Xuyu Qian yanked open an incubator door at the University of Pennsylvania to reveal rows of cylindrical tubes swirling, like shaken-up snow globes, with a strange and exotic flurry. The pale, peppercorn-sized spheres were lab-grown globules of human brain tissue, or, as Mr Qian occasionally refers to them, "minibrains."
"Minibrain" is a controversial nickname, loathed by some scientists who fear it conjures alarmist images of fully-functioning brains trapped in vats, while the reality today is balls of cells that can't think or feel.
But the term vividly evokes the aspirational goal of this fast-moving area of research: to mimic the complexity of the human brain and illuminate the biology of the human mind, one of science's darkest black boxes. As the technology, which scientists refer to in journal articles as "cerebral organoids," improves, the more the "minibrain" title fits.
Today, organoids that resemble different regions of the human brain are routinely spun up from stem cells in large batches in laboratories around the world. Researchers have refined their recipes since the technique was first described five years ago, but the process is surprisingly hands-off: after a few nudges from scientists, stem cells grow into spheres with about a million neurons through a naturally occurring choreography that mirrors early brain development in the womb. At Day 100, Mr Qian's minibrains resemble a portion of the prenatal brain in the second trimester of pregnancy.
"People are more worried about if they reach a certain level - if it's really like a human brain. We're not there; we're very far from there," said Hongjun Song, who leads the laboratory at Penn's Perelman School of Medicine, where Mr Qian works. "But the question people ask is, 'Do they have consciousness?' The biggest problem I have so far is I think, as a field, we don't know: What is consciousness? What is pain?"
At the moment, minibrains are far from anything approaching moral personhood in a dish and the technology may never come close. But the rapid pace of progress on organoids has led scientists and ethicists to call for a public ethical discussion that can move in tandem with the research.
The disembodied brain, after all, is a long-standing trope of cultural fascination and even philosophy, ranging from the serious metaphysical thought experiment called the "brain in a vat" to the screwball sci-fi comedy "The Man With Two Brains," in which Steve Martin finds himself falling in love with a charming woman's preserved brain.
"If, at the sunset of life, the brain is what you examine to know if someone has died, at the beginning of life is there a point where you might say, 'Look, the brain is at the beginning of life?'" said Insoo Hyun, a bioethicist at Case Western Reserve University. "Many people don't understand where the science is now, and where it could go in the future - including, I think, the researchers."
Organoids offer a powerful tool for scientists studying the mysteries of the brain, which by some estimates is the most complex object in the world. Unlike cancer, which researchers can study by growing cancer cells in a dish, the brain and its disorders have been largely off-limits, except through hard-to-get post-mortem tissue that offered only a snapshot or by trying to study much simpler animal brains.
More than a decade ago, scientists discovered it was possible to create stem cells by reprogramming a person's skin cells. They could use the procedure to create any cell type in the body and study the basic biology of specific diseases that afflict people, ranging from Down syndrome to diabetes.
Sergiu Pasca, a neuroscientist at Stanford University, dreamed as a medical student of understanding the biological basis of autism and schizophrenia. Now, his lab uses stem cells from people who have those conditions to grow cerebral organoids.
"This gives us aspects of human brain development that were previously inaccessible. Most of the work we're doing right now is to study really the hidden biology of the human brain," Mr Pasca said.
By creating organoids from people with a genetic disease that causes autism and epilepsy, he was able to watch how brain cells migrate during early development.
Mr Pasca and his colleagues saw clear differences - one type of neurons jumped around in an abnormal and inefficient way in the organoids from the patients, giving the researchers a window into a critical part of development that could have long-term consequences.
After the Zika virus was officially linked to brain defects in infants, Mr Song and his wife and close collaborator, Guo-li Ming, sent minibrains to collaborators, who exposed them to Zika. They saw the virus infect a kind of stem cell called a neural progenitor that generates new brain cells, wreaking havoc.
Ryan Salinas, a clinical fellow in Mr Ming's lab, grows organoids from patients with a deadly brain cancer, in the hopes they might provide a better tool for testing the effectiveness of drugs.
At the University of California at San Diego, stem-cell scientist Alysson Muotri is attempting to "Neanderthalise" a brain organoid as part of his larger search for the biological basis of modern humans' sophisticated social abilities. He used gene-editing technology to introduce a mutation found in Neanderthal genomes to a modern human stem cell. The early, unpublished results, he says, are an organoid that, instead of being smooth and spherical, is lumpy like popcorn - suggesting that the gene mutation significantly influences early brain development.
Five years ago, an ethical debate about organoids seemed to many scientists to be premature. The organoids were exciting because they were similar to the developing brain and yet they were incredibly rudimentary. They were constrained in how big they could get before cells in the core started dying, because they weren't suffused with blood vessels or supplied with nutrients and oxygen by a beating heart. They lacked key cell types.
Still, there was something different about brain organoids compared with routine biomedical research. Mr Song recalled that one of the amazing but also unsettling things about the early organoids was that they weren't as targeted to develop into specific regions of the brain, so it was possible to accidentally get retinal cells.
"It's difficult to see the eye in a dish," Mr Song said.
Now, researchers are succeeding at keeping organoids alive for longer periods of time. At a talk, Mr Hyun recalled one researcher joking that the lab had sung "Happy Birthday" to an organoid when it was a year old. Some researchers are implanting organoids into rodent brains, where they can stay alive longer and grow more mature. Others are building multiple organoids representing different parts of the brain, such as the hippocampus, which is involved in memory, or the cerebral cortex - the seat of cognition - and fusing them together into larger "assembloids."
Even as scientists express scepticism that brain organoids will ever come close to sentience, they're the ones calling for a broad discussion, and perhaps more oversight. The questions range from the practical to the fantastical. Should researchers make sure that people who donate their cells for organoid research are informed that they could be used to make a tiny replica of parts of their brain? If organoids became sophisticated enough, should they be granted greater protections, like the rules that govern animal research? Without a consensus on what consciousness or pain would even look like in the brain, how will scientists know when they're nearing the limit?
"The scientists often look at this tiny little piece, 2 millimetres cubed, and to think that represents the potential for say, consciousness, or some larger construct - as a scientist, you think of course that is not really a human brain," said Rusty Gage, president of the Salk Institute for Biological Studies. "On the other hand, I've been in this long enough to know it's not just the facts, it's the perception of it and how the public sees it."
As scientists implant organoids into animals, creating mixed creatures - chimeras - with some human brain cells, new questions emerge. So far, a human organoid in a mouse brain makes up only a tiny portion of the brain. Mr Gage has tested whether these mice have unusual cognitive abilities and found none.
But those questions will quickly change if an organoid is ever implanted into a primate brain. In October, the National Academies of Sciences, Engineering and Medicine will hold a workshop focused on the scientific opportunities and bioethical challenges that come either from altering genes or creating chimeras to model nervous system disorders in non-human primates. A draft policy from the National Institutes of Health proposed a steering committee that would advise on research in which human cells could make a "substantial contribution" to animal brains, excluding rodents.
And there are other, non-organoid ways to approach some of the same questions. Christof Koch, president of the Allen Institute for Brain Science, said that every few weeks, researchers on his team receive a small cube of human brain that was removed during surgery.
"The brain stays alive - this discovery we made is the brain stays alive much longer than a mouse brain," Mr Koch said before adding a philosophical caveat that often comes up when researchers are talking about this area of research: "We don't know what it means to stay 'alive' - the individual neurons remain responsive for three to four days."
For ethicists like Mr Hyun, the goal of scrutinizing the research at its early stages is to avoid a repeat of the controversy surrounding the cloning of "Dolly the Sheep." The public felt taken by surprise by cloning, triggering an ethical discussion in 1997 after the technology was out in the world.
The difference between this and other times when scientists have brushed up against an ethical frontier may be how much the scientists want to take part.
"I remember in the early days . . . I'd say, 'Oh, there's nothing to worry about. These are cells in a dish, the level of activity is quite immature,'” Mr Muotri said. "Now, you are asking this question today, and we have a very different picture."
The Washington Post
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