Effects of insulin on glycogen metabolism of serum-deprived BHK-21 fibroblasts

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Abstract
We have studied the effects of insulin on glycogen metabolism of cultured BHK‐21 fibroblasts. Addition of insulin to cells cultured in 0.5% serum stimulated incorporation of 5mM 14C‐glucose into glycogen and increased cellular glycogen content, without inducing proliferation. With serum‐deprived cells incubated for 2 hr, maximal stimulation of incorporation of glucose into glycogen was 3.5‐fold and the half‐maximal dose of insulin was 5nM. After culture for 24 hr with 100nM insulin, incorporation was increased 13‐fold and glycogen content was increased by 44%. Incorporation of glucose into glycogen was reduced by agents which elevate cellular adenosine 3,5‐cyclic monophosphate (cyclic AMP): 10μM prostaglandin E1 (PGE1), 0.25 mM 1‐methyl‐3‐isobutylxanthine (MIX), 10μM L‐epinephrine (L‐EPI), and 10μM L‐isoproterenol (L‐ISO). Culture with 100nM insulin for 2 hr stimulated incorporation three‐fold in the presence of any of these compounds, but insulin did not affect cellular cyclic AMP. At all times from 1 hr to 24 hr after addition of insulin to serum‐deprived cultures, the 1‐min uptake of 10μM 3H‐2‐deoxyglucose (3H‐2DG) was increased. There were small (30 to 40%) increases in glycogen synthase I activity at 15 and 30 min but not 4 hr after addition of insulin. After 24 hr with insulin, total glycogen synthase and phosphorylase activities were increased approximately two‐fold, without changes in their activation states. We conclude that insulin promotes glycogenesis in serum‐deprived BHK‐21 cells. This response is mediated principally by increased entry of glucose into cells, and is not mediated by a change in cellular cyclic AMP concentration.