Diabetes, Vol 40, Issue 5 612-620, Copyright © 1991 by American Diabetes Association

ARTICLES

Effect of Zn2+ on the proteolytic inhibitory action of insulin and biguanide antihyperglycemic drugs

DP Thorne and TD Lockwood
Department of Pharmacology, School of Medicine, Wright State University, Dayton, OH 45459.

The involvement of Zn2+ in the inhibitory action of insulin and phenformin on bulk proteolysis was studied in the Langendorff rat heart with a Zn(2+)-buffering perfusate (0.1 mM citrate, physiological complete amino acids and 0.2% albumin). Proteins were biosynthetically labeled in vitro for 10 min with [3H]leucine. Rapidly degraded proteins were eliminated during a 3-h preliminary degradation without insulin or added Zn2+ (2 mM nonradioactive leucine). Insulin (5 nM), the lysosomal inhibitor chloroquine (30 microM), and the biguanide antihyperglycemic agent phenformin (2 microns) each caused a sustained 35-40% inhibition of [3H]leucine release beginning within 1-2 min and reaching a maximum at 1-1.5 h. When these agents were combined, their simultaneous proteolytic inhibitory effects were not appreciably greater than the effect of chloroquine alone. Infusion of supraphysiological perfusate Zn2+ (greater than 15 microM) mimicked the inhibitory effect of insulin and chloroquine on lysosomal proteolysis. Infusion of supraphysiological Co2+, Mn2+, Fe2+, and Cr3+ (30 microM, 0.5 h) caused no change in proteolysis; however, 30 microM Cu2+ caused a slight inhibition. Presumptive chelation of the background (approximately 20 nM) Zn2+ by infusion of 3 microM CaNa2 EDTA caused no change in protein degradation over 1-2 h. The infusion of a physiological concentration of 1 or 5 microM Zn2+ (as ZnCl2) caused no change in protein degradation over 1-2 h. Biguanides are known to reversibly form a Zn2+ complex with affinity less than that of Zn2+ for EDTA. Prior infusion of 3 microM CaNa2 EDTA inactivated the proteolytic inhibitory effect of maximal (2 microM) phenformin over at least 1.25 h of concurrent infusion.(ABSTRACT TRUNCATED AT 250 WORDS)
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