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Original Studies |
Institute of Physiology, University of Lausanne School of Medicine, 1005 Lausanne, Switzerland
Address all correspondence and requests for reprints to: Dr. L. Tappy, Institut de Physiologie, 7 rue du Bugnon, 1005 Lausanne, Switzerland. E-mail: luc.tappy{at}iphysiol.unil.ch
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Abstract |
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Introduction |
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Increased free fatty acids can acutely produce endothelial dysfunction in healthy humans (10). It is therefore possible that elevated free fatty acid concentrations were responsible for the alteration of the metabolic and hemodynamic responses to mental stress observed in the obese patients. To further evaluate this hypothesis we assessed the effects of a lipid infusion on the hemodynamic and metabolic effects of mental stress in healthy humans.
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Subjects and Methods |
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Eight healthy female subjects were selected to participate in this study. Their mean age was 22.9 ± 2.5 yr, weight was 56.9 ± 2.5 kg, height was 1.6 ± 4.5 m, and body mass index was 23.4 ± 2.2 kg/m2. All were in good physical condition, had no known disease, and were not currently taking any medication, with the exception of oral contraceptive agents. The experimental protocol was approved by the ethical committee of Lausanne University School of Medicine, and every participant provided informed written consent.
General procedures
All experiments were performed during the follicular phase of each subject’s menstrual cycle and began in the morning after an overnight fast. The participants were instructed to consume an isocaloric diet containing 50–55% carbohydrate and to restrain from strenuous physical activities during the 2 days preceding the studies. On the morning of each study, the subjects reported to the metabolic investigation laboratory between 0700–0730 h. They had been fasting since 2200 h the night before. At their arrival, subjects were weighed and measured, and they voided before being transferred to a bed where they lay quietly during the 240 min of the studies. One indwelling venous cannula was inserted into an antecubital vein of the left arm for infusion of glucose, insulin, and 6,6-2H2 glucose. Another indwelling cannula was inserted into a wrist vein of the right arm to allow serial blood samples collection. The right hand was placed in a thermostabilized box heated to 56 C to achieve partial arterialization of venous blood. Throughout the study period, indirect calorimetry was performed using a ventilated hood as previously described (11). Each subject took part, in randomized order, in two study protocols, which took place at least 1 week apart.
Study 1. This experiment started between 0800–0830 h after subjects were prepared as described above. At 0 min, a primed (2 mg/kg BW) continuous infusion (20 µg/kg·min) of 6,6-2H2 glucose (Masstrace, Worcester, MA) was started and continued for 60 min. After 60 min, a primed continuous infusion of insulin was started at a rate of 0.4 mU/kg·min, whereas glucose was clamped at 5.6 mmol/L (12) by means of an infusion of 200 g/L dextrose. During the clamp, 6,6-2H2 glucose infusion was adjusted to that of unlabeled glucose to produce a 1.25% enrichment of exogenous glucose. At 210 min, a 30-min mental stress, consisting of 5-min periods of mental arithmetics alternated (6) with 5-min periods of Stroop’s color word conflict test, was applied (13). Indirect calorimetry was continuously performed, and the hyperinsulinemic euglycemic clamp was continued until 240 min. Blood samples were collected at 30-min intervals throughout the study and at 5, 10, 20, and 30 min after the beginning of the mental stress. Blood pressure was recorded every 30 min (using a Sentry HSD 400; Automated Screening Devices, Costa Mesa, CA) and for the mental stress every 5 min. Cardiac output was measured with thoracic bioimpedance (14) using a Bomed impedancemeter (Bomed NCCOM3-R7 CDDP system, Bomed Medical Manufacturing Ltd., Irvine, CA). Cardiac index was calculated as cardiac output/body surface area. Systemic vascular resistance was calculated as mean arterial pressure divided by cardiac output.
Study 2. This study was identical to study 1, except that an infusion of a 20% soybean lipid emulsion (20% Lipovenos, Fresenius, Stans, Switzerland) was infused at a rate of 1 mL/min from 0–210 min.
Analytical measurements
The plasma glucose concentration was measured enzymatically using a glucose analyzer II (Beckman Coulter, Inc., Fullerton, CA). Commercial RIA kits were used for determination of plasma insulin (Biochem Immunosystems GmbH, Freiburg, Germany) and glucagon (Linco Research, Inc., St. Charles, MO) concentrations. Plasma norepinephrine concentrations were determined by high pressure liquid chromatography with electrochemical detection (15). Plasma 6,6-2H2 glucose enrichments were determined by gas chromatography-mass spectrometry, as previously described (16). Plasma free fatty acid concentrations were measured by means of a calorimetric method, using a kit from Wako (Freiburg, Germany).
Calculations
The glucose rates of appearance and disappearance were calculated from 6,6-2H2 glucose dilution using the "hot" glucose infusion equations (17). Energy expenditure and net substrate oxidation rates were calculated from respiratory gas exchanges using the equations of Livesey and Elia (18). For this purpose, the urinary nitrogen excretion rate was calculated from urinary urea concentrations.
Statistics
All results are expressed as the mean ± 1 SEM unless stated otherwise. In each study protocol, the evolution of all parameters collected over time was assessed with repeated measures ANOVA. Comparison between studies 1 and 2 was performed by two-way ANOVA and paired t tests with Bonferonni’s adjustment.
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Results |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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). Plasma free fatty acid
concentrations were suppressed to 82% of their basal concentrations in
study 1 and remained elevated at an average value of 0.549 mmol/L in
study 2 as a consequence of lipid infusion (Fig. 1). After 2 h of
hyperinsulinemia, lipid infusion in study 2 resulted in a 16.5%
reduction in the glucose rate of disappearance and a 24% reduction in
net glucose oxidation compared with those in study 1 (Fig. 2). There was, however, no difference in
energy expenditure (Fig. 3) or in
endogenous glucose production (3.0 ± 0.9 µmol/kg·min in study
1 vs. 4.9 ± 0.8 µmol/kg·min in study 2; Fig. 2).
Hemodynamic parameters (Fig. 4) were also
comparable in both studies during this period.
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). It also stimulated energy expenditure, which increased by 15.8%
(Fig. 3), and enhanced the glucose rate of disappearance by 18.9%
(Fig. 2). There was no effect on endogenous glucose production, which
remained at 4.5 ± 1.0 µmol/kg·min (Fig. 2).
In study 2, lipid infusion did not alter the increases in plasma
norepinephrine (from 0.63 ± 0.04 to 0.94 ± 0.07 nmol/L;
P < 0.05), plasma epinephrine (from 371 ± 41 to
448 ± 33 pmol/L; P < 0.05), in heart rate (Fig. 4), and energy expenditure induced by mental stress (Fig. 3). However,
it totally prevented the increase in cardiac output and the decrease in
systemic vascular resistance (Fig. 4) as well as the stimulation of
glucose rate of disappearance (Fig. 2). Instead, it markedly enhanced
the rise in blood pressure, which amounted to 13 mm Hg, on the average
(P = 0.0005; Fig. 4). This increase in blood pressure
was 116.7% higher than that in study 1 (P <
0.01).
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Discussion |
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Lipid infusion prevented the drop in systemic vascular resistance and the increase in cardiac output normally elicited by mental stress. In contrast, it did not affect the stimulation of catecholamine release or the increase in heart rate, indicating that lipids did not interfere with sympathoadrenal stimulation during mental stress. Instead, these observations are consistent with a lipid-induced alteration of endothelial cell function, leading to inadequate NO release during mental stress. In support of this explanation, it has been demonstrated that the skeletal muscle vasodilation induced by mental stress could be inhibited by NO synthase inhibitors (2, 4), and that lipid infusion impaired the vasodilatory effects of intraarterial cholinergic agents, which require endothelial NO release (10).
As a consequence of an unaltered sympathetic stimulation but impaired
vasodilation, mental stress produced a substantial increase in arterial
pressure. A modest increase in cardiac output produced by the
chronotropic effect of mental stress together with an increase in
vascular resistance secondary to the predominant effect of sympathetic
activation of 
1-vascular receptor are likely
to explain this rise in blood pressure.
Mental stress is known to acutely increase insulin-mediated glucose disposal (5, 6, 7, 8). Although mental stress tended to transiently increase endogenous glucose production in experiments performed without lipids only, this effect was of slight magnitude and did not reach statistical significance. This observation is consistent with our previous report that stimulation of the sympathoadrenal nervous system elicited by mental stress fails to overcome insulin-induced inhibition of endogenous glucose production. Stimulation of insulin-mediated glucose disposal was also abolished by lipids. In contrast, the increase in energy expenditure was not altered, indicating that sympathetic activation of metabolically active tissues (19) was not impaired. We have proposed the hypothesis that the mechanism by which mental stress increases insulin sensitivity is through an increased blood flow to insulin tissue (8). There is indeed evidence that increased muscle blood flow may enhance insulin actions by increasing the amount of both glucose and insulin reaching insulin-sensitive tissues (reviewed in Ref. 20). Although we did not directly measure muscle blood flow in this study, our observation that lipids simultaneously prevents both the drop in systemic vascular resistance and the enhancement of insulin action during mental stress is entirely consistent with this hypothesis.
We previously observed that administration of ß-adrenergic
antagonists produce effects similar to those of lipids during mental
stress (8). Infusion of propranolol also inhibited the
decrease in systemic vascular resistance and the increase in
insulin-mediated glucose disposal during mental stress. Although still
consistent with the hypothesis that vasodilation is operative in
enhancing insulin actions, this observation suggests that the
hemodynamic effects of mental stress may involve simultaneous
ß-adrenergic and NO-ergic vasodilation and
1-adrenergic vasoconstriction.
We also reported that obese nondiabetic patients have a blunted decrease in systemic vascular resistance, an accentuated rise in blood pressure, as well as an absence of stimulation of insulin actions during mental stress (8). All of these alterations were reproduced in healthy lean females by lipid infusion, suggesting that increased fatty acid concentrations may be responsible for the alterations observed in obese patients. This hypothesis is consistent with the observation of Steinberg et al. that free fatty acids produce endothelial dysfunction in healthy human subjects (1). The endothelial dysfunction induced by lipids may have several important consequences. First, it may participate in the insulin resistance observed in obesity, in part by preventing regulatory increases in blood flow to insulin-sensitive tissues. Secondly, it may contribute to a rise in blood pressure observed in patients submitted to stressful conditions (21). Our present observations therefore suggest that fatty acid-induced endothelial dysfunction may be of importance in both the metabolic and cardiovascular complications of obesity.
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Footnotes |
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Received June 21, 2000.
Revised August 30, 2000.
Accepted September 11, 2000.
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). *, P < 0.05 or less vs.
control.
