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Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs
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Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs* M. De Silva, DVMa,*, A. Mihailovic, DVMb, M.B. Toaldo, DVM, PhDa,c a
Department of Veterinary Medical Sciences, Alma Mater Studiorum, University of Bologna, Italy b Bologna, Italy Received 21 August 2019; received in revised form 23 January 2020; accepted 24 January 2020
KEYWORDS Cavia porcellus; Exotics; Cardiology; Ultrasound; Heart Q3
Abstract Objectives: The objective of this study was to report normal echocardiographic values in healthy guinea pigs. Animals, materials, and methods: Twenty-two privately owned, apparently healthy, conscious guinea pigs underwent complete transthoracic echocardiography. Left ventricular (LV), right ventricular, left atrial, and aortic root dimensions were measured, as were forward flow velocities across the mitral, aortic, and pulmonic valves. The effects of age, body weight, sex, and heart rate on these variables were also investigated. Results: The median age (interquartile range) was 3.0 (1.8e4.0) years with a body weight of 902 (822e998) grams. Echocardiography was feasible in all conscious animals. Early and late diastolic transmitral flow waves were summated in 17 of 22 individuals. In the remaining five animals, the two waves were reversed (E waveto-A wave velocity less than 1.0). A positive correlation was detected between body weight and LV internal diameter at end-diastole and end-systole and left atrial diameter (P < .05). Heart rate was negatively correlated with LV internal diameter at end-systole (r ¼ .463, P ¼ .035). Age was positively correlated with LV posterior wall thickness at end-diastole and aortic diameter (P < .05). LV internal diameter at end-systole was larger in males than in females (P ¼ .012), while fractional shortening was lower (P ¼ .008).
* This work was done at the Department of Veterinary Medical Sciences, Alma Mater Studiorum e University of Bologna, Via Tolara di Sopra 50, 40,064, Ozzano Emilia, Italy. * Corresponding author. E-mail address: [email protected] (M. De Silva). c Current address is: Division of Cardiology, Clinic for Small Animal Internal Medicine, Vetsuisse Faculty University of Zu ¨rich.
https://doi.org/10.1016/j.jvc.2020.01.004 1760-2734/ª 2020 Elsevier B.V. All rights reserved.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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M. De Silva et al. Conclusions: Descriptive echocardiography ranges in apparently healthy awake guinea pigs have been provided and can be used for cardiac assessment in these pet animals. ª 2020 Elsevier B.V. All rights reserved.
Abbreviations A max Ao BW FS HR IQR LA LV LVIDs
peak velocity of late diastolic transmitral flow aorta body weight fractional shortening heart rate Interquartile range left atrium left ventricular left ventricular internal diameter at end-systole
Introduction Guinea pigs or cavies (Cavia porcellus) are hystricomorph rodents originally native to South America that have been historically used in biomedical research and that are now becoming increasingly popular as domestic pets in many parts of the world, such as Europe and North America [1,2]. Clinical cases of guinea pigs presented with signs related to cardiac disease have been reported in the literature [3e6]. Echocardiography is, therefore, becoming a useful tool to diagnose and treat cardiac morbidities in these pet animals [3e5]. However, the accuracy of transthoracic echocardiography in diagnosing cardiac diseases in the different species and animal breeds depends also on the availability of appropriate reference limits of normality for each specific population. To the best of the authors’ knowledge, in the scientific literature, there is a lack of normal echocardiographic reference values obtained under standardized conditions in nonanesthetized adult pet guinea pigs. Indeed, the normal echocardiographic values reported in the literature refer in most instances to poorly representative study populations, such as genetically similar laboratory Hartley male guinea pigs, of
unknown age, small body weight (BW), and subject to anesthetic agents [7]. Consequently, these data are not fully applicable to clinical practice in adult conscious pet animals, as the aforementioned extrapolation might be misleading. The objectives of the present study were (1) to assess the feasibility of two-dimensional, M-mode, and Doppler echocardiography in the evaluation of cardiac anatomy, function, and hemodynamics in conscious, apparently healthy pet guinea pigs; (2) to measure standard echocardiographic variables and establish normalreferenceintervalsforeach of them; and (3) to carry out a statistical analysis to investigate correlations between the echocardiographic variables and age, sex, BW, and heart rate (HR).
Animals, materials, and methods Twenty-two adult pet guinea pigs were brought to the Veterinary Teaching Hospital of the University of Bologna by private owners for the purpose of health consultation and cardiac assessment from April 2016 to November 2019. All animals were apparently healthy at clinical examination, with no preexisting history of systemic or cardiovascular illness as reported by the owners. Each animal underwent a full clinical examination carried out by a veterinarian with experience in exotic animal medicine (A.M.) and were auscultated by a boardcertified veterinary cardiologist (M.B.T.). All animals were unsedated and restrained on a standard echocardiographic table in left and right lateral recumbency by one trained exotic veterinarian (A.M.) and one trained assistant (M.D.S.). The region corresponding to the cardiac apex, between the third and fifth intercostal spaces, was shaved and a small amount of water-based, previously warmed ultrasound gel was applied to the area. Echocardiograms were performed with guinea pigs in lateral recumbency on a table with a cutout, contacting the thoracic body wall from below. Examination of all guinea pigs was possible without the use of sedation. This was achieved by gentle but confident handling by the pectoral
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Fig. 1 M-mode echocardiogram obtained from a right parasternal short-axis view at the level of the papillary muscles. A leading edge-to-leading edge method was used to measure right and left ventricular internal diameters and wall thickness. Dia: diastole; IVS: interventricular septum; LV: left ventricle; LVPW: left ventricular posterior wall; RV: right ventricle; Sys: systole.
girdle and the hind limbs. The procedure was well tolerated by all animals examined. Echocardiographic examinations were performed using a dedicated diagnostic ultrasound unitd equipped with a phased array probe of variable frequency (12-4 MHz) with an aperture of 9.78 mm, without continuous electrocardiographic monitoring. Standard transthoracic two-dimensional, M-mode, and Doppler ultrasound examinations were performed by a veterinary cardiologist (M.B.T.) following a standardized protocol as described in the literature for the dog [8], by means of right parasternal (long- and short-axis) and left apical (fourand five-chamber) views. Frame rates were maintained between 50 and 100 frames per second during two-dimensional echocardiography and more than 120 frames per second during M-mode studies. Images and video clips were acquired and analyzed at the end of the study using dedicated softwaree by the same operator (M.B.T.). Each measurement was derived from three consecutive cardiac beats, and then the mean value was calculated. Right ventricular and left ventricular (LV) internal diameters and wall thickness were measured from M-mode images obtained from a right
d e
iE33 ultrasound system, Philips Healthcare, Monza, Italy. OsiriX MD Software, Pixeo SARL, Geneva, Switzerland.
parasternal short-axis view at the level of the papillary muscles, with the cursor crossing the midventricle and using a leading edge-to-leading edge method (Fig. 1). Specifically, the following variables were measured: right ventricular internal diameter at end-diastole, interventricular septal thickness at end-diastole and at end-systole, LV internal diameter at end-diastole and at endsystole (LVIDs), and LV posterior wall thickness at end-diastole and at end-systole. Because electrocardiographic monitoring was not performed, the timing for end-diastole and end-systole was considered at the moment of maximal and minimal (apogee of the LV free wall) LV dimensions, respectively. Fractional shortening (FS), expressed as a percentage value, was then calculated. The diameters of the left atrium (LA) and of the aorta (Ao) at the level of the aortic annulus were obtained from a right parasternal short-axis view at the level of the cardiac base using an inner edge-to-inner edge method and following a technique proposed for dogs (Fig. 2) [9]. The LA:Ao ratio was then calculated by dividing the two diameters. Color Doppler examination was carried out at the level of each of the four cardiac valves to detect regurgitation, while pulsed wave spectral Doppler analysis was performed for forward flow velocity measurements only at the level of the Ao (Fig. 3), pulmonary valve, and mitral valve, using
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Fig. 2 Two-dimensional echocardiographic views and measurements. (A): Right parasternal long-axis four-chamber view; (B): right parasternal left ventricular outflow view; (C): right parasternal short-axis view at the level of the papillary muscles of the left ventricle; (D): right parasternal short-axis view at the heart base. The dashed lines indicate the measurements of the aortic root diameter and the left atrial diameter using an inner edge-to-inner edge method. Ao: aorta; LA: left atrium; LAu: left auricle; LV: left ventricle; RA: right atrium; RV: right ventricle.
Fig. 3 Pulsed wave spectral Doppler analyses of transaortic and transmitral flows. (A): Pulsed wave Doppler analysis of the monophasic transaortic systolic flow. The dotted line indicates the interval between two systolic flows used to calculate the heart rate. (B): Pulsed wave Doppler analysis of the transmitral biphasic flow showing separate early (E) and late (A) diastolic flow waves. Note the inverted ratio of their peak velocities. Ao: aorta.
a sample volume of 2 mm. In particular, peak aortic and pulmonary flow velocities were measured from a left apical five-chamber view and right parasternal short-axis view at the basilar level, respectively. Peak velocities of early (E max) and late (A max) diastolic transmitral flow were measured from a left apical four-chamber view. Valvular regurgitation was considered trivial if it extended minimally from the valvular plane (2e3 mm). The echocardiography-derived HR was measured from the aortic spectral flow profile.
Statistical analysis All statistical analyses were carried out using dedicated softwaref,g Normality of distribution of continuous variables was assessed by visual inspection of the plots. Since data were not normally distributed, all variables were expressed as
f Microsoft Excel; Microsoft Office 2011, Microsoft Corporation, Bellevue, WA. g Prism 7, GraphPad Software Inc., San Diego, CA.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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median (interquartile range [IQR]), range (minimumemaximum), and 90% upper and lower confidence intervals. Continuous variables were compared between sexes with a ManneWhitney test. The distribution of sexes between two classes of BW was tested with a Fisher’s exact test. Spearman’s correlation was used to evaluate the relationship between echocardiographic variables and age, BW, and HR. A P < .05 was considered significant.
Results The study population included 8 female and 14 male guinea pigs, weighing a median of 902 (822e998) grams, with a median age of 3.0 (1.8e4.0) years. Echocardiographic examinations were feasible in every animal without need for sedation. Each examination was of adequate quality to allow a clear visualization of the myocardium, valves, atria, and ventricles and to ensure accurate measurement for all variables. All data referring to the whole population are reported in Table 1. Color Doppler examination of one Table 1
male animal of 1 year of age revealed trivial tricuspid regurgitation, while another male guinea pig of 5 years of age had trivial aortic insufficiency. All other flow was judged to be laminar from spectral Doppler evaluation. Transmitral flow waves were fused in the majority of animals (17/ 22, 77%) because of the elevated HR. In the remaining five animals, A max exceeded E max (Fig. 3). Among the tested variables, LVIDs was larger in males (median ¼ 7.2 mm [IQR ¼ 6.5e7.8; 6.1e10.7 {minimumemaximum}]) than in females (median ¼ 6.1 mm [IQR ¼ 5.8e6.8; 4.5e7.4]) (P ¼ .012), while the FS value was lower in males (median ¼ 31% [IQR ¼ 26e33; 16e42]) than in females (median ¼ 33% [IQR ¼ 33e42; 33e50]) (P ¼ .008). Aortic flow peak systolic velocity was higher in females (median ¼ 91 cm/s [IQR ¼ 81e101; 73e132]) than in males (82 cm/s [IQR ¼ 69e87; 59e107]) (P ¼ .044). There was a positive correlation between age and LV posterior wall thickness at end-diastole (r ¼ .478, P ¼ .028) and aortic dimension (r ¼ .474, P ¼ .030). A negative correlation was found between HR and LVIDs (r ¼ .463, P ¼ .035). Finally, LV internal diameter at end-diastole
Demographic and echocardiographic data obtained from 22 healthy pet guinea pigs.
Variable Age (years) BW (grams) HR (bpm) RVDd (mm) IVSd (mm) LVIDd (mm) LVPWd (mm) IVSs (mm) LVIDs (mm) LVPWs (mm) FS (%) LA (mm) Ao (mm) LA:Ao EA max (cm/sec)a E max (cm/sec)b A max (cm/sec)b PV vel (cm/sec) Ao vel (cm/sec)
Median (IQR)
Range (minimumemaximum)
Lower 90% CI
Upper 90% CI
3.0 (1.8e4.0) 902 (822e998) 268 (238e287) 3.4 (2.8e4.2) 2.1 (1.9e2.3) 10.3 (9.3e10.8) 2.0 (1.8e2.1) 2.8 (2.6e3.1) 6.8 (6.1e7.4) 2.9 (2.7e3.2) 32 (29e38) 6.9 (6.4e7.6) 5.8 (5.3e6.1) 1.2 (1.1e1.4) 73 (65e89) 43 (38e58) 75 (51e84) 84 (64e96) 83 (72e93)
1.0e7.0 500e1200 168e316 2.2e5.5 1.7e2.4 8.7e12.7 1.6e2.4 2.3e4.1 4.5e10.7 2.4e4.2 16e50 5.3e8.6 4.6e7.3 .7e1.6 47e113 38e65 44e85 40e145 57e132
2.4 841 250 3.2 2.0 9.8 1.9 2.8 6.5 2.9 30 6.6 5.5 1.1 70 39 56 73 78
3.5 947 275 3.8 2.2 10.6 2.0 3.1 7.3 3.2 35 7.2 6.0 1.3 84 55 82 90 90
Ao: aorta; bpm: beats per minute; BW: body weight; CI: confidence interval; A max: peak velocity of late diastolic transmitral flow; E max: peak velocity of early diastolic transmitral flow; EA max: peak velocity of summated E and A transmitral waves; FS: fractional shortening; HR: heart rate; IQR: interquartile range; IVSd: interventricular septum thickness at end-diastole; IVSs: interventricular septum thickness at end-systole; LA: left atrium; LVIDd: left ventricular internal diameter at end-diastole; LVIDs: left ventricular internal diameter at end-systole; LVPWd: left ventricular posterior wall thickness in diastole; LVPWs: left ventricular posterior wall thickness in systole; PV: pulmonary valve; RVDd: right ventricular internal diameter in diastole; vel: velocity. a Number of animals ¼ 17. b Number of animals ¼ 5.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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CI: confidence interval; HR: heart rate; IQR: interquartile range; LA: left atrium; LVIDd: left ventricular internal diameter at end-diastole; LVIDs: left ventricular internal diameter at end-systole. a No difference between classes of body weight regarding sex distribution (Fisher’s exact test). b No difference between classes of body weight (ManneWhitney test).
4.4 281 11.5 8.2 7.7 2.7 230 10.5 6.7 7.0 3 (2.5e4) 262 (223e295) 10.8 (10.4e12) 7.2 (6.3e7.9) 7.6 (6.8e7.8) 3.2 281 9.9 6.8 6.7 1.6 267 9.1 5.9 6.0 1e5 231e300 8.7e10.7 4.5e7.4 5.3e7.2 2 (1e4) 273 (250e286) 9.4 (8.8e10.2) 6.7 (6e6.9) 6.6 (5.8e6.9)
Lower 90% CI Range (minimumemaximum)
1.3e7 168e316 9.8e12.7 6e10.7 6.6e8.6
Upper 90% CI Range (minimumemaximum)
Lower 90% CI
Age (years)b HR (bpm)b LVIDd (mm) LVIDs (mm) LA (mm)
Variable
Median (IQR)
Body weight 900gr (11 animals: 5 males, 6 females)a
Upper 90% CI
Median (IQR)
Body weight > 900gr (11 animals: 9 males, 2 females)a
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Selected echocardiographic data obtained from 22 healthy pet guinea pigs divided on the basis of body weight.
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Table 2
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(r ¼ .776, P < .001), LVIDs (r ¼ .443, P ¼ .044), and LA diameter (r ¼ .519, P ¼ .016) were positively correlated with BW. Owing to the effect of BW on some echocardiographic variables, descriptive reference intervals for two ranges of BW ( 900gr and >900gr) are provided in Table 2.
Discussion The present study showed that echocardiography in unsedated guinea pigs is feasible and allows clear visualization of all cardiac structures. This was likely facilitated by the use of a high-frequency transducer with adequate frame rate capability and by following the general procedural recommendations for small exotic mammals [10]. The study cohort comprised adult (>1 year of age) individuals of various BW and ages with a uniform distribution of both sexes, and therefore reflected a representative guinea pig population commonly seen in clinical practice. This makes the reported echocardiographic values useful for exotic animal specialists and cardiologists performing echocardiography in this species. A previous echocardiographic study included a population of genetically similar male laboratory guinea pigs of unknown age, smaller BW, and which were anesthetized with ketamine and xylazine [7]. Therefore, the aforementioned study is not fully applicable to clinical practice, as the extrapolation of the results may have limitations. As a matter of fact, measurements of cardiac dimensions obtained in our study have higher values than those reported in previously published studies [7]. Moreover, Doppler examination was not performed in the aforementioned study, while such data are available in the present one. A significant correlation between LV internal diameter at end-diastole, LVIDs, and LA with BW was detected, and this finding is consistent (except for LVIDs) with a previous echocardiographic study carried out on anesthetized guinea pigs [7]. The influence of BW on echocardiographic estimates of cardiac dimensions has already been observed in other species, such as dogs and cats [11,12]. Allometric scaling best describes the association between BW and some echocardiographic variables in these species, with curvilinear regression lines used for predicting reference ranges and with a scaling exponent usually set around .3 for both dogs and
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Echocardiography in healthy guinea pigs cats [11,12]. In the present study on guinea pigs, we did not perform regression analysis because of the small number of animals enrolled. Therefore, we preferred to present only descriptive values for two different ranges of BW. On the other hand, the LA:Ao ratio and LV FS were found to be independent from BW, as observed in dogs [13], cats [14], and rabbits [15]. In this study, aortic dimensions did not show any correlation with BW, as opposed to other species [11,12]. However, indexing LA to aortic diameter likely corrected for some BW effect, and therefore the LA:Ao ratio resulted as weight independent. It is not surprising that LV FS, being a functional parameter rather than a dimensional variable, is not influenced by the weight of the animal. HR showed a negative effect on LVIDs. This variable has been observed to decrease with increasing HR in cats [16], dogs [17], and people [18]. This behavior is likely due to a rate-related increase in myocardial contractility (Bowditch effect), where an increase in the HR (spontaneous or induced by artificial pacing) leads to an accumulation of intracellular calcium, with a subsequent increase in myofiber contraction and shortening. The augmented contractility produces a reduction of end-systolic volume and therefore LV systolic diameter [16]. Age was positively correlated with LV posterior wall thickness at end-diastole and aortic dimension, suggesting a possible progressive increase in cardiac dimensions with the aging process of the animal. Although the presence of subtle cardiomyopathy leading to increased LV wall thickness cannot be excluded, it is quite unlikely that this would be the case for this study, considering the concomitant increase of aortic root diameter, which is usually not observed in other species with myocardial disease. On color Doppler examination, a trivial estimated volume of tricuspid and aortic valve regurgitation was identified in one animal each. Both of the insufficiencies were of minor degree and not considered to be hemodynamically significant. Trace valvular insufficiencies, especially at the tricuspid valve, have been widely reported in healthy individuals of other species, such as cats, dogs, and horses, in which they are considered to be clinically irrelevant [19]. The assessment of diastolic transmitral flow velocities was limited because of the summation of the E and A waves in the majority of animals (77%) because of their rapid HR. In those five animals that showed separate E and A waves, peak
7 velocities of the two were reversed compared with what is considered normal in other species, with the A max having a higher value. The age of these animals was variable (1, 2, 2.5, 3, and 5 years), therefore this cannot be explained by the physiological aging process of myocardial fibers, which, as observed in other species, can lead to a delay in the recoiling of the left ventricle [20]. On the other hand, it has been described in dogs that a progressive increase in the HR produces a gradual reversal of the diastolic transmitral waves owing to decreased diastolic duration and time for rapid early LV filling [21]. Considering the elevated HR recorded in guinea pigs during echocardiography, this finding could at least partially explain the reversal of the expected E max:A max ratio. Finally, it could be that the animals experienced a transitory stress-induced hypertension, which could have temporarily altered the diastolic filling profiles of the LV, delaying its relaxation [19]. Finally, gender variations in the echocardiographic variables showed a difference only in LV systolic function, with males having reduced indices of systolic function (larger LVIDs and lower FS) compared with females. This difference is not explained by BW, as it has been observed in some canine breeds [22], but may be the result of a hormonal effect or of a different response to stressful events between the two sexes. The major limitation of the present study is the low number of guinea pigs enrolled. A larger number of individuals (>120) should be used to establish normal reference values [23]. Enrolling more animals could have allowed a clearer definition of the relationship between BW and echocardiographic variables and the creation of an equation useful to predict reference ranges for each BW [11,12]. However, considering the difficulty in recruiting exotic pets, a larger population could not be used for the present study. In conclusion, transthoracic echocardiography is feasible in conscious, manually restrained guinea pigs, provided that appropriate equipment and settings are used. Normal descriptive intervals for echocardiographic variables should be useful tools in the echocardiographic evaluation of guinea pigs.
Funding This research received no grant from any funding agency in the public, commercial, or not-for-profit sectors.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Conflicts of Interest Statement The authors have no conflicts of interest to report with regard to this study.
[11]
[12]
Acknowledgments Q4
The authors gratefully acknowledge guinea pig owners.
[13]
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Hannover: Schlu ¨tersche Verlagsgesellschaft; 2011. p. 188e223. Cornell CC, Kittleson MD, Della Torre P, Ha ¨ggstro ¨m J, Lombard CW, Pedersen HD, et al. Allometric scaling of Mmode cardiac measurements in normal adult dogs. J Vet Intern Med 2004;18:311e21. ˚ O, Falk T, Nilsfors L, OIsson U, Ha ¨ggstro ¨m J, Andersson A Kresken JG, et al. Effect of body weight on echocardiographic measurements in 19,866 pure-bred cats with or without heart disease. J Vet Intern Med 2016;30:1601e11. O’Grady MR, Bonagura JD, Powers JD, Herring DS. Quantitative cross-sectional echocardiography in the normal dog. Vet Radiol Ultrasound 1986;27:34e49. DeMadron E, Bonagura JD, Herring DS. Two-dimensional echocardiography in the normal cat. Vet Radiol Ultrasound 1985;26:149e58. Casamian-Sorrosal D, Saunders R, Browne WJ, Elliott S, Fonfara S. M-mode, two-dimensional and Doppler echocardiographic findings in 40 healthy domestic pet rabbits. J Vet Cardiol 2014;16:101e8. Sugimoto K, Fujii Y, Ogura Y, Sunahara H, Aoki T. Influence of alterations in heart rate on left ventricular echocardiographic measurements in healthy cats. J Feline Med Surg 2017;19:841e5. Jacobs G, Mahjoob K. Influence of alterations in heart rate on echocardiographic measurements in the dog. Am J Vet Res 1988;49:548e52. DeMaria AN, Neumann A, Schubart PJ, Lee G, Mason DT. Systematic correlation of cardiac chamber size and ventricular performance determined with echocardiography and alterations in heart rate in normal persons. Am J Cardiol 1979;43:1e9. Boon JA. Evaluation of size, function, and hemodynamics. In: Boon JA, editor. Veterinary Echocardiography. Chichester: Wiley-Blackwell; 2011. p. 153e266. Schober KE, Chetboul V. Echocardiographic evaluation of left ventricular diastolic function in cats: hemodynamic determinants and pattern recognition. J Vet Cardiol 2015; 17:S102e33. Schober KE, Fuentes VL. Effects of age, body weight, and heart rate on transmitral and pulmonary venous flow in clinically normal dogs. Am J Vet Res 2001;62:1447e54. Stephenson HM, Fonfara S, Lo ´pez-Alvarez J, Cripps P, Dukes-McEwan J. Screening for dilated cardiomyopathy in Great Danes in the United Kingdom. J Vet Intern Med 2012; 26:1140e7. Friedrichs KR, Harr KE, Freeman KP, Szladovits B, Walton RM, Barnhart KF, et al. ASVCP reference interval guidelines: determination of de novo reference intervals in veterinary species and other related topics. Vet Clin Pathol 2012;41:441e53.
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Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs* M. De Silva, DVMa,*, A. Mihailovic, DVMb, M.B. Toaldo, DVM, PhDa,c a
Department of Veterinary Medical Sciences, Alma Mater Studiorum, University of Bologna, Italy b Bologna, Italy Received 21 August 2019; received in revised form 23 January 2020; accepted 24 January 2020
KEYWORDS Cavia porcellus; Exotics; Cardiology; Ultrasound; Heart Q3
Abstract Objectives: The objective of this study was to report normal echocardiographic values in healthy guinea pigs. Animals, materials, and methods: Twenty-two privately owned, apparently healthy, conscious guinea pigs underwent complete transthoracic echocardiography. Left ventricular (LV), right ventricular, left atrial, and aortic root dimensions were measured, as were forward flow velocities across the mitral, aortic, and pulmonic valves. The effects of age, body weight, sex, and heart rate on these variables were also investigated. Results: The median age (interquartile range) was 3.0 (1.8e4.0) years with a body weight of 902 (822e998) grams. Echocardiography was feasible in all conscious animals. Early and late diastolic transmitral flow waves were summated in 17 of 22 individuals. In the remaining five animals, the two waves were reversed (E waveto-A wave velocity less than 1.0). A positive correlation was detected between body weight and LV internal diameter at end-diastole and end-systole and left atrial diameter (P < .05). Heart rate was negatively correlated with LV internal diameter at end-systole (r ¼ .463, P ¼ .035). Age was positively correlated with LV posterior wall thickness at end-diastole and aortic diameter (P < .05). LV internal diameter at end-systole was larger in males than in females (P ¼ .012), while fractional shortening was lower (P ¼ .008).
* This work was done at the Department of Veterinary Medical Sciences, Alma Mater Studiorum e University of Bologna, Via Tolara di Sopra 50, 40,064, Ozzano Emilia, Italy. * Corresponding author. E-mail address: [email protected] (M. De Silva). c Current address is: Division of Cardiology, Clinic for Small Animal Internal Medicine, Vetsuisse Faculty University of Zu ¨rich.
https://doi.org/10.1016/j.jvc.2020.01.004 1760-2734/ª 2020 Elsevier B.V. All rights reserved.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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M. De Silva et al. Conclusions: Descriptive echocardiography ranges in apparently healthy awake guinea pigs have been provided and can be used for cardiac assessment in these pet animals. ª 2020 Elsevier B.V. All rights reserved.
Abbreviations A max Ao BW FS HR IQR LA LV LVIDs
peak velocity of late diastolic transmitral flow aorta body weight fractional shortening heart rate Interquartile range left atrium left ventricular left ventricular internal diameter at end-systole
Introduction Guinea pigs or cavies (Cavia porcellus) are hystricomorph rodents originally native to South America that have been historically used in biomedical research and that are now becoming increasingly popular as domestic pets in many parts of the world, such as Europe and North America [1,2]. Clinical cases of guinea pigs presented with signs related to cardiac disease have been reported in the literature [3e6]. Echocardiography is, therefore, becoming a useful tool to diagnose and treat cardiac morbidities in these pet animals [3e5]. However, the accuracy of transthoracic echocardiography in diagnosing cardiac diseases in the different species and animal breeds depends also on the availability of appropriate reference limits of normality for each specific population. To the best of the authors’ knowledge, in the scientific literature, there is a lack of normal echocardiographic reference values obtained under standardized conditions in nonanesthetized adult pet guinea pigs. Indeed, the normal echocardiographic values reported in the literature refer in most instances to poorly representative study populations, such as genetically similar laboratory Hartley male guinea pigs, of
unknown age, small body weight (BW), and subject to anesthetic agents [7]. Consequently, these data are not fully applicable to clinical practice in adult conscious pet animals, as the aforementioned extrapolation might be misleading. The objectives of the present study were (1) to assess the feasibility of two-dimensional, M-mode, and Doppler echocardiography in the evaluation of cardiac anatomy, function, and hemodynamics in conscious, apparently healthy pet guinea pigs; (2) to measure standard echocardiographic variables and establish normalreferenceintervalsforeach of them; and (3) to carry out a statistical analysis to investigate correlations between the echocardiographic variables and age, sex, BW, and heart rate (HR).
Animals, materials, and methods Twenty-two adult pet guinea pigs were brought to the Veterinary Teaching Hospital of the University of Bologna by private owners for the purpose of health consultation and cardiac assessment from April 2016 to November 2019. All animals were apparently healthy at clinical examination, with no preexisting history of systemic or cardiovascular illness as reported by the owners. Each animal underwent a full clinical examination carried out by a veterinarian with experience in exotic animal medicine (A.M.) and were auscultated by a boardcertified veterinary cardiologist (M.B.T.). All animals were unsedated and restrained on a standard echocardiographic table in left and right lateral recumbency by one trained exotic veterinarian (A.M.) and one trained assistant (M.D.S.). The region corresponding to the cardiac apex, between the third and fifth intercostal spaces, was shaved and a small amount of water-based, previously warmed ultrasound gel was applied to the area. Echocardiograms were performed with guinea pigs in lateral recumbency on a table with a cutout, contacting the thoracic body wall from below. Examination of all guinea pigs was possible without the use of sedation. This was achieved by gentle but confident handling by the pectoral
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Fig. 1 M-mode echocardiogram obtained from a right parasternal short-axis view at the level of the papillary muscles. A leading edge-to-leading edge method was used to measure right and left ventricular internal diameters and wall thickness. Dia: diastole; IVS: interventricular septum; LV: left ventricle; LVPW: left ventricular posterior wall; RV: right ventricle; Sys: systole.
girdle and the hind limbs. The procedure was well tolerated by all animals examined. Echocardiographic examinations were performed using a dedicated diagnostic ultrasound unitd equipped with a phased array probe of variable frequency (12-4 MHz) with an aperture of 9.78 mm, without continuous electrocardiographic monitoring. Standard transthoracic two-dimensional, M-mode, and Doppler ultrasound examinations were performed by a veterinary cardiologist (M.B.T.) following a standardized protocol as described in the literature for the dog [8], by means of right parasternal (long- and short-axis) and left apical (fourand five-chamber) views. Frame rates were maintained between 50 and 100 frames per second during two-dimensional echocardiography and more than 120 frames per second during M-mode studies. Images and video clips were acquired and analyzed at the end of the study using dedicated softwaree by the same operator (M.B.T.). Each measurement was derived from three consecutive cardiac beats, and then the mean value was calculated. Right ventricular and left ventricular (LV) internal diameters and wall thickness were measured from M-mode images obtained from a right
d e
iE33 ultrasound system, Philips Healthcare, Monza, Italy. OsiriX MD Software, Pixeo SARL, Geneva, Switzerland.
parasternal short-axis view at the level of the papillary muscles, with the cursor crossing the midventricle and using a leading edge-to-leading edge method (Fig. 1). Specifically, the following variables were measured: right ventricular internal diameter at end-diastole, interventricular septal thickness at end-diastole and at end-systole, LV internal diameter at end-diastole and at endsystole (LVIDs), and LV posterior wall thickness at end-diastole and at end-systole. Because electrocardiographic monitoring was not performed, the timing for end-diastole and end-systole was considered at the moment of maximal and minimal (apogee of the LV free wall) LV dimensions, respectively. Fractional shortening (FS), expressed as a percentage value, was then calculated. The diameters of the left atrium (LA) and of the aorta (Ao) at the level of the aortic annulus were obtained from a right parasternal short-axis view at the level of the cardiac base using an inner edge-to-inner edge method and following a technique proposed for dogs (Fig. 2) [9]. The LA:Ao ratio was then calculated by dividing the two diameters. Color Doppler examination was carried out at the level of each of the four cardiac valves to detect regurgitation, while pulsed wave spectral Doppler analysis was performed for forward flow velocity measurements only at the level of the Ao (Fig. 3), pulmonary valve, and mitral valve, using
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Fig. 2 Two-dimensional echocardiographic views and measurements. (A): Right parasternal long-axis four-chamber view; (B): right parasternal left ventricular outflow view; (C): right parasternal short-axis view at the level of the papillary muscles of the left ventricle; (D): right parasternal short-axis view at the heart base. The dashed lines indicate the measurements of the aortic root diameter and the left atrial diameter using an inner edge-to-inner edge method. Ao: aorta; LA: left atrium; LAu: left auricle; LV: left ventricle; RA: right atrium; RV: right ventricle.
Fig. 3 Pulsed wave spectral Doppler analyses of transaortic and transmitral flows. (A): Pulsed wave Doppler analysis of the monophasic transaortic systolic flow. The dotted line indicates the interval between two systolic flows used to calculate the heart rate. (B): Pulsed wave Doppler analysis of the transmitral biphasic flow showing separate early (E) and late (A) diastolic flow waves. Note the inverted ratio of their peak velocities. Ao: aorta.
a sample volume of 2 mm. In particular, peak aortic and pulmonary flow velocities were measured from a left apical five-chamber view and right parasternal short-axis view at the basilar level, respectively. Peak velocities of early (E max) and late (A max) diastolic transmitral flow were measured from a left apical four-chamber view. Valvular regurgitation was considered trivial if it extended minimally from the valvular plane (2e3 mm). The echocardiography-derived HR was measured from the aortic spectral flow profile.
Statistical analysis All statistical analyses were carried out using dedicated softwaref,g Normality of distribution of continuous variables was assessed by visual inspection of the plots. Since data were not normally distributed, all variables were expressed as
f Microsoft Excel; Microsoft Office 2011, Microsoft Corporation, Bellevue, WA. g Prism 7, GraphPad Software Inc., San Diego, CA.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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median (interquartile range [IQR]), range (minimumemaximum), and 90% upper and lower confidence intervals. Continuous variables were compared between sexes with a ManneWhitney test. The distribution of sexes between two classes of BW was tested with a Fisher’s exact test. Spearman’s correlation was used to evaluate the relationship between echocardiographic variables and age, BW, and HR. A P < .05 was considered significant.
Results The study population included 8 female and 14 male guinea pigs, weighing a median of 902 (822e998) grams, with a median age of 3.0 (1.8e4.0) years. Echocardiographic examinations were feasible in every animal without need for sedation. Each examination was of adequate quality to allow a clear visualization of the myocardium, valves, atria, and ventricles and to ensure accurate measurement for all variables. All data referring to the whole population are reported in Table 1. Color Doppler examination of one Table 1
male animal of 1 year of age revealed trivial tricuspid regurgitation, while another male guinea pig of 5 years of age had trivial aortic insufficiency. All other flow was judged to be laminar from spectral Doppler evaluation. Transmitral flow waves were fused in the majority of animals (17/ 22, 77%) because of the elevated HR. In the remaining five animals, A max exceeded E max (Fig. 3). Among the tested variables, LVIDs was larger in males (median ¼ 7.2 mm [IQR ¼ 6.5e7.8; 6.1e10.7 {minimumemaximum}]) than in females (median ¼ 6.1 mm [IQR ¼ 5.8e6.8; 4.5e7.4]) (P ¼ .012), while the FS value was lower in males (median ¼ 31% [IQR ¼ 26e33; 16e42]) than in females (median ¼ 33% [IQR ¼ 33e42; 33e50]) (P ¼ .008). Aortic flow peak systolic velocity was higher in females (median ¼ 91 cm/s [IQR ¼ 81e101; 73e132]) than in males (82 cm/s [IQR ¼ 69e87; 59e107]) (P ¼ .044). There was a positive correlation between age and LV posterior wall thickness at end-diastole (r ¼ .478, P ¼ .028) and aortic dimension (r ¼ .474, P ¼ .030). A negative correlation was found between HR and LVIDs (r ¼ .463, P ¼ .035). Finally, LV internal diameter at end-diastole
Demographic and echocardiographic data obtained from 22 healthy pet guinea pigs.
Variable Age (years) BW (grams) HR (bpm) RVDd (mm) IVSd (mm) LVIDd (mm) LVPWd (mm) IVSs (mm) LVIDs (mm) LVPWs (mm) FS (%) LA (mm) Ao (mm) LA:Ao EA max (cm/sec)a E max (cm/sec)b A max (cm/sec)b PV vel (cm/sec) Ao vel (cm/sec)
Median (IQR)
Range (minimumemaximum)
Lower 90% CI
Upper 90% CI
3.0 (1.8e4.0) 902 (822e998) 268 (238e287) 3.4 (2.8e4.2) 2.1 (1.9e2.3) 10.3 (9.3e10.8) 2.0 (1.8e2.1) 2.8 (2.6e3.1) 6.8 (6.1e7.4) 2.9 (2.7e3.2) 32 (29e38) 6.9 (6.4e7.6) 5.8 (5.3e6.1) 1.2 (1.1e1.4) 73 (65e89) 43 (38e58) 75 (51e84) 84 (64e96) 83 (72e93)
1.0e7.0 500e1200 168e316 2.2e5.5 1.7e2.4 8.7e12.7 1.6e2.4 2.3e4.1 4.5e10.7 2.4e4.2 16e50 5.3e8.6 4.6e7.3 .7e1.6 47e113 38e65 44e85 40e145 57e132
2.4 841 250 3.2 2.0 9.8 1.9 2.8 6.5 2.9 30 6.6 5.5 1.1 70 39 56 73 78
3.5 947 275 3.8 2.2 10.6 2.0 3.1 7.3 3.2 35 7.2 6.0 1.3 84 55 82 90 90
Ao: aorta; bpm: beats per minute; BW: body weight; CI: confidence interval; A max: peak velocity of late diastolic transmitral flow; E max: peak velocity of early diastolic transmitral flow; EA max: peak velocity of summated E and A transmitral waves; FS: fractional shortening; HR: heart rate; IQR: interquartile range; IVSd: interventricular septum thickness at end-diastole; IVSs: interventricular septum thickness at end-systole; LA: left atrium; LVIDd: left ventricular internal diameter at end-diastole; LVIDs: left ventricular internal diameter at end-systole; LVPWd: left ventricular posterior wall thickness in diastole; LVPWs: left ventricular posterior wall thickness in systole; PV: pulmonary valve; RVDd: right ventricular internal diameter in diastole; vel: velocity. a Number of animals ¼ 17. b Number of animals ¼ 5.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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CI: confidence interval; HR: heart rate; IQR: interquartile range; LA: left atrium; LVIDd: left ventricular internal diameter at end-diastole; LVIDs: left ventricular internal diameter at end-systole. a No difference between classes of body weight regarding sex distribution (Fisher’s exact test). b No difference between classes of body weight (ManneWhitney test).
4.4 281 11.5 8.2 7.7 2.7 230 10.5 6.7 7.0 3 (2.5e4) 262 (223e295) 10.8 (10.4e12) 7.2 (6.3e7.9) 7.6 (6.8e7.8) 3.2 281 9.9 6.8 6.7 1.6 267 9.1 5.9 6.0 1e5 231e300 8.7e10.7 4.5e7.4 5.3e7.2 2 (1e4) 273 (250e286) 9.4 (8.8e10.2) 6.7 (6e6.9) 6.6 (5.8e6.9)
Lower 90% CI Range (minimumemaximum)
1.3e7 168e316 9.8e12.7 6e10.7 6.6e8.6
Upper 90% CI Range (minimumemaximum)
Lower 90% CI
Age (years)b HR (bpm)b LVIDd (mm) LVIDs (mm) LA (mm)
Variable
Median (IQR)
Body weight 900gr (11 animals: 5 males, 6 females)a
Upper 90% CI
Median (IQR)
Body weight > 900gr (11 animals: 9 males, 2 females)a
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Selected echocardiographic data obtained from 22 healthy pet guinea pigs divided on the basis of body weight.
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(r ¼ .776, P < .001), LVIDs (r ¼ .443, P ¼ .044), and LA diameter (r ¼ .519, P ¼ .016) were positively correlated with BW. Owing to the effect of BW on some echocardiographic variables, descriptive reference intervals for two ranges of BW ( 900gr and >900gr) are provided in Table 2.
Discussion The present study showed that echocardiography in unsedated guinea pigs is feasible and allows clear visualization of all cardiac structures. This was likely facilitated by the use of a high-frequency transducer with adequate frame rate capability and by following the general procedural recommendations for small exotic mammals [10]. The study cohort comprised adult (>1 year of age) individuals of various BW and ages with a uniform distribution of both sexes, and therefore reflected a representative guinea pig population commonly seen in clinical practice. This makes the reported echocardiographic values useful for exotic animal specialists and cardiologists performing echocardiography in this species. A previous echocardiographic study included a population of genetically similar male laboratory guinea pigs of unknown age, smaller BW, and which were anesthetized with ketamine and xylazine [7]. Therefore, the aforementioned study is not fully applicable to clinical practice, as the extrapolation of the results may have limitations. As a matter of fact, measurements of cardiac dimensions obtained in our study have higher values than those reported in previously published studies [7]. Moreover, Doppler examination was not performed in the aforementioned study, while such data are available in the present one. A significant correlation between LV internal diameter at end-diastole, LVIDs, and LA with BW was detected, and this finding is consistent (except for LVIDs) with a previous echocardiographic study carried out on anesthetized guinea pigs [7]. The influence of BW on echocardiographic estimates of cardiac dimensions has already been observed in other species, such as dogs and cats [11,12]. Allometric scaling best describes the association between BW and some echocardiographic variables in these species, with curvilinear regression lines used for predicting reference ranges and with a scaling exponent usually set around .3 for both dogs and
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Echocardiography in healthy guinea pigs cats [11,12]. In the present study on guinea pigs, we did not perform regression analysis because of the small number of animals enrolled. Therefore, we preferred to present only descriptive values for two different ranges of BW. On the other hand, the LA:Ao ratio and LV FS were found to be independent from BW, as observed in dogs [13], cats [14], and rabbits [15]. In this study, aortic dimensions did not show any correlation with BW, as opposed to other species [11,12]. However, indexing LA to aortic diameter likely corrected for some BW effect, and therefore the LA:Ao ratio resulted as weight independent. It is not surprising that LV FS, being a functional parameter rather than a dimensional variable, is not influenced by the weight of the animal. HR showed a negative effect on LVIDs. This variable has been observed to decrease with increasing HR in cats [16], dogs [17], and people [18]. This behavior is likely due to a rate-related increase in myocardial contractility (Bowditch effect), where an increase in the HR (spontaneous or induced by artificial pacing) leads to an accumulation of intracellular calcium, with a subsequent increase in myofiber contraction and shortening. The augmented contractility produces a reduction of end-systolic volume and therefore LV systolic diameter [16]. Age was positively correlated with LV posterior wall thickness at end-diastole and aortic dimension, suggesting a possible progressive increase in cardiac dimensions with the aging process of the animal. Although the presence of subtle cardiomyopathy leading to increased LV wall thickness cannot be excluded, it is quite unlikely that this would be the case for this study, considering the concomitant increase of aortic root diameter, which is usually not observed in other species with myocardial disease. On color Doppler examination, a trivial estimated volume of tricuspid and aortic valve regurgitation was identified in one animal each. Both of the insufficiencies were of minor degree and not considered to be hemodynamically significant. Trace valvular insufficiencies, especially at the tricuspid valve, have been widely reported in healthy individuals of other species, such as cats, dogs, and horses, in which they are considered to be clinically irrelevant [19]. The assessment of diastolic transmitral flow velocities was limited because of the summation of the E and A waves in the majority of animals (77%) because of their rapid HR. In those five animals that showed separate E and A waves, peak
7 velocities of the two were reversed compared with what is considered normal in other species, with the A max having a higher value. The age of these animals was variable (1, 2, 2.5, 3, and 5 years), therefore this cannot be explained by the physiological aging process of myocardial fibers, which, as observed in other species, can lead to a delay in the recoiling of the left ventricle [20]. On the other hand, it has been described in dogs that a progressive increase in the HR produces a gradual reversal of the diastolic transmitral waves owing to decreased diastolic duration and time for rapid early LV filling [21]. Considering the elevated HR recorded in guinea pigs during echocardiography, this finding could at least partially explain the reversal of the expected E max:A max ratio. Finally, it could be that the animals experienced a transitory stress-induced hypertension, which could have temporarily altered the diastolic filling profiles of the LV, delaying its relaxation [19]. Finally, gender variations in the echocardiographic variables showed a difference only in LV systolic function, with males having reduced indices of systolic function (larger LVIDs and lower FS) compared with females. This difference is not explained by BW, as it has been observed in some canine breeds [22], but may be the result of a hormonal effect or of a different response to stressful events between the two sexes. The major limitation of the present study is the low number of guinea pigs enrolled. A larger number of individuals (>120) should be used to establish normal reference values [23]. Enrolling more animals could have allowed a clearer definition of the relationship between BW and echocardiographic variables and the creation of an equation useful to predict reference ranges for each BW [11,12]. However, considering the difficulty in recruiting exotic pets, a larger population could not be used for the present study. In conclusion, transthoracic echocardiography is feasible in conscious, manually restrained guinea pigs, provided that appropriate equipment and settings are used. Normal descriptive intervals for echocardiographic variables should be useful tools in the echocardiographic evaluation of guinea pigs.
Funding This research received no grant from any funding agency in the public, commercial, or not-for-profit sectors.
Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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Conflicts of Interest Statement The authors have no conflicts of interest to report with regard to this study.
[11]
[12]
Acknowledgments Q4
The authors gratefully acknowledge guinea pig owners.
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Please cite this article as: De Silva M et al., Two-dimensional, M-mode, and Doppler echocardiography in 22 conscious and apparently healthy pet guinea pigs, Journal of Veterinary Cardiology, https://doi.org/10.1016/j.jvc.2020.01.004
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