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Adrenal glucocorticoids exert a permissive action on the glycogen phosphorylase cascade. Epinephrine activation of muscle phosphorylase and phosphorylase b kinase is depressed in adrenalectomized rats. Phosphorylase phosphatase activity is increased by steroid lack, and normal epinephrine inhibition of the enzyme does not occur. Phosphorylase b kinase phosphatase activity is also increased; epinephrine, however, does not inhibit activity in muscle from normal or adrenalectomized rats. Protein phosphatase inhibitor activity is depressed in boiled dialyzed preparations made from adrenalectomized rat muscle. Cortisol resplacement therapy restores protein phosphatase inhibitor activity, decreases increased protein phosphatase activity, and restores normal epinephrine-induced activation of phosphorylase b kinase and phosphorylase and epinephrine inhibition of phosphorylase phosphatase.

Protein phosphatase inhibitor-1 was purified from bovine adipose tissue. The protein had an apparent molecular mass of 32 kDa by SDS/PAGE and a Stokes' radius of 3.4 nm. It was phosphorylated by cAMP-dependent protein kinase on a threonyl residue; this phosphorylation was necessary for inhibition of protein phosphatase-1. Bovine adipose tissue inhibitor-1 was compared directly with rabbit skeletal muscle inhibitor-1 and with a 32000-Mr, dopamine- and cAMP-regulated phosphoprotein from bovine brain (DARPP-32), also an inhibitor of protein phosphatase-1. By the following biochemical and immunochemical criteria, bovine adipose tissue inhibitor-1 was found to be very similar and possibly identical to DARPP-32 and was clearly distinct from skeletal muscle inhibitor-1: molecular mass by SDS/PAGE; Stokes' radii; phosphorylation on threonine residues; Staphylococcus-aureus-V8-protease-generated peptide patterns analyzed by SDS/PAGE; tryptic phosphopeptide maps analysed by two-dimensional thin-layer electrophoresis/chromatography; elution on reverse-phase HPLC; chymotryptic peptide maps as analysed by reverse-phase HPLC; amino acid composition; antibody recognition by immunoprecipitation and immunoblotting; effect of cyanogen bromide cleavage on protein phosphatase inhibitor activity. Based on these results we conclude that bovine brain and adipose tissue contain an identical phosphoprotein inhibitor of protein phosphatase-1 (DARPP-32), which is distinct from that of skeletal muscle (inhibitor-1).