This study evaluated the effects of vagus nerve stimulation (VNS) therapy on cardiac function in canine models, both on its own and when combined with common heart failure drugs. The study found that a VNS frequency of 10 Hz was most effective at reducing heart rate and increasing heart rate variability, markers of improved autonomic function. When VNS was combined with drugs like beta blockers, ACE inhibitors, and ivabradine, it did not significantly alter VNS-induced heart rate responses. However, there were some differences in heart rate and variability depending on the specific drug combination used with VNS. Overall, the results suggest VNS therapy is safe to use along with standard heart failure medications and may provide

1. Modulatory Effect of Autonomic Nervous System Stimulation on
Cardiac Function is Maintained When Given in Combination with
Commonly Prescribed Heart Failure Drugs
Abstract
Progression of cardiac disease is associated with concurrent changes in heart muscle and the
nerves that innervate it. These include sympathetic nerves that make the heart beat faster and
stronger and parasympathetic nerves that have the opposite effect. During heart failure, activity
on sympathetic nerves increases with corresponding decreases in parasympathetic activity. While
beneficial in the short-term to help maintain cardiac output, excessive sympathetic activity
contributes to the deterioration of heart muscle. Standards of practice in heart failure include
treatment with drugs such as beta blockers, diuretics, and drugs targeted at angiotensin pathways.
While exerting a degree of protection in heart failure, such drug based approaches usually just
slow the deterioration process. Recent advances in bioelectronic medicine have opened the door
for novel therapies for heart disease that target the nervous system. Vagus nerve stimulation
(VNS) is one such approach, with multiple trials currently underway for treating heart failure.
Since such stimulation will on first pass be an add-on therapy, this project evaluated the effects
of commonly prescribed heart failure drugs on the efficacy of VNS in a canine model. Our
results show that VNS-induced heart rate responses are not altered significantly by drugs.
However, there are differences between the effects of the different drugs combined with VNS on
mean heart rate and heart rate variability. These results further suggest the safety of VNS when
given in combination with commonly prescribed heart failure drugs.
Keywords: Heart failure, vagal nerve stimulation, sympathetic nervous system, intrinsic cardiac
nervous system, holter monitoring, autonomic regulation therapy

2. 2
Introduction
Heart failure is a major health problem that causes one out of every nine deaths in the USA and
costs about $32 billion a year. There are many different types of this condition, but this report
will focus solely on left-sided heart failure. There are two main types of left sided heart failure,
heart failure with reduced ejection fraction (HFrEF) and heart failure with preserved ejection
fraction (HFpEF). The major problem in HFrEF is the reduction in the ejection fraction of the
left ventricle, which is the percentage of the blood in the left ventricle at the end of diastole that
is ejected out at the end of systole. HFrEF results in weak blood flow and circulation and a high
heart rate, as the cardiac nervous system tries to compensate for a failed systole by increasing the
heart rate. This is done by increasing activity on sympathetic nerves to the heart with
corresponding decreases in parasympathetic nerve activity. HFrEF often results from an ischemia
leading to myocardial infarction. Such disease processes change the sensory information arising
from the cardiovascular system and thereby reflexively alter the sympathetic and
parasympathetic nerve inputs to the heart (Buckley et al., 2015). In HFpEF, the main problem is
a very stiff ventricle and diastolic dysfunction, in which the heart can contract relatively
normally but cannot fully expand during diastole. Such pathologies are also associated with
altered activities on sympathetic and parasympathetic nerves (Munagala et al., 2005; Gladden et
al., 2014). This report will include data from both these types. Recently, a new treatment has
been gaining momentum called vagus nerve stimulation (VNS) that is part of a broader category
of autonomic regulation therapy (ART) (Buckley et al., 2015). ART has at its heart, the use of
targeted therapies, directed against specific nexus points of the cardiac nervous system to
stabilize reflex function and to thereby protect heart muscle. VNS aims to address a major
maladaptive response that the cardiac nervous system has to heart failure, namely excessive
sympathetic drive with the loss of parasympathetic restraint (Figure 1).

3. 3
Autonomic dysregulation and increased sympathetic drive. The main reactive
responses that the cardiac nervous system has in the presence of heart failure are an increase in
sympathetic drive, a decrease in vagal nerve activation, and overall restructuring of the cardiac
autonomic nervous system neuronal hierarchy. It increases sympathetic activation in order to
compensate for the decrease in heart output due to systolic dysfunction in HFrEF and diastolic
dysfunction in HFpEF. This is because the sympathetic nervous system is the part of the
autonomic nervous system (ANS) that generally increases activity in various human organs. The
cardiac sympathetic nervous system is adrenergic, meaning that with an increase in sympathetic
activation, there is also an increase in the net amount of norepinephrine innervation. While this
response may help deal with heart symptoms in the short term, it has various negative effects that
outweigh any benefits gained over the long term, such as the destruction of cardiomyocytes,
progression of heart failure and development of arrhythmias.
Sympathetic overactivation is due in a large part to local afferent inputs received after
heart failure that send excitatory stimuli to cause excess adrenergic innervation (Zucker et al.,
2012). Afferent neurites in the intrinsic cardiac ganglionated plexi (ICGP) send signals to their
local circuit neurons (LCNs), which in turn cause the postganglionic sympathetic efferent somata
in ICGPs to change their activity. These stimuli from neurites on diseased heart tissue may also
affect other levels of the cardiac neuraxis, including the intrathoracic extracardiac sympathetic
efferent somata, spinal cord, medullary regions, all the way up to the insular cortex, which is the
highest center for cardiac command. This response restructures and destabilizes the ANS by
creating a conflict between central command and peripheral local afferent inputs. This
dysregulation leads to the progression of cardiac disease and heart failure.

4. 4
This maladaptive response to heart failure is essentially an autonomic imbalance, where
one side of the system, the adrenergic, sympathetic side, is overstimulated while the other side,
the cholinergic, parasympathetic side, is understimulated. VNS in general targets the cervical
vagus nerve, which has two branches, the right and left sides, and is part of the parasympathetic
system. While it contains both efferent and afferent fibers, VNS essentially helps counterbalance
the downregulation of cholinergic fibers and stabilize the ANS. VNS also targets multiple levels
of the cardiac nervous system, which is necessary since it has been shown that targeting only one
area of the cardiac neuraxis does not have consistent, reliable effects because of the redundant
and overlapping way that cardiac neurons regulate cardiac regions and indices as well as the
interconnectedness of the different levels of the cardiac neuraxis (Dell'Italia, 2011; Florea &
Cohn, 2014; Fukuda et al., 2015). In this way it is better than ablation of particular ganglionated
plexi of the intracardiac nervous system with the intention of bradycardia induction because that
method does not address multiple areas and levels of the cardiac ANS.
One particularly relevant effect of increased sympathetic drive is the decrease in heart
rate variability (HRV). Normally, the heart rate varies significantly over the day. For example,
when in a state of activity, the heart rate is much higher than when the animal is in a state of rest
or sleep. Heart failure decreases HRV. This once again is a reflection of the increase in
sympathetic nerve activity and a decrease in parasympathetic activity (Florea & Cohn, 2014;
Fukuda et al., 2015).
ART, including VNS, has grown immensely as a field and so far there have been
numerous preclinical trials and a few clinical trials. These trials have generally shown VNS to be
an effective method of reducing sympathetic overdrive and helping with preserving heart
function. Among the clinical trials, the three major ones are called ANTHEM-HF, INOVATE-

5. 5
HF, and NECTAR-HF. ANTHEM-HF was a phase I-II trial that tested the safety, tolerability and
efficacy of VNS in heart failure patients (Premchand et al., 2014). INOVATE-HF is a phase II
trial (De Ferrari et al., 2011; De Ferrari, 2014) and NECTAR-HF is an ongoing phase III trial
(Zannad et al., 2015). INOVATE-HF failed to reach its primary endpoints likely because of the
stimulation parameters that were used. ANTHEM-HF, however, was a successful study that
showed the safety and tolerability of VNS, as well as improvements in heart failure symptoms
and left ventricular heart function.
For current state of practice, vagus stimulation is an add-on therapy used in conjunction
with pharmacological therapies including beta blockers, diuretic, and angiotensin pathway
inhibitors (Dell'Italia, 2011; Florea & Cohn, 2014). How these drugs can change the response to
vagus nerve stimulation is not well understood. The experiment being discussed in this proposal
consisted of 24-48 hour holter monitoring that tested the effects of VNS on cardiac function in a
canine model. From these continuous recordings of the heart, autonomic function can be
assessed using mean heart rate and heart rate variability (HRV), with and without the drugs of
interest. The overall objective of this study is to see if traditional HF-directed drug therapy
interferes with VNS.
Materials and Methods
Part 1. Holter monitoring of canine models to determine optimal stimulation parameters.
According to the design of this study (part 1), normal canines (n=8) were implanted with VNS
devices on the right and left cervical vagus nerves and programed to deliver chronic, intermittent
stimulation (14 s ON, 66 s OFF). Holter monitors recorded ECG continuously for 24 hours
during various combinations of stimulation frequency (5, 10, and 20 Hz) and pulse widths (250

6. 6
and 500 µs). VNS amplitude (mA) was programmed to whatever value obtained a 10% acute
decrease in heart rate; it averaged 1.75 mAmp.
Objectives and data analysis (part 1). The main objective of this study was to measure the
optimal frequency parameter to obtain the best stabilizing influence on mean heart rate and
HRV. HRV is done in both the time and frequency domains using 24 or 48 hours of continuous
electrocardiogram (ECG) recording. ECGs were recorded using DR200 Holter and event
recorders. Analysis was performed using LX Analysis Software. The primary endpoints were the
number of times per hour in which the change in consecutive normal sinus intervals exceeds
50ms (pNN50) and mean heart rate. In general, the lower the heart rate and the greater the
pNN50, the better the autonomic state is for the individual.
Part 2. Comparing heart rate responses of VNS and VNS+drugs. According to the design of
this study (part 2), four normal canines were implanted with VNS therapy systems on the right
cervical vagus (RCV) nerve and placed on medical therapy, either individually or in
combination. These animals were administered several common drugs prescribed for heart
failure along with the vagal nerve stimulation. These drugs were given twice a day orally for
successive 2 week phases. There were several combinations of drug doses:
1) Metoprolol, 12.5 mg, 2x daily;
2) Enalapril, 2.5 mg, 2x daily;
3) Ivabradine, 5mg, 2x daily;
4) Metoprolol, 12.5 mg and Enalapril, 2.5 mg, 2x daily;
5) Metoprolol, 12.5 mg, Enalapril, 2.5 mg, and Ivabradine, 5mg, 2x daily.

7. 7
Metoprolol is a beta blocker commonly given to patients to counter the effects of heart
failure caused by increased circulation of norepinephrine and catecholamines in general. It
lowers heart rate and helps the left ventricle relax and fill more completely. Enalapril is an ACE
(angiotensin-converting enzyme) inhibitor, which lowers the amount of angiotensin produced in
the body. Angiotensin is excessively produced as a response to heart failure. By reducing the
production of this substance, ACE inhibitors help blood vessels relax and widen, reducing the
toll on the heart. Finally, ivabradine is a funny channel blocker, and helps reduce the heart rate
by blocking the funny current of the cardioelectrical system. Together these drugs help to reduce
the autonomic imbalance associated with heart failure but reducing sympathetic/angiotensin
activation (metoprolol and enalapril) or by directly slowing heart rate (ivabradine).
In awake animals, heart rate responses to repeating cycles of 10 Hz RCV VNS (14-s ON-
time, 48-s OFF-time) were measured at the end of each 2-week therapy phase and compared to
baseline.
Objectives and data analysis (part 2). The main objective of this study was to determine how
drugs affected the efficacy of VNS when administered together. The primary endpoint was the
heart rate as a % of the baseline recorded values.
Part 3. Holter monitoring of animal models at 10 Hz VNS and drug therapy. According to
the design of this study (part 3), eight canines (n=8) were implanted with one bipolar electrode
on each of the right and left cervical vagus nerves. These electrodes were connected to separate
implanted VNS stimulators (Demipulse Model 103 Stimulator). This stimulation system was
programmed to deliver chronic, intermittent electrical stimulation of the vagal nerves. For these
animals, VNS was delivered at 10 Hz, 250 μs pulse width, stimulus intensity of ~2.00 mA and
with a 25% duty cycle (14 sec on and 48 sec off). Meanwhile, holter monitors were set to record

8. 8
EKG continuously for 24-48 hours, with and without relevant drug treatments. Holters were
obtained 10 days after start of drug treatment.
These animals were administered the same combination of drugs and VNS as in the
previous part of the study.
Objectives and data analysis (part 3). The main objective of this study was to measure the
effect of VNS and drug therapy on the mean heart rate and heart rate variability (HRV). HRV is
done in both the time and frequency domains using 24 or 48 hours of continuous
electrocardiogram (ECG) recording. ECG’s were recorded using DR200 Holter and event
recorders. Analysis was performed using LX Analysis Software. The primary endpoints were the
number of times per hour in which the change in consecutive normal sinus intervals exceeds
50ms (pNN50) and mean heart rate.
Statistical analysis. Data in figures are presented as the mean ± standard error. A repeated
measures mixed analysis of variance model was used for comparisons of mean current and
frequency curves generated in different manipulation conditions. Repeated measures analysis of
variance model with Tukey multiple comparison was used for analysis of threshold. P<0.05 was
considered to be statistically significant. Statistical analyses were performed using GraphPad
Prism version 6 and STATA 10.2.
Roles and accreditation. The author of this report worked as a data analyst in this study. The
study was overseen by the director of the lab. Laboratory technicians who are part of this lab
worked on the actual hands-on part of the experiment, such as implanting the bipolar electrodes
and setting up the holter monitoring systems.

9. 9
As a data analyst, the author worked to translate the data from the holter recordings into
data tables and charts. The author first translated and recorded the holter monitoring reports into
excel sheets. These excel documents and data were then further organized and translated into
easily understood tables and graphs with error analysis. Finally, the author worked on
interpreting this data in this report.
Results and Discussion
10 Hz frequency for VNS results in optimum cardiac influences. To determine the optimal
VNS frequency that provides the best stabilizing influence on mean heart rate and HRV, 8
normal canines were subjected to VNS at the stimulation frequencies of 5, 10 and 20 Hz, as
described above in study part 1 in Materials and Methods section. As summarized in Figure 3, a
10 Hz frequency parameter results in the best values for basal cardiac electrical function as
compared to animals that did not receive VNS (Figure 3).
Chronic RCV VNS (right-sided) resulted in a small increase in HRV and a small decrease
in heart rate with a stimulation frequency of 5 Hz. An increase in HRV and decrease in heart rate
is a marker of parasympathetic activity (Armour, 2008; De Ferrari, 2014). It resulted in a more
significant increase in HRV and a more significant decrease in heart rate with 10 Hz. However,
this effect did not occur with a frequency of 20 Hz, which did not increase HRV and did not
decrease the heart rate. Overall, lower frequencies of ART, especially 10 Hz, were deemed to
deliver the optimum level of cardiac control with minimal negative effects and therefore used in
the two subsequent studies. The reason that 10 Hz provided the optimum response is because this
is close to the natural frequency of the cardiac efferent neurons (Hardwick et al., 2014).
Stimulating at lower frequency does not fully exploit the system capabilities and stimulating at

10. 10
higher frequencies has many of the electrical impulses blocked at the synaptic junctions between
nerves.
Effective cardiac control is maintained by VNS in the presence of commonly prescribed
drugs for heart failure. In order to determine whether drugs that are currently used in patients
with heart diseases would affect the effectiveness of VNS, normal canines were subjected to
VNS and treated with various drugs singly or in combinations, as described above in Materials
and Methods section. The main result derived from this study is that commonly prescribed drugs
for heart failure do not reduce the effectiveness of vagal control over cardiac responses of heart
rate. These results are depicted in Figure 4.
Heart rate was reduced by up to 25% from the baseline values by control experiments. As
the intensity was increased, the heart rate was progressively decreased by RCV VNS. The
presence of funny channel blockade, ACE inhibition, and beta blockade in any combination did
not significantly alter heart rate responses to RCV VNS. Therefore, VNS retains effective control
over the cardiac nervous control despite the presence of these drugs.
Individual differences between the drugs with VNS. Study part 3 further investigated the
effects of RCV VNS in over the long term with 24-48 hour holter monitors. The same drugs
were used as in study 2. Results of part 3 study are shown in Figures 5 and 6. Metoprolol along
with VNS decreased the heart rate and HRV as compared to the VNS control (Figure 6). This is
likely due to the fact that metoprolol is a beta blocker and inhibits adrenergic activity in the
sympathetic efferent fibers in the cardiac neuraxis (Figure 2). The reason that HRV was not
increased, a marker of parasympathetic activity, is probably that metoprolol only impacts the
sympathetic nervous system, and does not directly induce parasympathetic excitation.

11. 11
Enalapril+VNS significantly increased mean heart rate and decreased HRV. This effect is likely
due to enalapril’s ACE inhibition. Circulating angiotensin II has an effect on both sympathetic
and parasympathetic fibers in the ICNS, the stellate ganglion and on beta receptors (Figure 2).
Angiotensin II potentiates the responses of IC neurons to both norepinephrine (sympathetic) and
bethanechol (parasympathetic) in normal animals (Girasole et al., 2011). The attenuation of this
effect by ACE inhibitors leads to an increase in mean heart rate and decrease in HRV. More light
can be shed on the effect of Enalapril+VNS on the ICNS by looking at the results that a
combination of Enal+Meto+VNS had on mean heart rate and HRV. Both mean heart rate and
HRV were reduced by this combination. This suggests that the attenuation of Ang. II had a larger
impact on parasympathetic fibers than sympathetic fibers. Because of this, when enalapril
decreased Ang. II circulation, it may be that parasympathetic potentiation was affected more than
sympathetic potentiation and mean heart rate increased, decreasing HRV. However, when that
sympathetic activity was blocked by beta blockers (metoprolol), heart rate decreased and HRV
was higher compared to Enal+VNS, but lower than VNS control values.
VNS, when given with ivabradine, the funny channel blocker, resulted in a decreased
heart rate and increased HRV compared to control values (Figures 5 and 6). This is because of
ivabradine’s blockade of the funny current. The funny current controls the rate of cardiac activity
(heart rate) because it is involved in the activation of diastolic depolarization (DD). The
parasympathetic system modulates the heart rate by slowing down DD, while the sympathetic
nervous system speeds up DD. By blocking this current, funny channel blockers slow down DD
activation and therefore the heart rate. This augments the parasympathetic effect and results in a
decrease in heart rate, therefore increasing HRV. There was no statistically significant effect on
heart rate when the VNS was combined with all three drugs (Metoprolol + Enalapril +

12. 12
Ivabradine + VNS) as compared to animals treated with VNS alone. The results described here
are novel and relevant because autonomic regulation therapy is a growing field and as it starts to
get used clinically, it will first be used as an add-on to this more traditional drug therapy.
Therefore, these conclusions support the hypothesis that VNS is effective and safe when
administered with these drugs.
Illustrations
Figure 1. Heart functions are controlled through a complex, hierarchical nervous system – CNS
neurons, peripheral extracardiac, and intrinsic cardiac neurons (Buckley et al., 2015). Vagus nerve
stimulation (VNS) target area shown. Sympath sympathetic, Parasym parasympathetic, LCN local circuit
neuron, DRG dorsal root ganglia, Aff. afferent, T1-T4 first to fourth level of thoracic cord, Ang
angiotensin, β beta adrenergic receptor, M muscarinic receptor, Gs and Gi G proteins, AC adenylate
cyclase, ATP adenosine triphosphate, cAMP cyclic adenosine monophosphate, Neurite sensory endings
embedded in the myocardium, Decent decentralization. VNS alters neural activity at multiple levels of
the cardiac nervous system, both central and peripheral. It has the net effect to make the heart more
electrically stable and to preserve heart function.

13. 13
Figure 2. Schematic summarizing proposed neural interactions within hierarchy for
cardiac control engaged by cervical vagus stimulation (VNS) (Buckley et al., 2015)(Ardell et
al., 2015). This figure demonstrates how three common drugs prescribed for heart failure,
metoprolol, ivabradine, and enalapril, act on the cardic neuronal hierarchy. Dashed lines indicate
preganglionic projections. Dot-dash lines indicate afferent projections. NTS – Nucleus tractus
solitaries; NA – Nucleus Ambiguus; BSRF – brainstem reticular formation; IML –
intermediolateral cell column, DRG – dorsal root ganglia; MCG – middle cervical ganglia; LCN
– local circuit neuron; β – beta adrenergic receptor; M2 – muscarinic receptor; Gs and Gi – g-
coupled proteins; AC – adenylate cyclase.
IvabradineMetoprolol
Circulating
Angiotensin II
Norepi/Epi
IKf
Enalapril

14. 14
Figure 3. Effects of low-intensity, continuously-cyclic, open-loop VNS on basal cardiac electrical
function as assessed by 24 hour holter monitor. Normal canines (n = 8) were implanted with VNS
devices on the right and left cervical vagus nerves and programed to deliver chronic, intermittent
stimulation (14 s ON, 66 s OFF). Holter monitors recorded ECG continuously for 24 hours during various
combinations of stimulation frequency (5, 10, and 20 Hz) and pulse widths (250 and 500 µs). VNS
amplitude (mA) was programmed to whatever value obtained a 10% acute decrease in heart rate; it
averaged 1.75 mAmp. Results are shown as the mean ± SE heart rate from eight canines. (#, *, +,
p<0.025, as explained below the right panel of the figure). Hz, hertz; PW, pulse width.

15. 15
Figure 4. Comparisons of cardiac function (heart rate responses) between VNS-only controls and
combinations of drugs + VNS. Four normal canines were implanted with VNS therapy systems on the
right cervical vagus (RCV) nerve and placed on medical therapy, either individually or in combination.
These drugs were given twice a day orally for successive 2 week phases. In awake animals, heart rate
responses to repeating cycles of 10 Hz RCV VNS (14-s ON-time, 48-s OFF-time) were measured at the
end of each 2-week therapy phase and compared to baseline. Results are shown as the mean ± SE heart
rate as a percent change from the baseline. Control animals that received VNS alone are shown in filled
circle and black line in each panel. Results from animals treated with VNS + a drug are shown in empty
circles, with each drug or drug combination represented with a different color line. Results show that
VNS is still just as effective in controlling cardiac function with drug therapy. Thus, commonly used
heart failure medications do not affect the cardiac responses to VNS.

16. 16
Figure 5. Effects of VNS in combination with various commonly prescribed heart failure drugs,
given singly or in combinations, on heart rate variability (HRV) over 24 to 48 hours. HRV was
measured through the parameter pNN50 (%). Eight canines (n=8) were implanted with one bipolar
electrode on each of the right and left cervical vagus nerves. These electrodes were connected to separate
implanted VNS stimulators (Demipulse Model 103 Stimulator). This stimulation system was programmed
to deliver chronic, intermittent electrical stimulation of the vagal nerves. For these animals, VNS was
delivered at 10 Hz, 250 μs pulse width, stimulus intensity of ~2.00 mA and with a 25% duty cycle (14 sec
on and 48 sec off). Meanwhile, holter monitors were set to record EKG continuously for 24-48 hours,
with and without relevant drug treatments. Results are shown as mean ± SE pNN50 (%). Enal, enalapril;
Ivab, ivabradine; Metop, metoprolol; VNS, vagal nerve stimulation
61.1
59.8 59.2
56.7
62.2
58.7 58.1
52.0
54.0
56.0
58.0
60.0
62.0
64.0
66.0
pNN50(%) Heart Rate Variability (pNN50 (%))

17. 17
Figure 6. Effects of VNS in combination with various commonly prescribed heart failure drugs,
given singly or in combinations, on heart rate (HR) over 24 to 48 hours. Eight canines (n=8) were
implanted with one bipolar electrode on each of the right and left cervical vagus nerves. These electrodes
were connected to separate implanted VNS stimulators (Demipulse Model 103 Stimulator). This
stimulation system was programmed to deliver chronic, intermittent electrical stimulation of the vagal
nerves. For these animals, VNS was delivered at 10 Hz, 250 μs pulse width, stimulus intensity of ~2.00
mA and with a 25% duty cycle (14 sec on and 48 sec off). Meanwhile, holter monitors were set to record
EKG continuously for 24-48 hours, with and without relevant drug treatments. Results are shown as mean
± SE HR. Enal, enalapril; Ivab, ivabradine; Metop, metoprolol; VNS, vagal nerve stimulation.
83.9
85.2
84.2
89.3
83.3 83.4
84.4
76.0
78.0
80.0
82.0
84.0
86.0
88.0
90.0
92.0
94.0
96.0
MeanHR(beats/minute)
Mean Heart Rate

18. 18
Conclusion and Future Work
This study for the first time, to the best of the student’s and his mentor’s knowledge,
demonstrates that commonly prescribed drugs for the treatment of heart failure only minimally
affect the normal cardiac responses to VNS therapy. Furthermore, it shows that the optimal heart
rate response to VNS is dependent on the frequency at which the nerves are stimulated. Overall,
lower frequencies (5 Hz and 10 Hz) had a more positive cardiac effect, while in higher
frequencies (20 Hz), this effect was attenuated. Finally, although overall the drugs tested did not
make VNS ineffective, there were individual differences between the different drugs based on
their respective effects on the cardiac nervous system.
Since the study was conducted in normal canines, the conclusions made are relevant to
normal physiological conditions. A natural progression for future work would be to conduct a
preclinical trial of these drugs with VNS in animal models of heart failure. It is unclear whether
certain developments in disease states will allow for the same results as with normal states. It
would also be important to investigate the effect of different doses of various drugs on VNS-
induced effects on cardiac function. Finally, the results in animal model must be translated onto
humans through clinical trials.

19. 19
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