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EFFECTS OF IVABRADINE, A NEW SELECTIVE If CURRENT INHIBITOR, ON HEART
RATE IN CATS
Master’s Thesis
Present in Partial Fulfillment of the Requirements for the Degree Master of Science in the
Graduate School of The Ohio State University
By
Richard E. Cober, DVM
Graduate Program in Veterinary Clinical Sciences
The Ohio State University
2010
Thesis Committee:
Karsten Schober, Advisor
John Bonagura
Robert Hamlin
Copyright by
Richard E. Cober
2010
ABSTRACT
Heart rate is one of the main determinants of ischemia, a pathophysiologic
characteristic of hypertrophic cardiomyopathy (HCM). Unwanted tachycardia may trigger
decompensation in previously asymptomatic cats with HCM. Beta blockers are the primary
medications used to reduce heart rate in cats, however, side effects or contraindications
sometimes limit their use. Ivabradine is a highly selective If current inhibitor. The drug
exerts negative chronotropic effects without significant effects on inotropy, lusitropy, or
dromotropy in multiple species. Ivabradine, has never been studied in cats. The purpose
of this study was to determine an effective oral dose of ivabradine to significantly reduce
heart rate in healthy cats.
In this single blinded, placebo controlled, randomized, fully-crossed study 8
healthy cats received placebo or one dose of ivabradine at 0.1 mg/kg, 0.3 mg/kg, or 0.5
mg/kg, PO. HR and blood pressure were monitored continuously for 24 hours via
radiotelemetry after each treatment. Response to stress was studied twice by 15-min
acoustic startle applied at baseline and 4 hours after drug. Statistical comparisons were
made using a linear mixed model and 2-way repeated measures ANOVA.
Peak negative chronotropic effect was observed 3 hours after ivabradine
administration. Heart rate (min-1) decreased significantly (p<0.05) in a dose-dependent
ii
manner (mean±SD for placebo: 144±20; ivabradine 0.1 mg/kg: 133±22; ivabradine 0.3
mg/kg: 112±20; ivabradine 0.5 mg/kg, 104±11). Heart rate (min-1) was still reduced
(p<0.05) at 12 hours after ivabradine (0.3 mg/kg: 128±18 and 0.5 mg/kg: 124±16)
compared to placebo (141±21). Heart rate response to acoustic startle was significantly
(p<0.01) blunted at all 3 doses of ivabradine. No effect of ivabradine on blood pressure
was identified and no clinically discernable side effects were observed.
These findings indicate that ivabradine at 0.3 mg/kg and 0.5 mg/kg PO predictably
lowers HR for at least 12 hours in healthy cats. Clinical studies in cats with HCM are
needed.
iii
VITA
August 24, 1979 . . . . . . . . . . . . . . . . . . Born – Annapolis, Maryland
2001 . . . . . . . . . . . . . . . . . . . . . . . . . . . B.S. Animal Science, Cornell University
2006 . . . . . . . . . . . . . . . . . . . . . . . . . . . DVM Veterinary Medicine, Kansas State University
2006-2007 . . . . . . . . . . . . . . . . . . . . . . Intern, Medicine and Surgery, Cornell University
Hospital for Animals, Cornell University
2007- present . . . . . . . . . . . . . . . . . . . . Resident, Cardiology, The Ohio State University
Veterinary Teaching Hospital, The Ohio State
University
Fields of Study
Major Field: Veterinary Clinical Sciences
iv
TABLE OF CONTENTS
Page
Abstract. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ii
Vita. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .iv
List of Tables. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vi
List of Figures. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vii
Chapters:
1.
Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
2.
Materials and Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .11
3.
Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .17
4.
Discussion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37
List of References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .46
v
LIST OF TABLES
Table
Page
1. Mean hourly heart rates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
2. P- values for mean hourly heart rates at hours 1, 2, 3, 4, 8, and 12 . . . . . . . . . . . . . . 22
3. Twenty two hour average heart rates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
4. Average 15 minute heart rates before and during Startle . . . . . . . . . . . . . . . . . . . . . . .23
5. Mean hourly systolic blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
6. Mean hourly diastolic blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
7. Mean hourly mean blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
8. Mean hourly rate-pressure product . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
9. Mean twenty two hour rate-pressure product . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
10. Mean twelve hour rate-pressure product. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27
vi
LIST OF FIGURES
Figure
Page
1. Mean hourly heart rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
2. Mean twenty two hour heart rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .29
3. Mean 15 minute heart rate before and during startle, prior to and after drug
administration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
4. Mean 15 minute heart rate during startle before and after drug administration . . . . . . 31
5. Mean hourly systolic blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32
6. Mean hourly diastolic blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
7. Mean hourly mean blood pressure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
8. Mean twenty two hour rate-pressure product . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
9. Mean twelve hour rate-pressure product. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36
vii
CHAPTER 1
INTRODUCTION
Hypertrophic cardiomyopathy (HCM) is the most common cardiac disease in cats and is
associated with significant mortality1-6. It is often detected during the asymptomatic stage of the
disease based on physical exam findings such as a cardiac murmur, a gallop sound, or an
arrhythmia1. The diagnosis is confirmed with transthoracic two-dimensional echocardiography
(2DE). The development of HCM in cats is most likely due to a genetic mutation of one or more of
the sarcomeric proteins that results in symmetric or regional concentric hypertrophy of the left
ventricle (LV) and/or papillary muscles2-4, 6. Additional histomorphologic hallmarks of the disease
include myofiber disarray leading to mechanical dysfunction and narrowing of intramural coronary
arteries leading to decreased myocardial perfusion, myocardial ischemia, necrosis, and
replacement fibrosis5, 7. Left ventricular hypertrophy may also result in a reduction of the LV
chamber size, slowed and incomplete ventricular relaxation, and increased wall stiffness all of
which impede diastolic filling. As a consequence, LV filling pressures (LVFP) rise leading to left
atrial enlargement, neurohormonal activation, arrhythmias, congestion, blood stasis, heart failure,
and sudden death. One half to two thirds of cats with HCM also develop dynamic obstruction of
the LV outflow tract from hypertrophy of the IVS and systolic anterior motion (SAM) of the mitral
1
valve. The latter further contributes to increased LV systolic pressure, a known trigger for more
hypertrophy, which may be associated with increased myocardial oxygen demand, worsening of
LV concentric hypertrophy, diastolic dysfunction, and left atrial enlargement.
Evidence of myocardial ischemia in cats with HCM has rarely been reported owing to the
difficulty of diagnosing the abnormality. ST segment elevation or depression on the ECG
suggestive of myocardial ischemia has diagnostic specificity but lacks sensitivity. In recent years,
serum cardiac troponin I (cTnI) has been used as a surrogate of myocardial ischemia and
subsequent cardiomyocyte injury in people8 as well as in cats9. Circulating cTnI is not detectable
or occurs only in traces in healthy cats, but is elevated in most cats with asymptomatic HCM, and is
even more elevated in cats with decompensated HCM. From 31 cats with asymptomatic HCM only
5 had ST segment abnormalities on the ECG but 28 had detectable serum cTnI (Schober et al.
recently unpublished data). Based on currently available data we believe that myocardial ischemia
is an essential abnormality in HCM in cats.
The progression of HCM is variable with most cats having a long asymptomatic period that
may end with the abrupt development of severe congestive heart failure (CHF), arterial
thromboembolism (ATE), or sudden cardiac death. Large scale studies involving more than 400
cats with HCM revealed that most cats with HCM will eventually die from their cardiac disease10-12.
Several risk factors have been identified as negative prognostic indicators including LV diastolic
function, left atrial size, and heart rate10, 13. Tachycardia is a major contributor to increased
myocardial oxygen consumption. High heart rates decrease myocardial perfusion by limiting
diastolic coronary blood flow, induce myocardial ischemia causing a further decrease in diastolic
chamber filling, and exacerbate LV outflow tract obstruction. Although the association between
progression of HCM and heart rate has not yet been fully elucidated, several studies in cats have
shown that advanced disease is accompanied by elevated heart rates. In a recent retrospective
2
study completed at our institution, 43 healthy cats, 49 cats with asymptomatic HCM, and 49 cats
with symptomatic HCM had mean heart rates taken during physical examination of 175 bpm, 191
bpm, and 201 bpm, respectively (P < 0.05; Schober et al, unpublished data). A study done with 56
cats with HCM found heart rate to be a prognostic indicator with a medium survival time after
diagnosis of 1,830 days if heart rate was < 200 bpm and a median survival time of 152 days if heart
rate was ≥ 200 bpm10. In another study with 27 normal cats and 29 cats with cardiomyopathy and
CHF, significant elevation of heart rate in the latter group (mean 182 bpm versus 217 bpm, P <
0.05) was reported14. While the association between heart rate and outcome or disease severity
has not yet, by itself, been proven to be causal in cats with HCM, human studies using logistic
regression analysis found heart rate to be an independent variable of outcome15, 16.
Although the pathogenesis of acute decompensation of otherwise clinically stable cats with
HCM is not fully understood, anecdotal evidence exists that stressful events leading to tachycardia
trigger cats with well compensated HCM to suddenly develop CHF11, 17. These events can include
but are not limited to routine examinations at a veterinary hospital, sudden changes in the cats
home environment, pain of any source, anesthesia, or elective surgery can. In people, physical or
emotional stress may lead to unwanted tachyarrhythmias that are known triggers for acute
decompensation of recently stable ischemic heart disease16. Also, heart rate not only is important
in the development of ischemia, but also influences whether ischemic episodes trigger serious
arrhthmias16. Therefore, pharmacologic modulation of heart rate aimed at lowering resting heart
rate and preventing sudden periods of uncontrolled tachycardia may reduce the risk of acute
decompensation of HCM, life-threatening arrhythmias during ischemic periods, and sudden cardiac
death.
3
In the past, treatment of cats with HCM has been directed at relieving obstruction of the LV
outflow tract, management of CHF, arrhythmia control, as well as the prevention and management
of ATE. The most common medications used include beta adrenergic blockers, calcium channel
antagonists, angiotensin converting enzyme inhibitors, venodilators, diuretics, and antiplatelet
drugs. Beta blockers (e.g., atenolol) and calcium channel blockers (e.g., diltiazem) are the most
frequently used agents in feline HCM. The rationale for their use is negative chronotropy and
inotropy leading to improvement in coronary perfusion, LV relaxation, and LV filling, as well as
reduction of LV outflow tract obstruction, and antiarrhythmic properties potentially lowering the risk
of sudden cardiac death. However, adverse effects induced by such drugs have been reported
including weakness, lethargy, salivation, weight loss, and reduced left atrial function18, 19. Beta
blockers may also be contraindicated in certain conditions such as CHF, allergic airway disease,
hypotension, and ATE. As a result, new medications have been developed to target specific ion
channels in the sinoatrial (SA) node to cause pure heart rate reduction in the absence of effects on
myocardial performance.
The SA node is located in the right atrium at the junction of the entry of the cranial and
caudal vena cava with the crista terminalis. The node is separated from the atrial myocytes by a
connective tissue barrier, primarily collagen and fibroblasts, with species-dependent existence
varying from as little as 50% in the rabbit to as much as 75-90% in the cat20, 21. The SA node
displays much heterogeneity from the center to the periphery. In the human, dog, and rabbit, the
center of the SA node contains characteristic nodal cells that are smaller, contain few
myofilaments, are poorly organized (interweaving), with fewer and smaller gap junctions compared
to atrial myocytes that are connexin 45 positive causing a slow conduction velocity ( 2-8 cm/s)20.
Nodal cells towards the periphery are at times termed “transitional cells” and begin to develop
characteristics similar to atrial myocytes21. These are larger, more organized, contain more
4
myofilaments (mitochondria and sarcoplasmic reticulum), and have larger and more functional gap
junctions made up of both connexin 45 and 43 (connexin 43 is present primarily in atrial and
ventricular myocardium) causing increased conduction velocities (36-50 cm/s)20. The leading
pacemaker site is located in the center of the node with conduction developing towards the
periphery and arriving at the crista terminalis in a broad wavefront21. There is also a block zone
located near the interatrial septum that blocks all conduction from the pacemaker site in that
direction. This block zone is thought to protect the SA node from reentry and invasion of action
potentials from the atrial myocardium21.
The action potential within the SA node of the rabbit and cat, like the nodal cells, is not
homogeneous. In the center, the action potential upstroke velocity is slow (<10 V/s), action
potential duration long (~150 ms), with a low overshoot (<10 mV), and low maximum diastolic
potential (-60 to -70 mV)21. From the center to the periphery towards the atrial myocytes the
upstroke velocity and overshoot increase, the maximum diastolic potential becomes more negative,
the action potential duration decreases, and spontaneous rate increases21. Within the SA node
there are also regional differences in the action potential in the superior and inferior directions.
Form superior to inferior, there is a decrease in upstroke velocity, increase in action potential
duration, and decrease in spontaneous depolarization rate21.
Not only does there appear to be a difference in the action potential from the periphery to
the center of the SA node, but there is also a difference in the intrinsic pacemaker activity20. In the
peripheral nodal cells, the pacemaker activity is greater than in the center. The mechanism behind
this phenomenon appears to be that the atrial myocardium exerts an electrotonic influence on the
peripheral cells. The peripheral cells as apposed to the central cells are connected to the atrial
myocytes through well developed gap junctions, and through these connections the depolarization
of the peripheral cells will be reduced by the more negative potentials and hyperpolarization
5
currents of the atrial cells21, 21. The central nodal cells, however, are less influenced by electrotonic
impedance due to the fact that there is no direct connection to the atrial myocardium, lesser
number and development of gap junctions. Therefore, the pacemaker activity in the right atrium is
governed by the pacemaker cells in the center of the SA node.
The SA node is the primary site of spontaneous pacemaker activity that produces the
electrical impulse causing cardiac automaticity. This electrical impulse is generated during phase
IV of the action potential and is termed diastolic depolarization. Diastolic depolarization occurs
when the negative membrane voltage at the end of repolarization is driven to a less negative
threshold potential that causes rapid depolarization and initiates the nodal action potential.
Diastolic depolarization is accomplished by the interplay of several ionic channels. The major ion
channels that appear to contribute to diastolic depolarization are the delayed rectifier potassium
current (IK,v), inward calcium channels (ICa-L and ICa-T)), and the inward “funny” current (If) 22.
The delayed rectifier potassium channel is a voltage and time dependent ion channel that
is activated by depolarization and contributes to phase III repolarization by causing an efflux of
potassium ions and an outward current that resets the resting membrane potential. It is thought
that this ion channel activity is time dependent and decays to allow a less negative membrane
potential to occur as well as enabling the inward currents to dominant and cause depolarization22.
In the rabbit, two types of voltage dependent calcium currents are involved in diastolic
depolarization, the long-lasting current (ICa-L) and the transient current (ICa-T)23. The long-lasting
calcium current classically has been thought to have a less negative threshold of activation (-40
mV), contributes to the fast depolarization involved in the action potential, and is considered
important for the effect of the sympathetic nervous system on heart rate control. The transient
current is activated at more negative thresholds (-60 to -50 mV) and is the major contributor to late
diastolic depolarization without being effected by the sympathetic nervous system or calcium
6
channel blockers23. More recently, different subtypes of the L-type calcium channel pore forming
alpha 1 subunit have been cloned with Cav 1.2 and 1.3 being expressed in the cardiovascular
system in mice23. The Cav 1.2 subunit was found to be associated with less negative threshold
potentials (- 40 mV), is activated at the action potential threshold and inactivated during
repolarization, indicating that it is primarily associated with the fast phase of depolarization with
little impact on diastolic depolarization but contributes mostly to myocardial contraction23. The Cav
1.3 subunit, however, was found to have more negative activation threshold potentials (- 50 to -55
mV) and contributed to both early and late diastolic depolarization, indicating that ICa-L, does
contribute to the ionic channels associated with diastolic pacemaking with Cav 1.3 being the major
contributor23. The T-type calcium channel α1 subunit has also been cloned with 3 distinct isoforms
discovered. The Cav 3.1 isoform appears to be the predominant determinant of Ica-T and
contributes to late diastolic depolarization23.
The inward “funny” current is also a voltage gated channel that becomes activated at
hyperpolarized potentials as opposed to typical depolarization activated voltage gated channels24.
Activation of this current is facilitated directly by cAMP, independent of protein kinase A
phosphorylation25. As a result, If is also referred to as the hyperpolarization-activated cyclic
nucleotide gated (HCN) channel. HCN channels are predominantly located in cardiac tissue and
neurons giving rise to the formation and regulation of pacemaker activity in the SAN, as well as the
rhythmic activity, resting membrane potential, dendritic integration, and synaptic transmission in
neuronal circuits25. Four HCN channel subunits exist (HCN 1-4), all have been detected in cardiac
tissue but HCN 2 and 4 appear to be the most abundant25. Within the SA node and conduction
system in all species studied (rabbit, guinea pig, mouse, and dog), HCN 4 is the dominant isoform
accounting for >80% of isoforms, while HCN 1 and 2 have been detected in the SA node and HCN
3 in the conduction system25. However, HCN2 predominates in the atrial and ventricular myocytes
7
in healthy myocardium. Within the family of HCN subunits the voltage dependent activation differs.
HCN 2 and 4 have midpoints of activation of – 95 mV and – 100 mV, while the midpoint of
activation for HCN 1 and 3 are -70 and -77 to – 95 mV25.
The HCN channel has a similar structure to other voltage gated pore forming channels,
with four transmembrane pore forming subunits with a cytosolic C-terminal domain. Each
transmembrane domain is composed of six alpha-helix segments. Within the six helices, the fourth
helix is positively charged and acts as the voltage sensor, while the pore forming properties of HCN
are in an ion-conduction loop between the 5th and 6th helices25. It is the ion-conducting loops that
determine the ionic permeability and conduction of HCN channels for sodium, potassium and
calcium, with the permeability ratio of 1:4 for sodium to potassium and a fractional calcium current
of only approximately 0.5 %25. The cytoplasmic C-terminal domain is composed of a 120 amino
acid cyclic nucleotide-binding domain (CNBD) and an 80 amino acid C-linker region. The CNBD is
the binding site of cAMP, which causes a shift towards more positive voltage dependent activation
(+10-25 mV) and increases channel opening25. The C-linker region connects the CNBD to the 6th
helix segment of the pore forming transmembrane domains.
The If pacemaker function occurs at end repolarization when the maximal diastolic
potential is most negative, causing activation of the If current, diastolic depolarization, and a less
negative membrane potential. When the membrane potential reaches the activation threshold of
the L-type calcium channels, a nodal action potential occurs26. The funny current has also been
implicated in modifying the heart rate response to the autonomic nervous system22, 27. When the
sympathetic nervous system’s neurotransmitter, norepinephrine, activates the beta adrenergic
receptor, the enzyme adenylyl cyclase becomes stimulated causing conversion of ATP to cyclicadenosine-monophosphate (cAMP). Cyclic AMP binds directly to If causing an increase in inward
current, less negative If activation potential, an increase in the slope of early diastolic
8
depolarization, and an increase in heart rate. Parasympathetic stimulation of the muscarinic
receptor by acetylcholine leads to a decreased formation of cAMP, causing a hyperpolarizing shift
in the If curve, inactivation of If current, a decrease in the slope of early diastolic depolarization, and
a decrease in heart rate. Blockade of the If current also causes a decrease in the slope of diastolic
depolarization and spontaneous heart rate.
Ivabradine is a highly selective If current inhibitor. It acts directly on the sinoatrial node to
induce a rapid, sustained, use- and dose-dependent reduction of heart rate by reducing the slope
of slow diastolic depolarization of cardiac pacemaker cells. This occurs without significant effects
on inotropy, lusitropy, or dromotropy in the rat, rabbit, and guinea pig28. The sole heart rate
reducing effects of ivabradine appear to be due to its high binding affinity for HCN4 within the SA
node, with minimal effects on the other ionic channels involved with spontaneous pacemaking,
where inhibition of the If channel at hyperpolarized states leads to a decrease in the heart rate
because of a reduction in the slope of diastolic depolarization29. The effects of ivabradine appear
to be localized to the SA node with minimal effects on the atrial or ventricular myocardium as well
as the remainder of the conduction system. These effects have been shown in electrophysiological
studies, where intravenous ivabradine was associated with a reduction in heart rate, an increase in
sinus node recovery time, and without a significant effect on the AV node, His-Purkinje system,
atrial or ventricular refractoriness30. Ivabradine’s heart rate reducing effects are also influenced by
the resting heart rate, with a greater magnitude of heart rate reduction occurring at higher resting
heart rates due to binding of ivabradine to HCN channel being restricted to open channel states29.
While HCN channels are abundant outside of cardiac tissue, there appears to be minimal
effect of ivabradine on other organ systems. Ivabradine does not cross the blood brain barrier,
therefore there are minimal to no effects on the central nervous system29. The most common side
effects are related to inhibition of HCN channels in the eye where HCN1 and HCN2 are responsible
9
for the Ih current in the retina which is involved in the response to light stimulus. Ivabradine,
appears to have no effect on retinal function nor does it cause retinal degeneration. However, it
does appear to result in luminous phenomena in people at therapeutic doses28, 31-33. Symptomatic
bradycardia appears to be an uncommon side effect of ivabradine with approximately 0.2% of
human patients compared with 0.4% treated with atenolol29l.
Ivabradine is currently the only If current inhibitor approved for the treatment of stable
ischemic heart disease in patients with normal sinus rhythm (marketed only in Europe). Ivabradine
has been shown to increase stroke volume and preserve cardiac output, increase coronary blood
flow, increase myocardial oxygen supply, and decrease oxygen consumption in failing myocardium
in people and experimental animals19, 31, 33-37. Ivabradine has also been shown in people to be
non-inferior to atenolol, diltiazem, and amlodipine for anti-anginal and anti-ischemic effects33. In a
large scale clinical trial in involving 10,917 patients with stable coronary artery disease, LV
dysfunction, and angina, treatment with ivabradine resulted in a reduction of primary endpoints of
cardiovascular mortality or hospitalization in patients with a resting heart rate greater than 70 min-1
32.
Due to Ivabradine’s unique selective heart rate-reducing properties and the lack of direct
negative inotropic or lusitropic effects, it may become a new therapeutic option for use in cats with
HCM. It should prolong diastolic filling time, decrease myocardial oxygen consumption, reduce
myocardial ischemia and its chronic consequence, fibrosis, relieve LV outflow tract obstruction, and
reduce tachycardia-induced crisis events. To the author’s knowledge, prospective studies on the
use of ivabradine in cats have not yet been published.
10
CHAPTER 2
MATERIAL AND METHODS
Animals – Eight domestic short hair cats, acquired from an established in house colony of healthy
cats (n=3) and cats with asymptomatic feline interstitial cystitis (FIC; n=5), were used for this study.
All cats were housed in individual stainless steel cages (69.9 X 78.1 X 74.9 cm). There were 5
spayed females and 3 neutered males cats. Ages ranged from 4 to 9 years (mean ± SD, 6.3 ± 1.7
years). Body weight ranged from 3 to 6.3 kg (mean ± SD of 5.1 ± 1.3 kg). All cats had previously
(2006) been instrumented with telemetric transmitter devices (PhysioTel D70PCTP, Data Science
International, St. Paul, MN) capable of measuring invasive blood pressure (systolic, diastolic, and
mean) and heart rate calculated from a continuous electrocardiogram (ECG) using Dataquest ART
3.1 software (Data Science International, St. Paul, MN). The ECG was stored digitally for later
review. Prior to enrollment, health status was determined in all cats based on a thorough physical
examination, resting ECG, 2DE, M-Mode and Doppler echocardiography, systolic blood pressure
measurement, and blood biochemical analyses including total T4 concentration in cats 6 years of
age or older. Exclusion criteria included the presence of a gallop sound or an arrhythmia, cardiac
disease based on established echocardiographic criteria38, blood biochemical abnormalities
11
suggestive of hyperthyroidism, renal disease or any other systemic disorder, and repeated blood
pressure measurements above 175 mmHg39. More specifically, echocardiographic exclusion
criteria were a maximum left atrial dimension > 16 mm measured from the right parasternal long
axis view, and/or LV wall thickness > 6mm measured at end diastole from the right parasternal long
axis view or short axis view at the level of the papillary muscles. Interstitial cystitis was judged to
be controlled based on review of daily clinical observation sheets indicating lack of pollakiuria,
dysuria, hematuria, inappetance or vomiting within the previous seven days. All screening
examinations were performed by the principal investigator (REC). The study protocol was
reviewed and approved by the Animal Care and Use Committee of The Ohio State University, and
all cats were treated in compliance with NIH Guidelines for the Care and Use of Laboratory
Animals.
Study Design – The study represented a repeated, multiple-crossover, randomized, singleblinded, placebo-controlled study design. The cats were randomized to receive either placebo or a
single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg). Randomization was
accomplished using a randomization table. All treatments were administered orally with a wash out
period of at least 48 hours. All cats were housed individually and remained in their respective
cages during the entire 24 hour data acquisition period with the exception of two stress periods
induced by acoustic startle. Telemetric devices were set to continuously record blood pressure
and heart rate at a sampling rate of two per minute. Baseline measurements were obtained over a
time period of 30 minutes with the cats resting quietly in their respective cages. Thereafter, the
cats were moved to a separate room were an acoustic startle period was performed for 15 minutes
to induce a stress response associated with an increase in heart rate. The cats were then moved
back to their cages and received the oral treatment 30 minutes after the baseline period was
completed and approximately 15-20 minutes after startle. Medications were given orally, hidden in
12
a “pill pocket,” and cats were monitored for five minutes afterward to assure swallowing of the drug.
After drug administration, the cats were left in their cages for approximately four hours. Thereafter,
they were moved to have another 15 minute period of acoustic startle performed. After the second
startle period, the cats were placed back in their cages for the remainder of the 24 hour testing
period. Due to limited availability of monitoring software only one cat per day was studied.
Acoustic Startle – Heart rate responses to acoustic startle were accomplished using a custom
built wooden chamber platform (35X70X2.5 cm) to which an open wire cage (20 X29X26cm) was
securely attached. The platform was secured with bolts to a plywood base at the corners with
heavy compression springs in between. The bolts were tightened to apply pressure to the corners
of the platform and base, leading to a highly damped connection.
Testing was performed in a quiet room with an ambient noise level of 60-65 dB. Labtec
speakers (Labtec, Vancouver, WA – range 70 Hz-20 kHz) were positioned approximately 12 cm
form each end of the cage. The speakers were connected to a 250 watt amplifier (Tandy Corp.,
Fort Worth, TX) and delivered a computer generated white noise acoustic pulse of varying intensity
(80 – 120 dB), with a 2.5 ms rise and fall time. The sound level inside the chamber was calibrated
before each session using a Radioshack digital sound level meter (Radioshack Corp., Fort Worth
TX).
For testing, cats were removed from their home cages placed into a small carrier
(53X30X36 cm), carried to the testing room and placed into the box on the platform. The transfer
was completed within five minutes of the subject leaving its home cage. The cats were given a five
minute acclimation period once positioned in the startle cage. A 118 dB pulse was delivered
randomly twice per minute for 15 minutes. At the end of the startle period, the cats were placed
back into the carrier and returned to their home cage.
13
For final data analysis, the two startle periods were divided into four periods: 1) baseline
before startle (Period 1), 2) baseline startle (Period 2), 3) treatment before startle (Period 3), and 4)
treatment startle (Period 4). Period 1 was defined as the 15 minute time period immediately prior
to moving the cats to the startle location at baseline. Period 2 was considered the 15 minute time
period during startle. Period 3 was the 15 minute time period immediately prior to the startle period
at four hours after drug administration, and Period 4 was the startle period four hours after drug
administration.
Heart Rate Analysis – Telemetric devices were set using the Dataquest ART 3.1 software to
average, record, and store heart rates in a Microsoft Office Excel (Microsoft Corporation,
Redmond, WA) worksheet. Heart rates were recorded every 30 seconds for the entire study period
of approximately 24 hours. Heart rate periods were then averaged into hourly heart rates, with the
exception of the first 30 minute baseline period and the 15 minute pre-startle and startle periods.
The baseline acquisition period was averaged over the first 30 minute period prior to startle so as
to acquire a true resting baseline heart rate. Heart rates during each startle period (Periods 2 and
4) were averaged for the fifteen minute time period during which startle was performed. The heart
rates for the fifteen minute pre-startle periods (Periods 1 and 3) were also averaged to be used as
a separate baseline for comparison to startle. The hourly heart rates were then averaged to obtain
a 24 hour average heart rate Periods 3 and 4.
Blood Pressure Analysis – Telemetric devices were set to record, store, and average invasive
systolic, diastolic, and mean arterial blood pressures and transfer data to a Microsoft Office Excel
worksheet every 30 seconds. As with heart rate, blood pressure was recorded every 30 seconds
for the entire study period. Blood pressure periods were then averaged into hourly blood pressures
including Periods 3 and 4 after drug administration.
14
ECG Analysis – All recordings were stored digitally for the entire study period for later analysis.
The principal investigator reviewed all ECG complexes of all studies (n= 32) to identify the total
number and type of ectopic complexes present during each treatment given to the cats.
Clinical Findings – Daily observation sheets were generated for each cat during each study
period. Quantitative and qualitative clinical variables evaluated were food and water intake, bowel
movements (including character and number), and urinations (including character and number). All
cats were also evaluated for vomiting, ocular or nasal discharge, shedding, grooming, coat quality,
vocalizations, weakness or signs of other medical or physical problems such as hypersalivation,
oral or skin lesions, itching, pain, or increased respiratory rate and effort.
Rate-Pressure Product - As a measure of myocardial oxygen consumption, the rate-pressure
product was calculated from the mean hourly heart rate and the mean hourly systolic blood
pressure40:
Rate-Pressure Product (RPP) = heart rate (HR) X systolic blood pressure (SBP)
Statistical Analysis – Statistical analyses were performed with commercially available software
(foot note). Descriptive statistics were calculated for all variables measured. Data were evaluated
for normality with the D’Agostino and Pearson test and are reported as mean±SD when normally
distributed or as median (range) when not normally distributed. Linear mixed-effects models were
used to analyze heart rate and blood pressure (systolic, diastolic, and mean) as the dependent
variables. The models included fixed effects of animal, treatment dose (0.1, 0.3, and 0.5 mg/kg),
treatment time (1h, 2h, 3h, 4h, 8h, and 12h), the interaction of dose and time, order of drug
administration, and the baseline measures as a covariate. A 2-way repeated measures ANOVA
and a Holm-Sidak post hoc test were used to evaluate differences between the treatments and prestartle and startle periods, and to determine if there were interactions between treatment and
startle period. Differences among treatment groups with regard to average 12 and 24 hour heart
15
rate, lowest hourly heart rate, and rate-pressure product were determined using a one-way
repeated measures ANOVA and a Tukey’s post hoc test. Values of P ≤ 0.05 were considered
significant for all analyses.
16
CHAPTER 3
RESULTS
No major adverse effects were seen during the study period. Three cats experienced a
total of seven days of a decreased appetite. Three of these days corresponded to placebo
administration while four days corresponded to administration of ivabradine. Of the four days of
decreased appetite associated with ivabradine, one occurred with the administration of 0.1 mg/kg,
two with 0.3 mg/kg, and one with 0.5 mg/kg. One cat had diarrhea and another cat vomited one
time. The diarrhea was seen in a cat on the day of administration of ivabradine at 0.3 mg/kg
whereas the vomiting occurred after placebo. A total of 14 single uniform VPCs were noted for all
eight cats at all doses administered. Three VPCs were associated with two cats given placebo,
nine with two cats given ivabradine 0.1 mg/kg, and two with one cat given 0.5 mg/kg. No other
arrhythmias were noted.
All cats had at least 22 hours of heart rate and blood pressure recorded for all doses with
the exception of 3 cats for a total of 6 doses (placebo, n=2; 0.1 mg/kg, n=1; 0.3 mg/kg, n=1; and
0.5 mg/kg, n=2). The lack of data recording was due to cats being able to move beyond the
maximal wireless connection between the telemetry transmitter and the recording plate of the data
17
acquisition system. All periods of lack of recording occurred sporadically between hours 18-24
corresponding with sleep activity.
Hourly Heart Rate Analysis
Among the study cats, administration of ivabradine induced a time-dependent negative
chronotropic effect compared to baseline and placebo that lasted for approximately 13 to 15 hours
with the lowest heart rates occurring approximately three hours after drug administration (Figure 1;
Tables 1 and 2). Heart rate was also affected in a dose-dependent manner, with a significantly
lower heart rate at higher dosages of ivabradine (Figure 1; Tables 1 and 2). Compared to placebo,
heart rate was significantly lower after ivabradine 0.1 mg/kg (at hours 2, 4, and 8), 0.3 mg/kg (at
hours 1, 2, 3, 4, 8, and 12), and 0.5 mg/kg (at hours 1, 2, 3, 4, 8, and 12; Tables 1 and 2).
Compared to ivabradine 0.1 mg/kg, heart rate was significantly lower after ivabradine 0.3 mg/kg
and 0.5 mg/kg at hours 2, 3, 4, 8 and 12 with the exception of dose 0.3 mg/kg at hour 12 (Tables 1
and 2). Ivabradine 0.5 mg/kg also significantly lowered heart rate compared to ivabradine 0.3
mg/kg at hours 3 and 4 (Tables 1 and 2).
Twenty Two Hour Average Heart Rate Analysis
Heart rates were decreased over the entire study period the heart rate with increasing
dose of ivabradine (Figure 2; Table 3). Average heart rate after ivabradine at 0.1 mg/kg (138 +/- 7
bpm), 0.3 mg/kg (132 +/- 10 bpm), and 0.5 mg/kg (130 +/- 13 bpm) were significantly (P<0.05)
decreased compared to placebo (148 +/- 7 bpm). Both ivabradine 0.3 mg/kg (132 +/- 10 bpm) and
0.5 mg/kg (130 +/- 13 bpm) significantly (P<0.05) decreased heart rate compared to ivabradine 0.1
mg/kg (138 +/- 7 bpm), but no significant (P>0.05) differences were found between heart rates after
ivabradine 0.3 mg/kg and 0.5 mg/kg.
18
Startle Analysis
No significant (P> 0.05) difference in heart rate was found at Period 1 between groups nor
was there a significant (P> 0.05) difference in heart rate Period 2 between groups (Figure 3, Table
4). In all groups, the fifteen minute baseline startle (Period 2) significantly elevated heart rate
compared to Period 1 (P<0.001) (Figure 3; Table 4). Heart rate decreased significantly for all
treatment groups between Period 2 and Period 3 (P<0.05) (Figure 3, Table 4). At Period 3, heart
rates after ivabradine 0.3 mg/kg and 0.5 mg/kg were significantly lower than heart rates after
placebo and ivabradine 0.1 mg/kg (P<0.001). However, there were no differences between
placebo and ivabradine 0.1 mg/kg and ivabradine 0.3 mg/kg and ivabradine 0.5 mg/kg (P=0.886
and P=0.683, respectively) (Figure 3; Table 1). Heart rates were significantly higher at Period 4
compared to the Period 3 for all treatments (P<0.001) (Figure 3, Table 4). However, heart rates at
Period 4 were significantly lower than during Period 2 for all treatments (P<0.01) (Figure 4; Table
4). Heart rates after placebo were not significantly different during Period 2 and Period 4
(P=0.941). During Period 4, heart rates were significantly lower after ivabradine 0.1 mg/kg, 0.3
mg/kg, and 0.5 mg/kg compared to placebo; and ivabradine 0.3 mg/kg and 0.5 mg/kg had
significantly lower heart rates than ivabradine 0.1 mg/kg (P<0.01 and P<0.01, respectively) (Figure
3; Table 1), whereas no difference between ivabradine 0.3 and 0.5 mg/k was found (P=0.171).
Blood Pressure and Rate-Pressure Product
Compared to placebo, no significant differences were found between systolic blood
pressure at hours 1, 2, 3, 4, 8, and 12 with the exception of dose 0.3 mg/kg at hour 12 (Figures 5;
Table 5). No significant differences were found between hourly diastolic blood pressure at hours
1,2, 3, 4, 8, and 12 with the exception of dose 0.3 mg/kg at hours 1 and 12 as well as dose 0.5
mg/kg at hour 12 compared to placebo (Figure 6; Table 6). No significant differences were found
between mean blood pressure at hours 1, 2, 3, 4, 8, and 12 with the exception of dose 0.3 mg/kg at
19
hour 12 compared to placebo (Figure 7; Table 7). Ivabradine 0.1 mg/kg (15148 +/- 1010), 0.3
mg/kg (14444 +/- 1603), and 0.5 mg/kg (14977 +/- 1932) significantly decreased the rate-pressure
product compared to placebo (P<0.0003) (16415 +/- 1132; Figure 8, Tables 7 and 8). There was
no significant difference in the rate-pressure product between ivabradine 0.1 mg/kg, 0.3 mg/kg, or
0.5 mg/kg (Figure 8, Tables 8 and 9).
20
TABLES
Time (h)
Baseline
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Placebo
147±21
156±27
146±19
144±20
157±24
167±29
156±30
157±29
157±26
148±20
146±16
146±25
141±21
143±18
145±17
143±17
149±25
149±25
142±21
139±16
141±16
141±22
142±19
0.1
153±8
152±24
133±16*
133±22
141±19*
139±18
133±20
139±18
144±16*
150±28
144±22
138±15
135±16
137±17
132±14
132±16
140±22
135±12
133±15
132 ±14
140 ±25
133±11
134±11
Ivabradine (mg/kg)
0.3
154±20
149±19*
119±18*
112±20*
120±13*
128±20
128±14
131±19
127±23*
135±11
124±11
125±15
128±18*
126±20
132±26
133±20
135±16
136±17
138±18
144±25
134±23
131±18
141±28
0.5
158±12
144±23*
109±15*
104±11*
116±14*
118±14
117±11
119±15
126±19*
130±26
129±22
128±19
124±16*
133±17
137±21
132±20
138±20
140±21
141±27
143±25
149±25
132±15
126±17
Table 1 – Mean +/- SD hourly heart rate (min-1) in eight cats after oral administration of placebo or
one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg). * indicates p < 0.05
compared to placebo. Data were compared statistically only at 1h, 2h, 3h, 4h, 8h, and 12h.
21
Ivabradine (mg/kg)
Hour
0.1
0.3
0.5
1
0.253‫٭‬
0.042‫٭‬
0.662†
0.049‫٭‬
0.312†
0.427‡
2
0.010‫٭‬
<0.001‫٭‬
0.009†
<0.001‫٭‬
0.002†
0.195‡
3
0.121‫٭‬
<0.001‫٭‬
0.009†
<0.001‫٭‬
0.003†
0.001‡
4
0.009‫٭‬
<0.001‫٭‬
0.001†
<0.001‫٭‬
0.002†
0.022‡
8
0.008‫٭‬
<0.001‫٭‬
0.022†
<0.001‫٭‬
0.004†
0.550‡
12
0.154‫٭‬
<0.001‫٭‬
0.335†
<0.001‫٭‬
0.010†
0.189‡
Table 2 – P- values for heart rates at hours 1, 2, 3, 4, 8, and 12. * Indicates statistical comparison
to placebo, † indicates statistical comparison to ivabradine 0.1 mg/kg, and ‡ indicates statistical
comparison to ivabradine 0.3 mg/kg.
Treatment
HR (min-1)
95% CI
Min
Max
Placebo
148±7
145, 151
139
167
0.1 mg/kg
138±7
136, 141
132
153
0.3 mg/kg
132±10
128, 136
112
154
0.5 mg/kg
130±13
125, 136
104
158
Table 3 - Twenty two hour heart rate (mean±SD, 95% Confidence Interval, Minimum and
Maximum) in eight cats after one oral dose of placebo or ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5
mg/kg). For analysis, only mean hourly heart rates were used to determine 95% CI and Range.
22
Treatment
Placebo
0.1 mg/kg
0.3 mg/kg
0.5 mg/kg
Period-1
140±22
144±17
155±25
146±17
Period-2
172±18
169±10
169±26
164±17
Change
32
25
14
18
Period-3
133±22
132±28
106±7
103±11
Period-4
172±21
145±16
127±20
117±11
Change
39
13
21
14
Table 4 – Average 15 minute heart rates (min-1; mean±SD) in eight cats during baseline before
startle (Period-1), during baseline startle (Period-2), and four hours after oral administration of
placebo or ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg) before startle (Period-3), and during
startle (Period-4).
Time (h)
Baseline
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Placebo
120±16
114±13
107±18
102±15
110±21
113±16
109±15
113±17
114±19
113±21
115±30
116±11
112±15
108±16
108±14
105±13
106±16
110±10
106±13
111±15
115±21
110±18
109±20
0.1
119±18
109±16
110±24
110±15
111±9
106±12
110±14
112±7
111±9
109±15
108±13
106± 9
109 ±11
104±18
104±12
109±8
108±6
107±6
110±13
111±18
114 ±19
107±19
n.d
Ivabradine (mg/kg)
0.3
124±16
115±19
109±9
110±11
111±12
111±17
106±14
112±13
119±17
112±18
107±14
104±11
104±11*
97.4±9
104±13
107±14
104±14
106±13
109±17
111±15
116±16
118±36
116±33
0.5
125±15
115±14
107±15
107±13
117±16
109±11
115±35
127±31
122 ±17
122±22
120±26
115±18
106±16
112±12
114±13
109±14
114±14
118±16
114±19
114±14
117 ±14
105±7
103±12
Table 5 – Mean±SD hourly systolic blood pressure (mmHg) in eight cats after oral administration of
placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, 0.5 mg/kg). n.d. = not determined.
* indicates p<0.05 compared to placebo. Data were compared only at 1h, 2h, 3h, 4h, 8h, and 12h.
23
Time (h)
Baseline
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Placebo
85±14
81±12
76 ±15
72±14
77±17
80±11
78±12
79±17
78 ±18
78±15
79±14
84±9
78±13
76±12
76±12
73±11
72±13
76±9
73±12
78 ±14
80±17
77±17
75±17
0.1
84±18
76±14
77±17
77±15
83±12
78±9
73±9
78±9
78±11
79±12
76±13
74±12
73±10
75±12
72±17
70±16
76±11
74±8
75±6
76±8
79±12
79±16
73±14
Ivabradine (mg/kg)
0.3
90±17
76±8*
71±9
69±10
73±13
74±17
68±17
73±12
79±15
82±18
69±18
72±13
67±10*
70±14
70±15
67±12
74±14
75±15
74±15
78±18
85±37
80±32
76±9
0.5
90±14
79±11
70±12*
68±12
75±16
71±10
80±39
90±34
82±16
80±18
75±15
74±9
71±10*
79±12
81±11
75±11
79±13
80±14
80±16
81±14
83±16
73±11
69±12
Table 6 – Mean±SD hourly diastolic blood pressure (mmHg) in eight cats after oral administration
of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, 0.5 mg/kg). * indicates p<0.05
compared to placebo. Data were compared only at 1h, 2h, 3h, 4h, 8h, and 12h.
24
Time (h)
Baseline
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Placebo
102±14
96±12
91±16
86±14
92±19
95±13
93±14
95±17
95±18
91±15
88±10
97±10
93±13
90±13
92±14
86±11
87±13
92±8
89±12
93±14
96±19
93±18
90±18
0.1
100±18
92±14
92±20
92±15
102±12
95±9
88±10
94±10
92±11
93±10
91±13
89±12
89±10
90±12
87±17
84±13
90±10
90±6
90±4
90±9
92±14
94±14
87±16
Ivabradine (mg/kg)
0.3
106±16
93±8
89±8
86±8
92±12
92±16
85±16
90±12
97±15
95±16
86±12
85±11
81±9*
83±12
87±16
84±12
90±13
93±14
90±15
93±15
87±11
84±11
92±12
0.5
107±14
97±13
88±12
86±11
96±13
89±10
84±9
97±12
100±16
95±17
93±11
92±8
88±13
94±11
95±10
91±12
95±14
97±15
95±17
97±13
99±15
88±9
85±12
Table 7 – Mean±SD hourly mean blood pressure (mmHg) in eight cats after oral administration of
placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, 0.5 mg/kg). * indicates p<0.05
compared to placebo. Data were compared only at 1h, 2h, 3h, 4h, 8h, and 12h.
25
Hour
Placebo
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
17640
17784
15622
14688
17270
18871
17004
17741
17898
16724
16790
16936
15792
15444
15660
14872
15794
16390
15052
15429
16215
15510
15478
0.1
18207
16568
14630
14630
15651
14734
14630
15568
15984
16350
15552
14628
14715
14248
13728
14388
15120
14445
14630
14652
15960
14231
n.d.
Ivabradine (mg/kg)
0.3
19096
17135
12971
12320
13320
14208
13568
14672
15113
15120
13268
13000
13312
12222
13728
14231
14040
14416
15042
15984
15544
15458
16356
0.5
19750
16560
11663
11128
13572
12862
13455
15113
15372
15860
15480
14720
13144
14896
15618
14388
15732
16520
16074
16302
17433
13860
12978
Table 8 – Mean hourly rate-pressure product (mmHg*min-1) in eight cats after oral administration of
placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg). n.d.= not
determined.
Treatment
Rate-Pressure Product
Placebo
16415±1132.2
0.1 mg/kg
15148±1010.8
0.3 mg/kg
14444±1603.3
0.5 mg/kg
14977±1932.6
Table 9 – Mean±SD 22-hour rate-pressure product (mmHg*min-1) in eight cats after oral
administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg).
26
Treatment
Rate-Pressure Product
Placebo
17081±1097
0.1 mg/kg
15594±1086
0.3 mg/kg
14483±1955
0.5 mg/kg
14628±2329
Table 10 – Mean±SD 12-hour rate-pressure product (mmHg*min-1) in eight cats after oral
administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg).
27
Heart Rate (min-1)
FIGURES
Placebo
0.1 mg/kg
0.3 mg/kg
0.5 mg/kg
160
140
120
100
1 ^ 3
5
7
9 11 13 15 17 19 21 23
Time (h)
Figure 1 – Mean hourly heart rate (min-1) in eight cats determined before (baseline) and after oral
administration of placebo or one dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg).
28
Heart Rate (min-1)
160
150
140
130
120
110
P
0.1
0.3
0.5
Ivabradine (mg/kg)
Figure 2 – Mean±SD of 22-hour average heart rates (min-1) in eight cats after oral administration of
placebo or one dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg). P = placebo, 0.1 =
ivabradine 0.1 mg/kg, 0.3 = ivabradine 0.3 mg/kg, and 0.5 = ivabradine 0.5 mg/kg. Other legends
as per Figure 1.
29
A.
Heart Rate (min-1)
200
180
160
140
120
100
Period-1
Period-2
Startle
B.
Heart Rate (min-1)
200
180
160
140
120
100
Period-3
Period-4
Startle
Figure 3 – Mean±SD of 15 minute average heart rates (min-1). A) Baseline before startle (Period-1)
and during baseline startle (Period-2) B) Treatment before second startle (Period-3) and during
second startle (Period-4). Baseline periods = prior to drug administration. Treatment periods are
four hours after oral administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3
mg/kg, and 0.5 mg/kg). Other legends as per Figure 1.
30
Heart Rate (min-1)
200
180
160
140
120
100
Period-2
Period-4
Startle
Figure 4 – Mean±SD of 15 minute average heart rates (min-1) during baseline (Period-2) and
treatment startle (Period-4). Baseline Startle is prior to drug administration and treatment Startle is
4 hours after oral administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg,
and 0.5 mg/kg. Other legends as per Figure 1.
31
Systolic Blood Pressure
(mmHg)
150
100
50
0
0 ^ 2
4
6
8 10 12 14 16 18 20 22 24
Drug Administration
Time (h)
Figure 5 – Mean hourly systolic blood pressure (mmHg) in eight cats determined after oral
administration of placebo or one dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg. Other
legends as per Figure 1.
32
Diastolic Blood Pressure
(mmHg)
150
100
50
0
0 ^ 2
4
6
8 10 12 14 16 18 20 22 24
Drug Administration
Time (h)
Figure 6 – Mean hourly diastolic blood pressure (mmHg) in eight cats determined after oral
administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg).
Other legends as per Figure 1.
33
Mean Blood Pressure
(mmHg)
150
100
50
0
0 ^ 2
4
6
Drug Administration
8 10 12 14 16 18 20 22 24
Time (h)
Figure 7 – Mean hourly mean blood pressure (mmHg) in eight cats determined after oral
administration of placebo or one single dose of ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg). See
Other legends as per Figure 1.
34
Rate-Pressure Product
(mmHg*min-1)
18000
16000
14000
12000
P
0.1
0.3
0.5
Ivabradine (mg/kg)
Figure 8 – Mean ±SD 22-hour rate-pressure product (mmHg*min-1) in 8 cats after oral
administration of placebo or one single dose of ivabradine 0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg.
Other legends as per Figure 1 and 2.
35
Rate-Pressure Product
(mmHg*min-1)
18000
16000
14000
12000
P
0.1
0.3
0.5
Ivabradine (mg/kg)
Figure 9 – Mean±SD 12-hour rate-pressure product (mmHg*min-1) in 8 cats after oral
administration of placebo or one single dose of ivabradine 0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg.
Other legends as per Figure 1 and 2.
36
CHAPTER 4
DISCUSSION
The results of this study indicate that a single dose of oral ivabradine effectively decreases
heart rate in a time-dependent and dose-dependent manner without clinically relevant effects on
systemic blood pressure in healthy cats. Ivabradine also blunts the ability to increase heart rate in
response to stress and may reduce myocardial oxygen consumption.
Ivabradine has been shown to have both time-dependent and dose-dependent effects on
heart rate in multiple species (rabbits, rats, dogs, and people). This effect is accomplished by
selective inhibition of If currents in the SA node leading to a decrease in the slope of diastolic
depolarization, reducing heart rate28-30, 41. The effects of ivabradine appear to be localized to the
SA node without effects on conductivity or dispersion of refraction in the atrium, AV node, HisPurkinje system, or ventricles, and without changing repolarization29. In all species studied thus
far, ivabradine has been shown to be devoid of concurrent negative inotropic, lusitropic or
dromotropic effects, is not inferior to atenolol and propanolol in reducing heart rate, and is similarly
effective compared to atenolol in people with stable angina.28-30, 34, 36, 41-44. To the author’s
knowledge, this is the first report on the use of ivabradine in cats. Our findings indicate that
ivabradine given orally once daily has a time dependent effect on heart rate with peak heart rate
37
reduction occurring approximately three hours after administration and duration of action lasting at
least 12 hours. Ivabradine also reduces heart rate in a dose dependent manner with higher doses
(0.3 and 0.5 mg/kg) causing a statistically significant and clinically relevant greater reduction of
heart rate compared to a lower dose of ivabradine (0.1 mg/kg).
The doses of ivabradine given in this study were clinically well tolerated without evidence
of significant adverse effects. The most commonly reported side effects in people treated with
ivabradine are visual symptoms (luminous phenomena) and bradycardia. Less commonly reported
were ventricular premature contractions, headache, and nausea29, 31, 33, 45, 46. The visual symptoms
in people are described as increases in brightness in limited areas of the visual field and are
related to ivabradine’s inhibition of another HCN channel (Ih) in the retina of people45. The visual
effects appear to be dose related and transient in nature. Changes in vision were not directly
examined in our cats, however, none of the cats appeared to have obvious visual impairment or
deficits during the study period. Sinus bradycardia with a HR < 55 min-1 occurred in 3.2% and < 40
bpm in 0.5% of people during long term safety and efficacy trials using oral ivabradine, however,
only 0.2% of people experienced clinically significant symptomatic bradycardia29. These results
were not different from patients treated with atenolol. While our cats heart rates were significantly
reduced after ivabradine there was no evidence of symptoms associated with severe bradycardia
such as weakness, fatigue, hypotension, or syncope. Although ventricular arrhythmias were
reported after administration of ivabradine in people, the incidence of the ventricular arrhythmias
was no different from placebo, amlodipine, or atenolol and most likely was caused by concurrent
disease rather than the drug administered29. Our cats experienced a total of 14 VPCs during the
entire study period and specific dose of ivabradine did not appear to cause VPCs compared to
placebo. Furthermore, the number of VPCs in this study is no different than the number of VPCs
over 24 hours previously published in normal cats47, 48. Nausea is a reported side effect in people
38
receiving ivabradine. The most common gastrointestinal sign in the cats of this study were
decreased appetite occurring in three cats for a total of seven (22%) study days. Inappetance,
however, was not only associated with administration of ivabradine but also placebo, with no
difference in frequency between the two treatments. Therefore, it is unlikely that the decreased
appetite observed was solely related to ivabradine.
The normal mean twenty four hour heart rate in healthy cats has been previously studied
in a total of 43 healthy cats based on mean heart rate determined from 24-hour Holter ECG
analysis47, 48. Ware et. al48 found an overall mean heart rate of 157 min-1 in 20 healthy cats with no
significant difference between younger(n=10; 1-4 years old) and older cats(n=10, 8-14 years old).
Hanas et al47 found a similar although slightly higher median 24-hour heart rate (165 min-1) in 23
healthy cats, but also reported on an effect of age with younger cats (1-6 years old) having a lower
median heart rate (167 min-1) compared to older cats (7-15 years old; HR 188 min-1). Results from
our study for the mean 22-hour heart rate in the placebo group (148 min-1) are in agreement with
the findings of the previous studies47, 48. Furthermore, compared to placebo and results of other
studies in cats48 ivabradine (0.1 mg/kg, 0.3 mg/kg, and 0.5 mg/kg) decreased mean 22-hour heart
rate in our study, indicating that ivabradine has a substantial negative chronotropic effect in healthy
cats.
MacGregor et. al49 and Quinones et. al50 evaluated the pharmacokinetic and
pharmacodynamic effects of orally administered atenolol on heart rate in healthy cats. These
studies revealed that atenolol induces a time-dependent effect on heart rate that lasts for at least
12 hours with a peak serum concentration and heart rate effect one to two hours after
administration49, 50. The study performed by Quinones et. al50 demonstrated that after oral
administration of atenolol, the atenolol blood concentration and heart rate were significantly
different from baseline at 2, 6, and 12 hours but not 24 hours post pill, which led to the conclusion
39
that atenolol should be administered twice daily for effective heart rate reduction in cats.
MacGregor et. al49 also found that oral administration of atenolol at 6.25 mg q 12h for one week
caused a reduction in median heart rate compared to baseline at 2 and 12 hours post
administration in cats. Our results revealed a similar effect of ivabradine on mean heart rate in
healthy cats with a slightly different peak effect, but a comparable duration of action. These finding
indicate that ivabradine given at doses of 0.3 and 0.5 mg/kg may have similar negative
chronotropic effects compared to atenolol and should therefore, be administered twice daily for
clinical use. A direct comparison between the studies, however, cannot be made as the
experimental design of the three studies was different. Our study did not evaluate the
pharmacokinetics of ivabradine, and heart rate was statistically analyzed at 1, 2, 3, 4, 8, and 12
hours. Further studies directly comparing the effects of atenolol and ivabradine in cats need to be
performed to determine comparability of efficacy.
Hypertrophic cardiomyopathy is the most common cardiac disease in cat, and a majority of
affected cats also develop systolic anterior motion (SAM) of the mitral valve causing dynamic LV
outflow tract obstruction referred to as hypertrophic obstructive cardiomyopathy (HOCM)11.
Multiple theories have been proposed to explain the pathophysiologic mechanism leading to the
development of SAM including a subaortic septal bulge casing narrowing of the outflow tract,
Venturi forces, abnormal papillary muscle location and dyssynchrony of contraction, an abnormal
anterior mitral valve leaflet, and flow drag, the pushing force of flow causing a dominant
hydrodynamic force on the mitral valve leaflets51. In all likelihood it is a combination of factors that
predispose people and cats with HCM to develop SAM. The most common medical therapy for
HOCM is beta adrenergic receptor blockers. Beta blockers decrease heart rate, increase diastolic
filling time, decrease inotropy, and decrease LV ejection acceleration causing a reduction or
abolishment of SAM52. Similar to a beta blocker, ivabradine is a negative chronotropic agent that
40
decreases heart rate response to sympathetic stimulation resulting in an increase in diastolic filling
time and decreased hydrodynamic force on the mitral valve and therefore may also be able to
reduce dynamic outflow tract obstruction due to SAM in cats with HOCM. However, ivabradine
lacks negative inotropic effects. Further studies on the effect of ivabradine on dynamic outflow
tract obstruction of the left ventricular outflow tract are needed.
A common adverse outcome of feline HCM is the development of arterial
thromboembolism (ATE) due to thrombus formation in the left atrium. While the exact mechanism
for the development of left atrial thrombi is unknown. Stagnation of blood flow, endothelial
damage, and activation of platelets and coagulation factors associated with left atrial dilation and
poor contractile function have been proposed as pathogenic factors favoring clot formation53.
Furthermore, the development of spontaneous echocardiographic contrast (SEC), associated with
ultrasonic backscatter from red blood cell aggregates, appears to be a marker of a prothrombotic
state and a risk factor for the development of ATE54. Schober et al55 determined that decreased
left atrial appendage (LAA) function assessed by LAA emptying velocities predicted the formation
of SEC. In people56 and cats, the use of beta blockers has been associated with decreased LAA
function and LAA emptying velocities. Recent observations from our laboratory indicate that
therapeutic doses of atenolol may effect left atrial and LAA function. Therefore, the use of beta
blockers for the treatment of HOCM in cats may predispose to the formation of SEC and ATE.
Ivabradine is a selective If current inhibitor with specificity for HCN4. The HCN4 channel is the
most abundant HCN channel in the SA node, while HCN2 is the predominant channel in the atrial
myocardium29. As ivabradine has no affinity for HCN2 it is unlikely to cause decreased LAA
emptying velocities and therefore may be superior to atenolol in the treatment of cats with HOCM.
In people, elevated heart rate is considered a risk factor for cardiovascular mortality in the
general population. A landmark study in people, the BEAUTIFUL trial, concluded that patients with
41
coronary artery disease, LV dysfunction, and 24-hour average heart rate >70 bpm had an
increased risk for cardiovascular death, admission to the hospital for heart failure, and admission
for myocardial infarction15, 32. Atkins et.al10 found in a retrospective study of 56 cats with HCM, that
a heart rate > 200 min-1 was a negative prognostic indicator for survival. A potential reason for the
devastating effect of dynamic heart rate elevation is that tachycardia decreases myocardial
perfusion by limiting diastolic coronary blood flow and, on the other hand, increases myocardial
oxygen consumption and thus, may induce myocardial ischemia. Ischemia aggravates diastolic
dysfunction, leads to Ca2+-overload, triggers apoptosis and arrhythmias, and finally induces
myocardial fibrosis. Myocardial oxygen consumption during exercise is well correlated to the ratepressure product in people 40, 57, 58. Ivabradine in conjunction with lowering heart rate also
decreases the rate-pressure product possibly indicating a reduction in myocardial oxygen
consumption. Hypertrophic cardiomyopathy is a disease characterized by LV hypertrophy with
myocardial ischemia due to dysfunction and narrowing of the intramural coronary arteries resulting
in decreased myocardial perfusion and increased myocardial oxygen consumption5, 7. Elevated
heart rate, and in particular unwanted sudden periods of tachycardia, may have detrimental effects
on myocardial function and survival in cats with HCM. The most common medications used
clinically in cats for the reduction of heart rate are beta blockers. Beta blockers, such as propanolol
and atenolol, reduce heart rate and may lead to reduced myocardial oxygen consumption but have
also been shown to possibly cause coronary arterial vasoconstriction at rest and during exercise in
experimental dogs and people with heart disease when given at high doses or intravenously,
potentially exacerbating ischemia35, 44. Ivabradine when compared to propranolol had similar
effects on heart rate reduction and surrogate measures of myocardial oxygen demand without
adverse effects of coronary vascular tone in dogs and people. A similar lack of coronary
vasoconstriction was observed in rats administered ivabradine34. Assuming comparable effects of
42
ivabradine on lusitropy, inotropy, and coronary vascular resistance in cats compared to other
species, ivabradine may be beneficial in the treatment of feline HCM.
Acoustic startle was used to induce stress associated with activation of the sympathetic
nervous system resulting in an increase of heart rate. This method, may mimic naturally-occurring
responses to stress as seen in cats around dogs, handled by children, or cats at the veterinarians
office. Previous studies evaluating pharmacologically induced or physical stress on heart rate in
normal cats revealed that isoproterenol, physical restraint, and hospitalization are effective in
significantly increasing heart rate50, 59, 60. The acoustic startle data during Periods 1 and 2 of our
study determined that heart rate significantly increased after startle for all cats supporting the
concept that acoustic startle acts as a stressor in cats and induces an increase in heart rate. The
results of the acoustic startle data during Periods 3 and 4 also indicate that ivabradine has the
ability to blunt the peak heart rate response to stress in healthy cats. Furthermore, while
ivabradine was able to blunt peak heart rate response to startle it did not completely nullify the
positive chronotropic responses to stress in these cats. Our findings are in accordance with a
study by Quinones et al50 where atenolol was able to reduce the isoproterenol-induced increase of
heart rate in cats, and likely indicates comparable effects of ivabradine to atenolol in their ability to
blunt tachycardic responses to stress. These results may be of clinical importance as ivabradine
may be useful in the prevention of sudden periods of tachycardia associated with routine
examinations at the veterinary hospital, stress experienced in the home environment, pain,
anesthesia, or elective surgery that may lead to acute decompensation of otherwise clinically
stable cats with HCM as reported elsewhere11, 17.
Experimental studies of ivabradine in people, dogs, and rats have shown that while
ivabradine causes a significant reduction of heart rate, it does not affect systemic arterial blood
pressure 34, 35, 44. Our study revealed similar results in healthy cats. Heart rate was significantly
43
reduced with all doses of ivabradine, but there was no general effect on systemic arterial blood
pressure. Although at dosages of 0.3 mg/kg and 0.5 mg/kg of ivabradine reduced the 12 hour BP,
the magnitude of reduction did not appear to be clinically relevant. As a selective If current
inhibitor, ivabradine has been shown to lack direct negative inotropic or lusitropic effects. While the
latter have not yet been directly studied in the feline it is likely that these effects are similar in cats.
As a consequence, no overall effect on cardiac output or blood pressure would be expected, and
was also observed in our study with regard to blood pressure. Ivabradine also does not have any
obvious indirect effects on vasoreactivity, therefore systemic vascular resistance is not affected as
found in experimental dogs and rats 34, 35. The latter is most likely also true in cats.
There are limitations to this study. First, to reduce the number of comparisons and to
maintain statistical power for detection of meaningful differences, statistical comparisons were only
performed for hours 1, 2, 3, 4, 8, and 12 h or were pooled for the entire 22-hour study period. As a
result, we may have missed significant changes for heart rate and blood pressure at other time
points. Due to the lack of analysis beyond 12 hours after the administration of ivabradine, the
exact duration of the effect of ivabradine cannot be stated based on the results reported. Another
limitation is the presence of cats in our study that had previously been diagnosed with feline
interstitial cystitis. Buffington et. al61 previously reported that cats with FIC have a significant
elevation in plasma norepinephrine concentration compared to healthy controls, indicating that cats
with FIC may have a higher level of baseline sympathetic tone compared to normal cats. People
with FIC have also been shown to have an exaggerated response to acoustic startle62. As a result,
even though the cats of our study population were not displaying clinical signs of FIC at the time of
study, their resting HR and response to acoustic startle may have been elevated, and the effects of
ivabradine may have been different compared to truly healthy cats. Furthermore, given the small
sample size of the study, FIC was not considered as a variable in our statistical analysis and we
44
cannot determine whether or not FIC had an effect on drug treatment. Finally, cats with HCM, the
disease in which ivabradine may potentially be clinically useful, were not studied. It is known from
people, that patients with HCM may have a different autonomic response to stress and exercise
compared to healthy individuals. Therefore, caution is advised in the extrapolation of our findings
to cats with naturally-acquired HCM.
In conclusion, our study indicates that ivabradine can be safely administered to healthy
cats at doses of 0.1, 0.3, and 0.5 mg/kg, PO. Ivabradine at 0.3 mg/kg and 0.5 mg/kg, appears to
be most effective in reducing overall heart rate and chronotropic response to physiologic stress and
will most likely have to be administered twice daily for clinical use. Further studies are warranted to
evaluate the pharmacokinetics and hemodynamic effects of ivabradine in healthy cats as well as
studies evaluating the effects of ivabradine in cats affected with HCM.
45
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