In 1935, French veterinarians observed a rabbit with a peculiar gait. Sometimes, when walking or running, the sauteur d’Alfort rabbit would lift its back legs over its head, scrambling along the ground on its forelimbs like a circus performer (see video, above).
Now, scientists have pinned down the genetic mutation that likely causes this breed to have this strange form of locomotion. The gene involved holds clues to how the spinal cord enables walking, hopping, and even hand-standing—a finding that dovetails with other work over the past decade on mice and horses. Together, the studies provide an emerging picture that may help explain how all vertebrates, including humans, move around.
The work could help scientists treat human motor deficits like Charcot-Marie-Tooth Disease, a nervous system disease characterized by muscle weakness, says Stephanie Koch, a neuroscientist at University College London who was not involved with any of the studies but has seen similar odd gaits in mice. The study’s results are “both surprising and exciting.”
Gait is complex. Left, right, front, and back limbs must move at the right times. Muscles need to contract just the right amount to bend, straighten, lift, and twist the legs appropriately. And the body has to be able to switch from, say, walking to running, or going forward to sideways, in an instant should the senses detect danger or obstacles.
A set of nerve cells in the spinal cord called the central pattern generator—not the brain—makes most of these decisions. But just how has been unclear, says Sónia Paixão, a neuroscientist at the Max Planck Institute of Neurobiology.
Researchers know nerve cells called interneurons, which relay sensory information from the rest of the body to the motor neurons that control muscles, play key roles. Several teams have been working to define classes of interneurons, often categorized by what genes are active in them. Then will come the hard work of figuring out what those neurons do. “The exact nature and function of relevant interneurons have been hard to pin down,” says Abdel El Manira, a neuroscientist at the Karolinska Institute (KI).
That’s where the sauteur, or jumper rabbit comes in. Geneticists Leif Andersson from Uppsala University (UU) and Miguel Carneiro from the University of Porto decided to try to track down the DNA behind the animal's strange gait after sequencing a rabbit genome in 2014. They mated jumper rabbits with another breed to create first- and second-generation animals with either the normal or hand-standing walk. Then the researchers compared DNA from affected and unaffected rabbits and pinned down one mutation in a gene called RORB. Working with UU developmental biologist Klas Kullander, they tracked down where and when this gene was active.
In these rabbits, the mutation causes aberrant versions—or sometimes none at all—of the RORB protein to be produced in a specific group of interneurons, the team reports today in PLOS Genetics. This protein is a transcription factor, meaning it controls the activity of many other genes. Developmental studies showed that the result of two defective RORB genes is those interneurons are completely missing, and in rabbits with one copy there are 25% fewer of them. These interneurons are inhibitory—they stop nerve cells from firing—and when they are missing, the rabbits flex certain muscles too much, lifting their hind legs more than they should.
“I was impressed that the authors were able to identify a single gene mutation,” says Jeremy Dasen, a neuroscientist at New York University. Because locomotion is such a complicated behavior, he expected multiple genes and multiple classes of interneurons would be involved. But this paper drives home that, like modular homes with independent sections put together to make a dwelling, locomotion is achieved through the combined efforts of individual classes of interneurons, he adds.
RORB also seems to control hind-limb coordination in mice: Rodents missing a functional RORB gene waddle like ducks. As a result, says KI neuroscientist Sten Grillner, “the importance of RORB applies most likely to all limbed animals” including humans. People with Charcot-Marie-Tooth disease also have atypical RORB proteins.
RORB is the second gene that Andersson’s team has pinpointed as important to gait. In 2012, he and colleagues linked a mutation in a protein called DMRT3, which helps researchers identify a subset of interneurons, to an unusual walking gait called toelt. In Icelandic horses that exhibit toelt, the hind limbs support more weight than the front limbs, making the gait more fluid. Andersson’s team confirmed the protein’s role by making the same mutation in mice. Breeders have selected for this horse mutation because the altered gait gives a very smooth ride. Some of the horses carrying this mutation can also trot and pace at high speed, which makes them excellent for harness racing. And a connection exists between this group of interneurons and the RORB defect, the researchers now report: Rabbits with the mutation make lots more of the DMRT3 interneurons. The researchers do not yet know why.
Understanding how all the nervous system components interact is a challenge, Paixao says. Advances like the rabbit paper illustrate the progress made possible by combining developmental, genetic, and behavioral studies. “We are now in a pivotal time to achieve these goals,” she adds. “It is an exciting time to see how all the pieces of motor control are coming together.”