Space Frogs: Ribbeting Research on STS-47

NASA’s Space Shuttle mission STS-47 had many accomplishments: the 50th Space Shuttle flight; Astronaut Mae Jemison became the 1st African-American female in space; Mamoru Mohri was the first Japanese astronaut to fly on the Space Shuttle; Marc Lee and Jan Davis became the first (and last) husband and wife to fly on the same mission; and the first and only Japanese sponsored Spacelab module (SL-J).

STS-47 Crew in Spacelab-J (CW from left): Mission Specialist 3 N. Jan Davis, Commander Robert L. Gibson, Pilot Curtis L. Brown, Jr.,  Mission Specialist 4 Mae C. Jemison, Japanese Payload Specialist 1 Mamoru Mohri, Mission Specialist 1 & Payload Commander Mark C. Lee and Mission Specialist 2 Jerome Apt in center. Photo: NASA.

Of the 43 experiments on board STS-47, one was particularly interesting to me: it was the first time frogs were handled in space. I know, I’m easily amused.
 
Mr. Toad’s Wild Ride
Because of their relatively small size, easy care and a several analogous physiological similarities, frogs have long been involved in biomedical research. It was therefore a natural leap to use these same advantages in helping us understand human adaptations to spaceflight.

Frogonaut history starts in 1961 with the Soviet satellite Korabl-Sputnik 4. I couldn’t find any information on what the frogs were doing on there research-wise, but since this was the penultimate test flight of the Vostok spacecraft before Yuri Gagarin’s famous flight, it was likely for basic survivability/life support testing.

Frogs were also aboard NASA’s Biosatellite I and II un-crewed program between 1966 and 1969. Biosatellite I failed to return due to a failed retro rocket. Biosatellite II had re-flight experiments from Biosatellite I and included frog embryos, similar to the experiment on STS-47. An ambitious NASA program launched in 1970 called “Orbiting Frog Otolith” tested the response of the gravity sensing organs called otoliths of two male bullfrogs (Rana catesbiana).

Orbiting Frog Otolith Experiment. Exterior (left) and diagram of components (right). You can kinda see the frogs in the center of the diagram on the right. Pictures or descriptions of space frog experiments prior to this are hard to come by. Photos: OFO Press Kit.

In 1990, Japan’s first ever citizen in space, cosmonaut/tourist/journalist Toyohiro Akiyama observed the behavior of six Japanese tree frogs (Hyla japonica) to see how they would adapt to microgravity.

Japanese Cosmonaut on Mir. Toyohiro Akiyama observes Japanese tree frog behavior living inside a disposable glove bag during his flight to Mir in 1990. The frogs typically experience “free fall” on Earth while jumping from tree to tree. They demonstrated a similar posture (left) while jumping in microgravity, but as the 8 day mission went on they reduced the amount of jumping, instead opting to crawl along surfaces. Photos: Frogs in Space PowerPoint.

 

The Great Leap Forward

Frogs have large, readily observable embryos that can be fertilized outside of their body, facilitating a commonly studied model of embryo development on Earth. Frogs from the genus Xenopus have been used in embryo development research since the early part of the 20th century.

An experiment aboard STS-47’s Spacelab module called “Effect of Weightlessness on Development of Amphibian Eggs” wished to understand if gravity is necessary for the normal development of amphibian embryos. 

Even if rotated on Earth, a frog embryo’s rapidly dividing animal cells and more slowly dividing vegetal cells always align with the normal gravity vector, such that the animal cells are “up” and the vegetal cells are “down.”

Diagram of Embryo Poles. Alignment of the animal and vegetal poles typically follow the gravity vector. Diagram: Wikipedia.

The location of these two cell types within the embryo are eventually responsible for their differentiation into the various tissues of an organism as well as dorsal and ventral body structures (i.e. limbs, fins). The researchers simply wanted to know if the lack of a gravity vector in the free fall of orbit would alter this polar alignment and therefore change the embryo’s normal development.

Embryo Development. The rapidly dividing animal cells on top can easily be seen by their tan color, while the slower developing yellowish vegetal cells can be seen on the bottom.

The method of project is where it gets interesting. Previous projects, such as that on Biosatellite II, had fertilized the embryos long before launch. In order to start the experiment while in a microgravity environment, the investigators flew four live female South African three-clawed frogs (Xenopus laevis). 18 hours after arriving in orbit, the crew subcutaneously injected the frogs with an ovulating hormone called chorionic gonadotropin. Then, 16 hours later the eggs were collected and placed in small incubation chambers and fertilized with sperm solution.

Experiment Hardware. Pic of the Frog Environmental Unit (FEU) (left) and the Egg Chamber Units (ECUs) (right). Photo: Life Sciences Data Archive at Johnson Space Center.

Diagram of the Frog Environmental Unit (FEU) from “A Decade of Life Sciences Experiment Unique Equipment Development for Spacelab and Space Station, 1990-1999”

If you’ve ever tried to catch a frog on Earth, it’s a slippery affair requiring quick reflexes and a large catalog of curses and obscenities. At least in 1g, the frog’s escape parameters are constrained roughly to jumping and squirming in a two dimensional coordinate system.

Give the frog a third dimension to squirm and jump in microgravity and handling it is about as much fun as it sounds. The gif below says it all. This had to be done multiple times!

Un-Hoppy Passenger. It’s uncertain if astronaut Marc Allen knew that his MIT degree and fighter pilot career would one day lead him to such an historic moment as the first frog wrangling in space, but it is certain those quick reflexes came in handy. I’m also sure his space to ground mic was turned off to spare the children watching. Animated gif made from STS-47 Mission Highlights Resource Tape.

Despite the squirmy start, the rest of the experiment hopped along well, with some of the embryos being placed in a 1g centrifuge and the rest staying in a microgravity incubator. Embryos were fixed at certain time-points to be sectioned and stained after the flight.

 

Results: Toadally Normal?

The successful fertilization of the embryos supported previous investigator findings that fertilization could properly occur in microgravity.

There were slight differences between the micro-g and 1g timepoint embryos, showing slight cell thickening in the micro-g blastocoel layer and a minor change in location of the vegetal cells in the micro-g samples. The researchers give several reasons for these differences, but notice that they were eventually corrected during development to the tadpole stage and “no significant differences in morphology were observed.”

Figure from PNAS paper.

As for the tadpoles, the only significant change was that the micro-g tadpoles had undersized lungs. On Earth, tadpoles typically come to the surface to fill their lungs with air within 2-3 days after hatching. The authors explain that since there were no gravitational clues, the tadpoles simply didn’t know where the surface was and therefore didn’t fill their lungs with air. This behavior was corrected after the tadpoles returned to Earth, but it’s unclear if this would have been detrimental to the frog reaching full maturity.

Overall, this project support previous observations that gravity is not necessary for the fertilization of embryos and, while there were some slight, unexplained differences during development, embryogenesis in microgravity proceeded without observable physiological effects.

STS-47 returned after a successful 8 day mission and the tadpoles and samples were safely returned to the researchers at Ames Research Center. The tadpoles that returned fully metamorphosed and matured normally, living long healthy lives in a nice pad in central California until they one day croaked.

And I’m officially out of puns.
 

Resources

Dorcas P. O’Rourke; Amphibians Used in Research and Teaching. ILAR J 2007; 48 (3): 183-187. doi: 10.1093/ilar.48.3.183

STS-47 Press Kit

Yamashita M, Izumi-Kurotani A, Mogami Y, Okuno M, Naitoh T, Wassersug RJ. “The Frog in Space (FRIS) experiment onboard Space Station Mir: final report and follow-on studies.” Biol Sci Space. 1997 Dec;11(4):313-20.

P. D. Savage, J. P. Connolly, B. J. Navarro “A Decade of Life Sciences Experiment Unique Equipment Development for Spacelab and Space Station, 1990-1999” 

Souza KA, Black SD, Wassersug RJ. “Amphibian development in the virtual absence of gravity.” Proceedings of National Academy of Science USA, vol. 92, March 1995, pp. 1975-1978.

STS-47 Mission Highlights Resource Tape

Life Sciences Data Archive at Johnson Space Center

Orbiting Frog Otolith Press Kit

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