Contact: Diana Kenney, Marine Biological Laboratory
508-289-7139; dkenney@mbl.edu
Longitudinal section of a lamprey spinal cord at 11 weeks post-injury, showing many regenerated axons (green) and a repaired central canal (blue tubelike structure). The original lesion site is in the center of the image. Credit: S. Allen and J. Morgan
In a new study, Marine Biological Laboratory (MBL) scientists report that lampreys recover and regenerate just as impressively after a second complete spinal cord injury at the same location. The study opens up a new path for identifying pro-regenerative molecules and potential therapeutic targets for human spinal cord injury.
“We’ve determined that central nervous system (CNS) regeneration in lampreys is resilient and robust after multiple injuries. The regeneration is nearly identical to the first time, both anatomically and functionally,” said senior author Jennifer Morgan, Director of the MBL’s Eugene Bell Center for Regenerative Biology and Tissue Engineering.
Morgan’s lab has been focusing on the descending neurons, which originate in the brain and send motor signals down to the spinal cord. Some of these descending neurons regenerate after CNS injury in lamprey, while others die.
“We are beginning to isolate individual descending neurons and look at their transcriptional profiles (gene activity) to see if we can determine what makes some of them better at regenerating than others,” Morgan said.
“The ‘good’ regenerators, for example, may express molecules that are known to promote growth during development. That’s one hypothesis,” she said.
Observing how the descending neurons respond to a second CNS injury can help the team tease out the factors required for repeated, resilient regeneration, which could have implications for designing better strategies for treatments aimed at promoting CNS re-growth after injury or disease.
Regeneration has been a core area of research at the Marine Biological Laboratory since its founding, particularly in the pioneering work of Nobel laureate Thomas Hunt Morgan, an embryologist and geneticist whose 1901 text "Regeneration" is a classic in the field.
Over a century later Morgan’s lab in particular has led many breakthroughs, including a 2018 study that found genes that aid spinal cord healing in lamprey are also present in mammals.
In 2010, the Eugene Bell Center for Regenerative Biology and Tissue Engineering was established at MBL in honor of Eugene Bell (1919-2007), a pioneer of tissue engineering and a valued member of the MBL scientific community. Scientists in the Bell Center, in collaboration with colleagues at the University of Chicago and the Argonne National Laboratory, are providing new insights into the basic mechanisms of tissue growth, repair and regeneration in all metazoans that will permit novel approaches to the understanding, treatment and prevention of human disease.
The present study, published in PLOS ONE, was conducted by first author Kendra L. Hanslik and other former research assistants in Morgan’s lab.
“These are all young scientists, many who have since gone on to graduate school,” Morgan said. “This paper was their labor of love. To go through two rounds of regeneration in the lamprey -- that’s nearly 6 months of waiting before they could collect the [spinal cord] tissue and begin the analysis. I’m really proud of their heroic efforts in pulling off this work.”
Citation:
Kendra L. Hanslik, Scott R. Allen, Tessa L. Harkenrider, Stephanie M. Fogerson, Eduardo Guadarrama, Jennifer R. Morgan (2019) Regenerative Capacity in the Lamprey Spinal Cord is Not Altered After a Repeated Transection. PLOS ONE doi: https://doi.org/10.1371/journal.pone.0204193
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The Marine Biological Laboratory (MBL) is dedicated to scientific discovery – exploring fundamental biology, understanding marine biodiversity and the environment, and informing the human condition through research and education. Founded in Woods Hole, Massachusetts in 1888, the MBL is a private, nonprofit institution and an affiliate of the University of Chicago.
Two pygmy zebra octopuses mating.
Credit: Bret Grasse
“It’s very exciting for us. Other scientists have successfully mated this species before, but nobody had achieved multigenerational culture until this,” Grasse said.
The MBL is interested in the pygmy zebra octopus as a contender for basic research on the cephalopods (octopus, squid, and cuttlefish). The cephalopods’ agile behavior, complex nervous system, skill at camouflage and regeneration, and other traits mark them as uniquely advanced, if very strange invertebrates indeed (see sidebar).
“Octopus breeding is not an easy feat. It takes a lot of attention to detail, and we are literally writing the book as we go,” Grasse said.” It’s really a testament to our team that second-generation pygmy zebra octopuses have been born in the lab for the first time.”
Mating: First Do No Harm
The first challenge for Grasse was importing male and female pygmy zebra octopuses (Octopus chierchiae) from their habitat in Nicaragua. Last year, he managed to get four wild-caught specimens from a wholesaler in Los Angeles, “who is the only person who has been able to get them into the country in the last several decades,” Grasse said. Once the exotic octopuses arrived at MBL, the next hurdle was to coax them to mate.
“Octopuses tend to be cannibalistic or, at the very least, aggressive toward one another. So just getting successful mating events without any sort of harm to the animals is difficult,” Grasse said.
“We kept the pygmy zebra octopuses well fed so they weren’t hungry during the mating trials,” Grasse continued. “Once together, we read their body language and if there were any negative interactions, we stopped the trial. If mating occurred and we believed the spermatozoa had been successfully passed, we separated them right away. We put the female in a nice, dark habitat and left her alone, so she didn’t feel threatened. Providing her with a laying refuge to encourage oviposition, or egg-laying, is important.”
Just over a week later, the team saw a clutch of tiny, pearl-white eggs appear. “We were really excited but we weren’t sure if they were fertilized,” Grasse said. They carefully tracked the eggs’ development using a microscope and later, a flashlight. After a few weeks they saw eye spots inside the eggs, “one of the first noticeable features that proves the embryo is viable. That was our big Eureka moment!” Grasse said.
After those first-generation embryos hatched, Grasse’s team cultured some of them to sexual maturity (about 4 months). They then mated those octopuses to create second- generation embryos. Those, too, hatched out just this week.
A pygmy zebra octopus (Octopus chierchiae) in the MBL's Marine Resources Center. An adult of this species is about the size of a table grape. Credit: Taylor Sakmar
Closing the Life Cycle and Beyond
Breeding multiple generations in the lab (called “closing the life cycle”) is critical in biology, as it allows researchers to study gene function and mutational effects from one generation to the next.
Yet crossing this major hurdle with the pygmy zebra octopus, Grasse says jokingly, “Was the easy part!” Given their aggressiveness, each hatched octopus needs a separate tank with a hiding spot and good water quality and flow. And the tanks must be secure, “so we don’t have octopuses escaping and running around,” Grasse said.
“And there are a lot of little, hungry mouths to feed on a day-to-day basis,” he said. Grasse and Taylor Sakmar, MBL Senior Cephalopod Culture Specialist, have trained a team of interns from Northeastern University’s co-op program to care for the species. “They have been a huge benefit,” Grasse said.
Grasse previously worked at Monterey Bay Aquarium, where he and others closed the life cycle on the flamboyant cuttlefish (Metasepia pfefferi) and the striped pyjama squid (Sepioloidea lineolata) for the first time. Those accomplishments attracted the attention of MBL leadership, which hired Grasse and his colleague Sakmar in 2017.
“Closing the life cycle of any cephalopod is no easy task,” Grasse said. “It takes a lot of effort and a lot of eyes on the animals. Having a great team is key! We are always reevaluating current practices, so we continue to improve as we learn more about the species. But collectively, we’ve reached that moment where we are breeding them consistently. We’re confident we can do it with very high survivorship of the offspring.”
Homepage photo:
A pygmy zebra octopus hatchling clings to the side of the tank. Notice how small it is compared to the salt crystals on the glass. Credit: Taylor Sakmar
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The Marine Biological Laboratory (MBL) is dedicated to scientific discovery – exploring fundamental biology, understanding marine biodiversity and the environment, and informing the human condition through research and education. Founded in Woods Hole, Massachusetts in 1888, the MBL is a private, nonprofit institution and an affiliate of the University of Chicago.
The MBL is testing several cephalopod species as potential “model systems” (laboratory organisms) for scientific research. It’s a big investment to breed, house, and develop genetic and imaging tools to study a cephalopod species, so “at some point we will need to narrow down our candidates,” said MBL Senior Scientist Joshua Rosenthal, director of the MBL’s Cephalopod Program.
Currently, the frontrunner is the Hawaiian bobtail squid (Euprymna scolopes). This animal’s life cycle was closed in 1997 at the MBL by Roger Hanlon and others. Today, MBL scientists are actively developing genetic tools for its study.
But the pygmy zebra octopus (Octopus chierchiae) has distinct advantages, including:
• O. chierchiae lays several clutches of 30-90 eggs over her reproductive period. Females of most octopus species lay just one clutch of eggs and then die (sometimes violently) after the eggs hatch. For pioneers like Grasse who are trying to breed these fickle creatures - and the MBL scientists who study them– several egg-laying events means more opportunities for deploying techniques, such as CRISPR-Cas9 gene editing, with the precious embryos.
• Adult pygmy zebra octopuses are small (about as big as a table grape). Since octopuses must be housed in separate tanks, smaller species are preferred in space-conscious labs.
“We still don’t know exactly which cephalopod species will rise to the top,” Grasse said.
A female rotifer (Brachionus), a model system for aging studies. Credit: Michael Shribak and Kristin Gribble
WOODS HOLE, Mass. —Why do we age? Despite more than a century of research (and a vast industry of youth-promising products), what causes our cells and organs to deteriorate with age is still unknown.
One known factor is temperature: Many animal species live longer at lower temperature than they do at higher temperatures. As a result, “there are people out there who believe, strongly, that if you take a cold shower every day it will extend your lifespan,” says Kristin Gribble, a scientist at the Marine Biological Laboratory (MBL).
But a new study from the laboratories of Gribble and MBL Director of Research David Mark Welch indicates that it’s not just a matter of turning down the thermostat. Rather, the extent to which temperature affects lifespan depends on an individual’s genes.
The MBL study, published in Experimental Gerentology, was conducted in the rotifer, a tiny animal that Gribble, Mark Welch, and colleagues have been developing as a modern model system for aging research. They exposed 11 genetically distinct strains of Brachionus rotifers to low temperature, with the hypothesis that if the mechanism of lifespan extension is purely a thermodynamic response, all strains should have a similar lifespan increase.
However, the median lifespan increase ranged from 6 percent to 100 percent across the strains, they found. They also observed differences in mortality rate.
The study clarifies the role of temperature in the free-radical theory of aging, which has dominated the field since the 1950s. This theory proposes that animals age due to the accumulation of cellular damage from reactive oxidative species (ROS), a form of oxygen that is generated by normal metabolic processes.
“Generally, it was thought that if an organism is exposed to lower temperature, it passively lowers their metabolic rate and that slows the release of ROS, which slows down cellular damage. That, in turn, delays aging and extends lifespan,” Gribble says.
Their results, however, indicate that the change in lifespan under low temperature is likely actively controlled by specific genes. “This means we really need to pay more attention to genetic variability in thinking about responses to aging therapies,” Gribble says. “That is going to be really important when we try to move some of these therapies into humans.”
Citation:
Gribble, Kristin E et al (2018) Congeneric variability in lifespan extension and onset of senescence suggest active regulation of aging in response to low temperature. Experimental Gerentology 114: 99-106, doi: 10.1016/j.exger.2018.10.023
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The Marine Biological Laboratory (MBL) is dedicated to scientific discovery – exploring fundamental biology, understanding marine biodiversity and the environment, and informing the human condition through research and education. Founded in Woods Hole, Massachusetts in 1888, the MBL is a private, nonprofit institution and an affiliate of the University of Chicago.
More Information
Rotifers are small, aquatic animals with about 1,000 cells, a brain and nervous system, and muscle, digestive and reproductive systems. They offer many advantages as a model organism for biomedical research, including aging studies:
- Rotifers have more genes in common with humans than do other popular invertebrate models (fruit flies and nematodes).
- Short lifespan (about two weeks), allowing for rapid lifespan assays.
- Easy to culture in laboratory.
- Rotifers are transparent, allowing for imaging of cellular processes.
- A great deal is known about rotifer ecology: how they reproduce and thrive in the natural world.
- A draft genome is sequenced and transcriptome is published.
Background:
Mark Welch, David. The Potential of Comparative Biology to Reveal Mechanism of Aging in Rotifers. In Conn’s Handbook of Models for Human Aging (Elsevier, 2018) doi: 10.1016/B978-0-12-811353-0.00037-3
Gribble, Kristin E and David B. Mark Welch (2017) Genome-wide transcriptomics of aging in the rotifer Brachionus manjavacas, an emerging model system. BMC Genomics, doi: 10.1186/s12864-017-3540-x.
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The Marine Biological Laboratory (MBL) is dedicated to scientific discovery – exploring fundamental biology, understanding biodiversity and the environment, and informing the human condition through research and education. Founded in Woods Hole, Massachusetts in 1888, the MBL is a private, nonprofit institution and an affiliate of the University of Chicago.