HAS anyone told you lately you’re electric? Well, you are. Your every pore oozes with the stuff. Must be all those ions you’ve been pumping. And we’re not just talking about nerve impulses here: every surface of your body, from your skin to your cell membranes, is humming with electrical activity.
Biologists have known for more than 200 years that nerve impulses are transmitted electrically. But only recently have they started eavesdropping on the electrical chatter of the rest of your body, and have discovered that electricity, in the form of electric fields, plays a vital role in numerous biological processes from embryonic development to cell division, nerve regeneration and wound repair. “The phenomenon is broadly applicable and I think we have only scratched the surface of something that is evolutionarily highly conserved and widely used,” says Colin McCaig of the University of Aberdeen, UK, who has been working on the biological effects of electric fields since the 1980s.
The first report that electric fields could influence the behaviour of individual cells came in 1920 when a Danish researcher, Sven Ingvar, showed that an externally applied field caused cultured chick neurons to grow in a particular direction. The years that followed saw a plethora of similar studies, but many were sloppily done and for years the interpretation of results was hampered by a lack of adequate recording techniques, contamination of cultures by electrode by-products, uncertainty about the magnitude of fields and sometimes by the complexity of the cells under study. The fact that some early researchers made exaggerated claims about the curative effects of electrical fields in nerve and limb regeneration hardly helped. Then, in the 1940s, Paul Weiss, a distinguished and influential biologist at the University of Chicago, said that he could not reproduce Ingvar’s findings and concluded that electric fields had no effect on cells.
Research into the biological effect of electrical fields gradually fell out of fashion and got left behind. “In the olden days, when people were really hot on this topic, they didn’t have molecular tools, and when molecular biology took off, these fascinating results were forgotten,” explains cell biologist Michael Levin of the Forsyth Institute in Boston, Massachusetts. Little wonder, then, that very few scientists today believe that electric fields have much physiological relevance and fewer still are doing any research in the area.
But in the past two decades, there has been a slow, steady revival in electric field research, and a small group of biologists is now presenting the case that they play an important role in biology.
Back in 1981, Kenneth Robinson of Purdue University in West Lafayette, Indiana, decided to redo Ingvar’s 1920 experiment. Instead of using large bundles of neurons containing thousands of cells, Robinson cultured a single frog neuron. He found that the nerve was acutely sensitive to the electric field – its long filament-like arms, called neurites, grew enthusiastically towards the negative electrode. “We showed that cells could respond to these small fields, which made it reasonable to imagine that during development or neuronal repair there might be natural electrical fields involved,” says Robinson, who has been studying electrical fields ever since. McCaig, who was part of Robinson’s team, adds: “I just could not imagine that such a profound effect did not have a physiological basis. And I was intrigued that most biologists dismissed the work as of little consequence.”
Countless studies have since confirmed that externally applied electric fields can affect the behaviour of cultured cells, influencing the way they migrate, develop and grow. There is also a revival of the idea that electric fields can have medical benefits. After successful trials in animals, for example, a team at the Centre for Paralysis Research at Purdue University, led by Richard Borgens, is trying to encourage the healing of spinal cord injuries with an external electric field.
But showing that externally applied fields have an effect on living material is not the same as showing that they are biologically relevant. To do that, you would need to show that an internally generated electric field was doing something important.
Internally generated electric fields are an inevitable product of biological systems. Cell membranes and epithelia – flat sheets of cells such as the outer layer of your skin and the lining of your gut – routinely pump ions from one side to the other, creating gradients in electrical potential. This makes them resemble charged batteries, with an excess of negative ions on one side and positive ions on the other. All it takes for current to flow is for a channel to open up linking the two sides, either through damage or deliberately. And where there is a current flowing, an electrical field inevitably follows (see Graphic).
Researchers have measured naturally occurring electric fields in organisms ranging from microbes to humans, and in biological systems ranging from cultured cells to embryos – in which an ion-separating epithelium is the first functional tissue to form. The strength of the field is typically between 10 and 100 millivolts per millimetre, but can sometimes reach 1600 millivolts per millimetre.
Until recently, no one had shown that such fields had any function. But this hurdle was overcome in 2002 when McCaig’s team showed that a naturally occurring field plays a vital role in wound healing in the cornea of rats (Proceedings of the National Academy of Sciences, vol 99, p 13577). In normal corneas, epithelial cells pump positively charged sodium and potassium ions inwards and push negatively charged chloride ions out, creating an electrical potential of about 40 millivolts. But if the epithelium is breached, current flows through the wound, setting up an electric field of about 40 millivolts per millimetre that stretches for half a millimetre across the surface of the cornea.
McCaig’s team has now shown that this electric field promotes healing, by influencing the behaviour of nearby cells. The actively dividing cells that patch up the damage extract important spatial information from the electric field. It causes them to divide along the plane perpendicular to the field, pushing new cells into the wound. If you nullify the field, the cells carry on dividing, but in random directions. Strengthen the field artificially, and cells further away from the wound also start dividing along this plane.
Neurons also use the cornea’s electric field to help re-establish themselves. McCaig’s team found that the natural field in a damaged cornea was able to stimulate nerves to grow towards the wound.
So how do cells sense and respond to electric fields? No one knows, but McCaig thinks that the electric fields may attract charged proteins or lipids in the cell membranes. “These are certainly required, but we do not yet know of a single molecule that is absolutely required. There may be several,” he says.
Another possibility, says Robinson, is that calcium channels embedded in the membrane, which are opened by changes in voltage, might convert electric fields into a signalling cascade. The field might cause calcium channels to open, allowing a rush of calcium into the cell. The calcium in turn activates a second messenger molecule and so on down a signalling chain. Scientists already know that calcium is important in cell responses to electric fields because when it is removed or calcium blockers are added to cells in culture, they no longer respond to electric fields.
Corneal repair is not the only system where innate electric fields have been shown to be important. In the past two years Levin has established that they have a role in embryonic development, particularly in the establishment of left-right asymmetry.
Look in the mirror and you will notice how symmetrical your body is: on each side there is an eye, an ear, a nostril, an arm, a hand, a leg and a foot. If you could peer inside, however, you would see that your organs, including your brain and lungs, are not symmetrical at all.
For a long time, biologists have known that the basis of this asymmetry is genetic: two developmental genes, Sonic hedgehog and Nodal, are expressed on the left side but not on the right. This leads some cells on the left to become heart muscle, while their neighbours on the right grow into the liver. It is also clear that these differences in gene expression are preceded and initiated by chemical gradients. But what is not known is what sets up the chemical gradient in the first place. The answer, Levin now believes, is an electric field.
Working with Robinson, Levin’s group studied chick embryos during the epiblast stage, when they consist of little more than a pouch of epithelium. This is the stage at which embryos start to show differences in gene expression across the “midline” that separates their right and left sides.
Levin first suspected electric fields might be involved when he found that tiny channels called gap junctions connecting the epithelial cells were necessary for proper asymmetry. When Levin blocked the gap junctions with drugs or antibodies, Sonic hedgehog and Nodal were no longer asymmetrically expressed. Instead, the two genes were expressed evenly on both sides of the midline.
Levin reasoned that there must be some molecule travelling through the gap junctions that sets up the chemical gradient, and he now has evidence that the driving force behind this molecule’s movement is an electric field. He soaked the embryos in a voltage-sensitive fluorescent dye and was able to map the electrical characteristics of the epithelium. He found that cells on the right side of the midline had a much larger membrane potential than those on the left – a difference of about 20 millivolts. This means there is an electric field across the midline.
Levin then discovered that the field is dependent on the uneven distribution of a membrane ion pump called hydrogen/potassium ATPase, which pumps hydrogen ions (H+) out of the cell in exchange for potassium ions (K+). The potassium ions then leak passively out of the cell, making the inside negatively charged relative to the outside. Cells on the right side of the midline have many more of these pumps that those on the left. When the pumps are artificially inactivated, not only does the electric field disappear but so does the lopsided gene expression, and the organs in these embryos grow randomly on either side (Cell, vol 111, p 77).
What remains unknown is the mystery molecule that responds to the embryo’s electric field. Whatever it is, it must be small enough to squeeze through the minuscule gap junctions, and it must carry an electric charge under normal body conditions. It is also unclear what establishes the asymmetry in the distribution of ATPase. But even so, Levin has made an important breakthrough by showing that an electric field plays a previously unknown, and crucial, role in amplifying a developmental signal.
Not everyone, though, is convinced. Clifford Tabin, a developmental biologist at Harvard University, says many biologists would agree that electric fields affect cell behaviour, but most are still sceptical that they are used as spatial cues. “I have not seen any convincing research to that effect,” he says. “Additionally, there is a long history of bad research in this area that discourages one from getting excited by new studies.”
“I don’t have any great problem with this,” responds Robinson. “The burden is on us to make the case.” One problem is that the vast majority of studies so far have been correlative: researchers have described electric fields and correlated them with important events, but have been unable to show that the fields were causal and not merely a by-product. The newer studies on corneal repair and left-right asymmetry are starting to show that electric fields do carry important spatial information. But even so, electric field supporters complain that the research continues to be overlooked. “Most developmental biologists have set this idea aside because they can’t fit it into existing ideas about how development works,” says Robinson.
What is needed, according to Robinson and others, is to combine work on electric fields with molecular genetics. “We must show what the targets of electric fields are. This is a major issue in understanding how individual cells sense and respond to electric fields. And we now have the tools to figure this out,” he adds.
That’s why Robinson has shifted his interest from frogs to fruit flies. So much of their developmental genetics is understood, he says, that it will be much easier to show that electric fields are a causal agent in development.
In his latest experiment, he applied electric fields to a fruit fly neuroblast – a small strip of embryonic tissue that develops into the nervous system – growing in a dish. With an electric field of only 2 millivolts per cell diameter, he was able to alter the front-to-back orientation of the neuroblast. In a living fly embryo, the neuroblast is surrounded by an epithelium with a potential of 30 to 50 millivolts across it. The results suggest that an electric field could be providing spatial cues to the developing neuroblast, says Robinson. His next step is to manipulate the electric potential in a living embryo to see what it does to nervous system development.
Meanwhile, Levin has turned to the flatworm Dugesia japonica to investigate the role of electric fields in regeneration. If you cut a flatworm in half, both halves regrow their lost head or tail and you end up with two flatworms. For a hundred years, scientists have been asking how the severed end knows whether to grow a tail or a head. In the 1950s, they discovered that the decision could be influenced by an externally applied electric field – apply it in one direction and the fragment always grows a head, reverse the polarity and a tail always grows. This led to speculation that innate electric fields might be involved in the worm’s regeneration.
Levin recently demonstrated that a natural voltage gradient exists between a severed worm’s tail and the place where its head once was. Six hours after cutting off the head, hydrogen/potassium ATPase ion pumps congregate at the site of the wound, and these are needed to form the voltage gradient. If the pumps are blocked, the voltage gradient disappears and the wound sprouts a second tail instead.
For now, most electric field researchers are struggling to put their discipline on an even footing with other areas of biology. But some have their eye on bigger prizes. In the long term, they say, the work could lead to important medical discoveries, such as drugs that speed up healing, or electrical therapies to aid wound closure or nerve regeneration. “I think understanding the bioelectrical energies involved in life processes will open up possibilities as great, or greater, than those resulting from the recent revolutions in molecular genetics,” says Levin. Now that’s an electrifying claim.