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Effect of prostaglandin E1 administration on the heart glycogen synthase and phosphorylase systems
EFFECT OF PROSTAGLANDIN E 1 ADMINISTRATION ON THE HEART GLYCOGEN SYNTHASE AND PHOSPHORYLASE SYSTEMS
Randall T. Curnow* and Frank Q. Nuttall
Medical Service and Endocrine-Metabolic Section, V.A. Hospital Minneapolis, Minnesota and Department of Medicine, University of Minnesota Minneapolis, Minnesota
*Present Address:
U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, Maryland
ABSTRACT The effects of intravenous administration of PGE I on the glycogen synthase and phosphorylase system in rat heart were studied. Unlike the consistent effects of PGE I on glycogen synthase in liver, the response in heart was variable. A significant decrease in the per cent synthase I occurred in fasted intact rats while a significant increase was seen in adrenalectomized hydrocortisone treated fasted rats. No significant effect was seen on the synthase system in either fed intact or fasted adrenalectomized rats. Phosphorylase a activity was increased significantly following PGE I administration in fed intact rats and slightly increased in adrenalectomized fasted rats. The phosphorylase system was not affected in fasted intact and fasted adrenalectomized rats given glucocorticoid replacement. With our present state of knowledge an adequate explanation for the response of these heart enzymes to PGE I under the various conditions of this study does not appear possible.
ACKNOWLEDGEMENTS This work was supported in part by a grant from the Twin Cities Diabetes Association. The authors gratefully acknowledge the excellent technical assistance of Mrs. Carol LaBresh. At the time this work was done Dr. Curnow was a Research and Education Trainee at the Minneapolis V. A. Hospital.
Accepted March 15
PROSTAGLANDINS APRIL 25, 1974
VOL. 6 NO. 2
115
PROSTAGLANDINS
INTRODUCTION Prostaglandin El (PGEl) has been shown to have insulin-like effects (1,2) and to alter cyclic AMP (CAMP) levels, either increasing or decreasing them in various tissues (3). Furthermore, PGEl has recently been reported to stimulate adenylate cyclase of guinea pig heart particles -in vitro (4). Therefore, the effect of this agent has been studied on two enzyme systems in rat heart which are known to be influenced by alterations in concentration of this cyclic nucleotide, and to be affected by insulin. These enzymes are glycogen synthase (glycogen transferase: UDP glucose o-1,4-glucan-glucosyl transferase, E.C. 2.4.1.11) and glycogen phosphorylase (a-1,4-glucan-orthophosphate glucosyl transferase, E.C. 2.4.1.1). GLycogen synthase is the primary control site for glycogen synthesis (5) while glycogen phosphorylase activity controls glycogen degradation (6). Both enzymes are known to exist in physiologically active and inactive forms and there is a rapid enzymic interconversion between the two forms. The chemical nature of the interconverting reactions of both glycogen synthase and phosphorylase has been shown to involve phosphorylation and dephosphorylation of the enzyme by a specific kinase and phosphatase respectively (7,8). The physiologically active or I form (glucose 6-phosphate independent) of glycogen synthase is the dephosphorylated form, while the phosphorylated or D form (glucose 6-phosphate dependent) is completely inactive under physiological conditions (9). Conversely, the physiologically active form of phosphorylase in heart is phosphophosphorylase (phosphorylase a> and the physiologically inactive form is dephosphophosphorylase (phosphorylase b). Injection of epinephrine (10) and glucagon (11) into the intact animal causes rapid simultaneous conversion of synthase from the I to D form and phosphorylase from dephospho- to phosphophosphorylase.- It is currently thought that these effects are mediated by activation of adenyl cyclase with resultant increase in intracellular CAMP (12). The nucleotide then initiates an enzyme cascade beginning with activation of protein kinase (8). Another potential control site for concomitant activation-inactivation of these enzymes is the phosphatase enzymes. Little is known concerning the hormonal control of these enzymes. Insulin stimulates an increase in the percent of synthase in the -I form (13,14) but the mechanism is not known. We previously reported that intravenous administration of prostaglandin El (PGEl) to anesthetized rats resulted in a rapid conversion of glycogen synthase I to glycogen synthase IJ in liver (15). This was independent of alterations in nutritional status, presence or absence of adrenal glands, and ganglionic or beta adrenergic blockade. Concomitant early increases in liver phosphophosphorylase activity occurred in fed and fasted intact animals. In fasted animals
116
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the effect on the phosphorylase system was completely inhibited by ganglionic blockade and partially inhibited by beta adrenergic blockade. These findings suggested that the effect of PGEl on the synthase system in liver was a direct effect, while the response of the phosphorylase system was dependent on the presence of adrenal glands and of an intact autonomic nervous system. In this paper the response of heart glycogen synthase and phosphorylase activities to PGEl administration in these same animals is presented. MATERIALS AND METHODS The materials and methods have been previous described (15). Briefly, male, Holtzman strain rats fed Purina Chow ad libitum were used in all experiments. Adrenalectomy, when done, was performed under ether-anesthesia by means of a bilateral dorsolumbar approach. Rats were used 3 days later. Adrenalectomized rats were provided with 0.1% NaCl for drinking water. One group of adrenalectomized rats also received 0.5 mg of hydrocortisone sodium succinate a single morning subcutaneous injection which was calculated to be a daily maintenance dose (F. Ungar, personal communication). All rats were anesthetized with 50 mg/kg of Seconal interperitoneally 15 to 30 minutes before use. Intravenous injections were made into the saphenous vein of anesthetized animals. In all studies, the hearts were quickly removed and immediately frozen by clamping in liquid Nz-cooled aluminum clamps (-196O) and stored at -65O. Blood for glucose analysis was obtained from the thoracic cavity after the heart was removed. Glycogen concentration, glycogen synthase activity and blood glucose determinations were done as previously described (15). Phosphorylase activity was also determined as previously described (15) except that phosphorylase a was assayed in the absence of added adenosine monophosphate (AMP), and total phosphorylase activity (phosphorylase 5 plus phosphorylase b) was measured in the presence of 3.3 mM AMP. Synthase activity is reported as the percent synthase in the I form and as units (micromoles glucose incorporated into glycogen per minute per g of wet tissue). Phosphorylase activity is reported as the percent phosphorylase in the a form and as units (micromoles of phosphate released from glucose-l-P per minute per g of wet tissue). RESULTS Following injection of 30 mg/kg PGEl intravenously into fed animals (Figure 1) there was a prompt statistically significant increase in the percentage phosphorylase in the a form at 30 seconds. The percentage phosphorylase in the a form subsequently remained variably increased until 3 minutes, thereafter returning to control levels. Total synthase and phosphorylase activities in controls were 1.13 + 0.05 units and 29.7 f. 1.6 units respectively (mean & S.E.). There was a significant increase in blood glucose occurring at 4 and 5 minutes following PGEl administration which probably represented stimulation of hepatic glycogenolysis by PGEl (15).
APRIL 25, 1974
VOL. 6 NO, 2
117
PROSTAGLANDINS
Fad Heart
Glycogen (mgm/gww)
4.0
8
4
.j
3.0
4
1
‘:$-y-y
Blood Glucose (mgm/looml)
03dll
2
3
4
5
Time (min.1 Figure 1. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase 5 (X Phos a), glycogen concentration, percent synthase I, (X GS-I) in heart and blood glucose of fed intact rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. PGEl promoted no appreciable change in total phosphorylase or phosphorylase 5 activity in fasted rats (Figure 2) nor was there any change in glycogen concentration. PGEl administration did, however, result in a rapid and significant decrease in the percent synthase in the I form even though the basal level was quite low. This effect is qualitatively similar to the effect of PGEl in the liver in the same rats. Total synthase and phosphorylase activities in controls were 1.19 + 0.11 units and 31.8 T 1.8 units respectively. Total synthase and phosphorylase activities were unchanged. Unlike the response of fed rats, PGEl caused no change in blood glucose.
118
APRIL 25, 1974
VOL. 6 NO. 2
PROSTAGLANDINS
Fasted Hearts
I
0307
I
I
1
I
I
2
3
4
5
Time (min.) Figure 2. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase a (X Phos a), glycogen concentration, percent synthase I (% GS-I) in heart and blood glucose of fasted intact rats. Vertical bars represent 5 the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate P~0.05 or less compared to controls at time zero. Following PGEl administration to fasted adrenalectomized rats, there was a small increase in percent phosphorylase in the 5 form (Figure 3). As in the previous experiments there was no change in total phosphorylase activity. As in fed animals, but in contrast to fasted animals, there was no significant change in percent synthase in the I form. Total synthase and phosphorylase in controls were 1.19 + 0.05-units and 24.6 + 2.6 units respectively. Again total synthase activity was unchanged by PGEl. No effect on cardiac glycogen concentration was noted. Blood glucose was again unchanged during the period of the study.
APRIL 25, 1974 VOL. 6 NO. 2
119
PROSTAGLANDINS
Adrwex
Fastad
Heart
Blood Glucosia (mgm/lOO ml 1 Tims (min.) Figure 3. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase 5 (% Phos a), glycogen concentration, percent synthase L (X GS-I) in heart and blood glucose of fasted adrenalectomized rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. In adrenalectomized, fasted, glucocorticoid replaced rats, PGEl administration resulted in no significant change in percent phosphorylase in the 5 form (Figure 4). Glycogen concentration was not affected. There was an increase in percent synthase in the I form that reached statistical significance at 6 minutes and returned gradually to the control level by 30 minutes. This was in contrast to the lack of effect noted in intact fed and adrenalectomized fasted rats and to the reduction found in fasted intact animals. Total synthase and phosphophosphorylase activities in controls were 1.00 + 0.04 units and 26.4 + 1.1 units respectively. Once again PGEl caused-no changes in total s-mthase or phosphorylase activities. Blood glucose was again unchanged.
120
APRIL 25, 1974 VOL. 6 NO. 2
PROSTAGLANDINS
Adrenex
I
Fasted
,
0
Glucocorticoid
I,
,
4
8
I
I,,
12
Replaced
I,,
16
20
Heart
I,
,
24
20
I
I
32
Tima (min.) Figure 4. Effect of a single 30 ng/kg PGEl IV injection on percent phosphorylase a (X Phos a), glycogen concentration, percent synthase I (% GS-I) in heart and blood glucose of fasted adrenalectomized hydrocortisone treated rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. DISCUSSION The effects of PGEl on the synthase and phosphorylase systems of the heart were more variable than those in the liver. PGEl caused no statistically significant effect on the heart synthase system in either the fed intact or adrenalectomized fasted rats. However, the mean percent synthase I activity was somewhat higher than controls in fed intact rats after FGEl administration. In heart from fasted intact animals there was a small but statistically significant decrease in percent synthase I activity. This was similar to the response in liver (15) and is similar to the effects noted previously in rat heart following epinephrine (9) and glucagon (10). It is the result one might expect if CAMP concentration had been increased by these agents, and in keeping with the reported activation by PGE of adenylate cyclase in isolated cardiac particles from guinea pig t4). In the,heart from fasted adrenalectomized rats given glucocorticoid
APRIL 25, 1974 VOL. 6 NO. 2
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PROSTAGLANDINS
replacement the synthase J activity was increased. Again the change was small but reached statistical significance h minutes following PGEl administration. This response is similar to that seen previously following insulin administration (10,16) and just opposite to the effect anticipated if CAMP levels were increased by PGE1. PGEl caused an increase in percent phosphorylase 5 in both fed intact and adrenalectomized fasted rats. The changes were statistically significance in the fed intact animals while in the adrenalectomized fasted animals the changes were small and not significant statistically. Again such changes are those expected if CAMP concentration had been increased by the prostaglandin. Following PGE administration no change occurred in the percent phosphorylase 8 in eitI! er the fasted intact or adrenalectomized fasted glucocorticoid replaced rats. The reason for the lack of response is not clear.
It is apparent that the nutritional status of the animals and the presence or absence of adrenal steroids and catecholamines influences the response of the synthase system in heart to PGE1. This is in contrast to the results in liver where a consistent lowering of the percent of synthase in the 1 form was observed. The reason for the variable response in heart is not known. The phosphorylase system in heart appears to be relatively insensitive to prostaglandin administration. A statistically significant increase in percent phosphorylase a only occurred in the fed intact animals and the increase was relatively small. Since a dose response curve was not done, it is possible that different results might have been obtained using a larger dose. However, this dose was considerably in excess of the required for a maximal effect on the liver synthase system. PGEl administration -in vivo has been reported to increase the secretion of several hormones including adrenocorticotrophic hormone (17), growth hormone (18), and insulin (19). It is, therefore, possible that the changes seen in these studies were mediated, at least in part, by PGEl-promoted alterations in the circulating hormonal milieu. With the present state of knowledge concerning the regulation of the synthase and phosphorylase enzyme systems, an adequate explanation for the results found does not appear possible.
122
APRIL 25, 1974
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REFERENCES 1.
Vaughn, M. In Prostaglandins, Proceedings of the Second Nobel Symposium (S. Bergstrom and B. Samuelson, Editors) Interscience Publishers, Inc., New York, 1967, p. 139.
2.
Bohle, E., H. H. Ditschuneit, R. Dobert and H. Ditschuneit. Dent. Ges. Inn. Med. 73:797, 1967.
3.
Ramwell, P. W. and J. E. Shaw. 1970.
4.
Klein, I.
5.
Villar-Palasi, C. and J. Larner. 1960.
6.
Sutherlund, E. W. and C. F. Cori.
7.
Larner, J., C. Villar-Palasi and N. E. Brown. Acta. 178:470, 1969.
8.
Krebs, E. G., R. J. DeLange, R. G. Kemp and W. D. Riley. Rev. 18:163, 1966.
9.
Piras, R., L. B. Rothman and E. Cabib.
and G. S. Levey.
Verh.
Recent Progr. Hormone Res.
Metabolism
26:139,
20:890, 1971.
Biochem. Biophys. Acta.
J. Biol. Chem.
39:171,
188:531, 1951.
Biochem. Biophy.
Biochemistry
Pharmacol.
7~56, 1968.
10.
Nuttall, F. Q. and W. J. Bergstrom.
Unpublished observations.
11.
Bergstrom, W. J. and F. Q. Nuttall. 146, 1972.
Biochem. Biophys. Acta.
12.
Butcher, R. W.
N. Engl. J. Med.
279:1378- 1968.
13.
Williams, B. J. and S. E. Mayer.
Mol. Pharmacol.
14.
Huijing, F., F. Q. Nuttall, C. Villar-Palasi and J. Larner. Biophys. Acta. 177~204, 1969.
15.
Curnow, R. T. and F. Q. Nuttall.
J. Biol. Chem.
247:1892, 1972.
16.
Williams, B. J. and S. E. Mayer.
Mol. Pharmacol.
2:454, 1966.
17.
Peng, T. C., K. M. Six and P. L. Munson.
18.
Kato, Y., J. Dupre and J. C. Beck.
19.
Bressler, R., M. Vargas-Cordon and H. F. Lebovitz. 17~617, 1968.
APRIL
25, 1974
VOL. 6 NO. 2
268:
2:454, 1966.
Endocrinology
Endocrinology
Biochem.
86:202, 1970.
93:135, 1973. Diabetes
123
Randall T. Curnow* and Frank Q. Nuttall
Medical Service and Endocrine-Metabolic Section, V.A. Hospital Minneapolis, Minnesota and Department of Medicine, University of Minnesota Minneapolis, Minnesota
*Present Address:
U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, Maryland
ABSTRACT The effects of intravenous administration of PGE I on the glycogen synthase and phosphorylase system in rat heart were studied. Unlike the consistent effects of PGE I on glycogen synthase in liver, the response in heart was variable. A significant decrease in the per cent synthase I occurred in fasted intact rats while a significant increase was seen in adrenalectomized hydrocortisone treated fasted rats. No significant effect was seen on the synthase system in either fed intact or fasted adrenalectomized rats. Phosphorylase a activity was increased significantly following PGE I administration in fed intact rats and slightly increased in adrenalectomized fasted rats. The phosphorylase system was not affected in fasted intact and fasted adrenalectomized rats given glucocorticoid replacement. With our present state of knowledge an adequate explanation for the response of these heart enzymes to PGE I under the various conditions of this study does not appear possible.
ACKNOWLEDGEMENTS This work was supported in part by a grant from the Twin Cities Diabetes Association. The authors gratefully acknowledge the excellent technical assistance of Mrs. Carol LaBresh. At the time this work was done Dr. Curnow was a Research and Education Trainee at the Minneapolis V. A. Hospital.
Accepted March 15
PROSTAGLANDINS APRIL 25, 1974
VOL. 6 NO. 2
115
PROSTAGLANDINS
INTRODUCTION Prostaglandin El (PGEl) has been shown to have insulin-like effects (1,2) and to alter cyclic AMP (CAMP) levels, either increasing or decreasing them in various tissues (3). Furthermore, PGEl has recently been reported to stimulate adenylate cyclase of guinea pig heart particles -in vitro (4). Therefore, the effect of this agent has been studied on two enzyme systems in rat heart which are known to be influenced by alterations in concentration of this cyclic nucleotide, and to be affected by insulin. These enzymes are glycogen synthase (glycogen transferase: UDP glucose o-1,4-glucan-glucosyl transferase, E.C. 2.4.1.11) and glycogen phosphorylase (a-1,4-glucan-orthophosphate glucosyl transferase, E.C. 2.4.1.1). GLycogen synthase is the primary control site for glycogen synthesis (5) while glycogen phosphorylase activity controls glycogen degradation (6). Both enzymes are known to exist in physiologically active and inactive forms and there is a rapid enzymic interconversion between the two forms. The chemical nature of the interconverting reactions of both glycogen synthase and phosphorylase has been shown to involve phosphorylation and dephosphorylation of the enzyme by a specific kinase and phosphatase respectively (7,8). The physiologically active or I form (glucose 6-phosphate independent) of glycogen synthase is the dephosphorylated form, while the phosphorylated or D form (glucose 6-phosphate dependent) is completely inactive under physiological conditions (9). Conversely, the physiologically active form of phosphorylase in heart is phosphophosphorylase (phosphorylase a> and the physiologically inactive form is dephosphophosphorylase (phosphorylase b). Injection of epinephrine (10) and glucagon (11) into the intact animal causes rapid simultaneous conversion of synthase from the I to D form and phosphorylase from dephospho- to phosphophosphorylase.- It is currently thought that these effects are mediated by activation of adenyl cyclase with resultant increase in intracellular CAMP (12). The nucleotide then initiates an enzyme cascade beginning with activation of protein kinase (8). Another potential control site for concomitant activation-inactivation of these enzymes is the phosphatase enzymes. Little is known concerning the hormonal control of these enzymes. Insulin stimulates an increase in the percent of synthase in the -I form (13,14) but the mechanism is not known. We previously reported that intravenous administration of prostaglandin El (PGEl) to anesthetized rats resulted in a rapid conversion of glycogen synthase I to glycogen synthase IJ in liver (15). This was independent of alterations in nutritional status, presence or absence of adrenal glands, and ganglionic or beta adrenergic blockade. Concomitant early increases in liver phosphophosphorylase activity occurred in fed and fasted intact animals. In fasted animals
116
APRIL
25. 1974
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PROSTAGLANDINS
the effect on the phosphorylase system was completely inhibited by ganglionic blockade and partially inhibited by beta adrenergic blockade. These findings suggested that the effect of PGEl on the synthase system in liver was a direct effect, while the response of the phosphorylase system was dependent on the presence of adrenal glands and of an intact autonomic nervous system. In this paper the response of heart glycogen synthase and phosphorylase activities to PGEl administration in these same animals is presented. MATERIALS AND METHODS The materials and methods have been previous described (15). Briefly, male, Holtzman strain rats fed Purina Chow ad libitum were used in all experiments. Adrenalectomy, when done, was performed under ether-anesthesia by means of a bilateral dorsolumbar approach. Rats were used 3 days later. Adrenalectomized rats were provided with 0.1% NaCl for drinking water. One group of adrenalectomized rats also received 0.5 mg of hydrocortisone sodium succinate a single morning subcutaneous injection which was calculated to be a daily maintenance dose (F. Ungar, personal communication). All rats were anesthetized with 50 mg/kg of Seconal interperitoneally 15 to 30 minutes before use. Intravenous injections were made into the saphenous vein of anesthetized animals. In all studies, the hearts were quickly removed and immediately frozen by clamping in liquid Nz-cooled aluminum clamps (-196O) and stored at -65O. Blood for glucose analysis was obtained from the thoracic cavity after the heart was removed. Glycogen concentration, glycogen synthase activity and blood glucose determinations were done as previously described (15). Phosphorylase activity was also determined as previously described (15) except that phosphorylase a was assayed in the absence of added adenosine monophosphate (AMP), and total phosphorylase activity (phosphorylase 5 plus phosphorylase b) was measured in the presence of 3.3 mM AMP. Synthase activity is reported as the percent synthase in the I form and as units (micromoles glucose incorporated into glycogen per minute per g of wet tissue). Phosphorylase activity is reported as the percent phosphorylase in the a form and as units (micromoles of phosphate released from glucose-l-P per minute per g of wet tissue). RESULTS Following injection of 30 mg/kg PGEl intravenously into fed animals (Figure 1) there was a prompt statistically significant increase in the percentage phosphorylase in the a form at 30 seconds. The percentage phosphorylase in the a form subsequently remained variably increased until 3 minutes, thereafter returning to control levels. Total synthase and phosphorylase activities in controls were 1.13 + 0.05 units and 29.7 f. 1.6 units respectively (mean & S.E.). There was a significant increase in blood glucose occurring at 4 and 5 minutes following PGEl administration which probably represented stimulation of hepatic glycogenolysis by PGEl (15).
APRIL 25, 1974
VOL. 6 NO, 2
117
PROSTAGLANDINS
Fad Heart
Glycogen (mgm/gww)
4.0
8
4
.j
3.0
4
1
‘:$-y-y
Blood Glucose (mgm/looml)
03dll
2
3
4
5
Time (min.1 Figure 1. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase 5 (X Phos a), glycogen concentration, percent synthase I, (X GS-I) in heart and blood glucose of fed intact rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. PGEl promoted no appreciable change in total phosphorylase or phosphorylase 5 activity in fasted rats (Figure 2) nor was there any change in glycogen concentration. PGEl administration did, however, result in a rapid and significant decrease in the percent synthase in the I form even though the basal level was quite low. This effect is qualitatively similar to the effect of PGEl in the liver in the same rats. Total synthase and phosphorylase activities in controls were 1.19 + 0.11 units and 31.8 T 1.8 units respectively. Total synthase and phosphorylase activities were unchanged. Unlike the response of fed rats, PGEl caused no change in blood glucose.
118
APRIL 25, 1974
VOL. 6 NO. 2
PROSTAGLANDINS
Fasted Hearts
I
0307
I
I
1
I
I
2
3
4
5
Time (min.) Figure 2. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase a (X Phos a), glycogen concentration, percent synthase I (% GS-I) in heart and blood glucose of fasted intact rats. Vertical bars represent 5 the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate P~0.05 or less compared to controls at time zero. Following PGEl administration to fasted adrenalectomized rats, there was a small increase in percent phosphorylase in the 5 form (Figure 3). As in the previous experiments there was no change in total phosphorylase activity. As in fed animals, but in contrast to fasted animals, there was no significant change in percent synthase in the I form. Total synthase and phosphorylase in controls were 1.19 + 0.05-units and 24.6 + 2.6 units respectively. Again total synthase activity was unchanged by PGEl. No effect on cardiac glycogen concentration was noted. Blood glucose was again unchanged during the period of the study.
APRIL 25, 1974 VOL. 6 NO. 2
119
PROSTAGLANDINS
Adrwex
Fastad
Heart
Blood Glucosia (mgm/lOO ml 1 Tims (min.) Figure 3. Effect of a single 30 ug/kg PGEl IV injection on percent phosphorylase 5 (% Phos a), glycogen concentration, percent synthase L (X GS-I) in heart and blood glucose of fasted adrenalectomized rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. In adrenalectomized, fasted, glucocorticoid replaced rats, PGEl administration resulted in no significant change in percent phosphorylase in the 5 form (Figure 4). Glycogen concentration was not affected. There was an increase in percent synthase in the I form that reached statistical significance at 6 minutes and returned gradually to the control level by 30 minutes. This was in contrast to the lack of effect noted in intact fed and adrenalectomized fasted rats and to the reduction found in fasted intact animals. Total synthase and phosphophosphorylase activities in controls were 1.00 + 0.04 units and 26.4 + 1.1 units respectively. Once again PGEl caused-no changes in total s-mthase or phosphorylase activities. Blood glucose was again unchanged.
120
APRIL 25, 1974 VOL. 6 NO. 2
PROSTAGLANDINS
Adrenex
I
Fasted
,
0
Glucocorticoid
I,
,
4
8
I
I,,
12
Replaced
I,,
16
20
Heart
I,
,
24
20
I
I
32
Tima (min.) Figure 4. Effect of a single 30 ng/kg PGEl IV injection on percent phosphorylase a (X Phos a), glycogen concentration, percent synthase I (% GS-I) in heart and blood glucose of fasted adrenalectomized hydrocortisone treated rats. Vertical bars represent + the standard error of the mean and the number of animals used is given above or below the bars. Asterisks indicate PcO.05 or less compared to controls at time zero. DISCUSSION The effects of PGEl on the synthase and phosphorylase systems of the heart were more variable than those in the liver. PGEl caused no statistically significant effect on the heart synthase system in either the fed intact or adrenalectomized fasted rats. However, the mean percent synthase I activity was somewhat higher than controls in fed intact rats after FGEl administration. In heart from fasted intact animals there was a small but statistically significant decrease in percent synthase I activity. This was similar to the response in liver (15) and is similar to the effects noted previously in rat heart following epinephrine (9) and glucagon (10). It is the result one might expect if CAMP concentration had been increased by these agents, and in keeping with the reported activation by PGE of adenylate cyclase in isolated cardiac particles from guinea pig t4). In the,heart from fasted adrenalectomized rats given glucocorticoid
APRIL 25, 1974 VOL. 6 NO. 2
121
PROSTAGLANDINS
replacement the synthase J activity was increased. Again the change was small but reached statistical significance h minutes following PGEl administration. This response is similar to that seen previously following insulin administration (10,16) and just opposite to the effect anticipated if CAMP levels were increased by PGE1. PGEl caused an increase in percent phosphorylase 5 in both fed intact and adrenalectomized fasted rats. The changes were statistically significance in the fed intact animals while in the adrenalectomized fasted animals the changes were small and not significant statistically. Again such changes are those expected if CAMP concentration had been increased by the prostaglandin. Following PGE administration no change occurred in the percent phosphorylase 8 in eitI! er the fasted intact or adrenalectomized fasted glucocorticoid replaced rats. The reason for the lack of response is not clear.
It is apparent that the nutritional status of the animals and the presence or absence of adrenal steroids and catecholamines influences the response of the synthase system in heart to PGE1. This is in contrast to the results in liver where a consistent lowering of the percent of synthase in the 1 form was observed. The reason for the variable response in heart is not known. The phosphorylase system in heart appears to be relatively insensitive to prostaglandin administration. A statistically significant increase in percent phosphorylase a only occurred in the fed intact animals and the increase was relatively small. Since a dose response curve was not done, it is possible that different results might have been obtained using a larger dose. However, this dose was considerably in excess of the required for a maximal effect on the liver synthase system. PGEl administration -in vivo has been reported to increase the secretion of several hormones including adrenocorticotrophic hormone (17), growth hormone (18), and insulin (19). It is, therefore, possible that the changes seen in these studies were mediated, at least in part, by PGEl-promoted alterations in the circulating hormonal milieu. With the present state of knowledge concerning the regulation of the synthase and phosphorylase enzyme systems, an adequate explanation for the results found does not appear possible.
122
APRIL 25, 1974
VOL. 6 NO. 2
PROSTAGLANDINS
REFERENCES 1.
Vaughn, M. In Prostaglandins, Proceedings of the Second Nobel Symposium (S. Bergstrom and B. Samuelson, Editors) Interscience Publishers, Inc., New York, 1967, p. 139.
2.
Bohle, E., H. H. Ditschuneit, R. Dobert and H. Ditschuneit. Dent. Ges. Inn. Med. 73:797, 1967.
3.
Ramwell, P. W. and J. E. Shaw. 1970.
4.
Klein, I.
5.
Villar-Palasi, C. and J. Larner. 1960.
6.
Sutherlund, E. W. and C. F. Cori.
7.
Larner, J., C. Villar-Palasi and N. E. Brown. Acta. 178:470, 1969.
8.
Krebs, E. G., R. J. DeLange, R. G. Kemp and W. D. Riley. Rev. 18:163, 1966.
9.
Piras, R., L. B. Rothman and E. Cabib.
and G. S. Levey.
Verh.
Recent Progr. Hormone Res.
Metabolism
26:139,
20:890, 1971.
Biochem. Biophys. Acta.
J. Biol. Chem.
39:171,
188:531, 1951.
Biochem. Biophy.
Biochemistry
Pharmacol.
7~56, 1968.
10.
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