoids) or was due to direct CoCl2-regulation of the TDO gene, rats we re adrenalectomized before CoCl2 administration. In adrenalectomized r ats, CoCl2 failed to induce TDO, but induction was completely restored on administration of the glucocorticoid hydrocortisone, but not of ad renaline. These findings reveal that CoCl2-mediated TDO induction is i ndirect and entails glucocorticoid participation. In addition, because CoCl2 lowered the % heme saturation of TDO [=100 (holoTDO activity/to tal (apo+holo)TDO activity] largely by increasing the apoTDO protein l evels rather than by affecting the basal holo-TDO levels (as expected from its inhibition of heme synthesis), these findings question the wi dely accepted use of the relative intrahepatic % heme saturation of TD O as a reporter of the hepatic ''free'' heme pool.