A cross-department collaboration headed by researchers at Harvard Medical School and Harvard University has witnessed the moment at which cells in the zebrafish embryo heart start beating in unison for the very first time. Co-led by Sean Megason, PhD, professor of systems biology at the Blavatnik Institute, HMS, the researchers used advanced optical electrophysiology tools to observe the first few beats of developing zebrafish hearts and to assess the underlying electrical excitability and connectivity patterns.

The team discovered that the heart cells start beating suddenly and all at once as calcium levels and electrical signals increase. Their findings also showed that each heart cell has the ability to beat on its own, without a pacemaker, and that the heartbeat can start in different places.

The researchers suggest that studying the basic biology of the heartbeat could help scientists better understand cardiac rhythm disorders in humans. “People place such importance on the heart beating that it’s been a focal point of investigations for a long time, but this is the first time we’ve been able to look at it in depth with so much resolution,” said Megason, who is co-senior author of the team’s published paper in Nature, titled “A bioelectrical phase transition patterns the first vertebrate heartbeats.”

The development out of just a handful of cells, of a complete organism with functioning tissues and organs is a highly synchronized process that requires cells to organize themselves in a precise manner, and begin working together. This cellular cooperation is especially dramatic in the heart, where static cells must start beating in perfect unison. As the authors wrote, “A first heartbeat is a once-in-a-lifetime event.”

Understanding the fundamental mechanisms underlying the heartbeat could also aid our understanding of what is happening when the cardiac system that regulates the heartbeat doesn’t develop properly, or begins to malfunction. But there are gaps in knowledge, the investigators noted. “The initial transition of the heart from silent to beating has never been characterized at the timescale of individual electrical events, and the structure in space and time of the early heartbeats remains poorly understood.” Co-senior author Adam Cohen, PhD, professor of chemistry and chemical biology and of physics at Harvard, added, “The heart beats about three billion times in a typical human lifetime, and it must never take a break,” said. “We wanted to see how this incredible machine first turns on.”

The researchers didn’t set out to study how the heart starts beating. Rather, they were casting about for a scientific question that would combine the Cohen lab’s expertise in imaging electrical activity with the Megason lab’s interest in studying how cells in developing zebrafish learn to communicate and cooperate. This then led them to consider the heart.

They realized that despite millennia of research on the developing heart, stretching all the way back to Aristotle’s observations in chicks, details about how heart cells start beating were still a mystery—one that they could potentially solve. “This scarcity of data means that the bioelectrical mechanisms for the emergence of organized cardiac function are still poorly understood,” the team commented. “We asked, how does the heart go from silent to regular beating? What are the intermediate activity states, and how do these states emerge from the ensemble of single-cell developmental trajectories.”

Megason further explained, “We wanted to answer a basic question: How do heart cells go from silent to beating? When your heart starts is a once-in-a-lifetime event, but it’s not obvious how that happens.”

As an exploratory study, the team didn’t know what they would find out. Maybe a few cells would start beating, and the beating area would slowly grow, they speculated, on different parts of the heart would start beating independently and eventually merge, or the heart would start with weak beats that would strengthen over time. Their results, in fact, indicated a very different scenario.

Using fluorescent proteins and high-speed microscope imaging, the researchers captured changes in calcium levels and electrical activity in heart cells of developing zebrafish embryos. “We sought to capture this event by means of calcium imaging as zebrafish embryos developed from 18–22 hours postfertilization,” they explained.

To the investigators’  surprise, they discovered that all the heart cells abruptly transitioned from not beating to beating—characterized by simultaneous spikes in calcium and electrical signals—and they immediately began beating in sync. “It was like somebody had flipped on a switch,” Cohen said.

Further experiments revealed that for each heartbeat, one region of the heart fires first, initiating a wave of electricity that rapidly flows through the rest of the cells and prompts them to fire. And interestingly, the heartbeats started from different spots in different zebrafish, suggesting that there’s nothing unique about the cells that fire first. This finding was counterintuitive because cells in adult hearts behave differently. As the authors noted in their paper, “The first few beats appeared suddenly, had irregular interbeat intervals, propagated coherently across the primordial heart and emanated from loci that varied between animals and over time.”

“Unlike the adult heart, where a specialized population of pacemaker cells drives the heartbeat, most cells in the embryonic heart have the ability to beat on their own, making it difficult to predict the location of the first beats,” said lead author Bill Jia, a joint graduate student in the Cohen and Megason labs. The investigators further commented, “Our work shows how gradual and largely asynchronous development of single-cell bioelectrical properties produces a stereotyped and robust tissue-scale transition from quiescence to coordinated beating.”

Because the heart cells start beating instantaneously, they must develop the ability to beat and sense their neighbors’ beating before their very first heartbeat—something Megason compares to an army that has to start marching in sync without practising first. “The heart first learns how to keep pace without a clock, and individual cells first learn to cooperate without agreeing on what their roles are,” Jia added. “It is very important for the heartbeat to be regular, but it is organized very quickly at the start of life from what seems to be a total mess.”

Developing zebrafish offer a convenient model for studying the heart because they are transparent, grow quickly—developing a heartbeat in only 24 hours—and can be imaged by the dozen.  However, Megason thinks the same developmental process may be conserved across species, including humans. This finding, the team noted, opens the door to learning more about the development of heartbeat across species, and may one day illuminate how cardiac irregularities such as arrhythmias arise in humans. “By looking at how the heart develops, we can see how different control mechanisms are layered on, which may tell us something about what happens if they break down,” Megason commented.

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