Further Evidence Implicating Diacylglycerol Generation and Protein Kinase C Activation in Agonist-Induced Increases in Glucose Uptake: Insulin-Like Effects of Phenylephrine in BC3H-1 Myocytes

Abstract

We have previously suggested that insulin effects on 2-deoxyglucose (2-DOG) uptake in BC3H-1 myocytes are due to increases in de novo phospholipid synthesis, diacylglycerol generation, and protein kinase C activation. To test this hypothesis further, we examined the effects of phenylephrine, an agonist that increases diacylglycerol and protein kinase C activity through phospholipase C activation. As evidence for phospholipase activation in BC3H-1 myocytes, we found that phenylephrine increased acute 32PO4 incorporation into phosphatidic acid and phosphatidylinositol, generation of [3H]inositol phosphates from prelabeled [3H]inositol phospholipids, cytosolic Ca2+, and membrane-bound protein kinase C. Phenylephrine also provoked dose-related increases in [3H]2-DOG uptake that were similar in magnitude and time course to those induced by insulin. As with insulin, phenylephrine effects on 2-DOG uptake were not apparent in myocytes that were maximally stimulated with 12-O-tetradecanoylphorbol-13-acetate, a diacylglycerol analogue that activates protein kinase C. These findings support our hypothesis that diacylglycerol generation and protein kinase C activation may be important in the stimulation of glucose uptake by agents such as phenylephrine and insulin that activate the phosphoinositide cycle.

Stimulatory effect of caffeic acid on α 1A -adrenoceptors to increase glucose uptake into cultured C 2 C 12 cells

“…Moreover, it has been documented that α 1 -adrenoceptor agonist, phenylephrine, can activate phospholipase C (PLC) in muscle cells (Farese et al 1986). Incubation with α 1 -adrenoceptor agonist releases inositol-1,4,5-phosphate into the cytosol and increases diacylglycerol (DAG) within membrane in white adipocytes of rats (Faintrenie and Géloén 1998).…”

“…Incubation with α 1 -adrenoceptor agonist releases inositol-1,4,5-phosphate into the cytosol and increases diacylglycerol (DAG) within membrane in white adipocytes of rats (Faintrenie and Géloén 1998). The accumulation of DAG and Ca 2+ as a result of activation of PLC causes the activation of protein kinase C (PKC; Farese et al 1986). Mediation of PLC-PKC pathway in the activation of α 1 -adrenoceptor was also characterized in C 2 C 12 cells depending on the blockade of caffeic acid-stimulated 2-DG uptake by the inhibitors specific for PLC or PKC.…”

“…Stimulatory effect of caffeic acid on α 1A -adrenoceptors to increase glucose uptake into cultured C 2 C 12 cells Activation of α 1 -adrenoceptor by phenylephrine, the specific agonist, is documented to increase phospholipase C (PLC)-mediated hydrolysis of inositol phospholipids in myocytes to result in an increase of protein kinase C (PKC) activity (Farese et al 1986). The present study is performed to characterize the subtype of α 1 -adrenoceptors presenting in C 2 C 12 cells.…”

Phorbol esters imitate in rat fat-cells the full effect of insulin on glucose-carrier translocation, but not on 3-O-methylglucose-transport activity

“…One piece of evidence supporting this hypothesis was the observation that phorbol esters, which can substitute for diacylglycerol as an activator of protein kinase C, show insulin-like effects in isolated adipocytes (Van de Werve et al, 1985;Kirsch et al, 1985) as well as in myocytes in culture (Farese et al, 1985). Whereas in the myocyte culture diacylglycerol and phorbol esters are able to elicit the full insulin response (Farese et al, 1986), only a partial insulin-like effect, usually not more than 30 % of the maximal insulin effect, can be induced in isolated rat 10 20…”

“…It is believed that they include activation ofthe intrinsic insulin-receptor kinase (Kasuga et al, 1983) and phosphorylation of cellular proteins (Hiring et al, 1987;White et al, 1985;Machicao et al, 1987), followed by as yet unknown intermediary steps, finally resulting in a translocation of glucose carrier sites from intracellular membranes to the plasma membrane (Wardzala et al, 1978;Cushman & Wardzala, 1980;Karnieli et al, 1981). Several lines of evidence have suggested that the steps linking insulin-receptor kinase and activation of the glucose-transport system might involve the phosphatidylinositol cycle (K6pfer- Hobelsberger & Wieland, 1984;Kirsch et al, 1985;Van de Werve et al, 1985;Nishizuka, 1986), release of diacylglycerol (Farese et al, 1985) and activation of protein kinase C (Farese et al, 1986). This speculation is partially based on the observation that tumour-promoting phorbol esters, which can substitute for diacylglycerol as stimulators of protein kinase C, are able to Abbreviation used: TPA, 12-0-tetradecanoylphorbol 13-acetate.…”