Hay_249-253.indd
INTRODUCTION
Six species of the plant family Rubiaceae, viz. Pachy-
stigma latifolium, Pachystigma pygmaeum, Pachy-
stigma thamnus, Pavetta harborii, Pavetta schu-
manniana and Fagodia homblei cause gou siekte, a
syndrome characterized by heart failure and sud-
den death, when ingested by ruminants (Theiler, Du
Toit & Mitchell 1923; Pretorius & Terblanche 1967;
Kellerman, Coetzer, Naudé & Botha 2005). A num-
ber of cardiodynamic changes (Pretorius & Ter-
blanche 1967) as well as myocardial lesions that
consist of a loss of myofilaments, replacement of
myocytes with collagenous tissue, lengthening of
sarcomeres and cardiac dilatation (Newsholme &
Coetzer 1984) have been reported. Apart from the
economic impact on farming with domestic rumi-
nants the plant has attracted the attention of re-
searchers for its possible use as a model for study-
ing heart failure.
One model that was successfully used in sheep was
that produced by addition of dried Pachystigma pyg-
maeum leaves to a normal diet (Schutte & Du Plooy
1990). A later study also succeeded in inducing
heart failure in rats by injecting crude extracts of Pa-
vetta harborii intraperitoneally (Hay, Pipedi, Schutte,
Turner & Smith 2001). Administration of dried leaves
or crude extracts to induce heart failure can, how-
ever, have limitations because of seasonal variations
in plant toxicity resulting in variations in the degree
of heart failure that develops. Theiler et al. (1923)
found that factors such as soil type and seasonal
climate changes caused variations in the toxicity of
Pachystigma pygmaeum (= Vangueria pygmaea)
249
Onderstepoort Journal of Veterinary Research, 75:249–253 (2008)
Cardiotoxic effects of pavetamine extracted from
Pavetta harborii in the rat
L. HAY1*, R.A. SCHULTZ2 and P.J. SCHUTTE1
ABSTRACT
HAY, L., SCHULTZ, R.A. & SCHUTTE P.J. 2008. Cardiotoxic effects of pavetamine extracted from
Pavetta harborii in the rat. Onderstepoort Journal of Veterinary Research, 75:249–253
Previous studies have shown that crude extracts from Pavetta harborii as well as dried plant material
have cardiotoxic effects on rats and sheep that can lead to heart failure. The active component has
since been isolated and identified. This substance has been named pavetamine. The aim of this study
was to determine whether pavetamine has cardiotoxic effects similar to those seen in previous re-
ports, when administered to rats intraperitoneally. Sprague Dawley rats received two doses, initially 4
mg/kg and then 3 mg/kg pavetamine respectively and were monitored for 35 days before cardiody-
namic parameters were measured by inserting a fluid-filled catheter into the left ventricle via the right
carotid artery. These values were compared to those of control rats that had received only saline.
Pavetamine significantly reduced systolic function and body mass in the treated rats, which indicates
that it has the potential to induce heart failure in this animal model.
Keywords: Cardiodynamics, cardiotoxic, pavetamine, Pavetta harborii, rat
* Author to whom correspondence is to be directed. E-mail:
lhay@ul.ac.za
1 Department of Physiology, University of Limpopo, Medunsa
Campus, P.O. Box 130, Medunsa, 0204 South Africa
2 Section of Toxicology, Private Bag X05, ARC-Onderstepoort
Veterinary Institute, Onderste poort, 0110 South Africa
Accepted for publication 17 June 2008—Editor
250
Cardiotoxic effects of pavetamine extracted from Pavetta harborii in the rat
and it should therefore be considered that this might
also be the case for other species of this family.
Fourie, Erasmus, Schultz & Prozesky (1995) suc-
ceeded in isolating and describing the active com-
pound which has become known as pavetamine. It
be came available in small quantities for further in-
vestigation.
Subsequent investigations have shown that paveta-
mine inhibits protein synthesis in the cardiac muscle
of rats but that it has no effects on other tissues
(Schultz, Fourie, Basson, Labuschagne & Prozesky
2001). The aim of this study was to investigate in
rats whether pavetamine has cardiodynamic effects
typical to those described previously for the dried
material of Pachystigma pygmaeum and crude ex-
tracts of Pavetta harborii given to sheep and rats
respectively.
METHODS
Experimental animals
The investigation conforms to the Guide for the care
and use of laboratory animals (NIH publication No.
85-23, revised 1996). Ethics approval was obtained
from the Animal Ethics Committee of the ARC-
Onder stepoort Veterinary Institute.
Healthy, young, male Sprague Dawley rats (n = 20)
of the same age were used and equally divided into
a control and a treated group. They were kept indi-
vidually in separate cages and had free access to
water and nutritionally balanced rat cubes (Epol Pty.
Ltd. SA). Pavetamine from dried P. harborii plant
material was extracted by the method described by
Fourie et al. (1995). The rats in the treated group
each received an intraperitoneal injection of 4 mg/
kg pavetamine in 0.5 mℓ saline on Day 1 and a sec-
ond injection of 3 mg/kg in 0.5 mℓ saline on Day 20.
The control rats only received similar quantities of
normal saline administered by the same route.
Cardiodynamic data were recorded for the individu-
al experimental animals on 1 day from Days 35–39.
These data were recorded for the individual rats
over a period of 5 days as time constraints on the
procedure limited the number of animals that could
be done in 1 day.
The animals were anaesthetized by administering
an intramuscular injection of 0.5 mℓ of a mixture of
ketamine (100 mg/mℓ) (Kyron Laboratories Pty Ltd.
Johannesburg, SA) and xylazine (2 %) (Premier
Pharmaceuticals Co Ltd. Johannesburg, SA) at a
ratio of 1:3. The anaesthesia was maintained
through out the procedure by further 0.1 mℓ injec-
tions at 20 min intervals or as needed. A constant
anaesthetic plane was continually assessed by
means of the tarsal pinch reflex.
The following surgical procedure and cardiodynamic
monitoring on each rat were performed in a sterile
environment on an operation table prewarmed to
maintain the body temperature of the animals. Each
rat was placed on its back and its right carotid artery
surgically exposed and separated from the accom-
panying vagus nerve. Care was taken not to damage
the nerve. A catheter (Cordis 2.5 F) was inserted into
the left ventricle of the heart via the exposed artery.
During this procedure care was taken to minimize
blood loss. The catheter was connected to a fluid-
filled Hewlett-Packard Quartz transducer which was
connected to a Hewlett-Packard 8 channel record-
ing system (HP7758) and the data obtained were
stored on a magnetic tape for later analysis. The
pressure recordings were monitored on an oscillo-
scope to confirm that the catheter was in the left
ventricle.
The following indices were monitored and comput-
ed: peak left ventricular pressure (LVSP), left ven-
tricular end-diastolic pressure (LVEDP), heart rate
(HR), the maximum rate of increase of left ventricu-
lar pressure (+LVdP/dtmax) indicating left ventricular
(LV) contractility, the time constant T calculated from
the LV pressure curve as the negative inverse of the
slope of the regression line of the LV pressure ver-
sus LV -dP/dt with a variable pressure asymptote,
which gives an indication of the rate of the left ven-
tricular isovolumic relaxation (Weisfeldt, Weis, Fre-
deriksen & Yin 1980), and the cardiac work (CW) that
was calculated by multiplying HR and SBP. Directly
after the last recordings the rats received a lethal
dose (500 mg/kg) of sodium pentobarbitone (Kyron
Laboratories (Pty) Ltd).
Statistical analysis was done with Gen Stat (2000)
and the student t-test was used to compare the
treated to the non-treated groups taking P < 0.05 as
significant.
RESULTS
Fig. 1 shows that the rats receiving pavetamine had
slower mass gains than those of control rats. After
the second administration of pavetamine the treated
rats showed no further mass gains up to the termi-
nation of the experiment.
Table 1 summarizes the effects of pavetamine on
cardiac function. Both T and LVEDP were not signif-
251
L. HAY, R.A. SCHULTZ & P.J. SCHUTTE
icantly affected by the pavetamine intervention. The
other parameters that were measured were all sig-
nificantly reduced. The reduction in dP/dtmax and
CW were identical at 30.9 % but since these two pa-
rameters are affected by different factors, this simi-
larity is probably purely coincidental.
DISCUSSION
It is not clear how the pavetamine caused a decrease
in the mass gain of the treated group. Previous stud-
ies did not reveal possible effects of these plant tox-
ins on the mass of treated animals. In a study inves-
tigating the affect of pavetamine on protein synthesis
in the different tissues of rats it was shown to have
a detrimental effect on cardiac tissue but other tis-
sues were not affected despite a loss in their mass
(2 % within the first 24 h) (Schultz et al. 2001).
In another study where Pavetta harborii extracts
were administered to rats, protein synthesis in car-
diac tissue was compromised to the extent that it
lead to a degeneration in the myofilaments (Hay et
al. 2001). A similar degeneration of myocardial fibres
was also seen in sheep to which dried Pachystigma
pygmaeum leaves had been administered (Schutte,
Els, Booyens & Pienaar 1984). These pathological
changes in the cardiac tissue could be the underly-
ing cause of the reduced cardiodynamic function
(systolic and diastolic) associated with the disease
as described by Hay et al. (2001) and by Schutte &
Du Plooy (1990). When a minimum critical dosage
of the plant material is administered to animals and
if enough time is allowed for the disease to progress
fully, it will eventually reach a stage where typical
clinical signs of congestive heart failure will manifest.
These signs were described in detail by Pre torius &
Terblanche (1967).
Our results show that the pavetamine significantly
reduced the systolic component of the cardiodynam-
ic function of the rats but did not affect the diastolic
TABLE 1 The cardiodynamic parameters of control rats and rats receiving pavetamine
Control Pavetamine %Δ P≤
T(ms)
LVEDP (mmHg)
dP/dtmax (mmHg.s
–1)
CW (mm Hg.beats.min–1)
LVSP (mmHg)
HR (beats.min–1)
52.11 ± 1.9
6.9 ± 1.3
2 746 ± 105
40 570 ± 1 799
158.4 ± 7.8
258 ± 11
52.10 ± 1.6
6.4 ± 1.4
1897 ± 107
28 010 ± 2 386
119.5 ± 6.8
231 ± 7
0
–7.2
–30.9
–30.9
–24.5
–10.5
NS
NS
0.001
0.001
0.002
0.05
See text for meanings of abbreviations
0
50
100
150
200
250
300
350
400
450
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39
Time (days)
M
a
s
s
(
g
ra
m
)
Control rats: A = 0.5 mℓ saline; B = 0.5 mℓ saline
A B
Pavetamine rats: A = 4 mg/kg; B = 3 mg/kg
FIG. 1 The effects of pavetamine administration on rat body mass over a period of 39 days
252
Cardiotoxic effects of pavetamine extracted from Pavetta harborii in the rat
component measured from T, nor did it influence
the LVEDP. This is in contrast to the significant
length ening in T that was found in rats to which
crude extracts of Pavetta harborii had been admin-
istered (Hay et al. 2001). A possible explanation for
this could be that the concentration of the paveta-
mine used in this study was too low or that the time
period over which the toxin was administered was
too short, or both, to cause these additional changes
associated with heart failure, namely a high LVEDP
and a reduced diastolic function.
All the indicators of systolic function in our study
were significantly reduced but the heart rate did not
increase significantly, suggesting that the clinical
signs usually associated with advanced congestive
heart failure (Pretorius & Terblance 1967) had not
yet fully developed.
Both CW and dP/dtmax were significantly decreased
and the substantial reduction in LVSP implies that
the reduction in dP/dtmax was sufficient to alter car-
diac effectiveness. It is known that altered loading
conditions of the heart can affect an index such as
dP/dtmax (Broughton & Korner 1980). In our study
only the afterload was significantly altered and this
would therefore suggest that a slight overestimation
of dP/dtmax would have occurred in the treated group.
This fact combined with the lower LVSP would there-
fore suggest that this factor should not detract from
our observations on dP/dtmax.
The HR was significantly reduced in the treated rats
and this was unexpected in the light of a diminished
contractile force. Similar observations were made in
rats receiving crude extracts (Hay et al. 2001). How-
ever, in goats administered pavetamine resulted in
tachycardia and ECG changes (Fourie et al. 1995).
Likewise, in the sheep studies heart rate was initi-
ally not affected but later arrhythmias appeared and
in the final stages of congestive heart failure tachy-
cardia occurred (Pretorius & Terblanche 1967).
Various changes in the ECG were described by Pre-
torius & Terblanche (1967). This would suggest that
the toxin contained in the dried plant material affect-
ed the electrophysiology of the pacemaker cells. The
authors of that paper suggested that ischaemia could
be responsible for the observed ECG changes (Pre-
torius & Terblanche 1967). T-wave inversion was a
common occurrence in sheep that received the dried
plant material of Pachystigma pygmaeum (Pretorius
& Terblanche 1967; Pretorius et al. 1973). T-wave
inversion is also a well known phenomenon associ-
ated with hypokalemia (Ganong 2001) and since
electrophysiological affects could also involve po-
tassium ions this aspect should be further investi-
gated.
Therefore a possible effect of the toxin on the ions
and the associated affect it might have on the mem-
branes of the pacemaker cells should not be disre-
garded when the HR observations are considered.
As mentioned before Pretorius et al. (1973) con-
firmed Ca2+ abnormalities in myocardial cells ex-
posed to this toxin. Ca2+ abnormalities can techni-
cally also impact on the cell membranes of damaged
myocardial cells via complex interactions in the
Ca2+/Na exchange system. This has been confirmed
to take place in cases where myocardial cells were
damaged in other ways than by this toxin (Pieske
1998).
In conclusion, the results suggest that pavetamine
administered to rats diminishes their systolic func-
tion, but the effects on diastolic function and HR sug-
gest that the rats were not in the advanced stages
of congestive heart failure associated with a high
LVEDP and compensatory tachycardia. Further dose
response studies need to be done to determine the
dose of pavetamine required to induce advanced
congestive heart failure similar to the clinical picture
described in sheep (Pretorius & Terblanche 1967;
Kellerman et al. 2005).
ACKNOWLEDGEMENTS
Our special thanks to Mr O.H.I. Majane (UL), Ms L.
La buschagne, Ms K.M. Basson (ARC-OVI) and Ms
M.F. Smith (ARC-Biometry Unit) for their valuable
contributions during this project. Funding was pro-
vided by the Gauteng Province (Department of Agri-
culture, Conservation and Environment) and the
North-West Province (Department of Agriculture,
Conservation, Environment and Tourism).
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/NLD (Gebruik deze instellingen om Adobe PDF-documenten te maken die zijn geoptimaliseerd voor prepress-afdrukken van hoge kwaliteit. De gemaakte PDF-documenten kunnen worden geopend met Acrobat en Adobe Reader 5.0 en hoger.)
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/UKR <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>
/ENU (Use these settings to create Adobe PDF documents best suited for high-quality prepress printing. Created PDF documents can be opened with Acrobat and Adobe Reader 5.0 and later.)
>>
/Namespace [
(Adobe)
(Common)
(1.0)
]
/OtherNamespaces [
<<
/AsReaderSpreads false
/CropImagesToFrames true
/ErrorControl /WarnAndContinue
/FlattenerIgnoreSpreadOverrides false
/IncludeGuidesGrids false
/IncludeNonPrinting false
/IncludeSlug false
/Namespace [
(Adobe)
(InDesign)
(4.0)
]
/OmitPlacedBitmaps false
/OmitPlacedEPS false
/OmitPlacedPDF false
/SimulateOverprint /Legacy
>>
<<
/AddBleedMarks false
/AddColorBars false
/AddCropMarks false
/AddPageInfo false
/AddRegMarks false
/ConvertColors /ConvertToCMYK
/DestinationProfileName ()
/DestinationProfileSelector /DocumentCMYK
/Downsample16BitImages true
/FlattenerPreset <<
/PresetSelector /MediumResolution
>>
/FormElements false
/GenerateStructure false
/IncludeBookmarks false
/IncludeHyperlinks false
/IncludeInteractive false
/IncludeLayers false
/IncludeProfiles false
/MultimediaHandling /UseObjectSettings
/Namespace [
(Adobe)
(CreativeSuite)
(2.0)
]
/PDFXOutputIntentProfileSelector /DocumentCMYK
/PreserveEditing true
/UntaggedCMYKHandling /LeaveUntagged
/UntaggedRGBHandling /UseDocumentProfile
/UseDocumentBleed false
>>
]
>> setdistillerparams
<<
/HWResolution [2400 2400]
/PageSize [595.276 841.890]
>> setpagedevice