Two aspects of study on metallothionein will be conducted: (1) isolated rat liver perfusion, and (2) in vitro reactivation of zinc and copper requiring metalloapoenzymes. In the former the hypothesis to be tested is whether metals other than zinc (Cd, Hg, Cu) can serve as inducers of metallothionein, or are they simply acting as displacers of zinc from pre-existing zinc thionein. Groups of animals to be studied will involve: normal (untreated), zinc and/or copper deficient, zinc and/or copper loaded. Perfusions then will be performed with Cd, Hg, and Cu in the perfusate. Also, zinc uptake and incorporation into metallothionein will be studied with perfusion of livers from rats which are zinc deficient or treated with Alpha- and Beta-adrenergic blockers to prevent the operation induced, epinephri mediated uptake of zinc by these livers. Analysis of hepatic total, cytosol, and metallothionein metal contents following these treatments will aid in the assessment of the processes involving metals, other than zinc, and metallothionein. The use of radioactive metals and amino acids in conjunction with cycloheximide will aid in the assessment of the requirement for protein synthesis in metal binding to metallothionein. In the latter, the hypothesis to be tested is whether metallothionein functions as a reservoir of zinc (and copper) intracellularly, providing it for newly synthesized metalloapoenzymes. In vitro reactivation of zinc apoenzymes by zinc thionein has been accomplished, encouraging us to pursue this hypothesis further. Specifically, the following types of experiments will be conducted: in vitro reactivation of copper apoenzymes with (Zn, Cu) - thionein; in vitro transfer of radioactive metals form metallothionein to zinc and copper apoenzymes; and in vivo and ex vivo uptake by rat liver in zinc and/or copper deficient rats of radiolabeled metallothionein, looking at reactivation of and label transfer to metalloenzymes. The effects of the presence of cadmium (and mercury) in the metallothionein used for reactivation will be studied to determine possible in vivo effects of these toxic metals on normal zinc (and copper) metabolism.