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Influence of nicotinic acid on the rates of turnover and oxidation of plasma glucose in man
Influence of Nicotinic Acid on the Rates of Turnover and Oxidation of Plasma Glucose in Man By E. 0. Balasse The effects of antilipolysis induced by nicotinic acid on the rates of turnover and oxidation of plasma glucose were studied in normal overnight fasted, obese overnight fasted, and obese starved subjects, using a “C-glucose infusion technique. Changes induced by nicotinic acid were similar whatever the nutritional state of the subjects. Plasma FFA levels and blood concentrations of glycerol and ketones decreased by about 60%; glycemia remained essentially unchanged but both the removal rate of plasma glucose and the hepatic glucose output increased by about 25%. Moreover, the fraction of glucose taken up by tissues and promptly oxidized and the fraction of expired COz derived from plasma glucose increased, respectively, by 16% and 33%. This enhancement
and M. A. Neef of glucose utilization occurred despite a small but significant decrease in plasma IRI concentration, indicating that nicotinic acid increased sensitivity to insulin. The above-mentioned results were obtained in 6 of the 10 patients studied. The remaining two subjects did not respond to the administration of nicotinic acid by any significant decrease in FFA nor in glycerol concentrations and showed no change in the rates of glucose turnover and oxidation. These data indicate that the effects of nicotinic acid on glucose metabolism may be partly mediated through changes in plasma FFA concentration and are consistent with the idea that the “glucose-fatty acid cycle” plays a significant role in the control of glucose metabolism in man.
Tp
HE “GLUCOSE-FATTY ACID CYCLE” theory proposed by Randle et al.’ almost 10 yr ago, has considerably stimulated further research on the interactions between lipid and carbohydrate metabolism in vivo. In an attempt to evaluate the possible role of plasma FFA in the regulation of glucose utilization, many authors have studied the effects of experimentally induced elevations or reductions in FFA levels on glucose tolerance.2-9 However, this approach has led to very conflicting results and the operation of the “glucosefatty acid cycle” as a factor regulating overall glucose metabolism in the intact organism is still questionable.‘O+” On the other hand, studies on the effects of FFA modifications on plasma glucose turnover and oxidation under normoglycemic conditions are limited in number’2T’3 and have never been performed in humans.
1
From the Metabolic Unit and the Laborator?, of Experimental Medicine, University of Brussels, Brussels, Belgium. Presented in part at the 8th Annual Meeting of the European Association for the Study of Diabetes held in Madrid, Spain. September 1972. Received for publication January 18. 1973. Supported by the Fonds de la Recherche Scientifque Medicale (Grant 20193) and by the Ministere Belge de la Politique Scientifique, within the framework of the Association Euratom- University of Brussels- University of Pisa. E. 0. Balasse, M.D., Ph.D.: Associate Professor in Medicine, Metabolic Unit and Laboratory of Experimental Medicine, University of Brussels, Brussels. Belgium. M. A. Neef, B.A.: Research Technician, Laboratory of Experimental Medicine, University of Brussels, Brussels, Belgium. Reprint requests should be addressed to E. 0. Balasse. Laboratory of Experimental Medicine, University of Brussels, 115. Bld de Waterloo, 1000 Brussels, Belgium. 01973 bv Grune & Stratton, Inc.
Metabolism, Vol. 22, No. 9 (September). 1973
1193
Subjects
109
150
176
181
OEC
RIN
MAH
VAN
cent change
‘On the left
l SEM
1.60
0.52
0.246
0.205
0.078
0.42
6.73
7.68
represent acetoacetate
3
72
74
79
72
64
3
87
92
83
89
91
so
75
>o 05
3.6
+ 1.8
84
77
83
76
85
83
84
90
83
77
ml
+ B-hydroxybutyrate.
in each test.
actid by a significant
value obtained
all subjects except VAH and HAN who did not respond to nicotinic
last expertmental
as above buf hgures correspond
Blood ketone conce”trat,ons
7 8.2
-73
7.02
6 18
571
4.26 4.22
7.06 6.47
t Same presentation
to the
3.9
-62.9
0.182
0.041
0.019
0.016
0.017
0.009
0.046
Plasma
mg/lOO
Glucose’
Concentration
6.7
-30.4
-
25
10.3
0.0
3.0
11.0
10.0
1.7
3 7
7
73
93
74
63
61
2
123
124
123
II8
128
118
127
98
73
94
85
158
139
149
139
150
151
Results)
52.2 47 5
50.0 39.3
13.3
19.0
19.8
32.5 29.8
45.4 30.1 3.4
co.01
3.6
+18X
34.0
32.4
36.5
48.8
37.9
. 4.9
b32.6
93
11.3
14.7
13.9
19.2
21 .o
27.2
25.2
25.3
32.9
values obtamed
14
Il.4
9.4
15.4
9.8
11.1
1.9
572
46.7
19.1
3.0
50.0
37.9
17.3
27.7
19.4
49.7
41 2
Glucose
from
Per cent co2
38.6
53.4
46.2
Oxidized
GlUCOSl3
Per cent
Glucose Oxldationt
(Individual
of the three expenmental
~0.001
4.6
+26.1
mg/mm
Rate
TUrflOVer
Glucose’
Metabolism
On the right. mean
3.3
6.4
11.2
1.3
13.0
0.0
2.1
8.8
5.7
16.0
15.0
4.7
5.0
6.3
PlaSma
fllJ/illl
5.0
Glucose
IRI’
and
drop in FFA nor in ketones.
during the last 60 mm of the control experiments.
0.013
0.141
0.164
0118
0.140
0.068
0.029
0.084
values obtamed
4.6
-60.3
1.31
I.23
0.50
0.032
0.082
0.032
0.019
0.089
0.120
0.004
0.029
0.087
0.273
0.027
Blood
0.1 19
0.174
0.052
0.087
0023
0.075 0.094
Blood
#mOleS/ml
#moles/ml
Metabolite
Ketones’
on Blood
Glycerol’
Acid
l This mean mcludes
with nicotintc acid
mean of the three experimental
from control t
per
SEM
mm of the experiments
P
14.7
1.18
1.31
190
HAN
14.2
1.26
1.20
0.05
0.10
122
VAH
149
16.7
1.1
0.20
0.88
12.1
0.80
0.35
0.83
149
0.26
0.75
14.5
12 1
0 50
f
129
DIE
0.13 0.30
0.87
0.86
0.58
Pl.3Sma
&moles/ml
15.7
16.2
18.9
of Nicotinic
FFA’
Effects
Mean
109
RAU
Starved
+ SEM
Meall
95
103
mg/kg
Body Weight
DES
Nicotinic Aod
of Ideal
GAU
fasted
Overnight
DOS?
1.
Per cent
Table
dunng
4.4
+12.9
10.52
6.99
7.23
9.92
13.08
1 1.52
10.39
9.19
9.80
8.19
the last 60
0.73
7.77
9.96
7.23
6.94
6.96
0.7
9.49
12.19
10.76
9.26
8.41
9.32
7.02
mmoles/mm
CO2 Production
L
NICOTINE
ACID AND
PLASMA
GLUCOSE
IN MAN
1195
The aim of the present work was to determine whether an acute reduction in plasma FFA levels induced by nicotinic acid modifies the rates of turnover and oxidation of circulating glucose in man, as measured by use of a “C-glucose infusion technique. These studies have been carried out in normal postabsorptive, in obese postabsorptive, and starved patients. These two last groups of subjects were included in the study because it has been suggested that elevated FFA levels might be responsible-at least partly-for the abnormalities in carbohydrate metabolism accompanying obesity and food deprivation.14 MATERIALS
AND
METHODS
Ten female subjects aged 17-34 yr were selected for the study. Their body weight varied between 95% and 190% of ideal weight (Table 1) and all had normal oral glucose tolerance. Six ambulatory subjects (three normal and three obese) were studied after an overnight fast of 15 hr and four obese hospitalized patients were studied after IO- I 1 days of complete starvation during which their intake consisted only of calorie-free beverages, vitamins, and minerals. For all subjects, only water was allowed during the 15 hr preceding the tests. The studies were conducted according to the following protocol (Fig. 1). After a 30-min basal NICOTINIC
OR
ACID
SALINE
o_
.
-0
-
11111111111 l_“C PLASMA
FFA
(pmoles /ml)
0.00 0.70
0.50 0.30
BLOOD GLYCEROL
IRI
(pU /ml) PLASMA / lOOmI)
PLASMA l.“C.GLUCOSE
GLUCOSE
S.A.
CO2
(mmoles/min)
cose
metabolism.
experiment fasted
antilipolysis acid
subject
(DES). CO2
glucose
specific
merator
was and
dpm&atoms
g C.
glu-
RESPIR.
‘4co2
specific activity
activity/ the in
denominator
~0 *O 3 600
I
500
I I I
D--
0
SO
-o__-o---o___0
600
10.0
9.0 1 0.50 0.40 0.30 0.20
INFUSED
overnight
14C
nudpm/ in
0.10 0
For calculat-
expressed the
on
Representative
in a normal
ing the ratio
pmoles
of
nicotinic
10 01
500 400 1
RESPIRATORY
Effect
GO5
300 I
(dpm/mg)
by
I
400
(dpm/m!)
1.
J_______.___._._o___~___~
100
GLUCOSE
Fig.
&-----
0 I
PLASMA
induced
OLUCOSE
0.10
(Pmolas/ml)
(mg
-
CO2 GLUCOSE
S.A. S.A
0.30 0.20 0.10
0 I
-30
120 MIN180
1196
BALASSE
AND
NEEF
period, I-‘4C~glucose (Amersham. England; specific activity 0.2 mCi/mmole), diluted in saline. was infused for 3 hr in an antecubital vein at the constant rate of about 0.15 pCi/kg. hr. A priming dose of radioactivity equivalent to the amount infused in 130 min was given at the start of the infusion. Blood samples were obtained at intervals from an indwelling plastic needle located in an antecubital vein of the opposite arm, transferred into heparinized tubes kept on ice, and processed promptly. Expired air was collected intermittently for 445 min periods in rubber bags through a low resistance valve and analyzed immediately. Each subject was studied on two separate occasions: (I) in the control state: (2) during the administration of nicotinic acid (Niacin, UCB. Belgium) which was injected as i.v. pulses of 100 or 150 mg every 20 min during the whole study, the first dose being given 30 min before the start of the 14C -glucose infusion. The highest dosage (150 mg) was used only in the two most obese patients (VAN and HAN). The total amount of nicotinic acid varied between 12.1 and 18.9 mg/kg (Table I .) In the control tests. nicotinic acid injections were replaced by saline injections. The control study always preceded the nicotinic acid study in order to avoid possible interference of the rebound in FFA levels which is known to occur after administration of this drug. The time interval between the two tests was I 3 days in the postabsorptive subjects who were instructed not to change their usual diet hetween the two successive studies. in the starved patients. the tests were always performed on 2 consecutive days so that closely similar nutritional conditions would prevail during the two successive experiments. Plasma concentration of glucose, FFA. and immunoreactive insulin (IRI): blood concentrations of glycerol. fl-hydroxyhutyrate. and acetoacetate; and specific activity of expired CO, were determined as indicated elsewhere.” Plasma I-‘4C~glucose was isolated as “C02 bv a fermentation method using Leuconortoc mesenteroides 16.17;the 14C02 was trapped in hyamine and counted For present in the plasma sample before fereach sample, the amount of 14C0, + 14Cbicarbonate mentation was determined on a separate aliquot and suhstracted from total 14C0, recovered after fermentation. The recovery of I-‘4C~glucose added to nonradioactive plasma was tested with each series of determinations and varied between 88.3”;, and 98.9”” (average 93 O”.,). rad’oactiv’ty in blood samples was corrected accordingly. For the test performed during nicotinic acid administration. a sample of expired a’r was collected before the start of the 14C~ glucose infusion in order to determine the residual “C02 concentration resulting from the preceding experiment and this residual activity was substracted from each determinat’on obtained during the 14C glucose infusion. Blood and expired-a’r analyses were ,111made in duplicate. Except for a cutaneous flush, nicotinic acid produced no discomfort for the pat’ents during the course of the studies. However, most of the subjects. especially the starved ones, felt nauseous a few hours after the termination of the experiments.
CALCULATIONS
Glucose Transport In both the control and the nicotinic acid studies, steady plasma concentrations of labeled and unlabeled glucose were obtained during the last 2 hr of the study allowing measurements of glucose kinetics. The rate of turnover of plasma glucose (i.e., the rate of influx or efflux of plasma glucose) was obtained as follows: F=
glucose
I specific activity’
where F is the glucose turnover rate (mg/min); I is the rate of infusion of labeled glucose (dpm/min); glucose specific activity is the mean of three determinations of plasma glucose specific activity (dpm/mg) obtained during the last 60 min of the “C-glucose infusion. The utilization of plasma glucose by tissues was expressed as glucose clear-
NICOTINIC
ACID AND
PLASMA
ante which represents plasma and the plasma
c, =
GLUCOSE
the ratio between the rate glucose concentration, F
glucose
1197
IN MAN
of efflux
of glucose
from
I
concentration
= i4C-glucose
concentration
’
where C, is the glucose clearance (ml/min); F is the turnover rate of glucose (mg/min); glucose concentration is the concentration of unlabeled glucose in plasma (mg/ml); I is the rate of infusion of labeled glucose (dpm/min); 14Cglucose concentration is the mean of three determinations of radioactive glucose concentration in plasma (dpm/ml) during the last 60 min of the study. Glucose Oxidation During I-r4glucose infusion, the efflux of i4C0, and the specific activity of expired CO1 rose in the form of exponential curves (Fig. 1) which did not completely attain their asymptotic values during the course of the studies. This is a general observation in this type of study and is related to a gradual increase in the labeling of intermediate metabolic pools, including the bicarbonate pool of the body. The fraction of plasma glucose converted to CO, and the per cent CO, derived from plasma glucose were calculated by the following equations: Per cent glucose
to CO* =
Efflux rate of i4C0, (dpm/min) Influx rate of *4C-glucose
Per cent CO, from glucose
=
x 100
(dpm/min)
’
CO, specific activity (dpm/pmole) x6x 100 Plasma glucose specific activity (dpm/pmole)
’
In these equations, the rate of efflux of i4C0, and the specific activity of CO, were the last values obtained during the studies. Since, as already mentioned, these values are lower than asymptotic values, the figures obtained from these equations slightly underestimate glucose oxidation, RESULTS
Representative
Experiment
Data obtained in a representative experiment on a normal weight overnight fasted subject (DES) are presented in Fig. 1. The FFA and glycerol levels measured during the nicotinic acid study amounted to about 40% of values observed in the control study. Nicotinic acid produced only slight changes in glycemia, in IRI concentration, and in cold CO, production. The plateaus of plasma 14Cglucose concentration and of glucose specific activity were about 25% lower than corresponding values observed in the basal state indicating that both the clearance of plasma glucose by tissues and the hepatic glucose output were stimulated in the presence of lowered FFA. Nicotinic acid also stimulated glucose oxidation so that the rate of 14C0, production increased by 20% and the ratio of CO, specific activity to plasma glucose specific activity increased by 46%. These results indicate that nicotinic acid enhanced both the conversion of glucose to CO, and the per cent CO, derived from plasma glucose.
BALASSE
1198
Individual
AND
NEEF
Data
Table 1 provides individual values of blood metabolite concentration, IRI, and glucose turnover and oxidation in the absence and in the presence of nicotinic acid. Control Studies Overnight fastedpatients. In this group, basal values were 0580.88 pmole/ ml for FFA, 0.075-0.099 pmole/ml for glycerol, 0.078-0.273 pmole/ml for ketones, and 75-92 mg/ 100 ml for glycemia. Basal IRI concentration was below 10 pU/ml except for two obese patients (RIN and MAH) whose levels were, respectively, 15.0 and 16.0 PI-I/ml. Small individual variations were observed among values of the glucose turnover rate (118-127 mg/min). The fraction of glucose taken up and promptly oxidized varied between 37.9% and 50.0% thereby providing 13.3x-27.7% of total CO, production. Unlabeled CO, output ranged between 7.02 and 12.19 mmole/min and tended to be positively correlated with excess body weight. Starved patients. In the starved patients, the concentrations of FFA (1.181.60 pmole/ml), glycerol (0.118-0.164 pmole/ml), and ketones (5.71-7.68 pmole/ml) were significantly higher (p < 0.001) than those observed in the overnight fasted group and glycemia (64-79 mg/ 100 ml) was significantly lower (0.001 < p < 0.01). The glucose turnover rate (61-93 mg/min) and the per cent CO, derived from glucose (9.4x-15.4%) were significantly reduced in the starved subjects as compared with the postabsorptive subjects (p values being, respectively, < 0.001 and < 0.02) but the per cent glucose converted to CO, and the unlabeled CO, production were similar in the two groups. Studies
With Nicotinic Acid
The administration of nicotinic acid strongly inhibited lipolysis in 8 of the 10 patients studied, including the six overnight fasted and two of the four starved patients (RAU and DIE). The average fall in FFA, glycerol, and ketone concentration observed in these subjects amounted, respectively, to 60.3% f 4.6x, 62.9% f 3.9x, and 73.7% + 8.2%. Glycemia did not change significantly but IRI decreased by 30.4% * 6.7%. The inhibition of lipolysis was accompanied in each subject by a rise in the rate of glucose turnover averaging 26.1% + 4.6%. The removal of plasma glucose was also systematically accelerated, the observed rise in plasma glucose clearance amounting to 24.0% f 4.1%. Nicotinic acid also significantly enhanced the oxidation of glucose: The fraction of glucose taken up and promptly oxidized increased by 18.6% f 3.6% and the fraction of total CO, derived from plasma glucose increased by 32.6% f 4.9%. Unlabeled CO, production was also slightly stimulated under these conditions (+12.9x f 4.4%). The two remaining starved patients (VAH and HAN) did not respond to nicotinic acid by a significant inhibition of adipose tissue lipolysis, although these subjects were given amounts of nicotinic acid similar to those given in the responders (Table 1). In subject VAH, there is even a slight rise in FFA and ketone concentration; under these conditions, the glucose turnover rate re-
NICOTINIC
ACID AND PLASMA
GLUCOSE
IN MAN
1199
mained unchanged and oxidation decreased. In the other patient (HAN), a small decrease in FFA and ketone levels was observed after nicotinic acid but blood glycerol rose slightly and the rates of glucose turnover and oxidation remained unaffected. Correlation
Between
Glucose Utilization
and Plasma FFA Levels
Figure 2 shows that a significant negative correlation has been found the various parameters of glucose metabolism (rate of turnover, per cose oxidized, and per cent CO2 derived from glucose) on one hand, responding plasma FFA concentration on the other hand. The data clude the figures obtained both in the absence and in the presence of acid.
between cent gluand corused innicotinic
DISCUSSION
Control values of the glucose turnover rate were higher in the overnight fasted patients than in the starved subjects and both groups of figures were of
60, so-
0
B Oo
\ 0
, .
.
LO_ : 30-
Fig. 2. Correlation between plasma FFA concentration and glucose turnover rate (panel A) or glucose oxidation (panels B and Cl in a group of patients including normal overnight fasted, obese overnight fssted. and obese starved subjects (see Table 1). Each subject was studied under control conditions (closed circles) end during administration of nicotinic acid (open circles). Statistical significance: (A), = - 0.78, A < 0.001; (6) r = -0.83. JJ< 0.001; (C) r = - 0.80, /J < 0.001.
0
. 00
0
0 0.2 0.6 I.0 I.6 1.L PLASMA FFA (pmoles/ml)
1200
BALASSE
AND
NEEF
the order of magnitude of those published by others using either isotope dilutions techniques’*-*’ or liver catheterization.*’ On the other hand, our estimates of glucose oxiation rates were about 20”/,-2%; lower than those published by Paul et al.*’ This difference is probably due to the fact that these authors used prolonged isotopic infusions (63 hr) and were therefore able to attain the true asymptotic value of the 14C0, curves. As mentioned earlier, this was not the case in our studies so that our results systematically underestimated glucose oxidation. The purpose of this study was to analyze the effects of a decrease in FFA concentration on these parameters of glucose metabolism. Antilipolysis was induced by a pharmacologic mean consisting of repeated nicotinic acid injections. In two of the four starved patients studied, nicotinic acid failed to decrease FFA levels thus indicating individual variations in the sensitivity to this drug. Obviously, the results obtained in these two patients do not provide any information regarding the influence of decreased FFA levels on glucose metabolism, and will therefore be analyzed separately. In the eight other patients (six overnight fasted and two starved), the administration of nicotinic acid resulted in a 60”/0 reduction in plasma FFA concentration and in a parallel decrease in ketone levels. The fall in ketones has to be taken into consideration especially in starved patients who rely on this substrate to a great extent for the provision of energy. After nicotinic acid, other fuels have to compensate for the reduced availability of FFA and ketones and our results demonstrate that circulating glucose plays a role of alternate fuel under these circumstances. Indeed. the clearance of plasma glucose by tissues was increased by about 24Y,, this change being entirely compensated by a corresponding rise in the hepatic glucose output so that, on an average, glycemia remained unaffected. Simultaneously, nicotinic acid induced a small but significant decrease in IRI concentration. This latter effect is presumably related to the fall in the level of plasma FFA which is known to promote insulin secretion in vivoi3.‘* and in vitro23; a direct inhibitory effect of nicotinic acid on P-cell secretion is unlikely according to in vitro data.24 The stimulation of glucose uptake in the presence of a decreased concentration of insulin in plasma indicates that the reduced availability of FFA enhanced insulin sensitivity. Our experiments provide no information on the type of tissue implicated in the increased glucose consumption. On the basis of in vitro data,’ cardiac and skeletal muscle are likely to be the primary site of this effect. Lassers et al.*’ have demonstrated that the antilipolytic effect of nicotinic acid in man is accompanied by an increase in the myocardial arterio- venous difference of glucose. Unfortunately, coronary blood flows were not measured in these experiments and absolute amounts of glucose extracted cannot be calculated. However, assuming that the coronary blood flow was 200 ml/min and was unaffected by nicotinic acid, it can be estimated from their data that the increase in myocardinal extraction of glucose after nicotinic acid amounted to 5--10 mg/min. This could account for about one-third of the increase in the total glucose utilization observed in our experiments. Skeletal muscle probably also participates in the increased glucose consumption. Have1 et a.26 have shown that glucose extraction by leg muscle is stimulated after administration of an anti-
NICOTINIC
ACID AND
PLASMA
GLUCOSE
IN MAN
1201
lipolytic compound but these results have been obtained during exercise and it is not known whether this applies to resting muscle. Finally, adipose tissue might also contribute to the increased uptake of glucose observed in our study, since nicotinic acid is known to stimulate glucose metabolism in this tissue.27 However, the participation of fat cells is likely to be small owing to the fact that adipose tissue participates very little in over-all glucose uptake.” If we assume that the increased uptake of glucose after nicotinic acid was essentially limited to cardiac and skeletal muscle, it can be calculated that the apparently small increase (24%) in glucose turnover rate corresponds in fact to a striking stimulation of glucose utilization by muscular tissues. Indeed, it is known that in the postabsorptive state, the uptake of glucose by cardiac and skeletal muscle does probably not exceed 30 g/24 hr.40 In our ove rnight fasted patients, this represents only a minor fraction of the over-all basal glucose consumption which approximated 177 g/24 hr. The major part of this glucose uptake is known to be accounted for by tissues for which glucose is an obligatory substrate (i.e., brain, blood cells, and renal medulla). Since the absolute increase in glucose turnover rate observed after nicotinic acid in our overnight fasted patients amounted to about 36 g/24 hr, it would appear that the uptake of plasma glucose by muscular tissues has approximately doubled during experimental antilipolysis. The increase in the hepatic glucose output observed in our experiments has also been observed in vitro on perfused livers from rats pretreated with nicotinic acid or related compounds. These liver preparations showed an increased rate of glycogenolysis29 and gluconeogenesi?’ as compared with livers from normal animals. This effect cannot be explained by a direct action of the drug on the liver since the addition of nicotinic acid to the perfusion medium of livers from normal rats is without effect3’ or even inhibits32 the rates of glycogenolysis and gluconeogenesis. It is also unlikely that the observed stimulation of hepatic glucose output after administration of nicotinic acid in vivo was related to the decrease in supply of FFA to the liver, since FFA have been shown to stimulate gluconeogenesis in the perfused liver preparation.33 All these reasons suggest that the enhancement of the hepatic glucose production observed was related to the hormonal changes associated with antilipolysis. One of these changes is the decrease in IRI concentration observed in these studies. Another possible factor is an increase in plasma glucagon levels. Luyckx et a1.34 have shown indeed that a rise in glucagon levels is observed in dogs whose FFA concentrations have been lowered with nicotinic acid. After nicotinic acid, the percentage of total CO, output derived from glucose oxidation was increased by 32.6%. This resulted from the combination of an augmented glucose turnover rate and an increased fractional conversion of glucose to CO,. There was also a concomitant rise in unlabeled CO, production which does probably not correspond to an augmented metabolic rate. Indeed, assuming that the oxygen consumption remained unchanged, an increase in CO, output was to be expected as a consequence of a rise in R.Q. reflecting the shift towards carbohydrate oxidation induced by nicotinic acid. Taken as a whole, our results indicate that plasma glucose can serve as alternate fuel when availability of FFA to tissues is reduced. However, this conclu-
BALASSE
1202
AND
NEEF
sion has to be taken cautiously with regard to the starved subjects, since only two of them could be adequately studied. Our interpretation of the data is based on the assumption that the effects of nicotinic acid on glucose metabolism is mediated through its antilipolytic properties and does not represent a direct pharmacologic effect of the drug. This concept is supported by the fact that in the two starved patients who did not respond to nicotinic acid by a fall in FFA concentration, no increase in the rates of turnover and oxidation of plasma glucose was observed. Circulating glucose is not the only substrate which is called upon by the body to meet the energy requirements in the presence of lowered FFA levels. It has been shown that muscle glycogen3sv36and stored triglyceride$’ are other possible sources of fuel under these conditions. Numerous studies have been devoted to the effects of antilipolytic substances on overall glucose metabolism in vivo. Most of them were based on the use of oral or i.v. glucose tolerance tests and yielded contradictory results.2-9 A few animal studies, however, have been conducted under normoglycemic conditions using an isotopic tracer. Paul et aLi using a constant infusion of r4C-glucose studied the effects of nicotinic acid on the rates of turnover and oxidation of plasma glucose in normal dogs and reached conclusions similar to ours; the observed stimulation of glucose turnover was of the same order of magnitude as that observed in our studies but enhancement of the rate of oxidation was more marked. Moreover, they observed the same negative correlation between parameters of glucose metabolism and plasma FFA levels. A stimulation of the removal rate constant and of the rate of oxidation of “C-glucose after nicotinic acid has also been shown in rats by Root et a1.35Finally, arguments for a regulatory role of plasma FFA on glucose metabolism in intact animals were also provided by the work of Seyffert et aLI who showed that experimental elevations of FFA levels inhibited both the peripheral glucose utilization and the hepatic glucose output in normal dogs. Recent studies from our laboratory3’ confirmed these findings by showing that increased FFA levels inhibited the uptake and the oxidation of infused glucose in insulinized dogs. In conclusion, our studies on the influence of antilipolysis on glucose kinetics and oxidation provide further support to the “glucose-fatty acid cycle” theory and demonstrate its applicability in man under various nutritional conditions. The difficulties encountered by many authors in attempting to approach this problem using glucose tolerance tests are probably related to the fact that changes in FFA levels interfere with the insulinic response to the glucose load, as discussed in an earlier paper from this laboratory.” ACKNOWLEDGMENT We wish to thank
D. Calay and A. Vanderborght
for their expert assistance.
REFERENCES 1. Randle PJ, Hales CN, Garland PB, et al: The glucose-fatty acid cycle. Its role in insulin sensitivity and the metabolic disturbances of diabetes mellitus. Lancet 1:785, 1963 2. Miettinen TA, Taskinen M, Pelkonen R,
et al: Glucose tolerance and plasma insulin in man during acute and chronic administration of nicotinic acid. Acta Med Stand 186247, 1969 3. Balasse E: Effects of heparin on lipacidemia and on the rate of glucose utilization in
NICOTINIC
ACID AND
PLASMA
GLUCOSE
IN MAN
normal subjects (in French). Rev Fr Etud Clin Biol (Paris) 11:79, 1966 4. Balasse E, Ooms HA: Effects of an acute elevation in FFA levels on glucose tolerance and on insulinic response to glucose in normal man (in French). Rev Fr Etud Clin Biol (Paris) 13:62, 1968 5. Felber JP, Vannotti A: Effect of fat infusion on glucose tolerance and insulin plasma levels. Med Exp (Basel) IO: 153, 1964 6. Btiber V, Felber JP, Vannotti A: Glucose tolerance improvement after acute medicamentous lowering of plasma free fatty acids (in German). Schweiz Med Wochenschr 98:7ll, 1968 7. Schalch DS, Kipnis DM: Abnormalities in carbohydrate tolerance associated with elevated plasma nonesterified fatty acids. J Clin Invest 44:2010, 1965 8. Pelkonen R, Miettinen TA, Taskinen M, et al: Effect of acute elevation of plasma glycerol, triglyceride and FFA levels on glucose utilization and plasma insulin. Diabetes 17:76, 1968 9. Stowers JM, Bewsher PD, Stein JM, et al: Studies on the effects of nicotinic acid given orally or intravenously on oral and intravenous glucose tolerance in man, in Gey KF, Carlson LA (eds): Metabolic Effects of Nicotinic Acid and Its Derivatives. Bern, Hans Huber, 1971, p 723 10. Renold AE: In Gey KF, Carlson LA (eds): Metabolic Effects of Nicotinic Acid and Its Derivatives. Bern. Hans Huber, 1971, p 1 I35 11. Ruderman NB, Toews CJ, Shafrir E: Role of free fatty acids in glucose homeostasis. Arch Intern Med 123:299,1969 12. Paul P, Issekutz B Jr, Miller HI: Interrelationship of free fatty acids and glucose metabolism in the dog. Am J Physiol 211:1313, 1966 13. Seyffert WA Jr, Madison LL: Physiologic effects of metabolic fuels on carbohydrate metabolism. I. Acute effect of elevation of plasma free fatty acids on hepatic glucose output, peripheral glucose utilization, serum insulin and plasma glucagon levels. Diabetes 16:765, 1967 14. Hales CN: The glucose fatty acid cycle and diabetes mellitus, in Bittar EE, Bittar N (eds): The biological basis of medicine, vol 2, New York, Academic Press, 1968, p 309 15. Balasse EO: Inhibition of free fatty acid oxidation by acetoacetate in normal dogs. Eur J CIin Invest 1:155, 1970 16. Bernstein IA, Leutz K, Maim M. et al: Degradation of gIucose-‘4C with Ieuconostoc
1203
alternate pathways and tracer patterns. J Biol Chem 215: 137, 1955 17. Demoss RS, Bard RC, Gunsalus JC: The mechanism of heterolactic fermentation. J Bacterial 62:499. 195 1 18. Franckson JRM, Malaisse W, Arnould Y, et al: Glucose kinetics in human obesity. Diabetologia 2:96, 1966 19. Cahill GF Jr, Herrera MG, Morgan AP, et al: Hormone-fuel interrelationships during fasting. J Clin Invest 45: 175 I, 1966 20. Paul P, Bortz WM: Turnover and oxidation of plasma glucose in lean and obese humans. Metabolism 18:570, 1969 21. Have1 RJ, Kane JP, Balasse EO, et al: Splanchnic metabolism of free fatty acids and production of triglycerides of very low density lipoproteins in normotriglyceridemic and hypertriglyceridemic humans. J Clin Invest 49:2017, 1970 22. Crespin SR, Greenough WB III, Steinberg D: Stimulation of insulin secretion by infusion of free fatty acids. J Clin Invest 48: 1934, 1969 23. Malaisse WJ, Malaisse-Lagae F: Stimulation of insulin secretion by non-carbohydrate metabolites J Lab Clin Med 72:438. 1968 24. Malaisse W, Malaisse-Lagae F, Mayhew D: A possible role for the adenylcyclase system in insulin secretion. J Clin Invest 46:1724, 1967 25. Lassers BW, Kaijser L, Wahlquist ML, et al: Relationship in man between plasma free fatty acids and myocardial metabolism of carbohydrate substrates. Lancet 2:448, 1971 26. Have1 RJ. Segal N, Balasse EO: Effects of 5-methylpyrazole-3-carboxylic acid (MPCA) on fat mobilization, ketogenesis and glucose metabolism during exercise in man, in Holmes WL, Carbon LA, Paoletti R (eds): Drugs Affecting Lipid Metabolism. New York, Plenum Press, 1969, p 105 27. Froesch ER: The physiology and pharmacology of adipose tissue lipolysis: Its inhibition and implications for the treatment of diabetes. Diabetologia 3:475, 1967 28. Rabinowitz D: Some endocrine and metabolic aspects of obesity. Ann Rev Med 32:241, 1970 29. Ammon HPT. Estler CJ, Heim F: Alteration of carbohydrate metabolism in liver, skeletal muscle and brain by nicotinic acid in mice, in Gey KF, Carbon LA (eds): Metabolic Effects of Nicotinic Acid and Its Derivatives. Bern, Hans Huber, 197 I, p 799 30. Hasselblatt A, Panten U, Poser W: Efmesenteroides;
1204
fects of antilipolytic compounds on hepatic and renal gluconeogenesis in the rat, in Gey KF, Carlson LA (eds): Metabolic Effects of Nicotinic Acid and Its Derivatives. Bern, Hans Huber, 1971, p 879 31. Exton JH, Lewis SB, Park CR: Examination of in vitro effects of nicotinic acid on basal and hormone-stimulated glycogenolysis. gluconeogenesis. ureogenesis and ketogenesis in the isolated perfused rat liver, in Gey KF. Carlson LA (eds): Metabolic Effects of Nicotinic Acid and Its Derivatives. Bern, Hans Huber, 1971, p 851 32. Walter P, Piquerez RJ: Effects of nicotinic acid and &pyridylcarbinol on the production of glucose and ketone bodies in the isolated perfused rat liver, in Gey KF, Carlson LA (eds): Metabolic EtTects of Nicotinic Acid and Its Derivatives. Bern, Hans Huber, 1971. p 869 33. Struck E, Ashmore J. Wieland 0: Effects of glucagon and long chain fatty acids on glucose production by isolated perfused rat liver. Adv Enzyme Regul4:219, 1966 34. Luyckx AS, Lefebvre PJ: Arguments for a regulation of pancreatic glucagon secretion
BALASSE AND
by circulating plasma free fatty Sot Exp Biol Med 133:524, 1970
acids.
NEEF
Proc
35. Root MA, Ashmore J: The hypoglycemic activity of nicotinic acid in rats. NaunynSchmiedeberg’s Arch Pharmakol Exp Pathol 248:117. 1964 36. Bergstrom J, Hultman E, Jorfeldt L, et al: Effect of nicotinic acid on physical working capacity and on metabolism of muscle glycogen in man. J Appl Physiol 26: 170, 1969 37. Carlson LA. Froberg SO, Nye ERj Acute effects of nicotinic acid on plasma, liver, heart and muscle lipids. Nicotinic acid in the rat. II. Acta Med Stand 180:57l, 1966 38. Balasse EO: Effect of free fatty acids and ketone bodies on glucose uptake and oxidation in the dog. Horm Metab Res 3403, 1971 39. Balasse EO. Ooms HA: Role of plasma free fatty acids in the control of insulin secretion in man. Diabetologia 9: 145. 1973 40. Cahill GF Jr., Owen OE: Some observations on carbohydrate metabolism in man. in Dickens F, Randle PJ, Whelan WJ (eds): Carbohydrate Metabolism and Its Disorders. London, Academic Press, 1968. p 497
and M. A. Neef of glucose utilization occurred despite a small but significant decrease in plasma IRI concentration, indicating that nicotinic acid increased sensitivity to insulin. The above-mentioned results were obtained in 6 of the 10 patients studied. The remaining two subjects did not respond to the administration of nicotinic acid by any significant decrease in FFA nor in glycerol concentrations and showed no change in the rates of glucose turnover and oxidation. These data indicate that the effects of nicotinic acid on glucose metabolism may be partly mediated through changes in plasma FFA concentration and are consistent with the idea that the “glucose-fatty acid cycle” plays a significant role in the control of glucose metabolism in man.
Tp
HE “GLUCOSE-FATTY ACID CYCLE” theory proposed by Randle et al.’ almost 10 yr ago, has considerably stimulated further research on the interactions between lipid and carbohydrate metabolism in vivo. In an attempt to evaluate the possible role of plasma FFA in the regulation of glucose utilization, many authors have studied the effects of experimentally induced elevations or reductions in FFA levels on glucose tolerance.2-9 However, this approach has led to very conflicting results and the operation of the “glucosefatty acid cycle” as a factor regulating overall glucose metabolism in the intact organism is still questionable.‘O+” On the other hand, studies on the effects of FFA modifications on plasma glucose turnover and oxidation under normoglycemic conditions are limited in number’2T’3 and have never been performed in humans.
1
From the Metabolic Unit and the Laborator?, of Experimental Medicine, University of Brussels, Brussels, Belgium. Presented in part at the 8th Annual Meeting of the European Association for the Study of Diabetes held in Madrid, Spain. September 1972. Received for publication January 18. 1973. Supported by the Fonds de la Recherche Scientifque Medicale (Grant 20193) and by the Ministere Belge de la Politique Scientifique, within the framework of the Association Euratom- University of Brussels- University of Pisa. E. 0. Balasse, M.D., Ph.D.: Associate Professor in Medicine, Metabolic Unit and Laboratory of Experimental Medicine, University of Brussels, Brussels. Belgium. M. A. Neef, B.A.: Research Technician, Laboratory of Experimental Medicine, University of Brussels, Brussels, Belgium. Reprint requests should be addressed to E. 0. Balasse. Laboratory of Experimental Medicine, University of Brussels, 115. Bld de Waterloo, 1000 Brussels, Belgium. 01973 bv Grune & Stratton, Inc.
Metabolism, Vol. 22, No. 9 (September). 1973
1193
Subjects
109
150
176
181
OEC
RIN
MAH
VAN
cent change
‘On the left
l SEM
1.60
0.52
0.246
0.205
0.078
0.42
6.73
7.68
represent acetoacetate
3
72
74
79
72
64
3
87
92
83
89
91
so
75
>o 05
3.6
+ 1.8
84
77
83
76
85
83
84
90
83
77
ml
+ B-hydroxybutyrate.
in each test.
actid by a significant
value obtained
all subjects except VAH and HAN who did not respond to nicotinic
last expertmental
as above buf hgures correspond
Blood ketone conce”trat,ons
7 8.2
-73
7.02
6 18
571
4.26 4.22
7.06 6.47
t Same presentation
to the
3.9
-62.9
0.182
0.041
0.019
0.016
0.017
0.009
0.046
Plasma
mg/lOO
Glucose’
Concentration
6.7
-30.4
-
25
10.3
0.0
3.0
11.0
10.0
1.7
3 7
7
73
93
74
63
61
2
123
124
123
II8
128
118
127
98
73
94
85
158
139
149
139
150
151
Results)
52.2 47 5
50.0 39.3
13.3
19.0
19.8
32.5 29.8
45.4 30.1 3.4
co.01
3.6
+18X
34.0
32.4
36.5
48.8
37.9
. 4.9
b32.6
93
11.3
14.7
13.9
19.2
21 .o
27.2
25.2
25.3
32.9
values obtamed
14
Il.4
9.4
15.4
9.8
11.1
1.9
572
46.7
19.1
3.0
50.0
37.9
17.3
27.7
19.4
49.7
41 2
Glucose
from
Per cent co2
38.6
53.4
46.2
Oxidized
GlUCOSl3
Per cent
Glucose Oxldationt
(Individual
of the three expenmental
~0.001
4.6
+26.1
mg/mm
Rate
TUrflOVer
Glucose’
Metabolism
On the right. mean
3.3
6.4
11.2
1.3
13.0
0.0
2.1
8.8
5.7
16.0
15.0
4.7
5.0
6.3
PlaSma
fllJ/illl
5.0
Glucose
IRI’
and
drop in FFA nor in ketones.
during the last 60 mm of the control experiments.
0.013
0.141
0.164
0118
0.140
0.068
0.029
0.084
values obtamed
4.6
-60.3
1.31
I.23
0.50
0.032
0.082
0.032
0.019
0.089
0.120
0.004
0.029
0.087
0.273
0.027
Blood
0.1 19
0.174
0.052
0.087
0023
0.075 0.094
Blood
#mOleS/ml
#moles/ml
Metabolite
Ketones’
on Blood
Glycerol’
Acid
l This mean mcludes
with nicotintc acid
mean of the three experimental
from control t
per
SEM
mm of the experiments
P
14.7
1.18
1.31
190
HAN
14.2
1.26
1.20
0.05
0.10
122
VAH
149
16.7
1.1
0.20
0.88
12.1
0.80
0.35
0.83
149
0.26
0.75
14.5
12 1
0 50
f
129
DIE
0.13 0.30
0.87
0.86
0.58
Pl.3Sma
&moles/ml
15.7
16.2
18.9
of Nicotinic
FFA’
Effects
Mean
109
RAU
Starved
+ SEM
Meall
95
103
mg/kg
Body Weight
DES
Nicotinic Aod
of Ideal
GAU
fasted
Overnight
DOS?
1.
Per cent
Table
dunng
4.4
+12.9
10.52
6.99
7.23
9.92
13.08
1 1.52
10.39
9.19
9.80
8.19
the last 60
0.73
7.77
9.96
7.23
6.94
6.96
0.7
9.49
12.19
10.76
9.26
8.41
9.32
7.02
mmoles/mm
CO2 Production
L
NICOTINE
ACID AND
PLASMA
GLUCOSE
IN MAN
1195
The aim of the present work was to determine whether an acute reduction in plasma FFA levels induced by nicotinic acid modifies the rates of turnover and oxidation of circulating glucose in man, as measured by use of a “C-glucose infusion technique. These studies have been carried out in normal postabsorptive, in obese postabsorptive, and starved patients. These two last groups of subjects were included in the study because it has been suggested that elevated FFA levels might be responsible-at least partly-for the abnormalities in carbohydrate metabolism accompanying obesity and food deprivation.14 MATERIALS
AND
METHODS
Ten female subjects aged 17-34 yr were selected for the study. Their body weight varied between 95% and 190% of ideal weight (Table 1) and all had normal oral glucose tolerance. Six ambulatory subjects (three normal and three obese) were studied after an overnight fast of 15 hr and four obese hospitalized patients were studied after IO- I 1 days of complete starvation during which their intake consisted only of calorie-free beverages, vitamins, and minerals. For all subjects, only water was allowed during the 15 hr preceding the tests. The studies were conducted according to the following protocol (Fig. 1). After a 30-min basal NICOTINIC
OR
ACID
SALINE
o_
.
-0
-
11111111111 l_“C PLASMA
FFA
(pmoles /ml)
0.00 0.70
0.50 0.30
BLOOD GLYCEROL
IRI
(pU /ml) PLASMA / lOOmI)
PLASMA l.“C.GLUCOSE
GLUCOSE
S.A.
CO2
(mmoles/min)
cose
metabolism.
experiment fasted
antilipolysis acid
subject
(DES). CO2
glucose
specific
merator
was and
dpm&atoms
g C.
glu-
RESPIR.
‘4co2
specific activity
activity/ the in
denominator
~0 *O 3 600
I
500
I I I
D--
0
SO
-o__-o---o___0
600
10.0
9.0 1 0.50 0.40 0.30 0.20
INFUSED
overnight
14C
nudpm/ in
0.10 0
For calculat-
expressed the
on
Representative
in a normal
ing the ratio
pmoles
of
nicotinic
10 01
500 400 1
RESPIRATORY
Effect
GO5
300 I
(dpm/mg)
by
I
400
(dpm/m!)
1.
J_______.___._._o___~___~
100
GLUCOSE
Fig.
&-----
0 I
PLASMA
induced
OLUCOSE
0.10
(Pmolas/ml)
(mg
-
CO2 GLUCOSE
S.A. S.A
0.30 0.20 0.10
0 I
-30
120 MIN180
1196
BALASSE
AND
NEEF
period, I-‘4C~glucose (Amersham. England; specific activity 0.2 mCi/mmole), diluted in saline. was infused for 3 hr in an antecubital vein at the constant rate of about 0.15 pCi/kg. hr. A priming dose of radioactivity equivalent to the amount infused in 130 min was given at the start of the infusion. Blood samples were obtained at intervals from an indwelling plastic needle located in an antecubital vein of the opposite arm, transferred into heparinized tubes kept on ice, and processed promptly. Expired air was collected intermittently for 445 min periods in rubber bags through a low resistance valve and analyzed immediately. Each subject was studied on two separate occasions: (I) in the control state: (2) during the administration of nicotinic acid (Niacin, UCB. Belgium) which was injected as i.v. pulses of 100 or 150 mg every 20 min during the whole study, the first dose being given 30 min before the start of the 14C -glucose infusion. The highest dosage (150 mg) was used only in the two most obese patients (VAN and HAN). The total amount of nicotinic acid varied between 12.1 and 18.9 mg/kg (Table I .) In the control tests. nicotinic acid injections were replaced by saline injections. The control study always preceded the nicotinic acid study in order to avoid possible interference of the rebound in FFA levels which is known to occur after administration of this drug. The time interval between the two tests was I 3 days in the postabsorptive subjects who were instructed not to change their usual diet hetween the two successive studies. in the starved patients. the tests were always performed on 2 consecutive days so that closely similar nutritional conditions would prevail during the two successive experiments. Plasma concentration of glucose, FFA. and immunoreactive insulin (IRI): blood concentrations of glycerol. fl-hydroxyhutyrate. and acetoacetate; and specific activity of expired CO, were determined as indicated elsewhere.” Plasma I-‘4C~glucose was isolated as “C02 bv a fermentation method using Leuconortoc mesenteroides 16.17;the 14C02 was trapped in hyamine and counted For present in the plasma sample before fereach sample, the amount of 14C0, + 14Cbicarbonate mentation was determined on a separate aliquot and suhstracted from total 14C0, recovered after fermentation. The recovery of I-‘4C~glucose added to nonradioactive plasma was tested with each series of determinations and varied between 88.3”;, and 98.9”” (average 93 O”.,). rad’oactiv’ty in blood samples was corrected accordingly. For the test performed during nicotinic acid administration. a sample of expired a’r was collected before the start of the 14C~ glucose infusion in order to determine the residual “C02 concentration resulting from the preceding experiment and this residual activity was substracted from each determinat’on obtained during the 14C glucose infusion. Blood and expired-a’r analyses were ,111made in duplicate. Except for a cutaneous flush, nicotinic acid produced no discomfort for the pat’ents during the course of the studies. However, most of the subjects. especially the starved ones, felt nauseous a few hours after the termination of the experiments.
CALCULATIONS
Glucose Transport In both the control and the nicotinic acid studies, steady plasma concentrations of labeled and unlabeled glucose were obtained during the last 2 hr of the study allowing measurements of glucose kinetics. The rate of turnover of plasma glucose (i.e., the rate of influx or efflux of plasma glucose) was obtained as follows: F=
glucose
I specific activity’
where F is the glucose turnover rate (mg/min); I is the rate of infusion of labeled glucose (dpm/min); glucose specific activity is the mean of three determinations of plasma glucose specific activity (dpm/mg) obtained during the last 60 min of the “C-glucose infusion. The utilization of plasma glucose by tissues was expressed as glucose clear-
NICOTINIC
ACID AND
PLASMA
ante which represents plasma and the plasma
c, =
GLUCOSE
the ratio between the rate glucose concentration, F
glucose
1197
IN MAN
of efflux
of glucose
from
I
concentration
= i4C-glucose
concentration
’
where C, is the glucose clearance (ml/min); F is the turnover rate of glucose (mg/min); glucose concentration is the concentration of unlabeled glucose in plasma (mg/ml); I is the rate of infusion of labeled glucose (dpm/min); 14Cglucose concentration is the mean of three determinations of radioactive glucose concentration in plasma (dpm/ml) during the last 60 min of the study. Glucose Oxidation During I-r4glucose infusion, the efflux of i4C0, and the specific activity of expired CO1 rose in the form of exponential curves (Fig. 1) which did not completely attain their asymptotic values during the course of the studies. This is a general observation in this type of study and is related to a gradual increase in the labeling of intermediate metabolic pools, including the bicarbonate pool of the body. The fraction of plasma glucose converted to CO, and the per cent CO, derived from plasma glucose were calculated by the following equations: Per cent glucose
to CO* =
Efflux rate of i4C0, (dpm/min) Influx rate of *4C-glucose
Per cent CO, from glucose
=
x 100
(dpm/min)
’
CO, specific activity (dpm/pmole) x6x 100 Plasma glucose specific activity (dpm/pmole)
’
In these equations, the rate of efflux of i4C0, and the specific activity of CO, were the last values obtained during the studies. Since, as already mentioned, these values are lower than asymptotic values, the figures obtained from these equations slightly underestimate glucose oxidation, RESULTS
Representative
Experiment
Data obtained in a representative experiment on a normal weight overnight fasted subject (DES) are presented in Fig. 1. The FFA and glycerol levels measured during the nicotinic acid study amounted to about 40% of values observed in the control study. Nicotinic acid produced only slight changes in glycemia, in IRI concentration, and in cold CO, production. The plateaus of plasma 14Cglucose concentration and of glucose specific activity were about 25% lower than corresponding values observed in the basal state indicating that both the clearance of plasma glucose by tissues and the hepatic glucose output were stimulated in the presence of lowered FFA. Nicotinic acid also stimulated glucose oxidation so that the rate of 14C0, production increased by 20% and the ratio of CO, specific activity to plasma glucose specific activity increased by 46%. These results indicate that nicotinic acid enhanced both the conversion of glucose to CO, and the per cent CO, derived from plasma glucose.
BALASSE
1198
Individual
AND
NEEF
Data
Table 1 provides individual values of blood metabolite concentration, IRI, and glucose turnover and oxidation in the absence and in the presence of nicotinic acid. Control Studies Overnight fastedpatients. In this group, basal values were 0580.88 pmole/ ml for FFA, 0.075-0.099 pmole/ml for glycerol, 0.078-0.273 pmole/ml for ketones, and 75-92 mg/ 100 ml for glycemia. Basal IRI concentration was below 10 pU/ml except for two obese patients (RIN and MAH) whose levels were, respectively, 15.0 and 16.0 PI-I/ml. Small individual variations were observed among values of the glucose turnover rate (118-127 mg/min). The fraction of glucose taken up and promptly oxidized varied between 37.9% and 50.0% thereby providing 13.3x-27.7% of total CO, production. Unlabeled CO, output ranged between 7.02 and 12.19 mmole/min and tended to be positively correlated with excess body weight. Starved patients. In the starved patients, the concentrations of FFA (1.181.60 pmole/ml), glycerol (0.118-0.164 pmole/ml), and ketones (5.71-7.68 pmole/ml) were significantly higher (p < 0.001) than those observed in the overnight fasted group and glycemia (64-79 mg/ 100 ml) was significantly lower (0.001 < p < 0.01). The glucose turnover rate (61-93 mg/min) and the per cent CO, derived from glucose (9.4x-15.4%) were significantly reduced in the starved subjects as compared with the postabsorptive subjects (p values being, respectively, < 0.001 and < 0.02) but the per cent glucose converted to CO, and the unlabeled CO, production were similar in the two groups. Studies
With Nicotinic Acid
The administration of nicotinic acid strongly inhibited lipolysis in 8 of the 10 patients studied, including the six overnight fasted and two of the four starved patients (RAU and DIE). The average fall in FFA, glycerol, and ketone concentration observed in these subjects amounted, respectively, to 60.3% f 4.6x, 62.9% f 3.9x, and 73.7% + 8.2%. Glycemia did not change significantly but IRI decreased by 30.4% * 6.7%. The inhibition of lipolysis was accompanied in each subject by a rise in the rate of glucose turnover averaging 26.1% + 4.6%. The removal of plasma glucose was also systematically accelerated, the observed rise in plasma glucose clearance amounting to 24.0% f 4.1%. Nicotinic acid also significantly enhanced the oxidation of glucose: The fraction of glucose taken up and promptly oxidized increased by 18.6% f 3.6% and the fraction of total CO, derived from plasma glucose increased by 32.6% f 4.9%. Unlabeled CO, production was also slightly stimulated under these conditions (+12.9x f 4.4%). The two remaining starved patients (VAH and HAN) did not respond to nicotinic acid by a significant inhibition of adipose tissue lipolysis, although these subjects were given amounts of nicotinic acid similar to those given in the responders (Table 1). In subject VAH, there is even a slight rise in FFA and ketone concentration; under these conditions, the glucose turnover rate re-
NICOTINIC
ACID AND PLASMA
GLUCOSE
IN MAN
1199
mained unchanged and oxidation decreased. In the other patient (HAN), a small decrease in FFA and ketone levels was observed after nicotinic acid but blood glycerol rose slightly and the rates of glucose turnover and oxidation remained unaffected. Correlation
Between
Glucose Utilization
and Plasma FFA Levels
Figure 2 shows that a significant negative correlation has been found the various parameters of glucose metabolism (rate of turnover, per cose oxidized, and per cent CO2 derived from glucose) on one hand, responding plasma FFA concentration on the other hand. The data clude the figures obtained both in the absence and in the presence of acid.
between cent gluand corused innicotinic
DISCUSSION
Control values of the glucose turnover rate were higher in the overnight fasted patients than in the starved subjects and both groups of figures were of
60, so-
0
B Oo
\ 0
, .
.
LO_ : 30-
Fig. 2. Correlation between plasma FFA concentration and glucose turnover rate (panel A) or glucose oxidation (panels B and Cl in a group of patients including normal overnight fasted, obese overnight fssted. and obese starved subjects (see Table 1). Each subject was studied under control conditions (closed circles) end during administration of nicotinic acid (open circles). Statistical significance: (A), = - 0.78, A < 0.001; (6) r = -0.83. JJ< 0.001; (C) r = - 0.80, /J < 0.001.
0
. 00
0
0 0.2 0.6 I.0 I.6 1.L PLASMA FFA (pmoles/ml)
1200
BALASSE
AND
NEEF
the order of magnitude of those published by others using either isotope dilutions techniques’*-*’ or liver catheterization.*’ On the other hand, our estimates of glucose oxiation rates were about 20”/,-2%; lower than those published by Paul et al.*’ This difference is probably due to the fact that these authors used prolonged isotopic infusions (63 hr) and were therefore able to attain the true asymptotic value of the 14C0, curves. As mentioned earlier, this was not the case in our studies so that our results systematically underestimated glucose oxidation. The purpose of this study was to analyze the effects of a decrease in FFA concentration on these parameters of glucose metabolism. Antilipolysis was induced by a pharmacologic mean consisting of repeated nicotinic acid injections. In two of the four starved patients studied, nicotinic acid failed to decrease FFA levels thus indicating individual variations in the sensitivity to this drug. Obviously, the results obtained in these two patients do not provide any information regarding the influence of decreased FFA levels on glucose metabolism, and will therefore be analyzed separately. In the eight other patients (six overnight fasted and two starved), the administration of nicotinic acid resulted in a 60”/0 reduction in plasma FFA concentration and in a parallel decrease in ketone levels. The fall in ketones has to be taken into consideration especially in starved patients who rely on this substrate to a great extent for the provision of energy. After nicotinic acid, other fuels have to compensate for the reduced availability of FFA and ketones and our results demonstrate that circulating glucose plays a role of alternate fuel under these circumstances. Indeed. the clearance of plasma glucose by tissues was increased by about 24Y,, this change being entirely compensated by a corresponding rise in the hepatic glucose output so that, on an average, glycemia remained unaffected. Simultaneously, nicotinic acid induced a small but significant decrease in IRI concentration. This latter effect is presumably related to the fall in the level of plasma FFA which is known to promote insulin secretion in vivoi3.‘* and in vitro23; a direct inhibitory effect of nicotinic acid on P-cell secretion is unlikely according to in vitro data.24 The stimulation of glucose uptake in the presence of a decreased concentration of insulin in plasma indicates that the reduced availability of FFA enhanced insulin sensitivity. Our experiments provide no information on the type of tissue implicated in the increased glucose consumption. On the basis of in vitro data,’ cardiac and skeletal muscle are likely to be the primary site of this effect. Lassers et al.*’ have demonstrated that the antilipolytic effect of nicotinic acid in man is accompanied by an increase in the myocardial arterio- venous difference of glucose. Unfortunately, coronary blood flows were not measured in these experiments and absolute amounts of glucose extracted cannot be calculated. However, assuming that the coronary blood flow was 200 ml/min and was unaffected by nicotinic acid, it can be estimated from their data that the increase in myocardinal extraction of glucose after nicotinic acid amounted to 5--10 mg/min. This could account for about one-third of the increase in the total glucose utilization observed in our experiments. Skeletal muscle probably also participates in the increased glucose consumption. Have1 et a.26 have shown that glucose extraction by leg muscle is stimulated after administration of an anti-
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lipolytic compound but these results have been obtained during exercise and it is not known whether this applies to resting muscle. Finally, adipose tissue might also contribute to the increased uptake of glucose observed in our study, since nicotinic acid is known to stimulate glucose metabolism in this tissue.27 However, the participation of fat cells is likely to be small owing to the fact that adipose tissue participates very little in over-all glucose uptake.” If we assume that the increased uptake of glucose after nicotinic acid was essentially limited to cardiac and skeletal muscle, it can be calculated that the apparently small increase (24%) in glucose turnover rate corresponds in fact to a striking stimulation of glucose utilization by muscular tissues. Indeed, it is known that in the postabsorptive state, the uptake of glucose by cardiac and skeletal muscle does probably not exceed 30 g/24 hr.40 In our ove rnight fasted patients, this represents only a minor fraction of the over-all basal glucose consumption which approximated 177 g/24 hr. The major part of this glucose uptake is known to be accounted for by tissues for which glucose is an obligatory substrate (i.e., brain, blood cells, and renal medulla). Since the absolute increase in glucose turnover rate observed after nicotinic acid in our overnight fasted patients amounted to about 36 g/24 hr, it would appear that the uptake of plasma glucose by muscular tissues has approximately doubled during experimental antilipolysis. The increase in the hepatic glucose output observed in our experiments has also been observed in vitro on perfused livers from rats pretreated with nicotinic acid or related compounds. These liver preparations showed an increased rate of glycogenolysis29 and gluconeogenesi?’ as compared with livers from normal animals. This effect cannot be explained by a direct action of the drug on the liver since the addition of nicotinic acid to the perfusion medium of livers from normal rats is without effect3’ or even inhibits32 the rates of glycogenolysis and gluconeogenesis. It is also unlikely that the observed stimulation of hepatic glucose output after administration of nicotinic acid in vivo was related to the decrease in supply of FFA to the liver, since FFA have been shown to stimulate gluconeogenesis in the perfused liver preparation.33 All these reasons suggest that the enhancement of the hepatic glucose production observed was related to the hormonal changes associated with antilipolysis. One of these changes is the decrease in IRI concentration observed in these studies. Another possible factor is an increase in plasma glucagon levels. Luyckx et a1.34 have shown indeed that a rise in glucagon levels is observed in dogs whose FFA concentrations have been lowered with nicotinic acid. After nicotinic acid, the percentage of total CO, output derived from glucose oxidation was increased by 32.6%. This resulted from the combination of an augmented glucose turnover rate and an increased fractional conversion of glucose to CO,. There was also a concomitant rise in unlabeled CO, production which does probably not correspond to an augmented metabolic rate. Indeed, assuming that the oxygen consumption remained unchanged, an increase in CO, output was to be expected as a consequence of a rise in R.Q. reflecting the shift towards carbohydrate oxidation induced by nicotinic acid. Taken as a whole, our results indicate that plasma glucose can serve as alternate fuel when availability of FFA to tissues is reduced. However, this conclu-
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sion has to be taken cautiously with regard to the starved subjects, since only two of them could be adequately studied. Our interpretation of the data is based on the assumption that the effects of nicotinic acid on glucose metabolism is mediated through its antilipolytic properties and does not represent a direct pharmacologic effect of the drug. This concept is supported by the fact that in the two starved patients who did not respond to nicotinic acid by a fall in FFA concentration, no increase in the rates of turnover and oxidation of plasma glucose was observed. Circulating glucose is not the only substrate which is called upon by the body to meet the energy requirements in the presence of lowered FFA levels. It has been shown that muscle glycogen3sv36and stored triglyceride$’ are other possible sources of fuel under these conditions. Numerous studies have been devoted to the effects of antilipolytic substances on overall glucose metabolism in vivo. Most of them were based on the use of oral or i.v. glucose tolerance tests and yielded contradictory results.2-9 A few animal studies, however, have been conducted under normoglycemic conditions using an isotopic tracer. Paul et aLi using a constant infusion of r4C-glucose studied the effects of nicotinic acid on the rates of turnover and oxidation of plasma glucose in normal dogs and reached conclusions similar to ours; the observed stimulation of glucose turnover was of the same order of magnitude as that observed in our studies but enhancement of the rate of oxidation was more marked. Moreover, they observed the same negative correlation between parameters of glucose metabolism and plasma FFA levels. A stimulation of the removal rate constant and of the rate of oxidation of “C-glucose after nicotinic acid has also been shown in rats by Root et a1.35Finally, arguments for a regulatory role of plasma FFA on glucose metabolism in intact animals were also provided by the work of Seyffert et aLI who showed that experimental elevations of FFA levels inhibited both the peripheral glucose utilization and the hepatic glucose output in normal dogs. Recent studies from our laboratory3’ confirmed these findings by showing that increased FFA levels inhibited the uptake and the oxidation of infused glucose in insulinized dogs. In conclusion, our studies on the influence of antilipolysis on glucose kinetics and oxidation provide further support to the “glucose-fatty acid cycle” theory and demonstrate its applicability in man under various nutritional conditions. The difficulties encountered by many authors in attempting to approach this problem using glucose tolerance tests are probably related to the fact that changes in FFA levels interfere with the insulinic response to the glucose load, as discussed in an earlier paper from this laboratory.” ACKNOWLEDGMENT We wish to thank
D. Calay and A. Vanderborght
for their expert assistance.
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