&#65279;This work was supported by research grants from the Fukuda Foundation for Medical Technology (to E.T. and G. N.) and the JSPS KAKENHI Scientific Research Grant-in-Aid 21K08047 (to E.T.).

This animal protocol was approved by the Institutional Animal Care and Use Committee and followed the regulations for animal experiments and related activities at the University of Tokyo. For the present study, 8-12 week old male C57/Bl6J mice were used. The animals were obtained from a commercial source (see the Table of Materials). A model of HFpEF was established by administering a high-fat diet for 15 weeks in conjunction with NG-nitro-L-arginine methyl ester, as described previously<sup class="xref"...

Exploring HR-Dependent Cardiac Performance Using an Atrial-Pacing Protocol

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We present a methodology to assess pressure-volume relationships with the application of transesophageal pacing. Exercise intolerance is one of the key characteristics of HFpEF, yet there are no techniques available for the evaluation of cardiac function in mice during exercise. Our pacing protocol offers a valuable tool for detecting diastolic dysfunction, which may not be apparent under resting conditions.
To achieve a PV loop of accurate and consistent quality, the following steps must be m...

Heart failure represents a leading cause of hospitalization and death across the globe, and heart failure with preserved ejection fraction (HFpEF) accounts for around 50% of all heart failure diagnoses. HFpEF is characterized by diastolic dysfunction and impaired exercise tolerance, and the associated hemodynamic abnormalities, such as diastolic dysfunction, can be clearly detected through exercise-stressed hemodynamic testing or MRI scans1,2.
In experimental models, however, available modalities for assessing the physiological abnormalities related to HFpEF are limited3,4. Treadmill exercise testing (TMT) is used to determine running time and distance, which might reflect exercise-stress cardiac hemodynamics; however, this method is susceptible to interference from extraneous variables such as the body weight, skeletal muscle strength, and mental status.
To circumvent these limitations, we have devised an atrial-pacing protocol that detects subtle but crucial changes in diastolic performance based on the heart rate (HR) and have validated its usefulness in a mouse model of HFpEF5. Several physiological factors contribute to exercise-related cardiac function, including the sympathetic nerve and catecholamine response, peripheral vasodilation, the endothelial response, and the heart rate6. Among these, however, the HR-pressure relationship (also called the Bowditch effect) is known as a critical determinant of cardiac physiological features7,8,9.
The protocol involves performing a conventional pressure-volume analysis at baseline to assess the systolic and diastolic function, including parameters such as the rate of pressure development (dp/dt), the end-systolic pressure-volume relationship (ESPVR), and the end-diastolic pressure-volume relationship (EDPVR). However, it should be noted that these parameters are influenced by the HR, which can vary between animals due to differences in their intrinsic heart rate. Additionally, the effects of anesthesia on the HR should also be considered. To address this, the HR was standardized by administering atrial pacing concomitantly with ivabradine, and cardiac parameter measurements were performed at incremental heart rates. Notably, the HR-dependent cardiac response distinguished HFpEF mice from the control group mice, while no significant differences were observed in the baseline PV loop measurements (using the intrinsic heart rate)5.
While this pacing protocol may seem relatively complicated, its success rate exceeds 90% when it is well understood. This protocol would provide a useful way to study the underlying mechanisms of diastolic dysfunction in HFpEF mouse models and help in the development of new treatments for this condition.

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	<tbody>
		<tr>
			<td>2-0 silk suture, sterlie</td>
			<td>Alfresa Pharma Corporation, Osaka, Japan</td>
			<td>62-9965-57</td>
			<td>Surgical Supplies</td>
		</tr>
		<tr>
			<td>2-Fr tetrapolar electrode catheter</td>
			<td>Fukuda Denshi, Japan and UNIQUE MEDICAL, Japan</td>
			<td>custom-made</td>
			<td>Surgical Supplies</td>
		</tr>
		<tr>
			<td>Albumin Bovine Serum</td>
			<td>Nacalai Tesque, Inc., Kyoto, Japan</td>
			<td>01859-47</td>
			<td>Miscellaneous</td>
		</tr>
		<tr>
			<td>C57/BI6J mouse</td>
			<td>Jackson Laboratory</td>
			<td></td>
			<td>animals</td>
		</tr>
		<tr>
			<td>Conductance catheter</td>
			<td>Millar Instruments, Houston, TX</td>
			<td>PVR 1035</td>
			<td></td>
		</tr>
		<tr>
			<td>Electrical cautery, Electrocautery Knife Kit</td>
			<td>ellman-Japan,Osaka, Japan</td>
			<td>1-1861-21</td>
			<td>Surgical Supplies</td>
		</tr>
		<tr>
			<td>Etomidate</td>
			<td>Tokyo Chemical Industory Co., Ltd., Tokyo Japan</td>
			<td>E0897</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Grass Instrument S44G Square Pulse Stimulator</td>
			<td>Astro-Med, West Warwick, RI</td>
			<td></td>
			<td>Pacing equipment</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Viatris Inc., Tokyo, Japan</td>
			<td>8803998</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Ivabradine</td>
			<td>Tokyo Chemical Industory Co., Ltd., Tokyo Japan</td>
			<td>I0847</td>
			<td>Miscellaneous</td>
		</tr>
		<tr>
			<td>LabChart software</td>
			<td>ADInstruments, Sydney, Australia</td>
			<td>LabChart 7</td>
			<td>Hemodynamic equipment</td>
		</tr>
		<tr>
			<td>MPVS Ultra</td>
			<td>Millar Instruments, Houston, TX</td>
			<td>PL3516B49</td>
			<td>Hemodynamic equipment</td>
		</tr>
		<tr>
			<td>Pancronium bromide</td>
			<td>Sigma Aldrich Co., St. Louis, MO</td>
			<td>15500-66-0</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>PE10 polyethylene tube</td>
			<td>Bio Research Center&#160; Co. Ltd., Tokyo, Japan</td>
			<td>62101010</td>
			<td>Surgical Supplies</td>
		</tr>
		<tr>
			<td>PowerLab 8/35</td>
			<td>ADInstruments, Sydney, Australia</td>
			<td>PL3508/P</td>
			<td>Hemodynamic equipment</td>
		</tr>
		<tr>
			<td>PVR 1035</td>
			<td>Millar Instruments, Houston, TX</td>
			<td>842-0002</td>
			<td>Hemodynamic equipment</td>
		</tr>
		<tr>
			<td>Urethane (Ethyl Carbamate)</td>
			<td>Wako Pure Chemical Industries, Ltd., Osaka, Japan</td>
			<td>050-05821</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Vascular Flow Probe</td>
			<td>Transonic, Ithaca, NY</td>
			<td>MA1PRB</td>
			<td>Surgical Supplies</td>
		</tr>
	</tbody>
</table>

a pacing-controlled procedure for the assessment of heart rate-dependent diastolic functions in murine heart failure models

Heart failure with preserved ejection fraction (HFpEF) is a condition characterized by diastolic dysfunction and exercise intolerance. While exercise-stressed hemodynamic tests or MRI can be used to detect diastolic dysfunction and diagnose HFpEF in humans, such modalities are limited in basic research using mouse models. A treadmill exercise test is commonly used for this purpose in mice, but its results can be influenced by body weight, skeletal muscle strength, and mental state. Here, we describe an atrial-pacing protocol to detect heart rate (HR)-dependent changes in diastolic performance and validate its usefulness in a mouse model of HFpEF. The method involves anesthetizing, intubating, and performing pressure-volume (PV) loop analysis concomitant with atrial pacing. In this work, a conductance catheter was inserted via a left ventricular apical approach, and an atrial pacing catheter was placed in the esophagus. Baseline PV loops were collected before the HR was slowed with ivabradine. PV loops were collected and analyzed at HR increments ranging from 400 bpm to 700 bpm via atrial pacing. Using this protocol, we clearly demonstrated HR-dependent diastolic impairment in a metabolically induced HFpEF model. Both the relaxation time constant (Tau) and the end-diastolic pressure-volume relationship (EDPVR) worsened as the HR increased compared to control mice. In conclusion, this atrial pacing-controlled protocol is useful for detecting HR-dependent cardiac dysfunctions. It provides a new way to study the underlying mechanisms of diastolic dysfunction in HFpEF mouse models and may help develop new treatments for this condition.

Author Spotlight: Investigating HR-Dependent Cardiac Function in Mouse Models Through a Novel Atrial-Pacing Approach 

A Pacing-Controlled Procedure for the Assessment of Heart Rate-Dependent Diastolic Functions in Murine Heart Failure Models

The heart rate-related relationship is important for determining cardiac characteristics, particularly, in HFpEF. This protocol aims to assess heart rate-dependent cardiac function. Any experimental models, available modalities for assessing heart rate-related cardiac function, are limited to treadmill exercise testing.The treadmill test, however, is influenced by variables such as body weight. To overcome these limitations, we have developed around atrial-pacing protocol to detect subtle, yet significant changes in the later performance that are linked to heart rate. Our protocol offers a variable tool for identifying cardiac dysfunction, manifested by heart rate changes.It also presents a number approach to studying diastolic dysfunction in mirroring heart failure models, particularly in HFpEF.

The baseline PV loop data are displayed in Figure 1 and Table 1. At baseline (in the absence of pacing), there were no significant differences in diastolic parameters such as the relaxation time constant (Tau), the minimum rate of pressure change (dP/dt min), and EDPVR between the control and HFpEF mice. However, the HFpEF mice exhibited higher blood pressure and arterial elastance (Ea), as shown in Figure 1, and demonstrated a typical mountain-...

Watch this Scientific Journal Video about Investigating HR-Dependent Cardiac Function in Mouse Models Through a Novel Atrial-Pacing Approach at JoVE.com

The present protocol describes obtaining the pressure-volume relationship through transesophageal pacing, which serves as a valuable tool in evaluating diastolic function&#160;in mouse models of heart failure.

Heart failure with preserved ejection fraction (HFpEF) is a condition characterized by diastolic dysfunction and exercise intolerance. While ...

a-pacing-controlled-procedure-for-assessment-heart-rate-dependent