Introduction

Flunitrazepam is a positive GABAA modulator of GABA neurotransmission that binds benzodiazepine sites on the GABA receptor and strongly enhances GABA receptor-mediated transmission [1,2,3]. Like all benzodiazepines, flunitrazepam mediates inhibitory neurotransmission in the brain producing sedative-hypnotic effects [4, 5]. This feature is greater than that of both diazepam [6, 7] and nitrazepam [8], as flunitrazepam induces sleep more quickly [9] and produces compelling anterograde amnesia [6, 7, 10, 11]. Flunitrazepam activity seems to be particularly significant in the limbic system [12, 13], thus, via the hypothalamus, influencing autonomic function and related cardiovascular changes [14, 15].

The cardiovascular effects of flunitrazepram [15] also seem to be dependent upon its primary peripheral vasodilatory effect. This distinctive action has been demonstrated in both experimental and human studies [16,17,18,19,20,21]. In isolated segments of the rat tail artery, flunitrazepam (not diazepam, however) produced marked relaxation by means of a 15% increase in diameter, even following maximum noradrenaline-induced constriction [16]. This vasodilatory effect has also been confirmed in human studies: a decrease of 15–24% in peripheral resistance was observed [17,18,19,20,21] following flunitrazepam administration. In healthy individuals, acute intravenous flunitrazepam administration consistently led to significant decreases in systolic blood pressure (SBP), diastolic blood pressure (DBP), and plasma noradrenaline (NE) [22,23,24,25]. The only study which evaluated the effect of a single oral flunitrazepam administration as a hypnotic agent in healthy subjects reported a significant fall in SBP values with no change in HR [26]. In patients with CHD, acute flunitrazepam administration produced a 15% reduction in blood pressure (BP) and a slight, transient increase in HR [21, 27]. A vasodilatory effect on the coronary vascular bed, together with a loss of coronary circulation autoregulation, was reported [21].

Flunitrazepam i.v. administration as a premedication in patients undergoing invasive diagnostic procedures, produced an increase in resting HR and a decrease in BP during bronchoscopy [28]. Conversely, no change was reported during gastroscopy [29, 30].

Use of flunitrazepam for induction of anesthesia in varying types of surgical operations constantly led to a significant reduction in BP [17,18,19, 31,32,33,34,35,36,37]; however, in only two studies was this reduction accompanied by an increase in HR [17, 34]. This lack of increase in HR in the presence of a decrease in BP might be related to the fact that, unlike diazepam, which has little effect on the baroreflex [38], flunitrazepam significantly attenuates baroreflex sensitivity [34, 36]. Interestingly, intravenous flunitrazepam administration (in patients who had already been anesthetized using other drugs) produced decreases in BP with unchanged HR [39, 40]. This suggests that these effects resulted from the direct vasodilatory action of flunitrazepam, and not from sleep induction and loss of consciousness.

The results of all these studies, however, are not conclusive, and comparison of results is not straightforward. This is mostly due to the varied nature of the study populations, distinct experimental and clinical settings, doses and administration routes, coupled with the relatively short duration of cardiovascular-effect monitoring. Moreover, to the best of our knowledge, to date, no study has evaluated the cardiovascular effects of flunitrazepam in healthy subjects who chronically consume the drug to induce sleep (the most frequent use of flunitrazepam).

In this context, the present study was undertaken to assess the effects of chronic evening oral administration of flunitrazepam on 24-h BP and HR in healthy young adults.

Materials and methods

Both male and female healthy volunteers, between 21 and 30 years, with a normal BP, a normal depression and anxiety evaluation and a normal BMI, ECG, and kidney function were eligible for enrolment in this randomized, double-blind cross-over study. Exclusion criteria were the following: history of smoking, shiftwork, diabetes mellitus, hepatic insufficiency, pregnancy, previous cardiovascular accidents, use of any type of drugs, and usual or occasional consumption of alcohol.

The study protocol was approved by the local Ethical Committee, and informed consent was obtained from all the subjects. Following an initial 2-week placebo period, subjects satisfying the inclusion criteria were randomized to flunitrazepam 1 mg or placebo, both administered once a day in the evening (22.00–23.00 h) for 4 weeks in two cross-over periods, each separated by a 2-week placebo period. This wash-out period was considered to be sufficient to prevent any carry-over effect [41]. At the end of each study period, 24-h non-invasive ambulatory BP and HR monitoring was performed using a validated device (Spacelabs 90207, Redmond, Washington) [42] that was programmed to measure BP every 15 min. Each recording was started in the morning and performed throughout a full 24-h period, during which subjects were allowed to follow their normal daily routine. The analysis of 24-h BP recordings were preceded by removal of artifacts, according to previously described editing criteria [43]. Recordings were excluded from the analysis when > 10% of all readings or more than one reading per hour was missed. Computed analysis of the individual recordings provided 24-h, night-time (23.00–07.00 h), morning-time (0.7.00–12.00 h), and afternoon/evening-time (12.00–23.00 h) mean values of SBP, DBP, and HR. At each visit, adverse events spontaneously reported were recorded.

Statistical analysis

The statistical analysis was performed using Analysis of Variance (ANOVA) and by means of the SPSS software package for Windows (version 11.0: Chicago, Illinois, USA); data are presented as means ± standard deviations. Paired tests were also used: a one-sample t test was used to compare values obtained after treatment administration; a two-sample t test was used to compare the change score (treatment-placebo) for a given parameter between the two groups.

Results

A total of 30 healthy young volunteers were recruited, but only 28 of them had the first ambulatory blood pressure recording complying with the quality criteria previously defined. Thus, only these 28 patients were randomized to receive placebo or flunitrazepam and included in the analysis. Their mean clinic SBP/DBP values were 125.2 ± 6.9/80.6 mmHg ± 3.3, and their mean resting HR was 75.4 ± 7.8 beats/min.

The main results of this study are shown in Table 1 and in Figs. 1, 2, and 3. Ambulatory monitoring data showed that 24-h SBP, DBP, and HR mean values were not significantly affected by flunitrazepam. Conversely, when analyzing separately the sub-periods of the day, different behaviors were observed. During nighttime (23.00–07.00 h), both treatments preserved the physiologic reduction of BP and HR. However, after flunitrazepam, SBP values were 6% and DBP 6.8% lower than those observed during placebo (both P < 0.05). Night-time HR mean values were unaffected.

Table 1 Ambulatory systolic blood pressure, diastolic blood pressure, and heart rate during placebo and flunitrazepam treatment
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Fig. 1
figure 1

Systolic blood pressure profile over the 24 h during placebo and flunitrazepam treatment. *P < 0.05

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Fig. 2
figure 2

Diastolic blood pressure profile over the 24 h during placebo and flunitrazepam treatment. *P < 0.05

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Fig. 3
figure 3

Heart rate profile over the 24 h during placebo and flunitrazepam treatment. *P < 0.05

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In the morning hours (07.00–12.00 h), i.e., during the 5 h following awakening, both the flunitrazepam and placebo administrations preserved the physiologic increase in HR and BP. Nonetheless, BP and HR mean values were significantly higher under flunitrazepam than under placebo: SBP and DBP were higher by 6.4% (+ 7.4 mmHg, P < 0.01) and by 4.6% (+ 3.4 mmHg, P < 0.05), respectively. HR increased by 5.3% (+ 3.9 beat/min, P < 0.05).

During afternoon/evening-time, SBP, DBP, and HR levels were not different in the two treatment groups.

With regard to the adverse effects reported by flunitrazepam-treated volunteers, 25 complained of anterograde amnesia before falling asleep. Twenty-two reported unusual very intense dreams: 11 subjects reported a prevalence of unpleasant dreams or nightmares with aggressive verbal or physical contents; 4 subjects reported a prevalence of pleasant dreams, with frequent sexual contents; 7 subjects did not remember the content of their dreams. Mild difficulty in waking up was reported by 4 subjects and 2 of them also complained of mild somnolence during the first hours of the day. On the contrary, 7 subjects referred to feel particularly active in the morning and more aggressive in dealing with everyday problems during treatment with flunitrazepam. Only 1 subject complained of palpitations in the morning hours.

Discussion

The first main finding of the present study was that chronic oral administration of flunitrazepam (taken in the evening as a hypnotic agent in healthy adults), although not interfering with the physiologic nocturnal fall in BP and HR, led to a significantly greater reduction in nocturnal BP values as compared to placebo. This finding is in accordance with observations made in all human studies to date, where flunitrazepam administration caused an immediate, rapid reduction in BP values [17,18,19,20,21]. However, the findings of these studies are not comparable with our results as those studies were carried out acutely, i.e., following intravenous administration of flunitrazepam to healthy subjects while awake or in patients under stressful conditions. These nocturnal reductions in BP, induced by flunitrazepam in the hours immediately following oral intake, are completely absent in oral administration of diazepam to the same category of subjects [44]. This does not seem to be related to central inhibitory effects [45, 46] but rather to direct vasodilatory effects, which are specific to flunitrazepam [16]. This aspect, however, had never been investigated during nighttime hours following oral administration of the drug.

It should be noted that any drug-related direct vasodilatory action produces an immediate reactive response through counter-regulatory mechanisms, with the activation of arginine, vasopressin, and renin. This is followed by a consequent increase in circulating angiotensin II and sodium retention, as reported with sodium nitroprusside, hydralazine, and nitroglycerine [47,48,49,50,51,52,53,54,55,56]. As a consequence, nocturnal BP behavior under flunitrazepam is likely to be the effect of a balance between the direct vasodilatory effect of flunitrazepam and counter-regulatory forces of activated neurohumoral factors.

Of particular interest is the behavior of nocturnal HR, whose values remained substantially unchanged and similar to placebo values, despite decreases in BP. This lack of HR adaptation to a fall in BP is likely to be related to reductions in baroreflex sensitivity (which is rapidly and greatly affected by flunitrazepam [34,35,36]). In this regard, flunitrazepam differs from diazepam, which does not significantly influence baroreflex sensitivity [38, 39].

The second main finding of this study is that physiological increases in BP and HR following awakening are significantly higher with flunitrazepam than with placebo. This increase in morning BP values has not been observed with diazepam, which, on the contrary, has been reported to cause decreases in SBP [44]. Therefore, increases in morning BP do not seem to be a class effect, but rather a characteristic and specific effect relating to flunitrazepam. The mechanisms underlying this effect are not easily explained; this is also due to a lack of previous similar observations in literature. We can only hypothesize that it could somehow be related to morning reductions in flunitrazepam plasma concentration with the consequent attenuation of direct vasodilatory effects and a prevalence of counter-regulatory neurohumoral responses. Indeed, it has been reported that, when direct vasodilator agents are discontinued, reflex counter-regulatory responses elicited by the agents do not cease immediately, but rather decline slowly over time [47,48,49, 51]. This regards the rebound activation of the sympathetic system, in particular, and of the renin angiotensin system (RAS) to an even greater degree, with consequent persistence of increased levels of angiotensin II (Ang II) and related effects on vascular smooth muscle cells and sodium retention [47,48,49, 51].

Increased values of morning HR could be related, in part, to the above-described counter-regulatory response to vasodilation, and, in part, to rebound activation of the baroreflex system (where reductions in flunitrazepam plasma concentrations might attenuate inhibitory effects of the drug on baroreflex sensitivity). Furthermore, we cannot exclude that, unlike what is found in the periphery, morning concentrations of flunitrazepam in the brain, although reduced, could be high enough to exert a GABA-mediated vagolytic effect, thus contributing to increases in HR.

Adverse events were found to be mild, predictable, and did not necessitate withdrawal of treatment.

To conclude, flunitrazepam, orally administered in the evening to induce sleep, causes a fall in BP which occurs rapidly and persists over the entire sleeping period, without concomitant changes in HR, followed by a transient rebound rise in BP and HR. The mechanisms underlying these cardiovascular effects remain unclear, and their understanding needs further and more detailed pathophysiologic study.