Anabolic androgenic steroids (AAS) administration has become a widespread strategy to increase physical performance especially in young athletes (12,30). However, the consumption of AAS has been associated with cardiac events and cardiovascular alterations (23). Some case reports link arrhythmias, myocardium infarction, and sudden death to abusive AAS self-administration in strength athletes (6,11,20,22). Cross-sectional studies associate AAS with cardiac hypertrophy, hypertension, and lipoprotein alterations (16,25,29). In addition, former AAS users maintain concentric left ventricular hypertrophy or high blood pressure, which seem to be related to time and type of AAS used (1,4,29).
Some investigators have shown that high doses of stanozolol increases vascular peripheral resistance in animals (5). In humans, AAS decreases flow-mediated dilatation, which seems to be associated with lipid alterations and a decrease in nitric oxide bioavailability (9). Unknown is whether the sympathetic nerve activity plays a role in vascular alteration in AAS users. The early observation that testosterone can increase the vascular response to norepinephrine (14) is suggestive of enhanced sympathetic activity in AAS users. In addition, previous findings (28) showing that sympathetic nerve activity reduces muscle blood flow and restrains endothelial-mediated muscle vasodilatation in humans with sympathetic hyperactivity strengthen the idea that the neurovascular control is altered in AAS users.
Accumulated evidences show that high blood pressure is due, at least in part, to increased peripheral vascular resistance (17,21). In addition, in a recent study, we found a significant association between high blood pressure and sympathetic nerve activity levels in hypertensive patients (19). Thus, it seems reasonable to raise the question that the augmented systemic blood pressure observed in AAS users is linked to augmented sympathetic outflow. In this study, we described the effects of AAS on muscle sympathetic nerve activity (MSNA) and blood flow in young AAS users. Further, we reported the association between 24-h blood pressure levels and sympathetic nerve activity levels in these individuals.
Our hypothesis was that AAS users would have increased MSNA and reduced muscle blood flow when compared with AAS nonusers. A second hypothesis was that there was an association between 24-h blood pressure levels and MSNA levels in individuals with chronic use of AAS.
Twelve male AAS users and nine age-matched male AAS nonusers were invited to participate in the study. All individuals were involved in strength training for at least 2 yr. The AAS users were self-administering AAS for at least 4 wk before the study (Table 1). In addition, they had been using AAS for at least 2 yr, two to four cycles per year. The AAS users were not taking other doping substances than the AAS. The use of AAS was confirmed by urine test (chromatography-mass spectrometry). The study protocol was approved by the Human Subject Protection Committee of the Heart Institute (InCor) and Clinical Hospital, University of São Paulo Medical School, and written consent was given by each individual.
Measurements and Procedures
Ambulatory blood pressure monitoring.
Ambulatory blood pressure monitoring was performed using Spacelabs 90270 (Spacelabs Medical, Cedex, France), which was previously programmed to perform measurements at 10-min intervals during the day time and at 20-min intervals during the night for 24 h. The cuff was adjusted to the size of patient's arm. The subjects were asked to document their activities, such as time in bed, time after retiring from work, time of awakening, and time on the bus or in the car. In addition, on both days, they were instructed not to exercise, not to take a shower, and not to sleep during the recording interval of daytime, and to relax and straighten out the arm during their waking hours. Three periods were taken into consideration for data analysis. First, the average time was obtained during the 24-h period. This included the next day, before the monitor was removed. Second was the daytime average, which included the period from the first measurement after the subject left the laboratory until the measurement was taken just before the subject went to bed. Third was the nighttime average, which included the period from the time when the subject went in bed, after retiring, and up to the moment of definite rising from bed in the morning.
Blood pressure and heart rate measures.
During the experimental procedure, blood pressure was monitored noninvasively and intermittently from an automatic and oscillometric cuff (DX 2710; Dixtal, Manaus, Brazil). Heart rate was monitored continuously through lead II of an ECG beat per beat.
Forearm blood flow.
Forearm blood flow was measured by venous occlusion plethysmography. The nondominant arm was elevated above heart level to ensure adequate venous drainage. A mercury-filled silastic tube attached to a low-pressure transducer was placed around the forearm and connected to a plethysmography (Hokanson, Bellevue, WA). Sphygmomanometer cuffs were placed around the wrist and upper arm. At 15-s intervals, the upper cuff was inflated above venous pressure for 7-8 s. Forearm blood flow (mL·min−1·100 mL−1 of tissue) was determined on the basis of a minimum of four separate readings. Forearm vascular conductance was calculated by dividing forearm blood flow by mean arterial pressure times 100 and expressed in arbitrary units. The reproducibility of forearm blood flow measured at different time intervals in the same individual expressed as milliliters per minute per 100 mL of tissue in our laboratory is r = 0.93.
Muscle sympathetic nerve activity.
MSNA was recorded directly from the peroneal nerve using the microneurography technique. Multiunit postganglionic muscle sympathetic nerve recordings were made using a tungsten microelectrode (tip diameter = 5-15 μm). The signals were amplified by a factor of 50,000-100,000 and band-pass filtered (700-2000 Hz). For recordings and analysis, nerve activity was rectified and integrated (time constant = 0.1 s) to obtain a mean voltage display of sympathetic nerve activity that was recorded on paper. All of the recordings of MSNA met previously established and described criteria. MSNA was quantified as burst frequency (bursts per minute) and burst incidence (bursts per 100 heartbeats). The reproducibility of MSNA measured at different time intervals in the same individual expressed as bursts per minute is r = 0.88 and expressed as bursts per 100 heartbeats is r = 0.91 (10).
Clinical laboratory measures.
A blood sample was collected for assessment of lipid profiles (total cholesterol and HDL cholesterol, LDL cholesterol fractions, and triglycerides) and glucose.
All studies were performed in a quiet, temperature-controlled (21°C) room in the morning period at approximately the same time of day. The participants were instructed to have the last meal 2 h before the experimental protocol and to avoid caffeine and high-fatty food intake 24 h before the measurements. They were instructed to restrain from exercise 48 h before the experimental protocol. After ECG leads were placed on the chest, the arm was positioned for venous plethysmography, and the left leg was positioned for blood pressure measure. On the right leg, a tungsten microelectrode was attached on the peroneal nerve. After that, the subject rested quietly for 15 min. MSNA, forearm blood flow, blood pressure, and heart rate were then recorded for 6 min. Blood pressure was monitored noninvasively and intermittently from an automatic and oscillometric cuff inflated every 30 s.
Data are presented as means ± SE. Data were tested for normality using the Shapiro-Wilk test. Data of demographic characteristics, baseline measurement, and lipid profile were compared with an unpaired Student t-test. The 24-h ambulatory blood pressure monitoring responses were subjected to two-way ANOVA with repeated measures. When significance was found, Scheffe's post hoc comparisons were performed. Pearson correlation coefficient was used to test the correlation between MSNA with 24-h mean blood pressure. Probability values of ≤0.05 were considered statistically significant.
Demographic and lipid characteristics.
The demographic and the lipid characteristics are shown in Table 2. There were no significant differences between AAS users and AAS nonusers in age, weight, and height. Blood glucose and triglycerides levels were similar between groups, but total cholesterol and LDL cholesterol were significantly higher and HDL cholesterol significantly lower in AAS users.
MSNA and muscle blood flow.
Data of MSNA are shown in Figure 1. Muscle sympathetic burst frequency and burst incidence were significantly higher in AAS users when compared with AAS nonusers (29 ± 3 vs 20 ± 1 bursts per minute, P < 0.01, and 43 ± 4 vs 30 ± 2 bursts per 100 heartbeats, P < 0.02). In contrast, forearm blood flow (1.92 ± 0.17 vs 2.77 ± 0.24 mL·min−1·100 mL−1, P < 0.01) and forearm vascular conductance (2.01 ± 0.17 vs 2.86 ± 0.31 U, P < 0.02) were significantly lower in AAS users (Fig. 2).
Ambulatory blood pressure.
Data of 24-h blood pressure and heart rate are shown in Figure 3. The 24-h blood pressure was significantly higher in AAS users compared with nonusers (systolic = 131 ± 4 vs 120 ± 3 mm Hg, P < 0.001; diastolic = 74 ± 4 vs 68 ± 3, P < 0.02; mean = 93 ± 4 vs 86 ± 3 mm Hg, P < 0.005). Similarly, awake and sleep systolic (awake = 134 ± 2 vs 123 ± 3 mm Hg, P < 0.01; sleep = 123 ± 3 vs 111 ± 2 mm Hg, P < 0.005), diastolic (awake = 77 ± 2 vs 71 ± 2 mm Hg, P < 0.05; sleep = 67 ± 2 vs 60 ± 2 mm Hg, P < 0.03), and mean blood pressure (awake = 96 ± 2 vs 89 ± 2, P < 0.02; sleep = 86 ± 2 vs 78 ± 2 mm Hg, P < 0.01) were significantly higher in users compared with nonusers. The 24-h heart rate was significantly higher in users compared with nonusers (74 ± 3 vs 68 ± 3 bpm, P < 0.02).
Association between MSNA and blood pressure.
Further analysis showed a significant correlation between MSNA and 24-h mean blood pressure levels (r = 0.75, P < 0.002; Fig. 4).
The main and new finding of this study is that AAS increases central sympathetic outflow in humans. MSNA directly measured from a multiunit postganglionic muscle sympathetic nerve is significantly increased in AAS users when compared with AAS nonusers. Evidence for increased sympathetic activity after administration of AAS in animals has been recently reported by other investigators. Pereira-Junior et al. found by spectral analysis that chronic treatment with supraphysiological doses of nandrolone decanoate increased cardiac sympathetic control in rats (26). The question that emerges from these findings is why AAS increases sympathetic nerve activity. The answer for this question is out of the scope of this study. However, previous observations provide some possibilities to explain such a puzzling physiological alteration. Impairment in arterial baroreflex control of heart rate has been found after chronic administration of stanozolol in rats (5). Because arterial baroreceptors reflexively modulate sympathetic nerve activity, it seems reasonable to think that alteration in baroreflex sensitivity contributes to the increase in sympathetic nerve activity in AAS users. However, we cannot rule out that the augmented sympathetic outflow after administration of AAS takes place in the central nerve system. First, the afferent arterial baroreceptors integrate in the central nerve system. Second, it has been demonstrated that steroids cross over the blood-brain barrier and act on specific androgen receptors in the central cardiovascular regulatory regions (27). In addition, there is evidence that supraphysiological steroid doses can selectively inhibit extraneuronal uptake of neuroamines and increase the vascular response to norepinephrine (14,15).
What are the hemodynamic implications for the augmented sympathetic outflow in AAS users? Our study shows at least two hemodynamic alterations that can be directly related to the augmented sympathetic outflow. Forearm blood flow was significantly lower and systemic blood pressure was significantly higher in AAS users compared with age-matched AAS nonusers. It has been well recognized that muscle blood flow depends upon the equilibrium between vasodilatory forces and vasoconstrictor forces. Increased sympathetic outflow in AAS users is consistent with disequilibrium to the vasoconstrictor forces. The early observation that testosterone increases the vascular response to norepinephrine (14) strengthens the idea that sympathetic exacerbation reduces forearm blood flow in AAS users. In addition, in pathological circumstances, as in patients with hypertension and heart failure in whom sympathetic nerve activity is substantially augmented, muscle blood flow is significantly reduced (19,24). We have also observed that intra-arterial infusion of phentolamine, an α1-adrenoceptor antagonist, increases forearm blood flow in patients with heart failure toward normal levels (28). Of course, there is the possibility that the reduced forearm blood flow in AAS users is mediated by alteration in nitric oxide production by endothelium. The lipid alterations reported in this study and by Ebenbichler et al. (9) may reduce nitric oxide bioavailability and, consequently, impair endothelial function. Previous studies clearly demonstrated that lipid disorders provoke endothelial dysfunction in humans (31).
In men, the increase in blood pressure during life span has been attributed to androgen effects (7). Therefore, someone could expect that supraphysiological doses of AAS anticipated the increase in blood pressure in young individuals. This hemodynamic alteration is in fact observed in this study. AAS users have greater 24-h blood pressure levels than AAS nonusers. Elevation in blood pressure at rest and during effort in AAS users has been previously reported by other investigators. In a cross-sectional study, Urhausen et al. (29) found that systolic blood pressure levels at rest and during exercise were higher in current AAS users when compared with non-AAS users. Palatini et al. (25) demonstrated that AAS users exhibited smaller reduction in blood pressure during the sleep period when compared with AAS nonusers. In addition, Grace et al. (13) reported increased blood pressure levels after the AAS cycle, although this alteration in blood pressure was no longer observed 6-8 wk after the AAS cycle. However, the mechanism by which AAS increases blood pressure is not fully understood. This study shows an association between augmented blood pressure levels and sympathetic nerve activity. A link between blood pressure levels and MSNA has been also observed in hypertensive patients (19). In that study, we found a significant association between blood pressure levels and MSNA, and between blood pressure levels and arterial baroreflex control of sympathetic nerve activity.
Another important piece of information in this study is the greater 24-h heart rate levels in AAS users. This finding is consistent with previous study in animal models in which alteration in autonomic control of heart rate was demonstrated. Pereira-Junior et al. (26) found a reduced parasympathetic index by spectral analysis in rats with administration of supraphysiological doses of nandrolone decanoate. In addition, these investigators observed a tendency toward an augmented sympathetic index in those rats.
We recognize limitations in our study. For ethical reasons, we could not conduct a controlled randomized study. However, the urine test confirmed that our individuals were in fact self-administrating AAS. The supraphysiological doses of AAS were studied at the end of the cycle of self-administration but not after discontinuation. Thus, we have no information regarding the possible recovery of MSNA and muscle blood flow levels after discontinuation of AAS in our study. The recovery of blood pressure levels after discontinuation of AAS is a controversial issue. Some investigators reported that blood pressure levels returned to baseline 6-8 wk after the AAS cycle (13). In contrast, others observed that delta day-night systolic blood pressure was decreased even after the period of AAS withdrawal (25). The fact that we have included only men in this study may limit the interpretation regarding the impact of AAS in humans. Similarly, we do not know the effects of AAS in older individuals.
This study provides important knowledge regarding physiological alterations provoked by AAS in humans. It shows that AAS increases sympathetic nerve activity and blood pressure and reduces muscle blood flow in humans. In addition, AAS causes a reduction and HDL cholesterol and an increase in LDL cholesterol. These findings have clinical implications that should be taken into consideration in medicine and sports practice. First, elevated MSNA is an independent predictor of mortality in patients with heart failure (2). Second, reduced forearm blood flow is associated with poor prognosis in those patients (2). Third, high blood pressure is considered one of the major risk factors for cardiovascular disease (8). Likewise, a reduction in HDL cholesterol and an increase in LDL cholesterol levels predispose to cardiovascular events in all age groups (3). The 24-h increases in heart rate associated with an imbalance between parasympathetic and sympathetic cardiac regulation may provide a key mechanism for arrhythmia and, in consequence, sudden cardiac death in AAS users. In contrast to our present study, other investigators did not find an increase in LDL cholesterol in AAS users (18). This controversy may be explained by the types of AAS used. In this study, the self-administration of AAS varied among individuals. This was not the case in other study, in which only two types of AAS were self-administered (18).
AAS causes abnormal neurovascular control in young men. The increased 24-h blood pressure levels in AAS users are associated with increased sympathetic nerve activity. These findings suggest that AAS increases susceptibility to cardiovascular disease in humans.
This study was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP grant no. 2005/59740-7), by the Conselho Nacional de Pesquisa (CNPq grant no. 474621/2004-9), and in part by the Fundação Zerbini. César Abreu Akiho was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP grant no. 03/09238-8), and Carlos E. Negrao and Maria U. Rondon were supported by the Conselho Nacional de Pesquisa (CNPq grant nos. 302146/2007-5 and 303518/2008-1, respectively).
The authors are indebted to the subjects who greatly contributed in this study.
The results of the present study do not constitute endorsement by the American College of Sports Medicine.
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Keywords:©2010The American College of Sports Medicine
ANABOLIC STEROIDS; BLOOD PRESSURE; SYMPATHETIC NERVE ACTIVITY; FOREARM BLOOD FLOW