## 2 Materials and Methods
### Study Protocol  
Each participant underwent a medical evaluation, including an electrocardiogram (ECG) and two-dimensional echocardiography at the time of enrollment.  
Medical examination and electrocardiogram. A full medical history was collected from each participant. A physical examination and a 12-lead ECG were performed. Any evidence of ischemic cardiomyopathy, arterial hypertension, atrial fibrillation, diabetes mellitus, and/or dyslipidemia was documented together with any history of chronic kidney disease and dialysis. New York Heart Association (NYHA) classes, anthropometric data, systolic and diastolic arterial pressures, and heart rhythms were also recorded. If available, the most recent routine blood chemistries were also documented.  
Echocardiographic evaluation. Two-dimensional echocardiographic imaging and Doppler acquisition were performed using an EPIQ CVx system (Philips, Amsterdam, The Netherlands) with an S5-1 transducer. At baseline, five cycles of ECG-guided standard parasternal long and short-axis, apical two-, three-, and four-chamber and subcostal views were acquired. LV end-diastolic diameter and the thicknesses of the interventricular septum and LV posterior wall were measured; the LV mass indexed for body surface area was calculated based on current recommendations [6]. A RV-focused four-chamber view was also obtained in order to analyze the tricuspid annular plane systolic excursion (TAPSE). LV end-diastolic volume (LVEDV), LV end-systolic volume (LVESV), and LVEF were calculated using Simpson's rule. Left atrial volume was also calculated using Simpson's rule and indexed for body surface area. Mitral (MR) and tricuspid regurgitation (TR) were evaluated semi-quantitatively using color Doppler and assigned arbitrary units ranging from 0 to 4. The maximum trans-tricuspid valve pressure gradient was assessed by continuous-wave Doppler. Pulmonary systolic arterial pressure (PASP) was estimated based on the diameter of the vena cava. We performed pulsed-wave Doppler and measured the diameter of the LV outflow tract according to recent guidelines [6]. Stroke volume, cardiac output, and cardiac index were also obtained from these measurements [6]. Peaks of early wave velocity at the mitral valve (E) were measured by pulsed Doppler. The early diastolic velocity peaks (e') at the level of the septal (e' s) and lateral (e' l) mitral annulus were also measured by pulsed and tissue Doppler imaging (TDI) to obtain the E/e' ratio [6]. The Tei index was calculated as the ratio between the sum of isovolumic relaxation and contraction times divided by the mitral valve closure time as measured by TDI at the level of the septal mitral annulus.  
The AutoStrain application of the Philips EPIQ CVx ultrasound systems was used for \"off-cart\" analysis of strain from stored examinations. Values for LVGLS were obtained from the analysis of two-, three-, and four-chamber views. The software automatically generated curves representing longitudinal strain; LVGLS was calculated by averaging values obtained in all segments. The operator can also modify the region of interest generated from the software which will then automatically recalculate this value. The RVGLS and RVfwLS were also calculated automatically from values obtained in an RV-focused four-chamber view. In this manuscript, reduced systolic strain is indicated by values that are less negative than those determined at baseline. Right ventricular arterial coupling was determined as the ratio of RVGLS/PASP and RVfwLS/PASP.  
Pacing protocol. HR was increased from each baseline value by programming the implantable devices with higher atrial stimulation frequencies to a maximum of 90 beats/min. The stability of RA pacing capture was established within the first minute of each newly established HR. Echocardiography was performed at each increase of 10 beats/min as previously described. Intermediate resting frequencies were estimated to the nearest integer for the calculation of the first paced 10 beat/min rate increase.