Effects of nitroglycerine on coronary flow velocity before and during adenosine provocation
Wittfeldt et al. Cardiovascular Ultrasound
Effects of nitroglycerine on coronary flow velocity before and during adenosine provocation
Ann Wittfeldt 0 1
Anders Jeppsson 1
Li-Ming Gan 1
0 Department of Cardiology, Sahlgrenska University Hospital , 41345 Gothenburg , Sweden
1 Dept of Molecular and Clinical Medicine, Institute of Medicine at Sahlgrenska Academy, University of Gothenburg , 41345 Gothenburg , Sweden
Background: Transthoracic echocardiography-assessed coronary flow velocity reserve (CFVR) evaluates coronary microvascular arterial function. Coronary flow velocity measurements at baseline and during hyperemia are used to calculate CFVR. Adenosine infusion induces hyperemia but it is not known if it causes a maximal response. We hypothesized that pre-treatment with nitroglycerine before adenosine provocation enhances hyperemia. Methods: Twenty-three healthy study subjects (mean age 27.5 ± 5.5, 35% women) underwent CFVR measurements before and after pretreatment with sublingual nitroglycerine (0.5 mg). Hyperemia was induced by adenosine infusion (140 μg/kg/min). In addition, the effect of nitroglycerin on left main coronary artery diameter was assessed. Results: Pretreatment with nitroglycerine increased median CFVR from 3.6 (range 2.8-4.3) to 5.0 (4.1-6.0), p = 0.002. The increase was caused by a marked reduction in baseline coronary flow velocity 17 (15-24) vs 27 (19-31) cm/s, p < 0.0001) while hyperemic velocity remained unchanged (90 (68-116) vs 93 (75-105) cm/s, p = 0.48). Nitroglycerin significantly dilated the left main coronary artery (from median 3.1 (2.7-3.6) mm to 3.8 (3.1-4.3) mm, p = 0.018). Conclusion: Pretreatment with nitroglycerine dilates coronary arteries and increases coronary flow velocity reserve indicating that adenosine alone causes a submaximal hyperemia.
Ultrasound; Non-invasive; Coronary flow velocity reserve; Nitroglycerine
Transthoracic color Doppler echocardiography-assessed
coronary flow velocity reserve (CFVR) is an emerging
non-invasive method to quantify coronary microvascular
function and evaluate volumetric coronary flow reserve
(CFR) [1–4]. CFVR has opened up new possibilities to
explore cardiovascular physiology and to evaluate
responses to treatment, due to its good reproducibility
and low method-dependent variability if performed by
well-trained operators [5–7], and it has been shown that
low CFVR predicts epicardial coronary artery stenosis
[8–10] and cardiovascular events in different patient
populations [11–13]. CFVR is suitable for repeated
measurements of coronary function since it is
noninvasive, without radiation, easily accessed with a clinical
ultrasound device and is associated with acceptable
discomfort to the patient. With modern ultrasound
platforms it is also possible to non- invasively evaluate
size and morphology of small structures like certain
segments of coronary arteries [14, 15].
Mean coronary flow velocity at baseline and during
maximum hyperemia are used to calculate CFVR.
Infusion of the vasodilator adenosine is an established
method to induce hyperemia  and studies has
suggested 140 μg/kg/min as suitable dose . Adenosine
acts mainly on small resistance vessels, but influences
also to a lesser degree epicardial arteries . Even small
changes in epicardial vessel diameter would result in
alterations of flow velocity . Thus, it is yet not
known whether CFVR due to adenosine-induced
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dilatation of epicardial vessels underestimate CFR, which
is a volumetric flow reserve.
It is possible that other vasodilators alone or in
combination with adenosine would induce an even higher
level of hyperemia . Nitroglycerin (NTG) is a
vasodilator mainly acting on epicardial arteries [20, 21].
NTG may be used in combination with adenosine
during measurements of flow fraction reserve in connection
to coronary angiogram in patients with coronary artery
disease. In this context NTG is in combination with
adenosine used to maximize blood flow and prevent wire
induced artery spasm.
We hypothesized that pre-treatment with nitroglycerine
before adenosine provocation in CFVR measurements in
healthy volunteers enhances the hyperemia further in
comparison to adenosine alone, due to maximum
dilatation of not only the resistance vessels but also epicardial
arteries. In addition, we explored the effects of
nitroglycerine on basal coronary flow and coronary artery diameter
and assessed the reproducibility of repeated CFVR
measurements using adenosine infusion with a short wash out
Twenty-three healthy volunteers (mean age 27.5 ±
5.5 years, 38% women) participated in the study. Inclusion
criteria were age 18–40 years, normal BMI and no
chronical or acute illness requiring medication. Exclusion criteria
were history of asthma, chronic obstructive pulmonary
disease, anxiety disorder, and ongoing infection. Study
subjects characteristics are presented in Table 1.
All study subjects were investigated on two consecutive
days. The study protocol on each study day included
Table 1 Study subjects characteristics. Number and percentage
or mean and standard deviation
Body mass index (kg/m2)
White blood cell count (×109/L)
Hemoglobin concentration (g/L)
Left ventricular ejection fraction (%)
Heart rate (beats per minute
two sets of coronary flow velocity measurements at
baseline and during adenosine infusion. On the study
days all subjects were fasting for four hours and were
caffeine free for at least 24 h before the investigation.
On day 1 a suitable vessel segment of mid left anterior
descending artery (LAD) regarding measurement quality
and breathing interference was identified with
transthoracic color doppler echocardiography. During the first
measurement (1:1), baseline coronary flow velocity was
recorded and thereafter adenosine infusion at a rate of
140 μg/kg/min was started. Hyperemic coronary flow
measurements were obtained during 5 min where the
highest recordings were registered. After completion of
the measurements, the adenosine infusion was paused.
After a ten minute washout period a new measurement
period was started (1:2) when new baseline
measurements were obtained before the adenosine infusion was
re-started at the same rate for new hyperemic
measurements. Heart rate and systolic blood pressure were
recorded every minute during adenosine infusion to
detect potential adverse reactions.
On day 2, two new measurements were performed
(2:1 and 2:2). The measurements were identical to the
procedures on day 1 with the exception that after five
minutes of the washout period between the two sets of
measurements 0.5 mg of sublingual nitroglycerine
(Recip, Jordbro, Sweden) was administrated to the study
subjects. Five minutes after nitroglycerine administration
new baseline measurements and adenosine–induced
hyperemic measurements were performed (period 2:2).
In a subgroup of nine subjects with good visualization of
the left main coronary artery, the diameter of the vessel was
measured before and after nitroglycerine administration.
Coronary blood flow velocity measurements
A Siemens Acuson platform equipped with a 4V1C
transducer with 3.5 MHz color Doppler frequency and
1.75 MHz spectral Doppler frequency (Siemens, Acuson
Sequoia 512, Mountainview) was used to measure coronary
flow velocity (CFV) in the mid to distal left anterior
descending coronary artery (LAD), in a slightly modified
apical two-chamber view. The same operator performed all
measurements to ensure minimum variability. CINE-loops
and Doppler images were stored for offline analysis using
Tomtec image analysis software (Image Arena 2.9.1,
Tomtec Imaging Systems, GmbH, Unterschleissheim,
Germany). CFVR was calculated as the ratio between mean
hyperemic CFV and mean baseline CFV. A typical
recording of coronary flow velocity at baseline without
nitroglycerine is shown in Fig. 1.
Diameter of the left main coronary artery
The left main coronary artery was visualized in a
modified short axis view at the level of aortic root and a
Fig. 1 Recording of coronary flow velocities in an individual study subject. The velocities were recorded at baseline (a), at adenosine-induced
hyperemia (b), at baseline with nitroglycerine (c), and at adenosine-induced hyperemia with nitroglycerine (d)
CINE-loop of a full cardiac circle with a frame rate up to
70 frames/s was stored for offline measurements. The
diameter was measured as close as possible to the R- wave
and at the same location both at baseline and after
No data for a sample size calculation was available
before study start. The number of included subjects
are based on own experiences and similar studies.
Data was not normally distributed. Data are thus
presented as median and range. Wilcoxon matched
paired test was used to compare mean CFV, CFVR
and artery diameter before and after adenosine and
nitroglycerine administration. Friedman’s test was
used to compare the two baseline measurements on
day 1 and the baseline measurement on day 2.
Baseline flow velocity
There was no significant difference in flow velocity
between the two baseline measurements on day one
(1:1 and 1:2) and the first baseline measurement on
day two (2:1), p = 0.82, Table 2. In contrast, baseline
flow velocity was significantly lower after nitroglycerine
administration (2:2) in comparison to the measurement
before nitroglycerine (2:1) and to the two measurements
on day 1, p < 0.001, Table 2.
Table 2 Coronary flow velocity at baseline and during
adenosine-induced hyperemia. Median and range
Day 1, measurement 1 (1:1)
Day 1, measurement 2 (1:2)
Day 2, measurement 1 (2:1)
Day 2, measurement 2
(after nitroglycerine) (2:2)
Coronary flow velocity during hyperemia
Maximum velocity during hyperemia was higher during
the second measurement period on day 1 (1:2) compared
to the first measurement the same day (1:1), p = 0.021),
Table 2. No significant differences in maximum velocity
during hyperemia were observed between the first
measurement on day 1 (1:1) and the first
measurement day 2 (2:1), p = 0.20. Maximum velocity during
nitroglycerine administration (measurement 2:2) did
not differ significantly compared with the
measurement without nitroglycerine on the same day (2:1) or
to the hyperemic measurements on the first day (1:1
and 1:2), p = 0.53, Table 2.
Coronary flow velocity reserve
CFVR during the different measurement periods are
shown in Fig. 2. There was no significant difference in
CFVR when comparing the first measurements on each
study day (measurement 1:1 and 2:1), 3.7 (2.3–7.2) vs.
3.6 (2.3–6.9), (p = 0.39). CFVR after nitroglycerine
administration (measurement 2:2) was significantly higher
than with only adenosine (measurement 2:1), 5.0 (2.4–9.2)
vs 3.6 (2.3–6.9), (p = 0.002) and significantly higher than at
the two measurements on day 1, 3.7 (2.3–7.2), (p < 0.001)
and 3.9 (2.5–7.3), (p = 0.007), respectively. CFVR was
significantly higher during the second measurement on
day 1 (1:2) than during the first measurement the same
day (1:1), (p = 0.018).
Effects of nitroglycerine on vessel diameter
Left main coronary artery dilated significantly after
nitroglycerine administration (from median 3.1 (2.7–3.6)
mm to 3.8 (3.1–4.3) mm, p = 0.018), Fig. 3.
The main findings of the present study were that
pretreatment with nitroglycerine before adenosine-induced
hyperemia significantly increased coronary flow velocity
In the present study we hypothesized that
pretreatment with nitroglycerine before adenosine
provocation in CFVR measurements would enhance the
hyperemia further in comparison to adenosine alone.
This hypothesis was based on the known vasodilating
effects of nitroglycerine on epicardial arteries and
clinical experiences from fractional flow reserve (FFR)
measurements and myocardial scintigraphy where pretreatment
with NTG may be used to enhance hyperemia [20–22].
After administration of NTG, there was a reduction in
baseline coronary flow velocity (before hyperemia) by
approximately 30%. This reduction in flow velocity is
more likely a result of a dilatation of the epicardial
arteries, than due to a reduction of total blood flow, since the
same amount of blood at a constant flow in a larger
vessel leads to a reduction in velocity. The effect on
vessel diameter is supported by the present study where
the diameter of the left main coronary artery increased
with approximately 20% after NG administration. Our
baseline results thus confirm the results shown by
Takagi et al.  where they, using the same
methodology, found that NTG administration caused a decrease
in coronary resting flow velocity and a dilatation of
Fig. 2 Coronary flow velocity reserve (CFVR) at baseline and during
adenosine-induced hyperemia. NTG = nitroglycerine. * = p < 0.05,
** = p < 0.01, *** = p < 0.001
Fig. 3 Left main coronary artery diameter before and after
During adenosine provocation after NTG
administration one would expect a reduction of coronary blood
flow velocity, compared to adenosine provocation
without NTG, since NTG dilates epicardial vessels. Contrary
to this hypothesis, there was no significant change in
hyperemic coronary blood flow velocity when compared
to the measurement without nitroglycerine. This may be
interpreted as an increase in total blood flow through
the myocardium after the combination of NTG and
adenosine administration, compared to only adenosine.
Thus, our results suggest that NTG pretreatment
increased adenosine-induced coronary blood flow in
healthy volunteers, supporting that epicardial vessel
dilation is essential to cause maximal hyperemic flow.
The reduced baseline velocity resulted in a
significantly increased CFVR even though nitroglycerine did
not increase hyperemic blood flow velocity (Table 2).
From a methodological point of view, CFVR is a good
estimate of CFR only when the epicardial vessel diameter
is maintained before and during adenosine infusion. In
case epicardial vessels dilate in response to increased
shear stress e.g., following inceased flow during
adenosine challenge, CFV increase measured at the same
epicardial vessel site as at baseline may be
underestimating the true flow increase. This indicates that
CFVR with nitroglycerine in combination with adenosine
might give a better volumetric coronary flow reserve
(CFR) estimation and a more stable measurement routine
than hyperemia induced by adenosine alone .
The results of the present study also demonstrate that
the reproducibility of flow velocity measurements is
satisfactory since the baseline measurements both at
the same day and between the two study days did not
differ significantly. This indicates that non.invasive
flow velocity measurements are stable and can be
used to detect even relatively small changes in flow
velocity over time. A low intraindividual variability
over time is a key factor for using this technique to
evaluate possible changes in coronary flow. However,
there was one potentially important exception. The
flow velocity during adenosine-induced hyperemia was
significantly higher during the second measurement
period on day 1 in comparison to the first
measurement the same day. The wash-out period between the
two measurements was only 10 min and it is possible
that the effects had not fully ceased, even though the
half life of adenosine is only a few seconds.
Alternative explanations include upregulation of adenosine
receptors or increased sympathicus drive. The latter
explanation may be less likely since basal flow and
heart rate was unaltered between the two
measurements. Independently of the reason, a longer washout
period than 10 min can be recommended if repeated
adenosine- induced hyperemic periods are required.
The present study had some limitations. The size of the
study population was limited. In particular, left main
diameter was measured only in eight patients. This may
reduce the validity of our observation. However, the 20%
difference between left main diameter before and after
nitroglycerine administration is within what previously
have been reported (8–25%) [21, 24]. A further
limitation is that the off line measurements were not blinded
to the operator. Finally, the study was performed in a
young, healthy population with healthy coronary arteries
and it is thus unclear whether the results can be applied
on patients with cardiac disease. A prospective study
with a larger number of subjects in a relevant patient
population with manifested coronary artery disease
would therefore be desirable.
Pre-treatment with nitroglycerine before adenosine
infusion could be a possible way to minimize the difference
between CFVR and CFR. It could also ensure maximal
coronary artery dilatation resulting in a more accurate
way of evaluating maximal coronary flow reserve.
CFR: Coronary flow reserve; CFV: Coronary flow velocity; CFVR: Coronary flow
velocity reserve; LAD: Left anterior descending artery; NTG: Nitroglycerine
This work was supported by University of Gothenburg. The sponsor had no
influence on the analysis and interpretation of data, in the writing of the
report, or in the decision to submit the paper for publication.
AW designed the study, collected and analyzed data and wrote the first draft
of the manuscript. AJ analyzed the data and revised the manuscript. LMG
designed the study, collected and analyzed data and revised the manuscript.
All authors read and approved the final manuscript.
Ethics approval and consent to participate
The study protocol was approved by the Regional Ethical Review Board in
Gothenburg, Sweden (number 449–06).
All participants signed a consent form after oral and written information.
All data generated or analysed during this study are included in this
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