Small animals support a wide range of pathological phenotypes and genotypes as versatile, affordable models for pathogenesis of cardiovascular diseases and for exploration of strategies in electrotherapy, gene therapy, and optogenetics. Pacing tools in such contexts are currently limited to tethered embodiments that constrain animal behaviors and experimental designs. Here, we introduce a highly miniaturized wireless energy-harvesting and digital communication electronics for thin, miniaturized pacing platforms weighing 110 mg with capabilities for subdermal implantation and tolerance to over 200,000 multiaxial cycles of strain without degradation in electrical or optical performance. Multimodal and multisite pacing in ex vivo and in vivo studies over many days demonstrate chronic stability and excellent biocompatibility. Optogenetic stimulation of cardiac cycles with in-animal control and induction of heart failure through chronic pacing serve as examples of modes of operation relevant to fundamental and applied cardiovascular research and biomedical technology.

The spontaneous rhythmic action potentials generated by the sinoatrial node (SAN), the primary pacemaker in the heart, dictate the regular and optimal cardiac contractions that pump blood around the body. Although the heart rate of humans is substantially slower than that of smaller experimental animals, current perspectives on the biophysical mechanisms underlying the automaticity of sinoatrial nodal pacemaker cells (SANCs) have been gleaned largely from studies of animal hearts. Using human SANCs, we demonstrated that spontaneous rhythmic local Ca2+ releases generated by a Ca2+ clock were coupled to electrogenic surface membrane molecules (the M clock) to trigger rhythmic action potentials, and that Ca2+–cAMP–protein kinase A (PKA) signaling regulated clock coupling. When these clocks became uncoupled, SANCs failed to generate spontaneous action potentials, showing a depolarized membrane potential and disorganized local Ca2+ releases that failed to activate the M clock. β-Adrenergic receptor (β-AR) stimulation, which increases cAMP concentrations and clock coupling in other species, restored spontaneous, rhythmic action potentials in some nonbeating “arrested” human SANCs by increasing intracellular Ca2+ concentrations and synchronizing diastolic local Ca2+ releases. When β-AR stimulation was withdrawn, the clocks again became uncoupled, and SANCs reverted to a nonbeating arrested state. Thus, automaticity of human pacemaker cells is driven by a coupled-clock system driven by Ca2+-cAMP-PKA signaling. Extreme clock uncoupling led to failure of spontaneous action potential generation, which was restored by recoupling of the clocks. Clock coupling and action potential firing in some of these arrested cells can be restored by β-AR stimulation–induced augmentation of Ca2+-cAMP-PKA signaling.

It is highly debated how cyclic adenosine monophosphate-dependent regulation (CDR) of the major pacemaker channel HCN4 in the sinoatrial node (SAN) is involved in heart rate regulation by the autonomic nervous system. We addressed this question using a knockin mouse line expressing cyclic adenosine monophosphate-insensitive HCN4 channels. This mouse line displayed a complex cardiac phenotype characterized by sinus dysrhythmia, severe sinus bradycardia, sinus pauses and chronotropic incompetence. Furthermore, the absence of CDR leads to inappropriately enhanced heart rate responses of the SAN to vagal nerve activity in vivo. The mechanism underlying these symptoms can be explained by the presence of nonfiring pacemaker cells. We provide evidence that a tonic and mutual interaction process (tonic entrainment) between firing and nonfiring cells slows down the overall rhythm of the SAN. Most importantly, we show that the proportion of firing cells can be increased by CDR of HCN4 to efficiently oppose enhanced responses to vagal activity. In conclusion, we provide evidence for a novel role of CDR of HCN4 for the central pacemaker process in the sinoatrial node.

Temporary postoperative cardiac pacing requires devices with percutaneous leads and external wired power and control systems. This hardware introduces risks for infection, limitations on patient mobility, and requirements for surgical extraction procedures. Bioresorbable pacemakers mitigate some of these disadvantages, but they demand pairing with external, wired systems and secondary mechanisms for control. We present a transient closed-loop system that combines a time-synchronized, wireless network of skin-integrated devices with an advanced bioresorbable pacemaker to control cardiac rhythms, track cardiopulmonary status, provide multihaptic feedback, and enable transient operation with minimal patient burden. The result provides a range of autonomous, rate-adaptive cardiac pacing capabilities, as demonstrated in rat, canine, and human heart studies. This work establishes an engineering framework for closed-loop temporary electrotherapy using wirelessly linked, body-integrated bioelectronic devices.

Rationale: Cardiac function is under exquisite intrinsic cardiac neural control. Neuroablative techniques to modulate control of cardiac function are currently being studied in patients, albeit with variable and sometimes deleterious results. Objective: Recognizing the major gaps in our understanding of cardiac neural control, we sought to evaluate neural regulation of impulse initiation in the sinoatrial node as an initial discovery step. Methods and Results: We report an in-depth, multi-scale structural and functional characterization of the innervation of the sinoatrial node (SAN) by the right atrial ganglionated plexus (RAGP) in porcine and human hearts. Combining intersectional strategies including tissue clearing, immunohistochemical and ultrastructural techniques, we have delineated a comprehensive neuroanatomic atlas of the RAGP-SAN complex. The RAGP shows significant phenotypic diversity of neurons while maintaining predominant cholinergic innervation. Cellular and tissue-level electrophysiologic mapping and ablation studies demonstrate interconnected ganglia with synaptic convergence within the RAGP to modulate SAN automaticity, atrioventricular (AV) conduction and left ventricular (LV) contractility. Using this approach, we comprehensively demonstrate that intrinsic cardiac neurons influence the pacemaking site in the heart. Conclusions: This report provides an experimental demonstration of a discrete neuronal population controlling a specific geographic region of the heart (SAN) that can serve as a framework for further exploration of other parts of the intrinsic cardiac nervous system (ICNS) in mammalian hearts and for developing targeted therapies.

Transparent microelectrodes have recently emerged as a promising approach for crosstalk‐free multifunctional electrical and optical biointerfacing. High‐performance flexible platforms that allow seamless integration with soft tissue systems for such applications are urgently needed. Here, silver nanowires (Ag NWs)‐based transparent microelectrode arrays (MEAs) and interconnects are designed to meet this demand. The nanowire networks exhibit a high optical transparency >90.0% at 550 nm, and superior mechanical stability up to 100,000 bending cycles at 5 mm radius. The Ag NWs microelectrodes preserve low normalized electrochemical impedance of 3.4–15 Ω cm2 at 1 kHz, and the interconnects demonstrate excellent sheet resistance (Rsh) of 4.1–25 Ω sq−1. In vivo histological analysis reveals that the Ag NWs structures are biocompatible. Studies on Langendorff‐perfused mouse and rat hearts demonstrate that the Ag NWs MEAs enable high‐fidelity real‐time monitoring of heart rhythm during co‐localized optogenetic pacing and optical mapping. This proof‐of‐concept work illustrates that the solution‐processed, transparent, and flexible Ag NWs structures are a promising candidate for the next‐generation of large‐area multifunctional biointerfaces for interrogating complex biological systems in basic and translational research.

Monitoring and control of cardiac function are critical for investigation of cardiovascular pathophysiology and developing life-saving therapies. However, chronic stimulation of the heart in freely moving small animal subjects, which offer a variety of genotypes and phenotypes, is currently difficult. Specifically, real-time control of cardiac function with high spatial and temporal resolution is currently not possible. Here, we introduce a wireless battery-free device with on-board computation for real-time cardiac control with multisite stimulation enabling optogenetic modulation of the entire rodent heart. Seamless integration of the biointerface with the heart is enabled by machine learning–guided design of ultrathin arrays. Long-term pacing, recording, and on-board computation are demonstrated in freely moving animals. This device class enables new heart failure models and offers a platform to test real-time therapeutic paradigms over chronic time scales by providing means to control cardiac function continuously over the lifetime of the subject.

OBJECTIVES This study sought to investigate the shift of leading pacemaker locations in healthy and failing mammalian hearts over the entire range of physiological heart rates (HRs), and to molecularly characterize spatial regions of spontaneous activity. BACKGROUND A normal heartbeat originates as an action potential in a group of pacemaker cells known as the sinoatrial node (SAN), located near the superior vena cava. HRs and the anatomical site of origin of pacemaker activity in the adult heart are known to dynamically change in response to various physiological inputs, yet the mechanism of this pacemaker shift is not well understood. METHODS Optical mapping was applied to ex vivo rat and human isolated right atrial tissues, and HRs were modulated with acetylcholine and isoproterenol. RNA sequencing was performed on tissue areas that elicited spontaneous activity, and comparisons were made to neighboring myocardial tissues. RESULTS Functional and molecular evidence identified and confirmed the presence of 2 competing right atrial pacemakers localized near the superior vena cava and the inferior vena cava—the superior SAN (sSAN) and inferior SAN (iSAN), respectively—which preferentially control the fast and slow HRs. Both of these regions were evident in non-failing rat and human hearts and maintained spontaneous activity in the rat heart when physically separated from one another. Molecular analysis of these 2 pacemaker regions revealed unique but similar transcriptional profiles, suggesting iSAN dominance when the sSAN is silent. CONCLUSIONS The presence of 2 spatially distinct dominant pacemakers, sSAN and iSAN, in the mammalian heart clarifies previous identification of migrating pacemakers and corresponding changes in P-wave morphology in mammalian species.