A cross-species validation of single-beat metrics of cardiac contractility

Mehdi Ahmadian; Alexandra M Williams; Joseph Mannozzi; Filip Konecny; Ryan L Hoiland; Liisa Wainman; Erin Erskine; Jennifer Duffy; Neda Manouchehri; Kitty So; Keerit Tauh; Javier A Sala-Mercado; Katelyn Shortt; Shera Fisk; Kyoung-Tae Kim; Femke Streijger; Glen E Foster; Brian K Kwon; Donal S O’Leary; Christopher R West

doi:10.1113/JP283319

. Author manuscript; available in PMC: 2023 Nov 1.

Published in final edited form as: J Physiol. 2022 Oct 5;600(22):4779–4806. doi: 10.1113/JP283319

A cross-species validation of single-beat metrics of cardiac contractility

Mehdi Ahmadian ^1,^2,³, Alexandra M Williams ^2,⁴, Joseph Mannozzi ⁵, Filip Konecny ^6,⁷, Ryan L Hoiland ^2,^4,⁸, Liisa Wainman ^2,^3,⁴, Erin Erskine ^2,^3,⁴, Jennifer Duffy ^2,^3,⁴, Neda Manouchehri ², Kitty So ², Keerit Tauh ², Javier A Sala-Mercado ⁵, Katelyn Shortt ², Shera Fisk ², Kyoung-Tae Kim ^2,⁹, Femke Streijger ², Glen E Foster ¹⁰, Brian K Kwon ², Donal S O’Leary ⁵, Christopher R West ^2,^3,^4,^*

¹School of Kinesiology, Faculty of Education, University of British Columbia, Vancouver, BC, Canada

²International Collaboration on Repair Discoveries, University of British Columbia, Vancouver, BC, Canada

³Centre for Chronic Disease Prevention and Management, University of British Columbia, Kelowna, BC, Canada

⁴Department of Cellular and Physiological Sciences, Faculty of Medicine, University of British Columbia, Vancouver, BC, Canada

⁵Department of Physiology, School of Medicine, Wayne State University, Detroit, MI 48009

⁶Transonic Scisense Inc., London, ON, Canada

⁷MaRS Centre Toronto Medical Discovery Tower, 3rd Floor, 101 College Street, M5G 1L7, Toronto, Ontario, Canada

⁸Department of Anesthesiology, Pharmacology and Therapeutics, Faculty of Medicine, Vancouver General Hospital, 899 West 12th Avenue, University of British Columbia, Vancouver, BC, Canada, V5Z 1M9

⁹Department of Neurosurgery, School of Medicine, Kyungpook National University Hospital, Daegu, South Korea.

¹⁰Centre for Heart, Lung, & Vascular Health, School of Health and Exercise Sciences, University of British Columbia Okanagan, 3333 University Way, Kelowna, BC, V1V1V7

Author contributions

All experiments were performed at three different universities across North America, including the University of British Columbia, Canada, the University of Health Network, Canada, and Wayne State University, USA. MA and CRW conceived the project. MA, AMW, JM, FK, LW, EE, NM, KS, KT, JAS, KS, SF, KK, and FS performed experiments or were involved in data collection. MA, CRW, and JD performed or contributed to data analysis. DSO, GEF, BK, and RLH contributed to the critical discussion. MA and CRW wrote the paper. All authors have approved the final version of the manuscript and agree to be accountable for all aspects of the work. All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed.

Correspondence: Dr. Christopher R. West, International Collaboration on Repair Discoveries – Blusson Spinal Cord Centre, 818 West 10th Avenue, Vancouver, BC, Canada, V5Z 1M9, Tel: 250-807-8891, chris.west@ubc.ca

PMCID: PMC9669232 NIHMSID: NIHMS1836892 PMID: 36121759

Abstract

The assessment of LV contractility in animal models is useful in various experimental paradigms, yet obtaining such measures is inherently challenging and surgically invasive. In a cross-species study using small and large animals, we comprehensively tested the agreement and validity of multiple single-beat surrogate metrics of left ventricular (LV) contractility against the field-standard metrics derived from inferior vena cava occlusion (IVCO). Fifty-six rats, 27 minipigs, and 11 conscious dogs underwent LV and arterial catheterization and were assessed for a range of single-beat metrics of LV contractility. All single-beat metrics were tested for various underlying assumptions required to be considered a valid metric of cardiac contractility, including load-independency, sensitivity to inotropic stimulation, and ability to diagnose contractile dysfunction in cardiac disease. Of all examined single-beat metrics, only LV maximal pressure normalized to end-diastolic volume (P_max-EDV), end-systolic pressure normalized to EDV (_LVESP-EDV), and the maximal rate of rise of the LV pressure normalized to EDV (_LVdP/dt_max-EDV) showed a moderate-to-excellent agreement with their IVCO-derived reference measure and met all the underlying assumptions required to be considered as a valid cardiac contractile metric in both rodents and large animal models. Our findings demonstrate that single-beat metrics can be used as a valid, reliable method to quantify cardiac contractile function in basic/preclinical experiments utilizing small and large animal models.

Keywords: cardiac inotropy, single-beat assessment, animal models

Graphical Abstract

The classical assessment of cardiac contractility in animal models requires caval occlusion that can be inherently challenging to perform reliably. In a cross-species study, we test whether multiple single-beat metrics of contractility can meet all the underlying assumptions required to be considered as a valid metric of contractility in small (male rats) and large animal (female pigs and dogs) models. We found that only LV maximal pressure normalized to end-diastolic volume (P_max-EDV), end-systolic pressure normalized to EDV (_LVESP-EDV), and the maximal rate of rise of the LV pressure normalized to EDV (_LVdP/dt_max-EDV) can be used in both small and large animal models as a reliable, alternative tool to quantify cardiac contractile function.

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INTRODUCTION

The assessment of cardiac contractility is integral for determining how different pathologies impact the heart and the efficacy of cardio-therapeutic interventions. Clinically, ejection fraction (EF) is the most widely utilized assessment of cardiac systolic function. Notwithstanding being a strong independent predictor of mortality in those with cardiovascular disease (Curtis et al., 2003; Weir et al., 2006), EF can fail to accurately identify myocardial contractile dysfunction. For instance, the ability of EF to detect left ventricular (LV) systolic dysfunction has recently been questioned in patients with septic shock (Boissier et al., 2017).

Direct and accurate measures of cardiac contractile function can be obtained in small and large animal preparations in vivo by performing LV pressure-volume (PV) catheterization (Pacher et al., 2008). In addition to LV hemodynamic parameters under steady-state conditions, a number of metrics associated with cardiac inotropy can be obtained during inferior vena cava occlusions (IVCO) (Burkhoff et al., 2005). These metrics include end-systolic elastance (E_es(IVC)), pre-load recruitable stroke work (PRSW_(IVC)), and the slope of the relationship between the maximal rate of rise of LV pressure (dP/dt_max) and EDV (dP/dt_max-EDV_(IVC)) (Suga et al., 1973; Glower et al., 1985; Little, 1985; Burkhoff et al., 2005). In small animals, however, IVC isolation and compression/occlusion requires additional surgical time, technical proficiency, has an associated risk of calamitous bleeding, and often causes post-occlusion hemodynamic instability. Although these limitations can be mitigated in large animals by placing occlusion catheters inside the vena cava, this approach requires additional time for surgery, and live fluoroscopic imaging. Moreover, whether the IVC is occluded externally or internally, it can be challenging to precisely control the degree/speed of occlusion, which increases the risk for impairing coronary perfusion, which in turn may elicit cardiomyocyte ischemic injury and arrhythmia (Barber, 1983; Heusch, 2019). Finally, a potential limitation with the interpretation of contractile indices obtained from an IVCO is that the IVCO itself can elicit reflex sympathoexcitation which increases cardiac contractility. Consequently, identifying methods to assess cardiac contractility that avoid IVCO would offer considerable advantages for the field.

Multiple single-beat metrics of LV contractility that do not require IVCO have been proposed/tested (Sagawa et al., 1977; Takeuchi et al., 1991; Kass & Beyar, 1991; Senzaki et al., 1996; Nakayama et al., 1998; Karunanithi & Feneley, 2000; Shishido et al., 2000; Abel, 2001; Chen et al., 2001; Blaudszun et al., 2013; García et al., 2019; Bombardini et al., 2021). Though these studies have generally reported promising findings, their interpretations are limited by relatively small sample sizes, variability in study populations and designs, or require an advanced mathematical pipeline for analyses that is not readily available in most settings (Sagawa et al., 1977; Takeuchi et al., 1991; Kass & Beyar, 1991; Senzaki et al., 1996; Nakayama et al., 1998; Karunanithi & Feneley, 2000; Shishido et al., 2000; Abel, 2001; Chen et al., 2001; Blaudszun et al., 2013; García et al., 2019; Bombardini et al., 2021). Most notably, these studies have typically validated only a single metric in one species and have generally failed to test the full range of assumptions required to be considered a valid measure of cardiac contractile function (i.e., load-independence, increases with beta-adrenergic stimulation, is reduced in animal models of with overt contractile dysfunction), which have perhaps contributed to the lack of widespread consensus on their use. Here, in a cross-species study, we comprehensively test whether the majority of previously reported/tested single-beat surrogates, together with two new proposed metrics (see methods), meet the assumptions required to be considered as valid/reliable metrics of contractile function.

In experiment 1, we assessed the agreement between single-beat metrics of cardiac contractility and IVCO-derived reference measures in rodents. In experiment 2, we assessed how our single-beat metrics behave in response to lower-body negative pressure (LBNP, i.e., reduced loading) manipulations in rats. In experiments 3 and 4, we examined whether these metrics can detect contractile dysfunction in two rodent models with overt contractile dysfunction, namely, a high-thoracic spinal injury model to mimic situations of lost bulbospinal sympathetic control over the heart, and a chronic myocardial infarction model. In experiments 5 and 6, we tested whether agreements observed in rodents can be translated to a porcine model while also testing the behavior of the examined metrics to an inotropic stimulus (i.e., dobutamine). Lastly, in experiments 7 and 8, using a conscious canine model we tested whether the examined metrics are capable of diagnosing weakened contractile function during transition to and following achievement of systolic heart failure (i.e., longitudinal tracking).

METHODS

Ethical approval

All experimental protocols and procedures were conducted in strict accordance with Canadian Council on Animal Care policies and the NIH Guidelines for the Care and Use of Laboratory Animals; ethical approval was obtained from the University of British Columbia Animal Care Committee (Rat = A18-0344; Pig = A16-0311), the Wayne State University Institutional Animal Care and Use Committee (IACUC, protocol number 19-11-1493), and the Institutional Animal Care and Use Committee of University Health Network (1999.1). The authors also confirm that they understand the ethical principles under which the journal operates and that their work conforms to the principles and regulations described in the Editorial by Grundy (2015).

Animals and experiments

For experiment 1, 30 adult (10 weeks) male Wistar rats (300–350g; Charles River Laboratories, Wilmington, MA, USA) were used to examine the agreement between metrics of interest. For experiment 2, eight rats from experiment 1 were subjected to LBNP manipulations to examine how these metrics behave in response to cardiac unloading. For experiment 3, 16 adult (10 weeks) male Wistar rats (300–350g; Charles River Laboratories, Wilmington, MA, USA) were used, of which eight were subjected to a chronic high-thoracic spinal cord injury (SCI) to induce marked contractile dysfunction via almost fully removing the major bulbospinal sympathetic outflow to the heart (Squair et al., 2017), and eight remained as uninjured controls. For experiment 4, 10 adult (10 weeks) male Sprague Dawley rats (300–370g; Charles River Laboratories, Wilmington, MA, USA) were used, in five of which myocardial infarction was induced by ligation of left anterior descending (LAD) coronary artery, and five remained as intact control. For experiment 5, 17 adult (2–3 months) female Yucatan mini-pigs (20–25kg; S&S Farms, Ramona, CA, USA) were used to test if the observed agreement in rodents can be translated to a large animal model. For experiment 6, 10 adult (2–3 months) female Yucatan mini-pigs (20–25kg; S&S Farms, Ramona, CA, USA) were used to test how the metrics of interest behave in the presence of a direct inotropic stimulus (i.e., dobutamine). For experiment 7, seven adult (1–2 years) mongrel dogs (1 male, 10 female) (20–25kg; Marshall BioResources, North Rose, NY, USA) were used to test the ability of examined metrics to reveal weakened cardiac contractile function following heart failure induction (dilated cardiomyopathy via sustained rapid ventricular pacing). For experiment 8, in four separate female adult (1–2 years) mongrel dogs (20–25kg; Marshall BioResources, North Rose, NY, USA) we longitudinally tested whether our single-beat metrics of contractility revealed weakened cardiac contractile function during transition to heart failure. Animals were provided with environmental enrichment, water ad libitum, and food. All of these animals were used in experiments designed for other purposes, but all underwent our standard 10-minute recording of baseline PV and hemodynamic indices, as well as IVCOs prior to any experimental manipulations.

Experiment 1: Examining the agreement between examined metrics in rats

General preparation

Rodents were anesthetized with urethane (1.6 ± 0.31 g/kg⁻¹ _I.P.). The hind-paw withdrawal and the corneal reflexes were employed to test for an adequate anesthesia level prior to commencing the surgery. The adequacy of anesthesia during surgery was further tested by monitoring acute changes in blood pressure (BP, increase/decrease in 10 mmHg) in response to noxious stimuli (tail or paw pinch). A solid-state pressure catheter (1.6F, Transonic, Ithaca, NY, USA) and a polyethylene catheter (PE-50 tubing, Intramedic™, BD) were then inserted in the femoral artery and vein to continuously record arterial blood pressure (BP) and infuse fluid, respectively. The animals were then further instrumented with a PV catheter to measure in vivo LV function. All animals were intubated and mechanically ventilated (VentElite, Harvard Apparatus, Holliston, MA, USA). Breathing frequency and tidal volume for each animal were calculated using previously described formulae⁸. Core temperature was maintained within a normal range (37.0 ± 0.5°C) using a servo-controlled heating pad (RightTemp® Temperature Monitor & Homeothermic Warming Control Module, Kent Scientific, Torrington, CT, USA). Upon experiment completion, animals were overdosed with an intraperitoneal/intravenous injection of chloral hydrate and euthanasia was confirmed by cessation of breathing and vital hemodynamic signals, followed by puncturing the diaphragm.

Cardiovascular catheterization and data acquisition

The LV catheterization was performed via a closed-chest, right common carotid artery approach using a PV admittance catheter (1.9F, Transonic, Ithaca, NY, USA), which is described in detail in our prior publications (Squair et al., 2018; Poormasjedi-Meibod et al., 2019). Optimal catheter placement was verified by high-frequency ultrasound imaging (Vevo 3100, Visualsonics, Canada). Following a 30-minute hemodynamic stabilization period, baseline PV indices were assessed. To derive reference measures of LV contractility we manipulated cardiac preload using an IVCO maneuver. To do so, a midline laparotomy was performed and the inferior vena cava isolated immediately caudal to the liver. Occlusions were then performed using either a cotton-tipped applicator or by tightening the suture already placed under the IVC. A solid-state pressure catheter (2F, Transonic, Ithaca, NY, USA) was also inserted in a femoral artery to allow for continuous monitoring of arterial BP. Transducers were connected in series to an analog-to-digital converter (1kHz/s sampling rate; PowerLab, ADInstruments, USA), which allowed for real-time monitoring of cardiovascular function via commercially available software (LabChart, Version 8.1; AD Instruments, USA). For each rat, we allowed for a 10-minute period of baseline hemodynamic recordings, the last 30 seconds of which was used to extract and calculate beat-by-beat PV as well as arterial indices. We additionally measured E_es(IVC), PRSW_(IVC) and dP/dt_max-EDV_(IVC) from the IVCO (Figure 1, see also section “Calculation of field-standard metrics of cardiac contractility derived from the IVCO”) using the PV analysis add-On in LabChart Pro software. From the baseline PV and arterial data, we then calculated single-beat surrogate metrics of cardiac contractility offline (see Table 1). All volume signals from the Transonic ADV500 PV system were calibrated with ultrasound-derived stroke volume (SV) obtained by tracing the borders of the LV in the long-axis parasternal imaging view.

Figure 1. — **(A)** E_es, end-systolic elastance, the slope of the end-systolic pressure-volume relationship (ESPVR); **(B)** *PRSW*, pre-load recruitable stroke work, the slope of the stroke work and end-diastolic volume (EDV) relationship; **(C)** *dP/dt*_max-EDV, the slope of the relationship between the maximal rate of rise of ventricular pressure (dP/dt_max) and EDV. Note that data were initially analyzed in LabChart and then reconstructed in GraphPad Prism.

Table 1.

Definitions and calculations of single-beat metrics examined.

Metric	Single-beat metric calculation	Definition and calculation of measurements used in each single-beat metric	Reference
*Surrogates for E* _es

E_es(SB1) E_es(SB2)	_LVESP normalized to ESV _ARTESP (0.9 × SBP) normalized to ESV	ESP is defined as the pressure at the point at which aortic valve closure occurs and ventricular relaxation initiates. In this manuscript, end-systole points were calculated as the maximum pressure/volume ratio during each cardiac cycle in LabChart using the PV loop-Add-On. Pressure and volume at this point were then used by the software to estimate ESP and ESV. Please note that the same method was used to estimate ESP for other contractility indices requiring ESP measurement.	Sagawa et al (1977), Grosu et al (2005)

*Surrogates for PRSW*

PRSW_(SB)	SW normalized to EDV	SW is the work performed by the ventricle to eject blood to the peripheral circulation during one cardiac cycle. In this manuscript, SW was calculated as the area enclosed by the pressure-volume loop using Green’s theorem in LabChart using the PV loop-Add-On. EDV is defined as the volume at end-diastole, which corresponds to the end of ventricular relaxation. In this manuscript, end-diastolic points were determined automatically from the pressure signal in LabChart using the PV loop-Add-On. Please note that the same method was used to estimate EDV for all the metrics examined.	Bombardini et al (2021)

PWR_max-EDV PWR_max-EDV²	PWR_max normalized to EDV PWR_max normalized to EDV²	Power is referred to the instantaneous product of pressure and rate of volume change. PWR_max is a measure derived during the ejection phase in each cardiac cycle and corresponds to the rate of ventricular work. In this work, PWR_max was calculated from each cardiac pressure-volume loop in LabChart using the PV loop-Add-On as the maximum value from the product of instantaneous ventricular pressure and rate of volume change (dV/dt).	Kass & Beyar (1991), Nakayama et al (1998)

LVEF_EA1 LVEF_EA2	$\sqrt{({}_{L V}{E S P} \times S V) /} E D V$	The methods used to calculate ESP and EDV for these metrics were analogous to previous metrics. SV was calculated in LabChart using the PV loop-Add-On via stable EDV minus ESV method, which is calculated as the mean volume during isovolumic contraction minus the mean volume during isovolumic relaxation.	García et al (2019)
LVEF_EA1 LVEF_EA2	$\sqrt{({}_{A R T}{E S P} \times S V}) / E D V$		García et al (2019)

*Surrogates for dP/dt* _max *-EDV*

_LVdP/dt_max-EDV _ARTdP/dt_max- EDV	LVdP/dt_max normalized to EDV _ARTdP/dt_max normalized to EDV	dP/dt_max is the maximal rate of rise of ventricular or arterial pressure. Left ventricular dP/dt_max in this manuscript was calculated in LabChart using the PV loop-Add-On as the maximum value of the LV pressure derivative. Arterial dP/dt_max was calculated as the average cyclic maximum derivative of arterial pressure using the Data Pad feature in LabChart.	Blaudszun et al (2013)
_LVdP/dt_max-EDV _ARTdP/dt_max- EDV			Garcia et al (2018)

P_max-EDV	P_max normalized to EDV	The term P_max is defined as the highest LV pressure during isovolumetric contraction. P_max in this work was calculated as the maximum pressure during each cardiac cycle in LabChart using the PV loop-Add-On. Note that this method for measuring LV P_max is somewhat different from the original metric proposed by Abel, where P_max was considered as the highest LV pressure during isovolumetric contraction.	Abel (2001)

_LVESP-EDV _ARTESP-EDV	_LVESP normalized to EDV _ARTESP normalized to EDV	The methods used to calculate ESP and EDV for these metrics were analogous to previous metrics.	Proposed in the present study; Please refer to the text

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E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), Arterial end-systolic pressure (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; LVEF_EA1, afterload adjusted LV ejection fraction using ventricular data; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data. PRSW_(SB), stroke work and end-diastolic volume (EDV) ratio; PWR_max-EDV, maximal power (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular (LV) pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max- EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure and EDV ratio; _LVESP-EDV₁, _LVESP and EDV ratio; _ARTESP-EDV₂, _ARTESP (0.9 × SBP) and EDV ratio; SV, stroke volume

Experiment 2: Testing the load dependency of examined metrics in rats

Eight rats from experiment 1 were placed in our custom-made LBNP chamber and subjected to four levels of LBNP manipulations that were standardized between animals based on reductions in mean arterial pressure (MAP). In vivo cardiovascular function was assessed at baseline, during stage 1 (−5mmHg MAP), stage 2 (−10mmHg MAP), stage 3 (−15mmHg MAP), and stage 4 (−20mmHg MAP) of LBNP manipulations (Figure 2). Each stage lasted for at least 60s and data analysis was performed over the last 30s of each stage. It should be noted that such manipulations are also expected to instigate mild reflexive sympathoexcitation through graded baroreceptor unloading (Goswami et al., 2019), which in turn elicits baroreflex-induced changes in ventricular contractility (Sala-Mercado et al., 2014). As such, leveraging this approach enabled us to test whether our selected surrogates of LV contractility are sensitive to loading and/or mild increases in cardiac inotropy. If the selected metric decreases during LBNP manipulation this implies it is sensitive to loading alternations. If a selected metric remains unchanged this suggests the metric is insensitive to both changes in loading and mild inotropic stimulation, and finally if the selected metric increases in the face of reducing preload then this suggests the metric is both insensitive to changes in loading and sensitive to mild changes in inotropic stimulation. The methods of anesthesia, euthanasia, and confirmation of euthanasia were analogous to experiment 1.

Figure 2. — **(A)**. An example of cardiac pressure and volume response to various stages of LBNP **(B)**. LV, left ventricle; *MAP*, mean arterial pressure; _LVdP/dt_max-EDV, the maximal rate of rise of LV pressure (dP/dt_max) normalized to end-diastolic volume (EDV). Some parts of this figure were prepared using BioRender (https://biorender.com/).

Experiments 3 and 4: Ability of the examined metrics to diagnose contractile dysfunction in animals with absent bulbospinal sympathetic outflow to the heart (3) and myocardial infarction (4)

Experiment 3, this experiment was conducted on 16 separate rats. Animals were randomly assigned to either naïve or with injury to the spinal cord (severe contusion injury at the T3 spinal level). All procedures associated with inducing injury and pre-and post-operative care are described in detail previously (Wainman et al., 2021). Briefly, rats were initially anesthetized using an inhalational anesthetic (5% isoflurane chamber induction, and maintenance on 1.5–2% isoflurane, Piramal Critical Care, Bethlehem, PA, USA) and received enrofloxacin (10 mg/kg; Bayer Animal Health, Shawnee, KS, USA), buprenorphine (0.5 mg/kg; Ceva Animal Health, Cambridge, ON, Canada) and warmed lactated ringer’s solution (5 mL subcutaneously; Baxter Corporation, Portland, OR, USA). The hind-paw withdrawal and the corneal reflexes were employed to test for an adequate anesthesia level prior to commencing the surgery. Following T3 laminectomy, rats were transported and mounted on a plastic staging platform where the T2 and T4 spinous processes were stabilized with curved tip clamps. Contusion injury was then performed using the custom impactor tip (3 mm; Infinite Horizons Impactor; Precision Systems and Instrumentation, Fairfax Station, VA, USA). Once injured, rats were recovered in an incubator for 30 min (37°C, 50% humidity) and received a subsequent 5 mL lactated ringer’s solution before they were returned to their home cages. The post-operative care for this experiment included administering subcutaneous lactated ringers (3× per day, 5 mL), buprenorphine (3× per day, 0.02 mg/kg), and enrofloxacin (1× per day, 10 mg/kg) up to for 4 days post-injury. Bladders were also manually expressed 4 times per day until spontaneous voiding was regained (4–6 days post-injury). During the recovery period, animals were further provided with a supportive diet consisting of Hydrogel (ClearH₂O), fruit, spinach, and cereal when necessary. For the present study, this injury level enabled us to survive the rodents whilst also removing practically all medullary sympathetic control over the heart (Squair et al., 2017). We have previously demonstrated that this time window (14 days), or even shorter (i.e., up to 4 hrs post-injury), and level of injury (i.e., T2–3) are sufficient to cause a marked decrement in cardiac contractile function, evidenced by a significant reduction in E_es in both rodent and porcine models (Squair et al., 2018; Poormasjedi-Meibod et al., 2019; Williams et al., 2020; Wainman et al., 2021; Fossey et al., 2022). Moreover, in a recent manuscript, describing a series of experiments in both rodents and individuals with SCI, we reported that SCI induces considerable contractile dysfunction post-injury that occurs due to the loss of bulbospinal sympathetic control over the heart (Fossey et al., 2022). Terminal in vivo cardiovascular function assessment (i.e., 14 days post-injury), as well as the methods of euthanasia and confirmation of death, was analogous to experiment 1. Experiment 4, this experiment was conducted on 10 separate rats. Rats were randomly assigned for either sham surgery (control) or LAD Ligation. Rats were pre-anesthetized and kept on 4% Isoflurane anesthesia. Pre-warmed saline along with buprenorphine (0.01–0.05 mg/kg) was given intraperitoneally. Adequate anesthesia was accompanied by loss of muscle tone and by loss of reflexes. For inducing the myocardial infarction, a suture was tied around the LAD coronary artery using a double throw and locked in position by using the third knot. The final knot location was based on a rat model published previously (Chen et al., 2013). Occlusion was confirmed by a transiently visible sudden change of anterior wall color to pale. At this stage, escaped blood in the area and in the chest cavity was absorbed using disposable surgical spears (Braintree Scientific, MA). The chest cavity was closed using a taper-point cutting 1/2 circle, 17mm needle with 4–0 nylon (N1245 Nurulon, Ethicon) suture. The chest was closed by bringing together the 5th and the 6th ribs (with pressure applied to the chest wall to reduce the volume of free air). A pneumothorax was eliminated by using a sterile catheter and syringe by further evacuation of the air through a small chest opening, later closed by a monofilament suture. The muscle layer and skin were closed with 6–0 absorbable and nylon sutures, respectively. About 15–20% mortality following this procedure was anticipated. Pre-warmed sterile isotonic fluids and analgesic (buprenorphine, 0.05 mg/kg _I.P.) were administered shortly after, then every 8 h for the next 48–72 h. The rat was placed in a quiet low-traffic area away from the operation room, and the core temperature was maintained using an infrared heating lamp. Rats were checked every 10–15 minutes and turned from side to side until fully recovered. Terminal LV hemodynamic measurement (~1-month post-myocardial infarction) was achieved by invasive PV catheterization using an open-chest surgery approach. Briefly, rats were pre-anesthetized in the induction chamber and kept on 2% of isoflurane anesthesia. Open chest surgery was performed while animals were secured in dorsal recumbence on the heating pad. A wide V-shape skin incision was made towards the xiphoidal process starting from the right lower thorax quadrant/upper abdomen area. The abdominal wall, skin, and abdominal muscles were carefully retracted and dissected to avoid any injury to other organs in the area. The diaphragm was cut through to expose the heart apex. The cardiac apex was positioned into the diaphragm opening while gently maneuvered using Q-tips. The LV cavity was accessed by means of an apical stab using the 24G needle with outer diameter OD=0.57 mm. The catheter was inserted using its tip to locate the apex stab. After a period of 20 minutes of steady state data collection, the surgical procedure was commenced to isolate IVC to perform temporary preload reduction maneuvers (i.e., occlusions) to obtain load-independent values of cardiac function (i.e., cardiac contractility). At the end of the experiment, animals were euthanized using an inhalational anesthetic overdose (5% isoflurane) and confirmed by a bilateral thoracotomy.

Experiments 5 and 6: Investigating whether any potential agreements observed between examined metrics in small rodents are translatable to large porcine models (5), and testing the behavior of examined metrics to direct inotropic simulation (6)

Experiment 5, Seventeen minipigs were anesthetized using a combined intramuscular injection of telazol (4–6 mg kg⁻¹), xylazine (1 mg kg⁻¹), atropine (0.02–0.04 mg kg⁻¹), and propofol (2 mg kg⁻¹). The adequacy of anesthesia was determined by the absence of jaw tone assessed by the veterinarian technicians. Next, animals were intubated and mechanically ventilated (10–12 breaths min⁻¹; tidal volume 12–15 mg kg⁻¹; Veterinary Anesthesia Ventilator model 2002, USA). All animals received intravenous continuous rate infusions of propofol (9–13 mg kg⁻¹ h⁻¹), fentanyl (10–15 mg kg⁻¹ h⁻¹), and ketamine (5–8 mg kg⁻¹ h⁻¹), as well as intravenous fluid to maintain hydration (7–10 ml kg⁻¹ h⁻¹, 2.5% dextrose + 0.9% NaCl) during the entire experiment. For all animals, core temperature was maintained at 38.5–39.5 °C with a heating pad (T/Pump, Gaymar Industries, Inc., USA) following a rectal temperature probe placement. All terminal in vivo cardiovascular assessments and data acquisition were analogous to the rats in experiment 1, except that a different LV PV catheter specific to a large animal model (5F; Transonic Scisense) was used to assess cardiovascular function (also via a closed-chest approach) and a balloon catheter was used to occlude the IVC (femoral access with placement in the IVC). Volume signals from the Transonic ADV500 PV system were corrected with SV obtained via a Swan-Ganz thermodilution catheter (7.5F; Edwards Lifesciences Canada Inc., Mississauga, ON) in the pulmonary artery. Experiment 6, ten minipigs were used to determine the sensitivity of examined metrics to a direct inotropic stimulation using constant dobutamine infusion (i.e., 10 μg/kg/min). The methods for anesthesia and ensuring the adequacy of the surgical plane, as well as all terminal in vivo cardiovascular assessments and data acquisition were identical to experiment 5. Animals from experiments 5 and 6 were euthanized with Pentobarbital (Euthanyl) administered intravenously (120 mg/kg) by a licensed animal care technician/veterinarian and confirmed by auscultation of the heart at the experimental end-point of the larger study.

Experiments 7 and 8: Investigating whether examined metrics are capable of diagnosing weakened contractile function in a conscious dog model of heart failure (7), as well as revealing a progressive decline in cardiac contractile function over time and during transition to heart failure (8)

Experiment 7, seven adult mongrel canines underwent a multi-staged surgical protocol (see below). The following anesthetic and analgesic regimen was used for each procedure. Thirty minutes prior to anesthetic induction, animals received a dose of acepromazine (0.2 mg/kg _I.M.). Anesthetic induction was achieved via intravenous administration of thiopental sodium (25 mg/kg) and was maintained pre and intra-operatively with isoflurane gas (1–3%). A 500 mg intravenous dose of prophylactic antibiotic cefazolin was started preoperatively and given throughout the procedure as required. Post-operatively, prophylactic antibiotics were continued for the duration of the study by oral administration of 30 mg/kg cephalexin twice daily. For analgesic maintenance, a fentanyl transdermal patch was applied and delivered a dose of 125–150 μg/h over a period of 3 days. During surgical recovery, doses of buprenorphine (0.015 mg/kg _I.V.), acepromazine (0.1 mg/kg _I.V.) and oral ketorolac (10–15 mg kg⁻¹ day⁻¹) were administered as needed for discomfort and sedation. For the surgical procedures, a blood flow transducer (Transonic Systems Ithaca, New York) was placed on the ascending aorta, via a left thoracotomy, to measure cardiac output. A telemetric pressure sensor (Data Sciences International St Paul, Minnesota) was placed in the LV through the apex of the heart for measures of LV pressure. Two pairs of sonomicrometry crystals (Sonometrics Corporation London, Ontario) were placed on the interior endocardial surface of the LV to measure anterior-to-posterior (short axis) and base-to-apex (long axis) dimensions which were used to estimate LV volume via a modified ellipsoid model (Cheng et al., 1992). Three stainless steel pacing leads (0-Flexon: Ethicon) were placed on the right ventricle free wall for induction of heart failure via rapid ventricular pacing. Vascular occluders (Holly Specialty Products LLC Petaluma, California) were placed on the vena cavae to induce transient reductions in cardiac preload. Following 2 weeks of recovery, additional catheters and transducers were placed on the circumflex artery (flow), renal artery (flow and pressure), and jugular vein (pressure), for purposes unrelated to the current study (Coutsos et al., 2013). Following a full recovery (14 days minimum) animals underwent experiments to produce PV loops at rest and during graded reductions in preload via partial inflation of the vena caval occluders during steady-state rest. These maneuvers were performed while animals were standing and fully conscious (i.e., closed-chest). PV loops were then repeated under the same state after induction of heart failure via rapid ventricular pacing (200– 230 bpm, ~ 30 days) to test the ability of the examined metrics to reveal weakened contractile function post-heart failure induction. All in vivo hemodynamic parameters were measured using a blood flow meter (Transonic Systems Ithaca, New York), and a Gould 6600 amplifier for pressure. Signals were acquired and plotted in real time in Windaq acquisition software (Dataq Instruments Akron, Ohio). Data was exported and transferred for analysis in Labchart Pro software. PV loops were generated by using the pressure signal derived from the LV pressure catheter, and employing long and short axis segment length to calculate volume by using the equation LV volume = (π/6) × D_SA² × D_LA, where D_SA is the anterior-to-posterior (short-axis) LV diameter and D_LA is the apex-to-base (long-axis) LV diameter (Cheng et al., 1992). At the cessation of the study, animals were euthanized by administration of a lethal dose of sodium pentobarbital (120 mg/kg _I.V.). Secondary assurance of death was performed by way of inducing pneumothorax through an incision between the 4^th and 5^th intercostal spaces after cessation of audible heart sounds and respiration. Experiment 8, for longitudinally tracking changes in examined metrics, four separate female adult mongrel canines underwent a two-part surgical protocol as previously described (Mannozzi et al., 2020, 2021). Pre, intra, and post-operative anesthesia, analgesia, and prophylactic antibiotic treatments were as follows. 30 minutes prior to surgery animals were sedated with an intramuscular injection of acepromazine (0.4–0.5 mg/kg) and provided long-term analgesics by way of subcutaneous injection with slow-release buprenorphine SR (0.03 mg/kg). Additional analgesia was provided preoperatively by way of intravenous administration of carprofen (4.4 mg/kg) Animals were induced by intravenous administration of ketamine (5 mg/kg) and diazepam (0.2–0.3 mg/kg) prior to intubation. Animals were anesthetically maintained using isoflurane gas (1–3%) for the duration of the surgical procedure. Post-operatively acepromazine (0.2–0.3 mg/kg _I.V.) and buprenorphine (0.03 mg/kg _I.M.) were administered as needed in conjunction with veterinary oversight. Pre- and post-operatively acute prophylactic antibiotic cephalexin (30 mg/kg _I.V.) was administered to prevent acute infection. Long-term prophylactic antibiotic cephalexin (30 mg/kg _{PO BID}) was administered for the duration of the study to prevent acute microbial infection. The first procedure was a left thoracotomy in which the intrathoracic space was entered through the 4th or 5th intercostal space based on palpated intercostal spaces to determine the location of the heart. The pericardium was opened to access the apex of the heart for placement of a telemetric pressure catheter (TA11 PA-D70, DSI) for measures of LV pressure and for placement of a telemetric positive ECG lead (TA11 PA-D70, DSI). Next, the upper section of the pericardium was retracted to expose the ascending aorta and pulmonary vein. The tissue surrounding the ascending aorta and adjacent to the pulmonary vein was dissected for placement of a blood flow probe around the ascending aorta (20 PAU, Transonic Systems) for measures of cardiac output. On the ventral chest wall of the thoracic cavity on the left side of the sternum the mammary artery was located and tracked into the muscle tissue. A small section of intercostal muscle was dissected to access the mammary artery for placement of a telemetric pressure sensor (TA11 PA-D70, DSI) for measures of systemic arterial pressure. Additionally, on the right ventral chest wall cranial to the heart a negative ECG lead (TA11 PA-D70, DSI) was sewn into the intercostal muscle in the 2nd intercostal space to be used with the positive ECG lead for real-time telemetric lead II ECG. On the right ventricular free wall four stainless steel pacing (0-Flexon, Ethicon) leads were placed for induction of heart failure via rapid ventricular pacing. The telemetry implant body (TA11 PA-D70, DSI) was placed on the left flank ventral to the floating rib and all telemetric cables were tunneled subcutaneously from the implant body through the 7th or 8th intercostal space into the chest cavity. The ribs were reapproximated and the tissue at the thoracotomy and implant body incisions was closed in layers and all cables and leads not attached to the implant body were tunneled subcutaneously and exited at the scapulae. In the second procedure no earlier than 14 days post-thoracotomy, devices unrelated to the current study were placed in the retroperitoneal space in the terminal aorta as previously described (Mannozzi et al., 2020, 2021). Briefly, a lumbar artery caudal to the renal artery was catheterized using a 19-gauge polyvinyl catheter (Tygon, S54-HL, Norton). Caudal to the catheter site a blood flow probe (10 PAU, Transonic Systems) and two hydraulic vascular occluders (DocXS Biomedical Products) were placed on the terminal aorta. Lastly, a flow probe (4 PSB, Transonic Systems) was placed on the left renal artery. The tissue was closed in layers and all cables, catheters and occluder lines were tunneled subcutaneously and exited at the scapulae. Animals were given a minimum of 14 days post-surgery prior to the performing of any experiments. For experimental protocol, animals were brought into the laboratory space and acclimated for 15–20 minutes prior to being directed to a motorized treadmill and acquisition equipment. All flow probe cables and catheters were connected to their respective equipment and the telemeter was turned on. Data acquisition was performed in Ponemah (DSI) at a sampling rate of 1000 samples per second. After three minutes of acclimatization to a standing rest, one minute of steady-state data was taken for analysis prior to additional experimental procedures unrelated to the current investigation. This experimental procedure was repeated after induction of heart failure via rapid ventricular pacing at 225–245 bpm for 3–4 weeks. Heart failure was determined by tachycardia, an approximate 50% reduction in dP/dt_max and dP/dt_min, as well as reductions in cardiac output and mean arterial pressure at rest after removal of the pacemaker. After conclusion of all experimental procedures, those included in and unrelated to the current study, animals were euthanized using an intravenous injection of pentobarbital (120mg/kg). Secondary assurance of euthanasia was performed by way of bilateral pneumothorax induced by an incision in the 4th-5th intercostal space at the cessation of any sign of respiration.

All analyses were analogous to previous experiments. However, in our longitudinal experiment, examined metrics in each dog were derived from PV loops constructed using aortic blood flow and LV pressure traces using a programming software (MATLAB R2018b, MathWorks®, USA) at baseline, post-pacing initiation (13.3±4.1 days), and following achievement of systolic heart failure. It also should be noted that given no absolute volume measurement was available/feasible in these animals, ESV was estimated (see below). EDV for each loop then was estimated via summation of SV and ESV. For estimating ESV we performed a linear regression of the ESV and SV relation from our previous dogs in experiment 7. Estimation of ESV from flow signal, rather than EDV, was preferred given this metric showed a lower standard deviation in animals from experiment 7, as well as the greater accuracy of sonomicrometry crystals to estimate ESV than EDV. Sam linear regression approach was conducted for estimating P_es based on P_max derived from LV pressure trace. It should also be noted that recordings in our conscious animals were performed when the pacing was turned off and left off for the duration of the experiment.

Calculation of field-standard metrics derived from the IVCO and single-beat surrogates

E_es(IVC) was measured as the slope of the linear relationship between ESP and ESV (Figure 1A)(Suga et al., 1973; Burkhoff et al., 2005). PRSW_(IVC) was measured as the slope of the linear relationship between SW (area of the PV loop) and EDV (i.e., preload) (Figure 1B)(Glower et al., 1985). Finally, we measured the slope of the linear relationship between dP/dt_max and EDV (dP/dt_max-EDV_(IVC)) (Figure 1C)(Little, 1985). In all instances, these 3 metrics of load-independent function were measured from the same IVCO. The definition of each examined surrogate metric is provided in Table 1. It should be noted that we examined only single-beat metrics that we could directly calculate from our LV PV and arterial recordings. Because of the considerable number of indices assessed in the present study, we also chose to only assess agreement between any single surrogate metric of LV contractility with (what we considered) the most relevant directly assessed IVCO reference measure, as opposed to comparing every single surrogate metric against all 3 IVCO reference measures. We chose this approach primarily for the brevity of results presentation but also because it is recognized that each of the 3 reference measures of LV contractility obtained from IVCO measure slightly different aspects of LV contractility and/or have varying underlying assumptions (i.e., some dependency on either pre-load or afterload) (Suga et al., 1973; Glower et al., 1985; Little, 1985; Burkhoff et al., 2005; Blaudszun & Morel, 2011; Blaudszun et al., 2013). In addition to previously tested single-beat surrogates of LV contractility, we also calculated preload-adjusted arterial dP/dt_max (_ARTdP/dt_max-EDV) as a surrogate for dP/dt_max-EDV_(IVC) since _ARTdP/dt_max has previously been demonstrated to behave in a similar manner as LV dP/dt_max (_LVdP/dt_max) when contractile state and afterload are manipulated (Garcia et al., 2018). Finally, we suggest using ESP normalized to EDV as a new surrogate metric of cardiac contractility. ESP is defined as the pressure at the point at which aortic valve closure occurs and ventricular relaxation initiates (Abel, 1981; Mada et al., 2015). Interestingly, ESP and dP/dt_max have been previously shown to behave analogously to each other when cardiac loading/contractile function is manipulated (Lionetti et al., 2013). Accordingly, since adjusting dP/dt_max to preload (i.e., EDV) has previously been demonstrated to be a reliable metric to quantify cardiac contractility (Blaudszun, et al, 2013), here we also postulate that normalizing ESP to EDV could be used to assess cardiac contractile function. In this work, end-systolic points were calculated as the maximum pressure/volume ratio during each cardiac cycle using an automatic detection approach in LabChart. With respect to any surrogate index that requires ESP, we calculated ESP both directly from the LV catheter (_LVESP), and indirectly (i.e., peripherally) as a product of 0.9 × SBP (_ARTESP) (Kelly et al., 1992).

Statistical analysis

The Shapiro-Wilk’s test was used to assess distribution characteristics. For experiments 1 and 4, the absolute agreement between metrics was examined using intra-class correlation coefficients (ICC) with a 2-way mixed model and interpreted as excellent (0.90 or higher), good (0.75 to 0.90), moderate (0.50 to 0.75), or poor (below 0.50) (Koo & Li, 2016). Bland-Altman analysis was also used to assess agreement between variables, as well as to visually depict dispersion patterns (Bland & Altman, 2012). The upper and lower limit of agreement (LOA) were defined as the mean bias ± 1.96 standard deviations. All agreement analyses for indices that used different units of measurement were performed on standardized z scores. For experiments 2 and 8, the responses of the examined metrics to LBNP manipulation in rats and during transition to heart failure in dogs were tested using either one-way repeated measure or Friedman test, with post-hoc comparisons conducted with Dunnett’s and Dunn’s tests for normally and non-normally distributed data, respectively. For experiments 3 and 4 between conditions (naïve vs. animal with absent bulbospinal sympathetic control; naïve vs. myocardial infarction), comparisons were performed with an independent t-test and Mann-Whitney tests, when appropriate. A paired t-test or Wilcoxon matched pairs test was also used to test the responses of the examined metrics to inotropic stimulation in pigs (experiment 6) and heart failure induction in dogs (experiment 7). Data analyses were conducted using R Studio (version 1.4.1106, R Studio Team, PBC, Boston, MA, USA) and figures prepared in GraphPad Prism (version 9.1.2, GraphPad Software, Inc., USA). Data are presented as means ± standard deviations (SD) in figures and tables (for non-normally distributed data, medians and interquartile ranges are provided). The alpha value was set at P < 0.05.

RESULTS

Rats

Agreement between single-beat metrics and those obtained from IVCO

Both E_es(SB1) (_LVESP/ESV) and E_es(SB2) (_ARTESP(0.9 × SBP)/ESV)) showed moderate agreement with E_es(IVC) (Figure 3, Section 1, A-B, Table 2). A moderate-to-good agreement was noted between PRSW_(SB)(SW/EDV), PWR_max-EDV, PWR_max-EDV², LVEF_EA1, and LVEF_EA2 with PRSW_(IVC) (Figure 3, Section 2, A-E, Table 3). Likewise, _ARTdP/dt_max-EDV, P_max-EDV, _LVESP-EDV, and _ARTESP(0.9 × SBP)-EDV all showed a good agreement with dP/dt_max-EDV_(IVC), whilst a moderate agreement was noted between _LVdP/dt_max-EDV and dP/dt_max-EDV_(IVC) (Figure 3, Section 3, A-E, Table 4). With respect to the agreement between examined indices calculated from baseline arterial and baseline LV data (Table 5), _ARTdP/dt_max-EDV showed a good-to-excellent agreement with _LVdP/dt_max-EDV, P_max-EDV, and _LVESP-EDV. Similarly, _ARTESP-EDV also showed an excellent agreement with _LVESP-EDV, _LVdP/dt_max-EDV, and P_max-EDV. _ARTESP also demonstrated good agreement with _LVESP. An excellent agreement was also noted between LVEF_EA1 and LVEF_EA2. For other metrics, only a poor-to-moderate agreement was noted (Table 5).

Table 2.

Intra-class correlation coefficients (ICC) and Bland-Altman results between end-systolic elastance (E_es(IVC)) obtained from inferior vena cava (IVC) occlusion and other single-beat surrogate metrics calculated from beat-by-beat baseline pressure-volume and arterial recordings in rats (n = 30).

	ICC				Bland-Altman, 95% LOA

	ICC	Size	95% CI	P	Lower	Upper
E_es(IVC) *vs. single-beat surrogates of E*_es(IVC)

E_es(SB1)	0.510^*	Moderate	−0.014–0.622	0.031	−3.531	3.011
E_es(SB2)	0.522^*	Moderate	0.012–0.771	0.024	−3.584	2E.86

E_es(IVC) *vs. other IVC occlusion-derived metrics*

PRSW_(IVC)	0.229	-	−0.669–0.468	0.251	−2.592	2.592
dP/dt_max-EDV_(IVC)	0.631	Moderate	0.213–0.825	0.005	−2.051	2.051

E_es(IVC) *vs. other single-beat surrogates of cardiac contractility*

PRSW_(SB)	−0.024	-	− 1.238–0522	0.524	−2.787	2.787
PWR_max-EDV	−0.078	-	−1.361–0.479	0.576	−2.821	2.821
PWR_max-EDV²	0.332	-	−.439–0.686	0.148	−2.488	2.488
LVEF_EA1	0.529^*	Moderate	−0.009–0.777	0.026	−2.232	2.232
LVEF_EA2	0.517^*	Moderate	−0.034–0.772	0.03	−2.25	2.25
_LVdP/dt_max-EDV	0.639^*	Moderate	0.232–0.829	0.004	−2.0341	2.0341
_ARTdP/dt_max-EDV	0.504^*	Moderate	−0.062–0.766	0.035	−2.269	2.269
P_max-EDV	0.643^*	Moderate	0.239–0.831	0.004	−2.027	2.027
_LVESP-EDV	0.525^*	Moderate	−0.016–0.776	0.027	−2.237	2.237
_ARTESP-EDV	0.534^*	Moderate	0.003–0.780	0.025	−2.223	2.223

E_es(IVC) *vs. load-dependent metrics*

_LVdP/dt_max	−0.21	-	−1.664–0.437	0.688	−2.895	2.895
_ARTdP/dt_max	−0.272	-	−1.809–0.408	0.732	−2.927	2.927
P_max	−0.909	-	−3.325–0.122	0.949	−3.159	3.159
_LVESP	−1.104	-	−3805–0.035	0.969	−3.208	3.208
_ARTESP (0.9 × SBP)	−2.342	-	−7.043−−0.500	0.998	−3.408	3.408
PWR_max	−0.719	-	−2.865–0.206	0.916	−3.103	3.103
SW	−2.087	-	−6.347−−0.392	0.997	−3.378	3.378

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E_es(IVC), end-systolic elastance obtained from IVC occlusion; E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; PRSW_(IVC), preload recruitable stork work obtained from IVC occlusion; dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVC occlusion; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted LV ejection fraction using ventricular data; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio; LOA; limit of agreement.

significant observation.

Table 3.

Intra-class correlation coefficients (ICC), and Bland-Altman results between preload recruitable stroke work (PRSW_(IVC)) obtained from inferior vena cava (IVC) occlusion and other single-beat surrogate metrics calculated from beat-by-beat baseline pressure-volume and arterial recordings in rats (n = 30).

	ICC				Bland-Altman, 95% LOA

	ICC	Size	95% CI	P	Lower	Upper
*PRSW_(IVC) vs. single-beat surrogates of PRSW*_(IVC)

PRSW_(SB)	0.631^*	Moderate	−0.194–0.868	< 0.001	−20.027	90.911
PWR_max-EDV	0.637^*	Moderate	0.227–0.929	0.005	−2.038	2.038
PWR_max-EDV²	0.699^*	Moderate	0.359–0.857	0.001	−1.903	1.903
LVEF_EA1	0.812^*	Good	0.603–0.911	< 0.001	−1.576	1.576
LVEF_EA2	0.799^*	Good	0.575–0.905	< 0.001	−1.621	1.621

*PRSW_(IVC) vs. other IVC occlusion-derived metrics*

dP/dt_max-EDV_(IVC)	0.653^*	Moderate	0.262–0.836	0.003	−2.005	2.005

*PRSW_(IVC) vs. other single-beat surrogates of cardiac contractility*

E_es(SB1)	0.447	-	−0.187–0.739	0.063	−2.35	2.35
E_es(SB2)	0.436	-	−0.210–0.734	0.069	−2.364	2.364
_LVdP/dt_max-EDV	0.611^*	Moderate	0.171–0.816	0.008	−2.089	2.0891
_ARTdP/dt_max-EDV	0.457^*	Poor	−.165–0.744	0.057	−2.336	2.336
P_maxEDV	0.656^*	Moderate	−0.268–0.837	0.003	−1.999	1.999
_LVESP-EDV	0.694^*	Moderate	0.350–0.855	0.001	−1.913	1.913
_ARTESP-EDV	0.699^*	Moderate	0.353–0.856	0.001	−1.91	1.91

*PRSW_(IVC) vs. load-dependent metrics*

_LVdP/dt_max	0.252	-	−0.617–0.649	0.226	−2.57	2.57
_ARTdP/dt_max	0.155	-	−0.835–0.604	0.332	−2.656	2.656
P_max	0.495^*	Poor	−0.081–0.762	0.039	−2.283	2.283
_LVESP	0.465	-	−0.246 −0.748	0.053	−2.325	2.325
_ARTESP (0.9 × SBP)	0.571^*	Moderate	0.083–0.797	0.015	−2.162	2.162
PWR_max	0.335	-	−0.433–0.687	0.146	−2.485	2.485
SW	−0.154	-	−1.535–0.462	0.643	−2865	2.865

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PRSW_(IVC), preload recruitable stork work obtained from IVC occlusion; PRSW_(SB), stroke work (SW) and end-diastolic volume (EDV) ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted left ventricular ejection fraction using ventricular data; LVEF_EA2, afterload adjusted left ventricular ejection fraction using arterial data; dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVC occlusion; E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP) and ESV ratio;; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio, _ARTESP-EDV, _ARTESP and EDV ratio; LOA; limit of agreement.

significant observation.

Table 4.

Intra-class correlation coefficients (ICC), and Bland-Altman results between dP/dt_max-EDV_(IVC) obtained from inferior vena cava (IVC) occlusion and other single-beat surrogate metrics calculated from beat-by-beat baseline pressure-volume and arterial recordings in rats (n = 30).

	ICC				Bland-Altman, 95% LOA

	ICC	Size	95% CI	P	Lower	Upper
*dP/dt_max-EDV_(IVC) vs. single-beat surrogates of dP/dt*_max-EDV_(IVC)

_LVdP/dt_max-EDV	0.700^*	Moderate	−0.213–0.907	< 0.001	−46.584	5.876
_ARTdP/dt_max-EDV	0.758^*	Good	0.486–0.885	< 0.001	−44.93	28.184
P_max-EDV	0.845^*	Good	0.673–0.927	< 0.001	−1.452	1.452
_LVESP-EDV	0.853^*	Good	0.689–0.930	< 0.001	−1.421	1.421
_ARTESP-EDV	0.822^*	Good	0.624–0.916	< 0.001	−1.54	1.54

*dP/dt_max-EDV_(IVC) vs. other single-beat surrogates of cardiac contractility*

E_es(SB1)	0.344	-	−0.414–0.691	0.138	−2.475	2.475
E_es(SB2)	0.313	-	−0.482–0.677	0.166	−2.509	2.509
PRSW_(SB)	0.413	-	−0.262–0.723	0.084	−2.394	2.394
PWR_max-EDV	0.437	-	−0.210–0.735	0.069	−2.364	2.364
PRW_max-EDV²	0.691^*	Moderate	0.343–0.854	0.001	−1.921	1.921
LVEF_EA1	0.782^*	Good	0.539–0.897	< 0.001	−1.675	1.675
LVEF_EA2	0.743^*	Moderate	0.455–0.878	< 0.001	−1.789	1.789

*dP/dt_max-EDV_(IVC) vs. load-dependent metrics*

_LVdP/dt_max	0.612^*	Moderate	0.173–0.817	0.008	−2.087	2.087
_ARTdP/dt_max	0.483^*	Poor	−0.107–0.756	0.044	−2.3	2.3
P_max	0.217	-	−0.696–0.632	0.263	−2.603	2.603
_LVESP	0.200	-	−0.734–0.624	0.282	−2.618	2.618
_ARTESP (0.9 × SBP)	0.061	-	−1.046–0.560	0.436	−2.729	2.729
PWR_max	−0.206	-	−1.656–0.438	0.685	−2.894	2.894
SW	−2.912	-	−8.653−−0.739	0.999	−3.464	3.464

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dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVC occlusion; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, LV end-systolic pressure (_LVESP) and EDV ratio, _ARTESP-EDV, Arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP) and EDV ratio; E_{es(SB1), LV}ESP and end-systolic volume (ESV) ratio; E_es(SB2), _ARTESP and ESV ratio; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted left ventricular ejection fraction using ventricular data; LVEF_EA2, afterload adjusted left ventricular ejection fraction using arterial data; LOA, limit of agreement.

significant observation

Table 5.

Intra-class correlation coefficients (ICC) and Bland-Altman results between metrics obtained from baseline ventricular and arterial beat-by-beat data in rats (n = 30).

Metrics	ICC				Bland-Altman, 95% LOA

	ICC	Size	95% CI	P	Lower	Upper
_LVdP/dt_max-EDV vs. _ARTdP/dt_max-EDV	0.838^*	Good	0.359–0.941	< 0.001	−16.575	42.537
_LVdP/dt_max-EDV vs. _ARTESP-EDV	0.948^*	Excellent	0.891–0.975	< 0.001	−0.881	0.881
P_max-EDV vs. _ARTdP/dt_max-EDV	0.913^*	Excellent	0.818–0.959	< 0.001	−1.122	1.122
P_max-EDV vs. _ARTESP-EDV	0.993^*	Excellent	0.986–0.997	< 0.001	−0.017	0.214
_LVESP-EDV vs. _ARTdP/dt_max-EDV	0.905	Excellent	0.800–0.955	< 0.001	−1.193	1.193
_LVESP-EDV vs. _ARTESP-EDV	0.985^*	Excellent	0.965–0.993	< 0.001	−0.115	0.074
E_es(SB1) vs. E_es(SB2)	0.993	Excellent	0.984–0.997	< 0.001	−0.651	0.448
LVEF_EA1 vs. LVEF_EA2	0.992^*	Excellent	0.980–0.996	< 0.001	−8.384	5.332
_LVdP/dt_max vs. _ARTdP/dt_max	0.311^*	Poor	−0.213–0.648	0.027	−1814.404	7369.039
_LVdP/dt_max vs. _ARTESP	0.654^*	Moderate	0.262–0.836	0.003	−2.004	2.004
P_max vs. _ARTdP/dt_max	0.541^*	Moderate	0.019–0.784	0.022	−2.211	2.211
P_max vs. _ARTESP	0.582^*	Moderate	−0.099–0.877	< 0.001	6.699	31.531
_LVESP vs. _ARTESP	0.896^*	Good	0.775–0.951	< 0.001	−20.356	13.757

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_LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular (LV) pressure (_LVdP/dt_max) and end-diastolic volume (EDV) ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _ARTESP-EDV, arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP) and EDV ratio; _LVESP-EDV, LV ESP (_LVESP) and EDV ratio; E_{es(SB1), LV}ESP and end-systolic volume (ESV) ratio; E_{es(SB2), ART}ESP and ESV ratio; LVEF_EA1, afterload adjusted left ventricular ejection fraction using ventricular data; LVEF_EA2, afterload adjusted left ventricular ejection fraction using arterial data; LOA, limit of agreement.

significant observation

Responses of single-beat metrics to LBNP manipulation

Efficacy of our LBNP manipulation was confirmed by demonstrating a graded reduction in load-dependent cardiac indices with advancing LBNP stage (Figure 4, Section 1, A-D, Table 6). E_es(SB1), E_es(SB2), LVEF_EA1, LVEF_EA2, and _ARTESP-EDV remained unchanged in response to LBNP suggesting they meet the assumption of load-independence (Figure 4, Section 2, A-B, Section 3, D-E, Section 4, E, Table 6). However, PRSW_(SB), PWR_max-EDV, and PWR_max-EDV² all decreased significantly from baseline during stages 3 and 4 of LBNP (Figure 4, Section 3, A-C, Table 6). In contrast, _LVdP/dt_max-EDV, _ARTdP/dt_max-EDV, P_max-EDV (P = 0.06), and _LVESP-EDV all increased from baseline during stage 3 and/or 4 of LBNP (Figure 4, section 4, A-D, Table 6).

Figure 4. — _LVdP/dt_max, the maximal rate of rise of the left ventricular (LV) pressure; P_max, maximal pressure; *PWR*_max, maximal power; SW, stroke work; E_es(SB1), LV end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; *PRSW*_(SB), SW and end-diastolic volume (EDV) ratio; *PWR*_max-EDV, PWR_max and EDV ratio; *PWR*_max-EDV², *PWR*_max and squared of EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio; *LVEF*_EA1, afterload adjusted LV ejection fraction using ventricular data; *LVEF*_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, _LVdP/dt_max and EDV ratio; _ARTdP/dt_max- EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, P_max and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio. Statistical analysis was performed using either one-way repeated measure or Friedman test, with post-hoc comparisons conducted with Dunnett’s and Dunn’s tests (n = 8). *P < 0.05, **P < 0.01, and ***P < 0.001. Actual p values are found in the Statistical Summary Document.

Table 6.

Behaviors of the single-beat surrogates of cardiac contractility during various levels of lower-body negative pressure (LBNP) manipulation in rats (n = 8).

Metrics	levels of LBNP

	Baseline	Stage 1	Stage 2	Stage 3	Stage 4
*Single-beat surrogates*

E_es(SB1) (mmHg/μl)	0.98±0.3	0.98±0.5	0.87±0.3	0.79±0.3	0.78±0.3
E_es(SB2) (mmHg/μl)	0.97±0.3	0.95±0.5	0.83±0.3	0.75±0.3	0.73±0.3
PRSW_(SB) (mmHg)	84(74–93)	76(65–85)	64(58–77)	51(50–71)^**	49(42–62)^***
PWR_max-EDV (mmHg/s)	6433±3675	5000±2954^*	3994±2336^*	3124±2017^*	2455±1711^*
PWR_max-EDV² (mmHg/s/μl)	20±10	18±9	15±7	12±7^*	10±6^*
LVEF_EA1 (%)	50±6	50±7	49±7	47±8	46±8
LVEF_EA2 (%)	50±6	49±7	48±7	46±8	44±8
_LVdP/dt_max-EDV (mmHg/s/μl)	35±10	37±10	40±12	41±14	42±13^*
_ARTdP/dt_max-EDV (mmHg/s/μl)	26±7	28±7	31±9	33±10^*	34±11
P_max-EDV (mmHg/μl)	0.41±0.1	0.43±0.1	0.45±0.1	0.47±0.1	0.48±0.1
_LVESP-EDV (mmHg/μl)	0.32(0.26–0.42)	0.36(0.26–0.4)	0.37(0.26–0.42)	0.38(0.27–0.43)	0.39(0.28–0.44)^**
_ARTESP-EDV (mmHg/μl)	0.35±0.1	0.35±0.1	0.36±0.1	0.37±0.1	0.36±0.1

*Systemic pressure and LV load-dependent metrics*

_LVdP/dt_max (mmHg/s)	10559±698	10023±986^*	9583±1003^***	9191±1016^***	8764±1152^***
_ARTdP/dt_max (mmHg/s)	7835±996	7631±983	7492±965	7309±990	7186±1368
P_max (mmHg)	120(119–131)	110(108–125)	105(103–117)^*	102(99–113)^***	98(91–110)^***
PWR_max (mmHg*μl/s)	2116063±1394497	1486788±1060859^*	1124138±887962^*	837188±703933^*	626485±583046^*
SW (mmHg/μl)	26928±8431	21085±5949^*	17109±5813^**	13659±4439^***	11498±4021^**
MAP (mmHg)	78±7	66±11^*	61±9^***	57±10^***	53±10^***
HR (bpm)	419±26	420±28	421±25	417±22	415±22
EDV (μl)	317±74	283±66^*	258±74^**	241±71^**	225±65^***
EF (%)	64±10	61±10	56±8^*	52±7^*	49±7^**

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Data are presented as means ± standard deviations for normally distributed data and as medians and interquartile ranges for non-normally distributed. E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × sytsolic blood pressure (SBP)) and ESV ratio; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted LV ejection fraction (EF) using ventricular data; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio; MAP, mean arterial pressure, HR; heart rate. Statistical analysis was performed using either one-way repeated measure or Friedman test, with post-hoc comparisons conducted with Dunnett’s and Dunn’s tests.

P < 0.05

^**

P < 0.01

^***

P < 0.001. Actual p values are found in the Statistical Summary Document.

Ability of single-beat metrics to detect contractile dysfunction in animals with absent bulbospinal sympathetic control and myocardial infarction

For diagnosing contractile dysfunction, all examined single-beat metrics, and field-standard indices were capable of revealing a significant reduction in cardiac contractile function in our spinal injury model with absent bulbospinal sympathetic outflow to the heart, except for E_es(SB1) and E_es(SB2) (Figure 5). Similarly, in the rat model of myocardial infarction, all examined indices revealed a marked cardiac contractile dysfunction (Figure 5).

Figure 5. — E_es(IVC), end-systolic elastance obtained from inferior vena cava occlusion (IVCO), E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; *PRSW*_(IVC), preload recruitable stork work obtained from IVCO; *PRSW*_(SB), stroke work (SW) and end-diastolic volume (EDV) ratio; *PWR*_max-EDV, maximal power (PWR_max) and EDV ratio; *PWR*_max-EDV², PWR_max and squared of EDV ratio; *LVEF*_EA1, afterload adjusted left ventricular ejection fraction using ventricular data; *dP/dt*_max-EDV_(IVC), the relationship between the maximal rate of rise of pressure and EDV obtained from IVCO; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio. Note that 2 control groups of animals were used as the myocardial infarction (MI) outcomes were assessed open-chest whereas in the animal with absent bulbospinal sympathetic control (SNX) the outcomes were assessed closed-chest. Statistical analysis was performed using either independent t-test or Mann-Whitney tests (SNX, n = 16; MI, n = 10). Note that given no peripheral catheterization was performed in the MI experiment, we have not calculated any indices of cardiac contractility from arterial data; therefore, only metrics derived from LV data are illustrated. Statistical findings for indices derived from arterial data in an animal model of spinal cord injury are found in the Statistical Summary Document.

Pigs

Agreement between single-beat metrics and those obtained from IVCO

PWR_max-EDV and PWR_max-EDV² both showed a moderate agreement (both approached statistical significance) with PRSW_(IVC) (Figure 6, A-B, Table 7). _LVdP/dt_max-EDV, P_max-EDV, _LVESP-EDV, and _ARTESP-EDV also all showed a moderate agreement with dP/dt_max-EDV_(IVC) (Figure 6, C-F, Table 7). No other single-beat surrogates showed agreement with their corresponding IVCO-derived reference measure (Table 7). Agreement between metrics calculated from baseline LV versus arterial data are presented in Table 7. _ARTdP/dt_max-EDV showed excellent and good agreement with P_max-EDV and _ARTESP-EDV, respectively. _ARTESP-EDV also showed an excellent agreement with _LVESP-EDV, _LVdP/dt_max-EDV, and P_max-EDV. _ARTESP (0.9 × SBP) showed a good agreement with _LVESP and an excellent agreement with P_max. An excellent agreement between LVEF_EA1 and LVEF_EA2 was also noted. For other metrics, only a poor-to-moderate agreement was observed (Table 7).

Table 7.

Intra-class correlation coefficients (ICC) and Bland-Altman results between examined metrics in pigs (n = 17).

Metrics	ICC				Bland-Altman, 95% LOA

	ICC	Size	95% CI	P	Lower	Upper
*Metrics obtained from IVC occlusion*

E_es(IVC) vs. PRSW_(IVC)	0.770^*	Good	0.347 −0.917	0.004	−1.728	1.728
E_es(IVC) vs. dP/dt_max-EDV_(IVC)	0.669^*	Moderate	0.051–0.882	0.020	−1.984	1.984
PRSW_(IVC) vs. dP/dt_max-EDV_(IVC)	0.743^*	Moderate	0.270–0.908	0.006	−1.803	1.803

E_es(IVC) *vs. single-beat surrogates of E*_es(IVC)

E_es(SB1)	0.444	-	−0.310–0.785	0.095	−2.083	1.222
E_es(SB2)	0.411	-	−0.270–0.763	0.075	−2.277	0.963

*PRSW_(IVC) vs. single-beat surrogates of PRSW*_(IVC)

PRSW_(SB)	0.084	-	−0.336–0.510	0.358	−10.856	40.519
PWR_max-EDV	0.570	Moderate	−0.245–0.847	0.058	−2.174	2.174
PRW_max-EDV²	0.537	Moderate	−0.386–0.831	0.083	−2.249	2.249
LVEF_EA1	0.033	-	−1.987–0.663	0.475	−2.749	2.749
LVEF_EA2	0.034	-	−1.984–0.663	0.474	−2.749	2.749

*dP/dt_max-EDV_(IVC) vs. single-beat surrogates of dP/dt*_max-EDV

_LVdP/dt_max-EDV	0.574^*	Moderate	−0.112–0.842	0.044	−24.773	17.654
_ARTdP/dt_max-EDV	0.284	-	−0.360–0.690	0.162	−12.673	31.666
P_max-EDV	0.649^*	Moderate	−0.007–0.875	0.026	−2.026	2.026
_LVESP-EDV	0.670^*	Moderate	0.055–0.882	0.020	−1.981	1.981
_ARTESP-EDV	0.585^*	Moderate	−0.199–0.852	0.050	−2.148	2.148

*Metrics obtained from ventricular vs. arterial beat-by-beat baseline data*

_LVdP/dt_max-EDV vs. _ARTdP/dt_max-EDV	0.373^*	Poor	−0.141–0.767	0.001	2.641	23.47
_LVdP/dt_max-EDV vs. _ARTESP-EDV	0.950^*	Excellent	0.860–0.982	< 0.001	−0.882	0.882
P_max-EDV vs. _ARTdP/dt_max-EDV	0.902^*	Excellent	0.727–0.965	< 0.001	−1.199	1.199
P_max-EDV vs. _ARTESP-EDV	0.986^*	Excellent	0.949–0.995	< 0.001	−0.206	0.112
_LVESP-EDV vs. _ARTdP/dt_max-EDV	0.885^*	Good	0.679–0.959	< 0.001	−1.287	1.287
_LVESP-EDV vs. _ARTESP-EDV	0.935^*	Excellent	0.570–0.988	< 0.001	−0.353	0.104
E_es(SB1) vs. E_es(SB2)	0.961	Excellent	0.674–0.990	< 0.001	−2.871	1.819
LVEF_EA1 vs. LVEF_EA2	0.955^*	Excellent	0.603–0.988	< 0.001	−10.403	3.185
_LVdP-dt_max vs. _ARTdP-dt_max	0.077	-	−0.085–0.355	0.148	236.704	1438.156
_LVdP/dt_max vs. _ARTESP	0.716^*	Moderate	0.191–0.898	0.010	−1.874	1.874
P_max vs. _ARTdP/dt_max	0.633^*	Moderate	−0.055.869	0.031	−2.058	2.058
P_max vs. _ARTESP	0.924^*	Excellent	0.756–0.974	< 0.001	−13.462	7.465
_LVESP vs. _ARTESP	0.809^*	Good	0.065–0.91	< 0.001	−22.690	6.664

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E_es(IVC), end-systolic elastance obtained from inferior venea cava (IVC) occlusion; PRSW_(IVC), preload recruitable stork work obtained from IVC occlusion; dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVC occlusion; E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted LV ejection fraction using ventricular data; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio.

significant observation

Responses of single-beat metrics to an inotropic stimulation

In our porcine model, all single-beat derived and field-standard (i.e., those obtained from caval occlusion) indices of contractility significantly increased under constant rate dobutamine infusion, except for PWR_max-EDV (P = 0.246) and PWR_max-EDV² (P = 0.057) (Table 8), suggesting that practically all indices meet the assumption of increasing in response to inotropic stimulation.

Table 8.

Behavior of examined metrics in response to inotropic stimulation in pigs (n = 10).

	Baseline	Dobutamine	P values
*IVC occlusion-derived metrics*

E_es(IVC) (mmHg/ml)	1.97(1.6–2.87)	6.26(3.98–8.99)	0.001^*
PRSW_(IVC) (mmHg)	58(47–73)	88(76–115)	0.013^*
dP/dt_max-EDV_(IVC) (mmHg/s/ml)	21±15	113±66	< 0.001^*

E_es(IVC) *surrogates*

E_es(SB1) (mmHg/ml)	2.23(1.98–2.74)	5.26(4.21–7.36)	0.001^*
E_es(SB2) (mmHg/ml)	2.6(2.27–2.9)	6.08(5.01–9.93)	0.001^*

*PRSW_(IVC) surrogates*

PRSW_(SB) (mmHg)	36 (32–39)	59(57–63)	0.001^*
PWR_max-EDV (mmHg/s)	119(106–217)	182(109–260)	0.246
PWR_max-EDV² (mmHg/s/ml)	2.51±1.34	3.87±2.47	0.057
LVEF_EA1 (%)	81(72–83)	107(97–124)	0.001^*
LVEF_EA2 (%)	85±12	121±17	< 0.001^*

*dP/dt* _max *-EDV* _(IVC) *surrogates*

_LVdP/dt_max-EDV (mmHg/s/ml)	23(17–25)	83(66–119)	0.001^*
_ARTdP/dt_max-EDV (mmHg/s/ml)	8(7–11)	38(30–49)	0.001^*
P_max-EDV (mmHg/ml)	1.34(1.13–1.49)	1.93(1.72–2.46)	0.001^*
_LVESP-EDV (mmHg/ml)	1.24±0.32	1.69±0.34	<0.001^*
_ARTESP-EDV (mmHg/ml)	1.37±0.26	2.06±0.45	<0.001^*

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E_es(IVC), end-systolic elastance obtained from inferior vena cava (IVC) occlusion; PRSW_(IVC), preload recruitable stork work obtained from IVC occlusion; dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVC occlusion; E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted LV ejection fraction using ventricular data; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio. Statistical analysis was performed using either a paired t-test or Wilcoxon matched pairs test (one-tailed).

significant observation.

Conscious dogs

The ability of single-beat metrics to detect contractile dysfunction in the setting of systolic heart failure

In our conscious canine model of systolic heart failure, all single-beat and field-standard indices were reduced significantly post-heart failure achievement, except for dP/dt_max-EDV_(IVC) (P = 0.084) and PWR_max-EDV (P = 0.064) (Table 9). With respect to tracking longitudinal changes in examined indices in conscious canines, the majority of single-beat metrics were able to reveal a progressive decline in cardiac contractile function from baseline to transition and during overt systolic heart failure (Figure 7).

Table 9.

Behavior of examined metrics in response to systolic heart failure (HF) induction in conscious canines (n = 7).

	Pre HF	HF	P values
*IVC occlusion-derived metrics (n =6)*

E_es(IVC) (mmHg/ml)	3.25±1.41	1.81±1.02	0.038^*
PRSW_(IVC) (mmHg)	84±23	63±17	0.002^*
dP/dt_max-EDV_(IVC) (mmHg/s/ml)	39±17	31±21	0.084

E_es(IVC) *surrogates*

E_es(SB1) (mmHg/ml)	2.24(1.59–3.65)	1.37(0.8–1.81)	0.007^*
E_es(SB2) (mmHg/ml)	2.21(1.53–3.73)	1.25(0.71–1.78)	0.007^*

*PRSW_(IVC) surrogates*

PRSW_(SB) (mmHg)	33±8	15±6	<0.001^*
PWR_max-EDV (mmHg/s)	205±131	115±30	0.064
PWR_max-EDV² (mmHg/s/ml)	2.13(1.41–2.75)	1.32(0.73–2.16)	0.007^*
LVEF_EA1 (%)	69±21	42±10	0.001^*
LVEF_EA2 (%)	67±23	40±10	0.002^*

*dP/dt* _max *-EDV* _(IVC) *surrogates*

_LVdP/dt_max-EDV (mmHg/s/ml)	32(25–50)	17(10–31)	0.007^*
_ARTdP/dt_max-EDV (mmHg/s/ml)	27(20–37)	8(7–21)	0.007^*
P_max-EDV (mmHg/ml)	1.44(1.16–2.22)	1.08(0.73–1.52)	0.007^*
_LVESP-EDV (mmHg/ml)	1.35(1.15–2.14)	1.05(0.72–1.5)	0.007^*
_ARTESP-EDV (mmHg/ml)	1.33(1.07–2.19)	0.96(0.64–1.47)	0.007^*

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significant observation.

Figure 7. — Note the down and right shift in estimated pressure and volume loops from baseline to transition (13.3±4.1 days post-pacing initiation) to systolic heart failure state in one of the canines studied. E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; E_es(SB2), Arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; *PRSW*_(SB), stroke work and end-diastolic (EDV) ratio; *PWR*_max-EDV, maximal power (PWR_max) and EDV ratio; *PWR*_max-EDV², *PWR*_max and squared of EDV ratio; *LVEF*_EA1, afterload adjusted left ventricular ejection fraction using ventricular data; *LVEF*_EA2, afterload adjusted LV ejection fraction using arterial data; _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular (LV) pressure (_LVdP/dt_max) and EDV ratio; _ARTdP/dt_max- EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio. B, baseline, T, transition, HF, heart failure. Statistical analysis was performed using either one-way repeated measure or Friedman test, with post-hoc comparisons conducted with Dunnett’s and Dunn’s tests (n = 4).

DISCUSSION

The major findings from the present study are 1) of all the metrics examined, only _LVESP-EDV, dP/dt_max-EDV, and P_max-EDV can be considered as valid, reliable single-beat metrics to quantify contractile function in experiments employing both small and large animal models; 2) in rodents the majority of the previously reported/tested metrics, as well as our proposed metrics, showed a moderate-to-excellent agreement with their IVCO-derived reference measure and were able to meet all the assumptions required to be considered as a valid cardiac contractile metric; 3) in large animal models only our proposed metric (_LVESP-EDV), together with dP/dt_max-EDV and P_max-EDV, demonstrated at least a moderate agreement with their IVCO-derived reference measure and met all the underlying assumptions studied; 4) arterial data-derived estimates of LV contractility showed a moderate-to-good agreement with their reference measure from IVCO and a good-to-excellent agreement with baseline LV indices, implying they can be used to quantify cardiac contractile function when cardiac catheterization is not available/feasible.

Single-beat metrics that can be used in both small and large animal models

Of all indices tested within this translational study, _LVESP-EDV, _LVdP/dt_max-EDV, and P_max-EDV were the only indices that showed at least a moderate agreement with their IVCO reference measure, exhibited load-independence, were sensitive to inotropic stimulation, and were able to detect contractile dysfunction in various animal models with marked contractile dysfunction (animals with absent bulbospinal sympathetic control, myocardial infarction, systolic heart failure) (Table 10). It should be noted that the calculation of P_max-EDV in the present study is somewhat different from the original metric proposed by Abel (2001), where P_max was considered as the highest LV pressure during isovolumetric contraction. Here, we measured P_max as the maximum pressure of LV during each cardiac cycle. Nonetheless, despite such difference, P_max-EDV, together with _LVESP-EDV and _LVdP/dt_max-EDV, met all the assumptions required for a valid metric of cardiac contractility. Arguably of most importance, _LVESP-EDV and _LVdP/dt_max-EDV were also able to reveal a progressive decline in cardiac contractile function from baseline to transition to overt systolic heart failure in our conscious canine model (Figure 7), suggesting they are capable of identifying longitudinal changes in LV contractility. To help aid researchers choose the best single-beat metric for their purpose we have summarized the performance of the best single-beat surrogates for E_es(IVC), PRSW_(IVC), dP/dt_max-EDV_(IVC), and arterial-derived estimates of LV contractility in Table 10.

Table 10.

List of the best metrics of cardiac contractility for each/all animal models studied.

Metrics	ANIMAL MODEL								Can be used in both small and large animal models

	RATS				PIGS AND DOGS

	Agreement^* with metric from IVCO (ICC)	Load-independency	Diagnosing contractile dysfunction	Confirm as a metric of cardiac contractility	Agreement^* with metric from IVCO (ICC)	Increase with inotropic stimulation	Decrease with HF	Confirm as a metric of cardiac contractility
E_es(IVC) *vs. single-beat surrogates of E*_es(IVC)

E _es(SB1)	YES (0.510)	YES	YES	YES	NO (0.193)	YES	YES	NO	NO

*PRSW_(IVC) vs. single-beat surrogates of PRSW*_(IVC)

PRSW _(SB)	YES (0.631)	NO	YES	NO	NO (0.084)	YES	YES	NO	NO
PWR _max -EDV	YES (0.637)	NO	YES	NO	YES (0.570)	NO	NO	NO	NO
PWR _max -EDV ²	YES (0.699)	NO	YES	NO	YES (0.537)	NO	YES	NO	NO
LVEF _EA1	YES (0.812)	YES	YES	YES	NO (0.330)	YES	YES	NO	NO

*dP/dt_max-EDV_(IVC) vs. single-beat surrogates of dP/dt*_max-EDV

_LV dP/dt _max -EDV	YES (0.700)	YES	YES	YES	YES (0.574)	YES	YES	YES	YES
P _max -EDV	YES (0.845)	YES	YES	YES	YES (0.649)	YES	YES	YES	YES
_LV ESP-EDV	YES (0.822)	YES	YES	YES	YES (0.670)	YES	YES	YES	YES

*Metrics obtained from arterial beat-by-beat baseline data*

E _es(SB2)	YES (0.522)	YES	YES	YES	NO (0.284)	YES	YES	NO	NO
_ART dP/dt _max -EDV	YES (0.758)	YES	YES	YES	NO (0.411)	YES	YES	NO	NO
_ART ESP-EDV	YES (0.822)	YES	YES	YES	YES (0.585)	YES	YES	YES	YES
LVEF _EA2	YES (0.799)	YES	YES	YES	NO (0.034)	YES	YES	NO	NO

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E_yes(IVC), end-systolic elastance obtained from inferior vena cava occlusion (IVCO); PRSW_(IVC), preload recruitable stork work obtained from IVCO; dP/dt_max-EDV_(IVC), the relation between the maximal rate of rise of pressure and end-diastolic volume (EDV) obtained from IVCO; E_es(SB1), left ventricular (LV) end-systolic pressure (_LVESP) and end-systolic volume (ESV) ratio; PRSW_(SB), stroke work (SW) and EDV ratio; PWR_max-EDV, power max (PWR_max) and EDV ratio; PWR_max-EDV², PWR_max and squared of EDV ratio; LVEF_EA1, afterload adjusted LV ejection fraction using ventricular data _LVdP/dt_max-EDV, the maximal rate of rise of the left ventricular pressure (_LVdP/dt_max) and EDV ratio; P_max-EDV, maximal pressure (P_max) and EDV ratio; _LVESP-EDV, _LVESP and EDV ratio; E_es(SB2), arterial ESP (_ARTESP, 0.9 × systolic blood pressure (SBP)) and ESV ratio; _ARTdP/dt_max-EDV, the maximal rate of rise of the arterial pressure (_ARTdP/dt_max) and EDV ratio; _ARTESP-EDV, _ARTESP and EDV ratio; LVEF_EA2, afterload adjusted LV ejection fraction using arterial data.

at least moderate agreement with IVCO-derived reference measure was considered for this validation step.

Performance of single beat metrics within species

As there is no single animal model that perfectly resembles the human cardiovascular system (Milani-Nejad & Janssen, 2014; Lelovas et al., 2014) and/or its clinical pathologies, researchers inevitably rely on employing various small and large animal models across different species to address questions concerning the pathophysiology and treatment of cardiovascular diseases. In the present study, using both small and large animal models with different cardiovascular pathologies (myocardial infarction, systolic heart failure, lack of bulbospinal sympathetic control) we were able to determine the best single-beat metric for each animal model studied and have also provided this information in our summary table (Table 10). In rodents, the majority of the previously reported/tested metrics, as well as our proposed metric (_LVESP-EDV), showed a moderate-to-excellent agreement with their IVCO-derived reference measure and met all the assumptions required to be considered as a valid cardiac contractile metric. As we transition to our large animal models, the number of valid single-beat estimates was much lower. Indeed, we found that only _LVESP-EDV, dP/dt_max-EDV, and P_max-EDV were valid single-beat metrics of LV contractility in both rats and pigs, as well as capable of revealing a progressive decline in cardiac contractility longitudinally in our conscious canines (only _LVESP-EDV and dP/dt_max-EDV). These findings clearly imply that when one considers employing single-beat metrics as a tool to quantify cardiac contractility in various experimental paradigms, the animal model studied should be taken into account.

Can arterial pressure be used as a surrogate for LV pressure and estimate cardiac contractility?

Another intriguing finding of the present study is the good-to-excellent agreement observed between single-beat metrics of cardiac contractility derived from baseline arterial data against either their corresponding IVCO-derived reference measure or single-beat metrics obtained from baseline LV data. Indeed, in addition to demonstrating a remarkably strong agreement with metrics obtained from LV baseline data, E_{es(SB2), ART}dP/dt_max-EDV, _ARTESP-EDV, and LVEF_EA2 derived from arterial baseline data showed a moderate-to-good agreement with their corresponding reference measure from IVCO in rats (see, Tables 2–5, 10). These metrics were also able to pass all validation steps considered in the present study. However, in pigs, only _ARTESP-EDV (see, Tables 7, 8, 10) appeared valid, suggesting that this metric is the only arterial data-derived single-beat metric that can be used in both small and large animal models to quantify cardiac contractile function. Accordingly, these intriguing observations confirm that single-beat surrogates derived from arterial data, in either animal model, can act as useful means of quantifying cardiac contractility when LV catheterization is infeasible. In theory, these indices all could be generated from a combination of telemetric arterial blood pressure recordings with echocardiography to measure cardiac volumes which would potentially allow for measuring/monitoring cardiac inotropic function longitudinally from within the same animal.

Potential factors mediating species differences

There are several plausible explanations as to why we found species differences in the usefulness of various indices of contractility. Perhaps one of the most relevant explanations for the species differences observed in the present study is the differences that exist in the cardiac contractile kinetics between species. It has been demonstrated in ex vivo preparations that species-related differences (i.e., mouse, rat, rabbit, canine, human) occur in cardiomyocyte excitation, calcium handling, and myofilament protein isoforms (Janssen & Periasamy, 2007). It is also possible that the anesthetics used for each individual animal might have influenced our findings. Indeed, such differences could be explained by the known impacts of various anesthetics on cardiovascular function and myocardial contractility (Vatner, 1978; Rusy & Komai, 1987; Barker et al., 1987). We tried to obviate such impacts by selecting anesthetics that have a minimal depressive effect on sympathetic tone and cardiac inotropy; however, it is still possible that the anesthetics per se exerted different effects on cardiac inotropic action across animals studied, presumably through inhibiting the influx of Ca²⁺ (Rusy & Komai, 1987) or influencing the supraspinal centers located in the brainstem that are associated with cardiovascular function. Moreover, the choice of experimental design employed to manipulate cardiac inotropic function differed between the species studied. It is plausible that these differences in design could be responsible for why we found species-related differences with respect to the performance of various single-beat metrics. For instance, in our small animal model (rodent) we included experiments testing the load-dependency of the examined metrics via LBNP, as well as their ability to identify impaired inotropic function in animal models with overt contractile dysfunction (i.e., myocardial infarction, high-thoracic SCI), whereas in our large animal model we tested the sensitivity of the examined metrics to inotropic stimulation using dobutamine (porcine) and their ability to diagnose/longitudinally track contractile dysfunction in a conscious animal model of systolic heart failure (canine). Lastly, the sex of the animals could be another factor underlying our observed species differences. However, whilst the rodents utilized in the present study were all males, our large animals were all females. Since we found at least some metrics that tracked well across all species we suspect any influence of sex on the validity of single-beat indices is limited. Nevertheless, important sex differences in a number of cardiac metrics do exist and as such it is reasonable that sex may have influenced some metrics more than others. For example, relative to males, smaller and shorter contractions have been reported for females in previous studies employing isolated cardiomyocytes (Parks & Howlett, 2013).

Clinical implications

Identifying contractile dysfunction in the clinic would be useful in a number of cardiovascular disease states. Although procedures such as caval occlusion and LV-PV catheterization are gaining traction in the clinic, such approaches are yet to be widely implemented. Moreover, such assessments require a cath lab and cannot be performed at the ‘bedside’ or in research laboratories that are interested in human cardiac physiology. The findings from the present series of experiments, validating various single-beat surrogates of cardiac contractility, are clinically intriguing since at least one of these surrogates can be estimated non-invasively using a combination of echocardiographic techniques and peripheral blood pressure assessment. Moreover, when LV catheterization is clinically indicated, it is typically performed with a ‘pressure-only’ catheter. By utilizing either a synchronous echocardiography or magnetic resonance imaging-derived measure of end-diastolic/-systolic volume, the clinician would be able to calculate almost every metric we validated in this manuscript. Of note is the metric P_max-EDV. Indeed, in a healthy heart and vasculature with no overt pathology (e.g., aortic valve disease, aortic stenosis) in which the slight difference in maximum systolic pressure between LV and peripheral artery (i.e., systolic brachial cuff pressure) can be disregarded, this metric may be of significant clinical interest given it can be assessed non-invasively at the bedside. In this regard, we hope that these findings will eventually allow for improved diagnostic capabilities in human cardiac disease.

Limitations and future directions

In the present study, we have used healthy animals to examine the agreements between metrics of interest and we only employed animal models of heart diseases to validate their ability in diagnosing contractile dysfunction. Future studies exploring the agreements between these indices across different experimental conditions in the same animal model of heart disease (e.g., only MI or systolic heart failure) and sex (male/female) are warranted. Another factor that needs to be taken into consideration when interpreting the findings of the present study is the matter of linearity or non-linearity of the measures derived from IVCO. In the current study, we used linear regression to construct end-systolic PV relationship (ESPVR) in order to be consistent with the majority of the contemporary literature, as well as between animals species studied; however, there is evidence reporting curvilinearity of ESPVR, especially in large animal models (Burkhoff et al., 1987; Kass et al., 1989; Su & Crozatier, 1989; Sato et al., 1998). This curvilinear relationship, however, appears to precipitate only when the first 5–6 beats of an IVCO are included in the analyses along with beats below a critical LV systolic pressure of 60–70mmHg. Whilst this study was not designed to further investigate the concept of ESPVR linearity this point is important as it could imply that a change in resting LV systolic pressure following an experimental intervention (i.e., SCI, MI, systolic heart failure) could alter the starting point from which an IVCO is performed and therefore place the animal on the steeper portion at the “bottom” of the typical LV IVCO curve, which would result in an “inflated ESPVR”. Whilst we acknowledge that three of our interventions did reduce LV systolic pressure (i.e., SCI, MI, systolic heart failure), peak LV systolic pressure was still well within the normal physiological range (80–100mmHg) and all interventions produced the expected reduction in ESPVR. As such, we believe it is unlikely that potential non-linearity in the gold-standard IVCO-derived measures of contractility impacted our results. Lastly, for indirect estimation of ESP (i.e., 0.9 × SBP) we acknowledge that whilst this estimation may be applicable in non-systolic disease it may not hold true in clinical situations like systolic dysfunction or systemic hypertension. This needs to be taken into consideration when interpreting data associated with metrics calculated using this formula in the current investigation.

Conclusions

Our primary finding is that P_max-EDV, _LVESP-EDV, and _LVdP/dt_max-EDV are the best 3 single-beat estimates to measure cardiac inotropic function in both small and large animal models. If cardiac catheterization is not available/feasible, then _ARTESP-EDV obtained from baseline beat-by-beat arterial data can be employed to quantify cardiac contractile function in either animal model. The latter could provide future utility for longitudinal studies when telemetric blood pressure measures are paired with ultrasound measures of EDV.

Supplementary Material

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NIHMS1836892-supplement-supinfo2.xlsx^{(74.7KB, xlsx)}

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NIHMS1836892-supplement-supinfo1.pdf^{(532.4KB, pdf)}

Key Points.

Validating and comparing indices of cardiac contractility that avoid caval occlusion would offer considerable advantages for the cardiovascular field.
We comprehensively test the underlying assumptions of multiple single-beat indices of cardiac contractility in rodents and translate these findings to pigs and conscious dogs.
We show that when performing caval occlusion is infeasible, single-beat metrics can be utilized to accurately quantify cardiac inotropic function in basic and preclinical research employing various small and large animal species.
We document that P_max-EDV, _LVESP-EDV, and _LVdP/dt_max-EDV derived from baseline hemodynamic recording are the best 3 single-beat metrics to measure cardiac inotropic function in both small and large animal models.

Acknowledgments

The authors gratefully acknowledge the veterinary staff at the UBC Center for Comparative Medicine (CCM), who support the complex in vivo porcine model experiments and who provide care for the animals involved in the studies.

Funding

Studies in rodents were supported by grants from the Blusson Integrated Cures Partnership (Grant number: GR016954) and NSERC (Grant number: RGPIN-2020-06240). The porcine studies were supported by a grant from The United States Department of Defense (Grant number: SC160098). The canine studies were supported by the National Heart, Lung and, Blood Institute Grants HL-055473 and HL-126706. Research in the laboratory of Dr. West is supported by an infrastructure grant from the Canadian Foundation for Innovation and the BC Knowledge Development Fund. Dr. Kwon is the Canada Research Chair in Spinal Cord Injury and holds the Dvorak Chair in Spine Trauma. Dr. Foster is a Michael Smith Foundation for Health Research Scholar.

ABBREVIATIONS AND ACRONYMS

EF: Ejection fraction
LV: Left ventricular
EDV: End-diastolic volume
PV: Pressure-volume
IVCO: Inferior vena cava occlusion
E_es(IVC): End-systolic elastance obtained from vena cava occlusion
PRSW(IVC): Pre-load recruitable stroke work obtained from vena cava occlusion
dP/dt_max-EDV(IVC): The relationship between the maximal rate of rise of ventricular pressure and end-diastolic volume obtained from vena cava occlusion
LBNP: Lower-body negative pressure
SV: Stroke Volume
LVdP/dt_max: Maximal rate of rise of ventricular pressure
ARTdP/dt_max: Maximal rate of rise of arterial pressure
ARTdP/dt_max-EDV: The maximal rate of rise of the arterial pressure and end-diastolic volume ratio
LVESP: End-systolic pressure obtained from ventricular pressure
ARTESP: End-systolic pressure obtained from arterial pressure
LVdP/dt_max-EDV: The maximal rate of rise of the left ventricular pressure and end-diastolic volume ratio
MAP: Mean arterial pressure
SBP: Systolic blood pressure
SCI: Spinal cord injury
LAD: Left anterior descending coronary artery
ICC: Intra-class correlation coefficients
LOA: Limit of agreement
SD: Standard deviations
E_es(SB1): Left ventricular end-systolic pressure and end-systolic volume ratio
E_es(SB2): Arterial end-systolic pressure and end-systolic volume ratio
PRSW_(SB): Stroke work and end-diastolic volume ratio
PWR_max-EDV: Maximal power and end-diastolic volume ratio
PWR_max-EDV2: Maximal power and squared of end-diastolic volume ratio
LVEF_EA1: Afterload adjusted left ventricular ejection fraction using ventricular data
LVEF_EA2: Afterload adjusted left ventricular ejection fraction using arterial data
LVdP/dt_max-EDV: The maximal rate of rise of the left ventricular pressure and end-diastolic volume ratio
P_max-EDV: Left ventricular maximal pressure and end-diastolic volume ratio
LVESP-EDV: Left ventricular end-systolic pressure and end-diastolic volume ratio
ARTESP-EDV: Arterial estimated end-systolic pressure and end-diastolic volume ratio
DSA: Anterior-to-posterior (short-axis) LV diameter
DLA: Apex-to-base (long-axis) LV diameter
ESPVR: End-systolic pressure volume relationship

Biography

graphic file with name nihms-1836892-b0009.gif

Mehdi Ahmadian is currently a PhD candidate in the Translational Integrative Physiology Laboratory (West lab) at the University of British Columbia, Canada. Mehdi has a keen interest in exploring factors affecting cardiovascular autonomic behavior in various health and disease scenarios, with the view of developing effective medical tools, procedures, and therapies for the field of cardiovascular (patho)physiology.

Footnotes

Additional information

Competing interests

The authors declare that they have no competing interests.

Data availability statement

All data are reported in the published article.

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Nakayama M, Chen C-H, Nevo E, Fetics B, Wong E & Kass DA (1998). Optimal preload adjustment of maximal ventricular power index varies with cardiac chamber size. Am Heart J 136, 281–288. [DOI] [PubMed] [Google Scholar]
Pacher P, Nagayama T, Mukhopadhyay P, Bátkai S & Kass DA (2008). Measurement of cardiac function using pressure–volume conductance catheter technique in mice and rats. Nat Protoc 3, 1422–1432. [DOI] [PMC free article] [PubMed] [Google Scholar]
Parks RJ & Howlett SE (2013). Sex differences in mechanisms of cardiac excitation–contraction coupling. Pflugers Arch 465, 747–763. [DOI] [PMC free article] [PubMed] [Google Scholar]
Poormasjedi-Meibod MS, Mansouri M, Fossey M, Squair JW, Liu J, McNeill JH & West CR (2019). Experimental Spinal Cord Injury Causes Left-Ventricular Atrophy and Is Associated with an Upregulation of Proteolytic Pathways. J Neurotrauma 36, 950–961. [DOI] [PubMed] [Google Scholar]
Rusy BF & Komai H (1987). Anesthetic depression of myocardial contractility: a review of possible mechanisms. Anesthesiology 67, 745–766. [DOI] [PubMed] [Google Scholar]
Sagawa K, Suga H, Shoukas AA & Bakalar KM (1977). End-systolic pressure/volume ratio: A new index of ventricular contractility. Am J Cardiol 40, 748–753. [DOI] [PubMed] [Google Scholar]
Sala-Mercado JA, Moslehpour M, Hammond RL, Ichinose M, Chen X, Evan S, O’Leary DS & Mukkamala R (2014). Stimulation of the cardiopulmonary baroreflex enhances ventricular contractility in awake dogs: a mathematical analysis study. Am J Physiol - Regul Integr Comp Physiol 307, R455–R464. [DOI] [PMC free article] [PubMed] [Google Scholar]
Sato T, Shishido T, Kawada T, Miyano H, Miyashita H, Inagaki M, Sugimachi M & Sunagawa K (1998). ESPVR of in situ rat left ventricle shows contractility-dependent curvilinearity. Am J Physiol - Hear Circ Physiol 274, H1429–H1434. [DOI] [PubMed] [Google Scholar]
Senzaki H, Chen C-H & Kass DA (1996). Single-beat estimation of end-systolic pressure-volume relation in humans: a new method with the potential for noninvasive application. Circulation 94, 2497–2506. [DOI] [PubMed] [Google Scholar]
Shishido T, Hayashi K, Shigemi K, Sato T, Sugimachi M & Sunagawa K (2000). Single-beat estimation of end-systolic elastance using bilinearly approximated time-varying elastance curve. Circulation 102, 1983–1989. [DOI] [PubMed] [Google Scholar]
Squair JW, DeVeau KM, Harman KA, Poormasjedi-Meibod M-S, Hayes B, Liu J, Magnuson DSK, Krassioukov AV & West CR (2018). Spinal cord injury causes systolic dysfunction and cardiomyocyte atrophy. J Neurotrauma 35, 424–434. [DOI] [PMC free article] [PubMed] [Google Scholar]
Squair JW, West CR, Popok D, Assinck P, Liu J, Tetzlaff W & Krassioukov AV (2017). High thoracic contusion model for the investigation of cardiovascular function after spinal cord injury. J Neurotrauma 34, 671–684. [DOI] [PubMed] [Google Scholar]
Su JB & Crozatier B (1989). Preload-induced curvilinearity of left ventricular end-systolic pressure-volume relations. Effects on derived indexes in closed-chest dogs. Circulation 79, 431–440. [DOI] [PubMed] [Google Scholar]
Suga H, Sagawa K & Shoukas AA (1973). Load Independence of the Instantaneous Pressure-Volume Ratio of the Canine Left Ventricle and Effects of Epinephrine and Heart Rate on the Ratio. Circ Res 32, 314–322. [DOI] [PubMed] [Google Scholar]
Takeuchi M, Igarashi Y, Tomimoto S, Odake M, Hayashi T, Tsukamoto T, Hata K, Takaoka H & Fukuzaki H (1991). Single-beat estimation of the slope of the end-systolic pressure-volume relation in the human left ventricle. Circulation 83, 202–212. [DOI] [PubMed] [Google Scholar]
Vatner SF (1978). Effects of anesthesia on cardiovascular control mechanisms. Environ Health Perspect 26, 193–206. [DOI] [PMC free article] [PubMed] [Google Scholar]
Wainman L, Erskine EL, Ahmadian M, Hanna TM & West CR (2021). Development of a Spinal Cord Injury Model Permissive to Study the Cardiovascular Effects of Rehabilitation Approaches Designed to Induce Neuroplasticity. Biology (Basel) 10, 1006. [DOI] [PMC free article] [PubMed] [Google Scholar]
Weir RAP, McMurray JJ V & Velazquez EJ (2006). Epidemiology of heart failure and left ventricular systolic dysfunction after acute myocardial infarction: prevalence, clinical characteristics, and prognostic importance. Am J Cardiol 97, 13–25. [DOI] [PubMed] [Google Scholar]
Williams AM, Manouchehri N, Erskine E, Tauh K, So K, Shortt K, Webster M, Fisk S, Billingsley A, Munro A, Tigchelaar S, Streijger F, Kim K-T, Kwon BK & West CR (2020). Cardio-centric hemodynamic management improves spinal cord oxygenation and mitigates hemorrhage in acute spinal cord injury. Nat Commun 11, 5209. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

supinfo2

NIHMS1836892-supplement-supinfo2.xlsx^{(74.7KB, xlsx)}

supinfo1

NIHMS1836892-supplement-supinfo1.pdf^{(532.4KB, pdf)}

Data Availability Statement

All data are reported in the published article.

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[R56] Williams AM, Manouchehri N, Erskine E, Tauh K, So K, Shortt K, Webster M, Fisk S, Billingsley A, Munro A, Tigchelaar S, Streijger F, Kim K-T, Kwon BK & West CR (2020). Cardio-centric hemodynamic management improves spinal cord oxygenation and mitigates hemorrhage in acute spinal cord injury. Nat Commun 11, 5209. [DOI] [PMC free article] [PubMed] [Google Scholar]

PERMALINK

A cross-species validation of single-beat metrics of cardiac contractility

Mehdi Ahmadian

Alexandra M Williams

Joseph Mannozzi

Filip Konecny

Ryan L Hoiland

Liisa Wainman

Erin Erskine

Jennifer Duffy

Neda Manouchehri

Kitty So

Keerit Tauh

Javier A Sala-Mercado

Katelyn Shortt

Shera Fisk

Kyoung-Tae Kim

Femke Streijger

Glen E Foster

Brian K Kwon

Donal S O’Leary

Christopher R West

Abstract

Graphical Abstract

INTRODUCTION

METHODS

Ethical approval

Animals and experiments

Experiment 1: Examining the agreement between examined metrics in rats

General preparation

Cardiovascular catheterization and data acquisition

Figure 1. An example of invasive cardiac contractility metrics calculated from the left ventricle pressure-volume loop after preload manipulation via vena cava occlusion in a rat and a pig.

Table 1.

Experiment 2: Testing the load dependency of examined metrics in rats

Figure 2. Experimental preparation used for lower body negative pressure (LBNP) manipulation.

Experiments 3 and 4: Ability of the examined metrics to diagnose contractile dysfunction in animals with absent bulbospinal sympathetic outflow to the heart (3) and myocardial infarction (4)

Experiments 5 and 6: Investigating whether any potential agreements observed between examined metrics in small rodents are translatable to large porcine models (5), and testing the behavior of examined metrics to direct inotropic simulation (6)

Experiments 7 and 8: Investigating whether examined metrics are capable of diagnosing weakened contractile function in a conscious dog model of heart failure (7), as well as revealing a progressive decline in cardiac contractile function over time and during transition to heart failure (8)

Calculation of field-standard metrics derived from the IVCO and single-beat surrogates

Statistical analysis

RESULTS

Rats

Agreement between single-beat metrics and those obtained from IVCO

Figure 3. Agreement between metrics in rodents.

Table 2.

Table 3.

Table 4.

Table 5.

Responses of single-beat metrics to LBNP manipulation

Figure 4. Responses of the metrics to lower-body negative pressure manipulations in rats.

Table 6.

Ability of single-beat metrics to detect contractile dysfunction in animals with absent bulbospinal sympathetic control and myocardial infarction

Figure 5. Ability of the metrics to detect cardiac dysfunction in rats (mean ± SD).

Pigs

Agreement between single-beat metrics and those obtained from IVCO

Figure 6. Agreement between metrics in pigs.

Table 7.

Responses of single-beat metrics to an inotropic stimulation

Table 8.

Conscious dogs

The ability of single-beat metrics to detect contractile dysfunction in the setting of systolic heart failure

Table 9.

Figure 7. Ability of the metrics to detect cardiac dysfunction longitudinally in conscious dogs.

DISCUSSION

Single-beat metrics that can be used in both small and large animal models

Table 10.

Performance of single beat metrics within species

Can arterial pressure be used as a surrogate for LV pressure and estimate cardiac contractility?

Potential factors mediating species differences

Clinical implications

Limitations and future directions

Conclusions

Supplementary Material

Key Points.

Acknowledgments

Funding

ABBREVIATIONS AND ACRONYMS

Biography

Footnotes

Data availability statement