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Extreme sensitivity of obese hyperglycemic mice to caffeine and coffee
Extreme
Sensitivity of Obese Hyperglycemic Mice to Caffeine and Coffee
By DUBRAVKOM. KUFTINEC AND JEAN MAYER Mice with the hereditary obese hyperglycemic syndrome are unusually sensi-
and pyrimidines other than alloxan (shown previously to cause in this syn-
tive to the toxic action of caffeine. A single injection of caffeine, or the replacement of drinking water by coffee for 1 week causes a drastic and prolonged (2 or more months) elevation of blood glucose. Administration of purines
drome prolonged lowering of blood glucase and regranulation of beta cells) did not lead to striking results. No explanation for this unexpected effect of caffeine is offered.
T
HE HEREDITARY RECESSIVE HYPERGLYCEMIC SYNDROME in mice is characterized inter alia by extreme adiposity, moderate hyperphagia but extreme inactivity, a high blood glucose despite high pancreatic and circulating insulin-like activity, hypercholesterolemia, extreme sensitivity to cold, and insulin resistance.l The adipose tissue of animals with this syndrome shows enormously increased capacity for lipogenesis from acetate despite decreased glucose uptake. 2 Fat mobilization in these animals is not increased by epinephrine or F.M.S .3 The syndrome may be due to the fact that the adipose tissue of these animals contains glycerokinase.4 Because these animals show a paradoxic response to alloxan5 with blood glucose decreased and appearance of massive granulation in beta cells of the islets of Langerhans, we decided to investigate the action of substances related chemically to alloxan-purines, pyrimidines and one alkaloid-caffeine. The effect of caffeine was so striking that it seems important to report it, particularly in view of our inability to find any study of the effect of this substance, or of coffee, in human beings with obesity and maturity onset diabetes, a condition which in many ways resembles the hereditary hyperglycemic syndrome. &SULTS
Toxicity Obese hyperglycemic mice were found to be much toxic effects of caffeine than normal animals. The L.D. 42 obese and 28 nonobese animals. Under conditions it was found to be 10 mg./animal for the lean animals weight) and 3.5 mg. for obese hyperglycemic animals From
the Department
of Nutrition,
Harvard
University
more sensitive to the 50 was determined in of ad libitum feeding (400 mg./Kg. of body (80 mg./Kg. of body
School
of Public
Health,
Bos-
ton, Massachusetts. This study was supported in part by a grant in aid from the National Institute of Arthritis and Metabolic Disease (AM-2911), National Institutes of Health, Bethesda, May&&; and by The Fund for Research and Teach&g of the Depatiment of Nut&ion, Harvard University School of Public Health, Boston, Massachusetts. Received
for publication
Apr.
13, 1964. 1369 METABOLISM, VOL. 13, No. 11 (NOVEMBER), 1984
1370
KUFTINEC
AND
MAYER
SENSITIVITY
OF OBESE HYPERGLYCEMIC
MICE
TO CAFFEINE
1371
weight). Unlike the situation with alloxan which was better tolerated after 24-48 hours of fasting, 5 deprivation of food increased toxicity of caffeine. In obese mice, survival rate, following administration of 2 mg. caffeine/animal after 24 hours fasting, was 72 per cent ( 18 of 25 injected animals survived); after 48 hours of food deprivation with the same dose of caffeine, only 10 per cent of obese animals survived (2 out of 20 injected animals). All lean caffeine-injected animals survived the injection of caffeine after 24 and 48 hours of fasting (lean animals were injected with 5 mg. caffeine/animal). Effect of Cafleine
on Blood Glucose
The effect of caffeine on blood glucose levels was remarkable. Sixty obese animals were injected with 2 mg. of caffeine in 0.3 ml. of distilled water, 40 lean animals with 5 mg. of caffeine in the same volume of distilled water. An obese-untreated group of 30 animal. F and nonobese-untreated group of 30 animals were injected with the same volume (0.3 ml/animal) of distilled water. Glucose determinations were done in duplicate by Somogyi-Nelson method modified for ultramicro determinations after 3, 6, 9, 12, 24 hours and 2, 4, 6, 10, 14, 30, 45, and 60 days following injection. Lean animals showed an initial drop from an average level of 132 mg./lOO ml. 2 14 to a lower level of 59 mg./lOO ml. * 12 after 6 hours and a minimum level of 50 mg./lOO ml. * 13 after 9 hours. Blood sugar values were back to normal after 24 hours and hereafter did not significantly differ from those of untreated animals. In obese animals the blood glucose level rose from 293 t 32 mg./lOO ml. to 487 * 85 mg./lOO ml. after 6 hours, 539 r+ 88 mg./lOO ml. after 12 hours, and reached a peak of 601 * 112 mg./lOO ml. after 4 days. From there on, the blood glucose of the caffeine-injected obese animals remained, with minor fluctuations, in the range of 500-600 mg./lOO ml. until the termination of the experiment, 60 days after injection. Blood sugar value was at that time 521 * 119 mg./lOO ml. Glucose and Insulin Tolerance
after Caffeine
Treatment
A glucose tolerance test was conducted by injecting 50 mg. of glucose/ animal intraperitoneally on the tenth day after injection of caffeine in 15 treated and 10 nontreated obese animals. At that time the treated group had blood sugars of 523 * 92 and the untreated group 287 * 42. Blood sugars failed to rise much in the caffeine-treated group (the 30 minute level was 595 2 87, the 60 minute level was 574 -t 91, and the 120 minute level, 544 * 98), while the nontreated group showed characteristically elevated glucose level curve. This failure of the caffeine-treated group to show a rise in blood sugar levels can probably be explained by the fact that in animals already having very high levels of glucose, additional glucose is very quickly spilled over in urine (a fact confirmed by urinalysis). Insulin tolerance tests were conducted on the fifteenth day in 15 fed and 15 fasted (24 hour) obese mice and 10 fed and 10 fasted untreated animals; one unit of regular crystalline insulin was the dose used. Results confirmed
1372
KUFTINEC
AND
MAYER
Table 2.-Glucose Tolerance Test in Cafjein-Treated and in Untreated Obese Hyperglycemic Mice -.. ~___.
__~
0
___ OH Exptl.
30'
____
120'
60'
523 s 92
595 xk 87
574 t 91
544 zk 108
287 t
425 k 78
462 r+ 92
303 rt 67
(15) OH Contr.
42
(IO) Figures following -t are standard deviations. Figures in parentheses are number of animals. 50 mg. glucose administered to each animal.
Table X--Insulin Tolerance in Nonfasted Cafleine-Treated and Mice _Untreated Obese Hyperglycemic _~____ -._
--____
-
6"'
30'
0 OH Exptl.
487 rt_ 71
412 I+ 72
493 zk 87
(15) OH Contr.
304 + 67
220 f
297 _” 56
(IO)
--___~.
..~
46
~__
-___
Table 3a.--Insulin Tolerance in Caffeine-Treated and Untreated Hyperglycemic Mice after 24-Hour Fast ~_____~ _~ --____ --. .-
60'
HO'
0 OH Exptl.
336 +- 74
325 t
82
(15) OH Contr.
128 F 32
65 t
17
(16)
____ -___ ..___~_ _.__~ Figures following t are standard deviations. Figures in parentheses are number of animals. 1 I.U. reg. insulin/animal.
Obese
290 zk 62 55 f
6
__ ~_
the previously established? resistance to inuslin of obese hyperglycemic mice, and do not indicate increased sensitivity to insulin after caffeine treatment. Histologic
Study
A group of 4 caffeine-treated and 4 nontreated obese animals was sacrificed on the day after caffeine injection. Histologic studies did not reveal the cause of the hyperglycemic effect of caffeine. The livers, adrenals, kidneys and pituitaries did not show any significant change. The pancreatic islets were even more degranulated than are those of the untreated obese hyperglycemic mice.* (We are indebted to Dr. S. B. Andrus for this histological examination of these animals.) The use of C-14 labeled cafieine (label in position 2) did not reveal incorporation of C-14 compound in the livers, pancreata, adrenals. pituitaries or kidneys of treated animals. “The
striking and apparently
reduction of blood glucose previous1y.a
paradoxic
effect of alloxan in this syndrome
and regranulation
of pancreatic
beta cells)
(permanent
has been reported
SENSITIVITY
OF OBESE HYPERGLYCEMIC
MICE
TO CAFFEINE
1373
Eflect of Cofee Finally, in order to find out whether or not the ingestion of coffee would have effects similar to those of caffeine injection, 30 obese and 20 nonobese mice were given coffee as their exclusive source of fluid. Coffee was prepared daily from “instant” powder to insure greater reproducibilty than could be obtained by percolation. The solution contained 1.33 Gm. coffee and 2 Cm. sugar/100 ml. of distilled water corresponding to 0.33 mg. of caffeine/ml.* The animals were maintained on ad libitum intake of coffee and laboratory (Purina) chow for 7 days. Daily intake of coffee was measured. Daily intakes of coffee ranged from 2.5 to 14.0 ml. with an average of 6.2 * 3.1 ml. Weekly intakes ranged from 21 to 58 ml. with an average of 43.5 z+ 12.1 ml. After 7 days, the drinking fluid was changed back to water. Blood glucose determinations were carried out daily during the coffee ingestion and 3, 6, 10, 14, 21, 30, 45, 60, 75, 90, 92, 94, 96, 98 and 100 days after the animals stopped drinking coffee. The blood sugar levels of the nonobese animals were not affected (average in the range of 100 to 135 mg./lOO ml. for both water and coffee group). The blood glucose of the obese animals rose gradually while they were drinking coffee from 285 -C 29 mg./lOO ml. to 395 * 52 mg./lOO ml. on the sixth day. Coffee was discontinued after 7 days and the animals were allowed to rest for 3 days in order to excrete most of the caifeine to enable us to establish the “late” effect of coffee intake. Blood sugar values continued to rise steadily to reach their peak 7 days after coffee had been discontinued. At that time, the value was 537 2 82 mg./lOO ml. Blood sugar values remained elevated, with minor fluctuations, in the range of 480-550 mg./lOO ml. until the 75th day of the experiment (or 68 days after coffee had been discontinued). The next blood glucose determination was done on the 90th day of the experiment and at that time, we found that blood sugar values had fallen to a very low level (128 2 37 mg./lOO ml.) From that day on, blood sugar values were checked every 2 days through day 100. At that time, the experiment was discontinued because in the last 5 days of the experiment, 18 of the remaining 24 animals had died. The average blood sugar level on day 100 was 108 * 54 mg./lOO ml. It might be of interest to report that the blood sugar levels in the animals drinking coffee appeared to rise in direct proportion to the amounts of coffee they had drunk (and hence of the caffeine they ingested), p being less than 0.01.
Other Compounds In order to find out whether or not this specific glucotropic effect of alloxan and/or caffeine is due to their specific chemical structure, we tested, on 10 animals each, other alloxan-like and caffeine-like compounds (uric acid, lactin and lactan, adenine, cytosine, guanine, hypoxanthine, uracil, thymine ‘We are grateful to Mr. Nevis Cook, of the Food and Drug Administration for the determination of the caffeine content of the type of instant coffee used in this experiment.
of replacement
by coffee.
108 F 32 279 2 34
94
294 2 48
of water
277 i
318 z!z 72
64
135 _’ 42 304 2 59 ~___~ ~~ -___.
128 + 37 286 I+ 42
509 i: 102
51
527 +- 99
Figures following f are standard deviations. Figures in parentheses are numbers of animals. 0 day is the beginning of the experiment, and the beginning ducted before any coffee was drunk.
OH Expd. OH Contr.
248 t
92
288 A 60
-
14
537 -c 82
10
of Water Mice
417 -c 76
90
265 + 39
6
395 -t 52
3
on Blood Glucose Levels of Replacement Blood Glucose of Obese Hyperglycemic
360 * 47
Effect
15
60
285 & 29
(20)
(30) OH Corm.
0
285 + 29
OH Exptl.
DWS
Table 4.-Long-Term
61
_____ 97
The
45
98
__~___. 100
was con-
108 & 54 302 rt 60
determination
114 I+ 34 308 -c 48 -__
-__
298 + 47
504 f. 72
initial glucose
96 + 47 299 f 59
96
.~_____
302 *
486 t
30
by Co%ee for 1 Week on the
sn~srrrvr~y
0~ OBESEHYPERGLYCEMICMICE TOCAFFEINE
1375
and xanthine) but found no significant long-term changes in blood sugar levels with any of these compounds. The only one to show some temporary rise in blood sugar levels was hypoxanthine where blood sugars rose from an initia1 302 * 47 mg./lOO ml. to 397 -+ 54 mg./lOO ml. on the third day after 4.2 mg./animal injection of hypoxanthine. Blood sugar levels rose further on the fifth day to 422 + 68 mg./lOO ml., but after 7 days after injection, they were back to normal, i.e., 310 I+ 24 mg./lOO ml. and remained in that range for 3 more weeks, at the end of which period, the experiment was discontinued.
DISCUSSION It is intriguing to note that in obese-hyperglycemic mice, caffeine in many ways has an action opposite to that previously described for alloxan. Further work is obviously necessary to establish the mechanism of this unexpected action of caffeine and the possible relevance of these findings to the management of maturity-onset diabetes in obese human subjects. The word possible shouId be underlined: we know at present of no evidence bearing on the effect of caffeine or coffee in human diabetes, but in view of the large consumption of coffee, this is obviously a subject which deserves further study. REFERENCES 1. Mayer, J.: The obese hyperglycemic syndrome of mice as an example of “metabolic” obesity. Am. J. Clin. Nutrition 8:712, 1960. 2. Christophe, J., Jeanrenaud, B., Mayer, J., and Renold, A.: In vitro metabolism of adipose tissue in obese hyperglycemic and goldthioglucose treated mice. II. Metabolism of pyruvate and acetate. J. Biol. Chem. 236:648, 1961. 3. Leboeuf, B., Lochaya, S., Leboeuf, N., Wood, F., Mayer, J., and Cahill, G.: Glucose metabolism and mobilization of fatty acids by adipose tissue from
obese mice. Am. J. Physiol. 201:19, 1961. 4. Treble, D., and Mayer, J.: Glycerokinase Activity in white adipose tissue of obese hyperglycemic mice. Nature 260~363, 1963. 5. Solomon, J. B., and Mayer, J.: Effect of alloxan on obese hyperglycemic mice. Nature 192:135, 1962. Ultra6. Natelson, C. C.: Nelson-Somogyi micro method in: Micro Techniques in Clinical Chemistry, 2nd ed. Springfield, Ill., Thomas, 1961, pp. 218-219.
Dubravko M. Kutfinec, M.D., Research Fellow in Nutrition, Department of Nutrition, Harvurd University School of Public Health, Boston, Mass. lean Mayer, Ph.D., D.Sc., Associate Professor of Nutrition, Department of Nutrition, Harvard University School of Public Health, Boston, Mass.
Sensitivity of Obese Hyperglycemic Mice to Caffeine and Coffee
By DUBRAVKOM. KUFTINEC AND JEAN MAYER Mice with the hereditary obese hyperglycemic syndrome are unusually sensi-
and pyrimidines other than alloxan (shown previously to cause in this syn-
tive to the toxic action of caffeine. A single injection of caffeine, or the replacement of drinking water by coffee for 1 week causes a drastic and prolonged (2 or more months) elevation of blood glucose. Administration of purines
drome prolonged lowering of blood glucase and regranulation of beta cells) did not lead to striking results. No explanation for this unexpected effect of caffeine is offered.
T
HE HEREDITARY RECESSIVE HYPERGLYCEMIC SYNDROME in mice is characterized inter alia by extreme adiposity, moderate hyperphagia but extreme inactivity, a high blood glucose despite high pancreatic and circulating insulin-like activity, hypercholesterolemia, extreme sensitivity to cold, and insulin resistance.l The adipose tissue of animals with this syndrome shows enormously increased capacity for lipogenesis from acetate despite decreased glucose uptake. 2 Fat mobilization in these animals is not increased by epinephrine or F.M.S .3 The syndrome may be due to the fact that the adipose tissue of these animals contains glycerokinase.4 Because these animals show a paradoxic response to alloxan5 with blood glucose decreased and appearance of massive granulation in beta cells of the islets of Langerhans, we decided to investigate the action of substances related chemically to alloxan-purines, pyrimidines and one alkaloid-caffeine. The effect of caffeine was so striking that it seems important to report it, particularly in view of our inability to find any study of the effect of this substance, or of coffee, in human beings with obesity and maturity onset diabetes, a condition which in many ways resembles the hereditary hyperglycemic syndrome. &SULTS
Toxicity Obese hyperglycemic mice were found to be much toxic effects of caffeine than normal animals. The L.D. 42 obese and 28 nonobese animals. Under conditions it was found to be 10 mg./animal for the lean animals weight) and 3.5 mg. for obese hyperglycemic animals From
the Department
of Nutrition,
Harvard
University
more sensitive to the 50 was determined in of ad libitum feeding (400 mg./Kg. of body (80 mg./Kg. of body
School
of Public
Health,
Bos-
ton, Massachusetts. This study was supported in part by a grant in aid from the National Institute of Arthritis and Metabolic Disease (AM-2911), National Institutes of Health, Bethesda, May&&; and by The Fund for Research and Teach&g of the Depatiment of Nut&ion, Harvard University School of Public Health, Boston, Massachusetts. Received
for publication
Apr.
13, 1964. 1369 METABOLISM, VOL. 13, No. 11 (NOVEMBER), 1984
1370
KUFTINEC
AND
MAYER
SENSITIVITY
OF OBESE HYPERGLYCEMIC
MICE
TO CAFFEINE
1371
weight). Unlike the situation with alloxan which was better tolerated after 24-48 hours of fasting, 5 deprivation of food increased toxicity of caffeine. In obese mice, survival rate, following administration of 2 mg. caffeine/animal after 24 hours fasting, was 72 per cent ( 18 of 25 injected animals survived); after 48 hours of food deprivation with the same dose of caffeine, only 10 per cent of obese animals survived (2 out of 20 injected animals). All lean caffeine-injected animals survived the injection of caffeine after 24 and 48 hours of fasting (lean animals were injected with 5 mg. caffeine/animal). Effect of Cafleine
on Blood Glucose
The effect of caffeine on blood glucose levels was remarkable. Sixty obese animals were injected with 2 mg. of caffeine in 0.3 ml. of distilled water, 40 lean animals with 5 mg. of caffeine in the same volume of distilled water. An obese-untreated group of 30 animal. F and nonobese-untreated group of 30 animals were injected with the same volume (0.3 ml/animal) of distilled water. Glucose determinations were done in duplicate by Somogyi-Nelson method modified for ultramicro determinations after 3, 6, 9, 12, 24 hours and 2, 4, 6, 10, 14, 30, 45, and 60 days following injection. Lean animals showed an initial drop from an average level of 132 mg./lOO ml. 2 14 to a lower level of 59 mg./lOO ml. * 12 after 6 hours and a minimum level of 50 mg./lOO ml. * 13 after 9 hours. Blood sugar values were back to normal after 24 hours and hereafter did not significantly differ from those of untreated animals. In obese animals the blood glucose level rose from 293 t 32 mg./lOO ml. to 487 * 85 mg./lOO ml. after 6 hours, 539 r+ 88 mg./lOO ml. after 12 hours, and reached a peak of 601 * 112 mg./lOO ml. after 4 days. From there on, the blood glucose of the caffeine-injected obese animals remained, with minor fluctuations, in the range of 500-600 mg./lOO ml. until the termination of the experiment, 60 days after injection. Blood sugar value was at that time 521 * 119 mg./lOO ml. Glucose and Insulin Tolerance
after Caffeine
Treatment
A glucose tolerance test was conducted by injecting 50 mg. of glucose/ animal intraperitoneally on the tenth day after injection of caffeine in 15 treated and 10 nontreated obese animals. At that time the treated group had blood sugars of 523 * 92 and the untreated group 287 * 42. Blood sugars failed to rise much in the caffeine-treated group (the 30 minute level was 595 2 87, the 60 minute level was 574 -t 91, and the 120 minute level, 544 * 98), while the nontreated group showed characteristically elevated glucose level curve. This failure of the caffeine-treated group to show a rise in blood sugar levels can probably be explained by the fact that in animals already having very high levels of glucose, additional glucose is very quickly spilled over in urine (a fact confirmed by urinalysis). Insulin tolerance tests were conducted on the fifteenth day in 15 fed and 15 fasted (24 hour) obese mice and 10 fed and 10 fasted untreated animals; one unit of regular crystalline insulin was the dose used. Results confirmed
1372
KUFTINEC
AND
MAYER
Table 2.-Glucose Tolerance Test in Cafjein-Treated and in Untreated Obese Hyperglycemic Mice -.. ~___.
__~
0
___ OH Exptl.
30'
____
120'
60'
523 s 92
595 xk 87
574 t 91
544 zk 108
287 t
425 k 78
462 r+ 92
303 rt 67
(15) OH Contr.
42
(IO) Figures following -t are standard deviations. Figures in parentheses are number of animals. 50 mg. glucose administered to each animal.
Table X--Insulin Tolerance in Nonfasted Cafleine-Treated and Mice _Untreated Obese Hyperglycemic _~____ -._
--____
-
6"'
30'
0 OH Exptl.
487 rt_ 71
412 I+ 72
493 zk 87
(15) OH Contr.
304 + 67
220 f
297 _” 56
(IO)
--___~.
..~
46
~__
-___
Table 3a.--Insulin Tolerance in Caffeine-Treated and Untreated Hyperglycemic Mice after 24-Hour Fast ~_____~ _~ --____ --. .-
60'
HO'
0 OH Exptl.
336 +- 74
325 t
82
(15) OH Contr.
128 F 32
65 t
17
(16)
____ -___ ..___~_ _.__~ Figures following t are standard deviations. Figures in parentheses are number of animals. 1 I.U. reg. insulin/animal.
Obese
290 zk 62 55 f
6
__ ~_
the previously established? resistance to inuslin of obese hyperglycemic mice, and do not indicate increased sensitivity to insulin after caffeine treatment. Histologic
Study
A group of 4 caffeine-treated and 4 nontreated obese animals was sacrificed on the day after caffeine injection. Histologic studies did not reveal the cause of the hyperglycemic effect of caffeine. The livers, adrenals, kidneys and pituitaries did not show any significant change. The pancreatic islets were even more degranulated than are those of the untreated obese hyperglycemic mice.* (We are indebted to Dr. S. B. Andrus for this histological examination of these animals.) The use of C-14 labeled cafieine (label in position 2) did not reveal incorporation of C-14 compound in the livers, pancreata, adrenals. pituitaries or kidneys of treated animals. “The
striking and apparently
reduction of blood glucose previous1y.a
paradoxic
effect of alloxan in this syndrome
and regranulation
of pancreatic
beta cells)
(permanent
has been reported
SENSITIVITY
OF OBESE HYPERGLYCEMIC
MICE
TO CAFFEINE
1373
Eflect of Cofee Finally, in order to find out whether or not the ingestion of coffee would have effects similar to those of caffeine injection, 30 obese and 20 nonobese mice were given coffee as their exclusive source of fluid. Coffee was prepared daily from “instant” powder to insure greater reproducibilty than could be obtained by percolation. The solution contained 1.33 Gm. coffee and 2 Cm. sugar/100 ml. of distilled water corresponding to 0.33 mg. of caffeine/ml.* The animals were maintained on ad libitum intake of coffee and laboratory (Purina) chow for 7 days. Daily intake of coffee was measured. Daily intakes of coffee ranged from 2.5 to 14.0 ml. with an average of 6.2 * 3.1 ml. Weekly intakes ranged from 21 to 58 ml. with an average of 43.5 z+ 12.1 ml. After 7 days, the drinking fluid was changed back to water. Blood glucose determinations were carried out daily during the coffee ingestion and 3, 6, 10, 14, 21, 30, 45, 60, 75, 90, 92, 94, 96, 98 and 100 days after the animals stopped drinking coffee. The blood sugar levels of the nonobese animals were not affected (average in the range of 100 to 135 mg./lOO ml. for both water and coffee group). The blood glucose of the obese animals rose gradually while they were drinking coffee from 285 -C 29 mg./lOO ml. to 395 * 52 mg./lOO ml. on the sixth day. Coffee was discontinued after 7 days and the animals were allowed to rest for 3 days in order to excrete most of the caifeine to enable us to establish the “late” effect of coffee intake. Blood sugar values continued to rise steadily to reach their peak 7 days after coffee had been discontinued. At that time, the value was 537 2 82 mg./lOO ml. Blood sugar values remained elevated, with minor fluctuations, in the range of 480-550 mg./lOO ml. until the 75th day of the experiment (or 68 days after coffee had been discontinued). The next blood glucose determination was done on the 90th day of the experiment and at that time, we found that blood sugar values had fallen to a very low level (128 2 37 mg./lOO ml.) From that day on, blood sugar values were checked every 2 days through day 100. At that time, the experiment was discontinued because in the last 5 days of the experiment, 18 of the remaining 24 animals had died. The average blood sugar level on day 100 was 108 * 54 mg./lOO ml. It might be of interest to report that the blood sugar levels in the animals drinking coffee appeared to rise in direct proportion to the amounts of coffee they had drunk (and hence of the caffeine they ingested), p being less than 0.01.
Other Compounds In order to find out whether or not this specific glucotropic effect of alloxan and/or caffeine is due to their specific chemical structure, we tested, on 10 animals each, other alloxan-like and caffeine-like compounds (uric acid, lactin and lactan, adenine, cytosine, guanine, hypoxanthine, uracil, thymine ‘We are grateful to Mr. Nevis Cook, of the Food and Drug Administration for the determination of the caffeine content of the type of instant coffee used in this experiment.
of replacement
by coffee.
108 F 32 279 2 34
94
294 2 48
of water
277 i
318 z!z 72
64
135 _’ 42 304 2 59 ~___~ ~~ -___.
128 + 37 286 I+ 42
509 i: 102
51
527 +- 99
Figures following f are standard deviations. Figures in parentheses are numbers of animals. 0 day is the beginning of the experiment, and the beginning ducted before any coffee was drunk.
OH Expd. OH Contr.
248 t
92
288 A 60
-
14
537 -c 82
10
of Water Mice
417 -c 76
90
265 + 39
6
395 -t 52
3
on Blood Glucose Levels of Replacement Blood Glucose of Obese Hyperglycemic
360 * 47
Effect
15
60
285 & 29
(20)
(30) OH Corm.
0
285 + 29
OH Exptl.
DWS
Table 4.-Long-Term
61
_____ 97
The
45
98
__~___. 100
was con-
108 & 54 302 rt 60
determination
114 I+ 34 308 -c 48 -__
-__
298 + 47
504 f. 72
initial glucose
96 + 47 299 f 59
96
.~_____
302 *
486 t
30
by Co%ee for 1 Week on the
sn~srrrvr~y
0~ OBESEHYPERGLYCEMICMICE TOCAFFEINE
1375
and xanthine) but found no significant long-term changes in blood sugar levels with any of these compounds. The only one to show some temporary rise in blood sugar levels was hypoxanthine where blood sugars rose from an initia1 302 * 47 mg./lOO ml. to 397 -+ 54 mg./lOO ml. on the third day after 4.2 mg./animal injection of hypoxanthine. Blood sugar levels rose further on the fifth day to 422 + 68 mg./lOO ml., but after 7 days after injection, they were back to normal, i.e., 310 I+ 24 mg./lOO ml. and remained in that range for 3 more weeks, at the end of which period, the experiment was discontinued.
DISCUSSION It is intriguing to note that in obese-hyperglycemic mice, caffeine in many ways has an action opposite to that previously described for alloxan. Further work is obviously necessary to establish the mechanism of this unexpected action of caffeine and the possible relevance of these findings to the management of maturity-onset diabetes in obese human subjects. The word possible shouId be underlined: we know at present of no evidence bearing on the effect of caffeine or coffee in human diabetes, but in view of the large consumption of coffee, this is obviously a subject which deserves further study. REFERENCES 1. Mayer, J.: The obese hyperglycemic syndrome of mice as an example of “metabolic” obesity. Am. J. Clin. Nutrition 8:712, 1960. 2. Christophe, J., Jeanrenaud, B., Mayer, J., and Renold, A.: In vitro metabolism of adipose tissue in obese hyperglycemic and goldthioglucose treated mice. II. Metabolism of pyruvate and acetate. J. Biol. Chem. 236:648, 1961. 3. Leboeuf, B., Lochaya, S., Leboeuf, N., Wood, F., Mayer, J., and Cahill, G.: Glucose metabolism and mobilization of fatty acids by adipose tissue from
obese mice. Am. J. Physiol. 201:19, 1961. 4. Treble, D., and Mayer, J.: Glycerokinase Activity in white adipose tissue of obese hyperglycemic mice. Nature 260~363, 1963. 5. Solomon, J. B., and Mayer, J.: Effect of alloxan on obese hyperglycemic mice. Nature 192:135, 1962. Ultra6. Natelson, C. C.: Nelson-Somogyi micro method in: Micro Techniques in Clinical Chemistry, 2nd ed. Springfield, Ill., Thomas, 1961, pp. 218-219.
Dubravko M. Kutfinec, M.D., Research Fellow in Nutrition, Department of Nutrition, Harvurd University School of Public Health, Boston, Mass. lean Mayer, Ph.D., D.Sc., Associate Professor of Nutrition, Department of Nutrition, Harvard University School of Public Health, Boston, Mass.