GROWTH-HORMONE INCREASES CALCIUM-UPTAKE IN RAT FAT-CELLS BY A MECHANISM DEPENDENT ON PROTEIN-KINASE-C

Growth hormone (GH; 500 ng/ml) rapidly doubled cytosolic free Ca2+ con centration ([Ca2+](i)) in rat adipocytes as determined with the Ca2+ i ndicator fura 2. No response was seen in Ca2+-free medium, suggesting that the increase in [Ca2+](i) was due to Ca2+ influx. GH also doubled the influx of Mn2+ as inferred from the rate of fluorescence quenchin g. Depolarization with 30 mM K+ also increased [Ca2+](i), and the incr ease in [Ca2+](i) due to either GH or 30 mM K+ was blocked by 100 nM n imodipine, suggesting that GH increases [Ca2+](i) by activating voltag e-sensitive L-type Ca2+ channels. GH increased [Ca2+](i) even when Kt channels were blocked, suggesting that activation of Ca2+ uptake was n ot secondary to closure of K+ channels and consequent depolarization. A diacylglycerol (DAG) analogue, 1,2-dioctanoyl-sn-glycerol (50 mu M), duplicated, and the protein kinase C (PKC) inhibitors calphostin C (1 00 nM), chelerythrine (1 mu M), and bis-indolylmaleimide (250 nM) inhi bited the effects of GH on [Ca2+](i). Xanthogenate tricyclodecan-9-yl (D609), a specific inhibitor of phospholipase C (PLC), abolished the i ncrease in [Ca2+](i) due to GH but not to DAG. The results suggest tha t GH increases [Ca2+](i) by activation of PLC, release of DAG, and act ivation of a Ca2+-independent isoform of PKC. PKC-catalyzed phosphoryl ation of either the Ca2+ channels or a protein that regulates them may account for the influx of Ca2+ produced by GH.