Lipolytic Hormones and Glucose Metabolism in Fat CeUs
445
insulin could interact at SM receptors in the target mals. In view of the fact that circulating insulin letissue to stimulate growth in the younger animals or vels are high during this period it is possible that it may act synergistically with SM in these younger ani- is active as a growth stimulus at this time.
MarshalI, R.N., L.E. Underwood, S.J. Voina, D.B. Fouchee, J.J. Van Wyk: Characterisation of the insulin and somatomedin C receptors in human placental ceU membranes. J.Clin.Endocrinol.Metab. 39: 283-292 (1974) Miller, E., c.J. Head, S. Levine, J.H. Holmes, H. Elrick: The use of radioisotopes to measure body fluid consti· tuents. I. Plasma Sulphate. J.Lab.Clin.Med. 58: 656-661 (1961) Rieutort, M.: Pituitary content and plasma levels of growth hormone in fetal and weanling rats. J.Endocr. 60: 261268 (1974) Tato, L., M. v.L. Du Caju, C. Prevot, R. Rappaport: Early variations of plasma somatomedin activity in the new· born. J.Oin.EndocrinoI.Metab. 40: 534-536 (1975) Uthne, K.: Human somatomedins purification and some studies on their biological actions. Acta Endocrinolog. Suppt. 175 (1973) Van den Brande, J.L., M. V.L. Du Caju: An improved tech· nique for measuring somatomedin activity in vitra. Acta Endocrinol. 75: 233-242 (1974) Van Wyk, J.J., L.E. Underwood, R.L. Hintz, D.R. Clemmons, S.J. Viona, R.P. Weaver: The somatomedins: A family of insulinlike hormones under growth hormone control. Rec.Progr.Horm.Res_ 30: 259-318 (1974) Yde, H.: A simplified technique for the determination of growth hormone dependent sulphation factor using intact animals. Acta Endocrinol. 57: 557-564 (1968)
Requests for reprints should be addressed to: Dr. M.C. Stuart, Garvan Institute of Medical Research, St. Vincent's Hospital, Sydney, 2010 .05). In contrast, for 1 hr at 37 , in Krebs Ringer bicarbonate-albumin medium with and without glucose, in the abepinephrine produced a modest increase in glucose sence or presence of epinephrine (0.5 Ilg/mI), or inmetabolism in the cells of Group A (mean of 19%, sulin (1 mU/mI). The results were expressed as IlEq p> .05), but a more marked and statistically significant increase in those of Groups Band C (mean in- of intracellular FFA per 10 7 fat cells, measured at the end of 1 hour incubation, and are shown in crements of 97% and 99% respectively; p < .01 and Table 2. In absence of glucose or hormones, IFFA < .05 respectively). levels were progressively higher as the fat-cell volAs seen in the control group of Table 1, the amounts umes increased. These increases in mean IFFA vaof glucose converted, in absence of hormones, to CO 2 ' lues from small cells to intermediate, and to large glyceride glycerol or glyceride fatty acids varied with cells, were statistically significant (p < 0.02). of the hormone which produced a statistically significant increase above baseline control values.
Downloaded by: NYU. Copyrighted material.
Insulin (1 mU/ml) and epinephrine (0.5 JJg/ml) were added at the beginning of I hr incubation to the hormone-containing flasks. ·Total is the sum of the amounts of glucose converted to CO 2 , glyceride glycerol and glyceride faUy acids.
448
M. DiGirolamo and J.L Owens
Table 2. Intracellular free fatty acid levels in isolated fat cells. Effects of glucose, insulin and epinephrine. Without Glucose Number Mean fat cell volume of animals
Control
small (48 pi)
6
intermediate (185 pI) large (608 pI)
With Glucose +INS
+EPI
0.46 ± 0.14
0.52 ± 0.12
1.12 ± 0.69
1.80 ± 0.40
1.18 ± 0.19
1.20 ± 0.24
0.84 ± 0.30
6.24 ± 1.38
4.16 ± 1.20
4.14 ± 1.44
5.05 ± 1.54
+INS
+EPI
0.44 ± 0.10
0.56 ± 0.16
2.82 ± 0.83
7
1.24 ± 0.29
1.26 ± 0.30
5
4.72 ± 1.31
4.41 ± 1.42
Control
Values are means ± SE of duplicate observations in 5-7 rats for each group. Intracellular free fatty acids were expressed as "Eq/10 7 fat cells and were measured at the end of 1 hr incubation at 37 0 C, in absence or presence of 6 millimolar glucose, 1 mU/ml bovine insulin (INS), or 0.5 P8/ml epinephrine (EPI). Sre text Cor details.
Figure 1 shows the effects of the two hormones on glucose conversion to (C0 2 + glyceride glycerol + glyceride fatty acids). In contrast to the declining insulin effect previously observed with increasing adipocyte size, the effect of these lipolytic hormones on glucose metabolism was progressively and markedly greater in the large cells than in the small ones.
.,
EPINEPHRINE
~
ACTH
';;:+300
~+200
SMALL FAT CELLS (52pll
+
8-+100 .~ ~
0 -50
LARGE FAT CELLS (6661'1)
~+300 G .!ö+200
~
..2
+100
U
O~.~~~~~~~--~~~~~·~~~·~
o
Q",
~O "'0 ~~~~ 0
Q","'o "'0
~~~'\
Downloaded by: NYU. Copyrighted material.
Addition of insulin, glucose, or glucose and insulin did not produce significant changes in intracellular FF A values in any of the 3 groups studied, and thus did not modify the relationship between fat-cell size and IFFA levels. In absence of glucose, addition of epinephrine to the incubation medium increased the intracellular FFA concentration in the small fat cells by 541%, in the intermediate-size cells by 45%, and in the large cells by 32%. The effect of epinephrine was statistically significant for the small fat cells (p < .05) but not so for the intermediate- and largesize cells (p > .05). In presence of glucose, however, the effect of epinephrine on the intracellular FF A levels was markedly curtailed (see last column of Table 2) and did not produce statistically significant changes from control values in any of the 3 groups. These results show a relative stability of the intracellular FF A levels, even at a time when Iipolysis is markedly stimulated by Iipolytic hormones, provided glucose is present in the incubation medium. Effects of variable concentrations of epinephrine and ACTa on rate and pattern of glucose metabolism in small and /arge fat cells. In a third series of experiments, the effects of graded doses of epinephrine and ACTH were compared in selected populations of small fat cells (mean volume 52 pI) from growing 6 wk-old rats and large fat cells (mean volume 666 pI) from adult, moderately obese, 10-14 month-old rats. Isolated fat cells were incubated, in triplicate sampIes, in KRB-albumin medium at 6 millimolar glucose concentration, in the absence and presence of progressively increasing concentrations of the hormones (from 0.5 ng/ml to 50 pg/ml).
Hormone Concentration (ngAnl)
Fig. 1. Dose-response of epinephrine and ACTH on stimulation of glucose metabolism in small and large fat cells. The results are expressed as percent change from baseline con· trol values (0%) produced by a given hormone concentration. Tbe height of the bars represents the mean of duplicate observations in 4 experiments for each cell size; the vertical line above or below the mean bars is ± 1 SE. Tbe aso terisks show the probability that the difference between mean values (Iarge fat cells - small fat cells) and zero, at a .01; given hormone concentration, is due to chance; *p "p .001.
445
insulin could interact at SM receptors in the target mals. In view of the fact that circulating insulin letissue to stimulate growth in the younger animals or vels are high during this period it is possible that it may act synergistically with SM in these younger ani- is active as a growth stimulus at this time.
MarshalI, R.N., L.E. Underwood, S.J. Voina, D.B. Fouchee, J.J. Van Wyk: Characterisation of the insulin and somatomedin C receptors in human placental ceU membranes. J.Clin.Endocrinol.Metab. 39: 283-292 (1974) Miller, E., c.J. Head, S. Levine, J.H. Holmes, H. Elrick: The use of radioisotopes to measure body fluid consti· tuents. I. Plasma Sulphate. J.Lab.Clin.Med. 58: 656-661 (1961) Rieutort, M.: Pituitary content and plasma levels of growth hormone in fetal and weanling rats. J.Endocr. 60: 261268 (1974) Tato, L., M. v.L. Du Caju, C. Prevot, R. Rappaport: Early variations of plasma somatomedin activity in the new· born. J.Oin.EndocrinoI.Metab. 40: 534-536 (1975) Uthne, K.: Human somatomedins purification and some studies on their biological actions. Acta Endocrinolog. Suppt. 175 (1973) Van den Brande, J.L., M. V.L. Du Caju: An improved tech· nique for measuring somatomedin activity in vitra. Acta Endocrinol. 75: 233-242 (1974) Van Wyk, J.J., L.E. Underwood, R.L. Hintz, D.R. Clemmons, S.J. Viona, R.P. Weaver: The somatomedins: A family of insulinlike hormones under growth hormone control. Rec.Progr.Horm.Res_ 30: 259-318 (1974) Yde, H.: A simplified technique for the determination of growth hormone dependent sulphation factor using intact animals. Acta Endocrinol. 57: 557-564 (1968)
Requests for reprints should be addressed to: Dr. M.C. Stuart, Garvan Institute of Medical Research, St. Vincent's Hospital, Sydney, 2010 .05). In contrast, for 1 hr at 37 , in Krebs Ringer bicarbonate-albumin medium with and without glucose, in the abepinephrine produced a modest increase in glucose sence or presence of epinephrine (0.5 Ilg/mI), or inmetabolism in the cells of Group A (mean of 19%, sulin (1 mU/mI). The results were expressed as IlEq p> .05), but a more marked and statistically significant increase in those of Groups Band C (mean in- of intracellular FFA per 10 7 fat cells, measured at the end of 1 hour incubation, and are shown in crements of 97% and 99% respectively; p < .01 and Table 2. In absence of glucose or hormones, IFFA < .05 respectively). levels were progressively higher as the fat-cell volAs seen in the control group of Table 1, the amounts umes increased. These increases in mean IFFA vaof glucose converted, in absence of hormones, to CO 2 ' lues from small cells to intermediate, and to large glyceride glycerol or glyceride fatty acids varied with cells, were statistically significant (p < 0.02). of the hormone which produced a statistically significant increase above baseline control values.
Downloaded by: NYU. Copyrighted material.
Insulin (1 mU/ml) and epinephrine (0.5 JJg/ml) were added at the beginning of I hr incubation to the hormone-containing flasks. ·Total is the sum of the amounts of glucose converted to CO 2 , glyceride glycerol and glyceride faUy acids.
448
M. DiGirolamo and J.L Owens
Table 2. Intracellular free fatty acid levels in isolated fat cells. Effects of glucose, insulin and epinephrine. Without Glucose Number Mean fat cell volume of animals
Control
small (48 pi)
6
intermediate (185 pI) large (608 pI)
With Glucose +INS
+EPI
0.46 ± 0.14
0.52 ± 0.12
1.12 ± 0.69
1.80 ± 0.40
1.18 ± 0.19
1.20 ± 0.24
0.84 ± 0.30
6.24 ± 1.38
4.16 ± 1.20
4.14 ± 1.44
5.05 ± 1.54
+INS
+EPI
0.44 ± 0.10
0.56 ± 0.16
2.82 ± 0.83
7
1.24 ± 0.29
1.26 ± 0.30
5
4.72 ± 1.31
4.41 ± 1.42
Control
Values are means ± SE of duplicate observations in 5-7 rats for each group. Intracellular free fatty acids were expressed as "Eq/10 7 fat cells and were measured at the end of 1 hr incubation at 37 0 C, in absence or presence of 6 millimolar glucose, 1 mU/ml bovine insulin (INS), or 0.5 P8/ml epinephrine (EPI). Sre text Cor details.
Figure 1 shows the effects of the two hormones on glucose conversion to (C0 2 + glyceride glycerol + glyceride fatty acids). In contrast to the declining insulin effect previously observed with increasing adipocyte size, the effect of these lipolytic hormones on glucose metabolism was progressively and markedly greater in the large cells than in the small ones.
.,
EPINEPHRINE
~
ACTH
';;:+300
~+200
SMALL FAT CELLS (52pll
+
8-+100 .~ ~
0 -50
LARGE FAT CELLS (6661'1)
~+300 G .!ö+200
~
..2
+100
U
O~.~~~~~~~--~~~~~·~~~·~
o
Q",
~O "'0 ~~~~ 0
Q","'o "'0
~~~'\
Downloaded by: NYU. Copyrighted material.
Addition of insulin, glucose, or glucose and insulin did not produce significant changes in intracellular FF A values in any of the 3 groups studied, and thus did not modify the relationship between fat-cell size and IFFA levels. In absence of glucose, addition of epinephrine to the incubation medium increased the intracellular FFA concentration in the small fat cells by 541%, in the intermediate-size cells by 45%, and in the large cells by 32%. The effect of epinephrine was statistically significant for the small fat cells (p < .05) but not so for the intermediate- and largesize cells (p > .05). In presence of glucose, however, the effect of epinephrine on the intracellular FF A levels was markedly curtailed (see last column of Table 2) and did not produce statistically significant changes from control values in any of the 3 groups. These results show a relative stability of the intracellular FF A levels, even at a time when Iipolysis is markedly stimulated by Iipolytic hormones, provided glucose is present in the incubation medium. Effects of variable concentrations of epinephrine and ACTa on rate and pattern of glucose metabolism in small and /arge fat cells. In a third series of experiments, the effects of graded doses of epinephrine and ACTH were compared in selected populations of small fat cells (mean volume 52 pI) from growing 6 wk-old rats and large fat cells (mean volume 666 pI) from adult, moderately obese, 10-14 month-old rats. Isolated fat cells were incubated, in triplicate sampIes, in KRB-albumin medium at 6 millimolar glucose concentration, in the absence and presence of progressively increasing concentrations of the hormones (from 0.5 ng/ml to 50 pg/ml).
Hormone Concentration (ngAnl)
Fig. 1. Dose-response of epinephrine and ACTH on stimulation of glucose metabolism in small and large fat cells. The results are expressed as percent change from baseline con· trol values (0%) produced by a given hormone concentration. Tbe height of the bars represents the mean of duplicate observations in 4 experiments for each cell size; the vertical line above or below the mean bars is ± 1 SE. Tbe aso terisks show the probability that the difference between mean values (Iarge fat cells - small fat cells) and zero, at a .01; given hormone concentration, is due to chance; *p "p .001.
