Human organoids filled more than 90 percent of the cortex in mice bred without one

The Nature paper "Developmental xenocortication using human-derived organoids in mice" establishes a transplantation platform using a genetic strategy to deplete glutamatergic neurons from mouse neocortex and hippocampus (apallial) and neonatally engraft the cortical cavity with human stem-cell-derived cortical organoids, generating xenocortical (XCX) mice.

Researchers report the work on September 16 in Nature; Sergiu Pasca is a neuroscientist at Stanford University.

Over three months the human tissue expanded nearly fivefold in volume in these mice, ultimately filling more than 90 percent of the cortex.

Pasca said the mice engineered to lack more than 90 per cent of their cortex only experience some memory and movement deficits, since their brain adapts to compensate for the missing cortex.

The mouse cortex and neighbouring hippocampus are two structures involved in movement, learning and memory that together account for about half of the brain's total volume.

The researchers subjected grafted animals to oxygen deprivation to model cerebral palsy; afterward the human tissue showed cellular signs of injury and the mice developed problems with gait and limb coordination, features of the disability in people.

The transplantation of human brain organoids into the cavity successfully took hold in 25 of 29 attempts.

Gabriel Balmus of the University of Cambridge said mice have evolved to be highly resilient to low oxygen levels, so it is hard to replicate hypoxia responses in a normal mouse, and that the grafted animals could provide a better way to study cerebral palsy.

The technique "is certainly bold," said H. Isaac Chen, a neurosurgeon at the University of Pennsylvania who studies brain organoids but was not involved in the research; Chen notes that the extent of humanization raises questions about the ethical boundaries of such experiments.

In about half of the engineered mice the team surgically implanted four human brain organoids, generated by bathing human stem cells in chemicals for about 40 days, filling the cavity in each skull to produce XCX mice.

Pasca consulted an outside panel of experts led by Insoo Hyun, a bioethicist at the National University of Singapore Yong Loo Lin School of Medicine; Hyun says there is little evidence these mice have crossed an ethical line, that what matters is not how much human tissue an animal contains but what that tissue enables it to do, and "From a secular ethical point of view, it boils down to cognitive capacity."

Grafts occupied most of the cortical volume and generated a diversity of human cortical cell types including layer 5 extratelencephalic projection neurons; human cortical neurons integrated with the mouse nervous system, and in vivo cortical graft-wide calcium imaging and electrophysiological analyses revealed patterns of organized activity resembling developing circuits.

Pasca told Live Science: "Even when they're put in an animal, in a mouse or in a rat, they will still develop about 20 times slower than the mouse or the rat."

Pasca explained that the host's brain cells quickly grow and form new connections while human cells lag behind and get outcompeted, limiting the space human cells can take up, and that rodent neurons become myelinated faster, with that fat creating a physical barrier the human cells struggle to penetrate.

Transplanted human cortical organoids can occupy up to a third of a rat's cortical hemisphere in the 2022 newborn-rat model, where human neurons develop in the context of extant rat connections.

Knoblich said of concerns about consciousness: "The data so far doesn't say that that putting [a human organoid] in makes the mouse smarter or more conscious than a normal mouse."

25 of 29 successful engraftments is about 86 percent.

Unsupervised machine learning applied to spontaneous mouse behaviour using motion sequencing revealed distinct behavioural repertoires in control and apallial mice, with the XCX group positioned intermediate between, yet distinct from, both groups.

"This could be used down the line to test many therapeutics that are being considered for cerebral palsy," Pasca said; he plans to study frontotemporal dementia and genetic forms of autism with the mice, using organoids made from patient-derived cells.

The Nature abstract and main-text excerpt describing the behavioural analyses do not state the number of animals in each group.

Apallial mice showed deficits on tasks requiring neocortex and hippocampus (fine motor coordination in the CatWalk and working memory in the Y maze), a pattern that was only partially observed in XCX mice.

New Scientist reported that the apallial mice performed worse than normal mice in a maze memory test, but that this deficit was almost completely reversed in the XCX mice, whose gait fell between that of apallial and normal mice.

On tests of fine motor control and memory, the grafted mice's performances fell somewhere between that of mice with intact brains and those lacking a cortex.