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Regulation of glocagon-stimulatedproduction of glucose in rat liver by guanosine 3',5'-cyclic phosphate RICHARD M. EPAND AND CONNIE PROSSER Bepnrttnent of Bioc.lr~t?li.\lry,McMnster Ilniversity, Harniltorz, Om., Cannda L8S 459

Received February 16, 1976

EPAND, R. M., and PROSSER, C. 1976. Regulation of glucagon-stimulated production of glucose in rat liver by guanosine 3',5'-cyclic phosphate. Can. J . Physiol. Pharmacol. 54, 834-837. Exogenous cGMP can inhibit both basal and glucagon-stimulated production of glucose in liver slices from fed rats. Thus, cAMP and cGMP have opposite effects on the production of glucose in rat liver. Acetylcholine, an activator of guanylate cyclase (EC 4.6.1.2) in other ~ystems,also inhibits the glucagon-stimulated production of glucose. N o effect on glucose production was observed with secretin or exogenous GTP.

EPAND,R . &Ie et PROSSER,C. 1976. Regulation of glucagon-stimulated production of glucose in rat liver by guanosine 3',5'-cyclic phosphate. Can. J. Physiol. Pharmacol. 54, 834-837. Ee cGMP exopkne peut inhiber la prodnction d e glucose, ri la fois avant et apr6s stimulation par le glucagon, dans les tranches de foie de rat? alimentCs. Le cAMP et le cGMP ont donc des actions contraires sur la production du glucose dans le foie de rat. L9ac6tylcholine, qui est un activateur de la grlanylate cyclase (EC 4.6.1.2) dans d'autres systkmes, inhibe Cgalement la production de glucose sous l'influence du glucagon. Aucun effet sur la production de glucose n'est observe avec le GTP exogkne ou la secrbtine. [Traduit par le journal]

Introduction A unifying theory to explain the action of cyclic nucleotides on n~etabolicregulation, the Yin-Yang hypothesis, proposes that regulation of cellular function occurs by two counterbalancing forces represented by the concentration of cAMP and cGMP (Goldberg et al. 1974). This type of regulation may come into play in the hepatic production of glucose. In this system, glucagon, which causes a rise in cAMP levels (Exton, Robison pt al. 1 97 1 ) , increases the rate of glucose production, whereas insulin, which causes a rise in cGMP levels (Illiano et ul. 1973), opposes this effect. Exogenous cAMP can mimic the effect of glucagon but under the conditions tested exogenous cCMP was found to have the same effects as cAMP (Glinsmann et al. 1969; Conn and Kipnis 1969; Exton, Hardman et nl. 1971; Conn et ul. 197 1;Helderman et a!. 1972), although one recent report has showrl an inhibition of glucose production by exogenous cGMP in isolatcd liver cells from fasted rats (Fain ef al. 1975). We have examCAMP, adenosine 3',5'-cyclic phosABBREVIATIONS: phate; cGMP, guanosine 3',S'-cyclic phosphate; GTP, guanosine 5'4riphosphate.

ined this question further by measuring the effects of exogenous cGMP on glucagon-stimulated rates and on rates glucose Productioll in liver from fed rats. Materials and Methods The procedure for measuring glucose production in rat liver slices is similar to that previously described (Wheeler and Epand 1975). Livers were obtained from male Woodlyn-Wistar rats (150-200 g) fed ad libitum, and were perfused with Krebs-Ringer-phosphate buffer, pH 7.4. Slices (75-125 mg) were made using a Stadie-Riggs tissue slicer (A. H. Thomas Company). Tissue slices were washed with buffer and then incubated at 37 " C in a reciprocating water bath in plastic tubes containing 2 ml of Krebs-Ringerphosphate buffer. pH 7.4, with 4 % defatted bovine serum albumin and 10 m M pyruvate. The absence of pyruvate did not affect the rate of glucose production. 7 he tubes were incubated for 30 min, after which time a \ample of the supernate was taken for deproteinization and subsequent glucose analysis by the glucose oxidase method. During this time only 10% of the total lactic dehydrogenase (EC 1.1.1.27) content of the slice appeared in the extracellular fluid (Ryan, E. A. and Epand, K. M., unpublished observation), indicating the viability of the preparation. In addition, the basal rate of glucose production was linear with time over 1 h. The basal rate of glucose production was 102.7 1+ 7.4 ~ m o per l gram wet weight of liver per hour. Basal and glucagon-stimulated rates of glucose

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EPAND A N D PROSSER

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ImM 3pM 50OpM 50pM 5pM 0.5pM 0.05pM ACh Secretin cGMP cGMP cGMP cGMP cGMP

FIG.1 . Effect of acetylcholine, secretin, and c G M P on the giucagon-stimulated production of glucose in rat liver slices. Results are plotted as the measured rate in the presence of both 10 pg/ml glucagon and the added drug minus basal rate divided by the rate in the prescnce of 10 pg/ml glucagon minus basal rate and are shown as the mean + one standard deviation. 1 m M ACh-1 m M acetylcholine chloride, 15 determinations, P 0.02; 3 p M secretin-15 determinations, P not significant; 500 p M cGMP- 17 determinations, P = 0.05; 50 p M cGh4P-35 determinations, P = 0.03; 5 p M cGMP-47 determinations. P = 0.002; 0.5 p M cGMP-23 determinations, P = 0.02; and 0.05 p M cGMP-11 determinations, P not significant.

Regulation of glucagon-stimulated production of glucose in rat liver by guanosine 3',5'-cyclic phosphate.

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