Smart from the start

Animal embryos get some respect for their survival skills

Karen Warkentin speaks admiringly of the eggs of red-eyed tree frogs because, for one thing, they know what’s shaking.

SMART FROM THE START Three-day-old embryos of red-eyed tree frogs position their big, branching gills near the oxygen-rich egg surface. K. Warkentin/Boston Univ. and Smithsonian Tropical Research Institute

PRENATAL ESCAPE A female pale spitting spider can’t spit at predators while carrying an egg mass. In danger zones, the eggs hatch earlier than when mom and offspring are safe. Richard Seaman

EMBRYOS FEEL THE VIBES | Frog embryos know when to cut and run. As a cat-eyed snake bites into fertilized eggs of a red-eyed tree frog in Costa Rica (left), embryos that don’t go down the snake’s throat early in the attack escape by hatching and dropping off the leaf. In tests of hatching cues, researchers mimicked vibrations from a parrot snake attack (below, top) and a rainstorm (below, bottom). Embryos fled from the simulated snake but not from the storm, apparently assessing risk based on a combination of the duration, frequency and spacing of shake-ups. K. Warkentin/Boston Univ. and Smithsonian Tropical Research Institute

Masses of these glistening eggs hang on leaves that dangle over tropical ponds, and the eggs stay put even when branches thrash in storms. A hungry snake biting into one end of an egg mass can make the embryos’ home dip and dance too. But at this jouncing, older embryos flee. They can’t run, but they can hatch. A sudden burst of emergency hatching sends a rain of new tadpoles into the water, often saving some 80 percent of a clutch.

Pretty sophisticated for a glob of goo.

It turns out that frog eggs and other embryonic blobs possess a rather advanced repertoire for coping with the earliest stages of life. Embryos can react to perils both inside their eggs and out. Even while still within their shelter, they’re learning lessons about eating. And being eaten. These and other recent findings are forcing researchers to expand their ideas of how capably and subtly these half-baked beings can act.

Maybe deft dodges and finely judged trade-offs should be expected: Life for the small and underpowered is dangerous, and plenty of good embryos die young. “That creates opportunities for natural selection to shape a response,” says Warkentin, a biologist at Boston University. But there’s still the lingering old view of early life stages as clods. Warkentin says: “We think, ‘It’s a frog egg, it’s a snail embryo — what can it do?’ That’s what surprises us.”

Even from inside its egg, a remarkably young embryo can do a thing or two to get what it needs.

Among red-eyed tree frogs, Agalychnis callidryas, embryos develop big feathery gills for extracting oxygen from the watery world inside their eggs. But just where an embryo’s head-to-be lies within its egg makes a difference in oxygen availability. The part of an egg closest to the air typically carries twice the oxygen concentration of the deep interior squeezed among neighboring eggs, Warkentin and her student Jessica Rogge found. When Rogge prodded embryos so their gills fell into the low-oxygen zone, most of the 3-day-olds twitched themselves back into place within 15 seconds.

Rogge even tested embryos just a day old. “They don’t have blood. They don’t have a heart,” Warkentin says. They move only by beating a fuzz of tiny hairlike projections. Yet when Rogge nudged embryo front ends toward the downside of the oxygen gradient, more than half of the embryos she tested worked their fuzz to chug at least halfway back to the sweet spot within five minutes, she and Warkentin reported in 2008 in the Journal of Experimental Biology.

Red-eyed tree frogs, of course, could just have exceptionally smart babies. But research published last summer suggests pond snails also can fix a bad situation.

Masses of Helisoma trivolvis eggs stick to plants or other surfaces in freshwater streams. Like the youngest frog embryos, these snail embryos also depend on hairlike projections to shift around in the egg. Sometimes the developing snails rotate slowly like little turning planets, and sometimes they wind up for a brisk spin. Researchers know that some of the snails’ developing neurons respond to dissolved gases, so Jeffrey Goldberg of the University of Calgary in Canada proposed that the turning movements improve the aeration of fluid inside the eggs. In the Journal of Experimental Biology in 2008, he called the idea the “stir-bar hypothesis.”

Baby bailouts

What’s stirring outside the egg also can provoke embryos to action. Scientists are learning that embryos are pretty savvy when it comes to deciding when the coast is clear for them to hatch — and what to do when it’s not.

Warkentin remembers facing a skeptical audience at a scientific meeting in the early 1990s when she was a grad student presenting her idea that some frog eggs panic-hatch to escape attacks — but then face extra perils as puny preemies in the water. Some scientists doubted that the little embryos could hatch fast enough to do themselves any good or that plunking into the water several days early would make much practical difference. Both hypotheses had grounding in the field of behavioral ecology, but the time periods involved were short and, come on, these were just frog eggs.

Her first field experiment still stands out in her memory. “It rained a lot, and frogs were bouncing into the ponds — it was very intense,” she says. She let a snake bite a mass of eggs and saw them respond so strongly that she said to herself, “OK, I have a dissertation.”

Today, looking for changes in hatching time as a reaction to danger is becoming a cottage industry among biologists, Warkentin says. She and her team have now shown that red-eyed tree frog embryos can shorten their time in the egg by some 30 percent if attacked by snakes, wasps or killer molds. And embryos of at least 17 other species of amphibians can split open their eggs for an early escape in a crisis, according to various studies. So can two species of fish as well as a lizard.

Predator-sensitive hatching has even shown up in an animal where mom looks after the eggs: the spitting spider Scytodes pallida. She carries the eggs around in grasping jaw parts but has to put them down if she needs to defend herself by spitting goo at an attacker. Daiqin Li of the National University of Singapore threatened egg-carrying moms with a predatory Portia labiata jumping spider or its silk. Egg clutches of these imperiled females hatched sooner than eggs of mothers in safer spots, he reported in 2002 in Proceedings of the Royal Society B. But eggs exposed to spider scent without her care didn’t hatch appreciably early. Just what hurry-up message mom sends to the eggs or how she fast-forwards their development is a question for future research.

Staying in the egg’s safe haven can be a good idea too. In 1993, the year after Warkentin started wading around in snaky wetlands to test her idea, Andrew Sih, now at the University of California, Davis,  published evidence of the reverse strategy, where embryos delay hatching in dangerous places.

Water with the mere scent of  predatory flatworms convinced streamside salamander embryos to linger in their eggs, Sih and Robert Moore, both then at the University of Kentucky in Lexington, reported in The American Naturalist. Phagocotus gracilis flatworms feast on the smallest salamander hatchlings, but larger hatchlings usually escape. Staying in the egg longer means growing larger, and salamanders that delayed their debut into the free-swimming world indeed improved their chances of survival.

Other hatching delays are turning up too. Embryos of two frog species just stay in the egg extra-long and grow to maximum size when leeches or egg-gulping fish are about.
Species-to-species differences in what embryos can do pose rich evolutionary puzzles. In 2008, Warkentin and her colleagues reported in Ecological Monographs on tests of crisis-hatching in relatives of the talented red-eyed tree frogs. As long as embryos had reached a minimum age, all the Agalychnis tree frog species tested hatched in a hurry if dunked in water. If they didn’t hatch, they would have drowned in their eggs, despite their large gills.

Even though these frog embryos can burst out of their eggs in the face of one menace, some species just sit there and perish when confronted by another danger, the researchers found. Embryos of one frog species that lives in the same snake-infested forests where Warkentin studied the red-eyed tree frog just lie on their leaf as a snake picks off mouthful after mouthful. This snake-bait species coats its eggs in a tougher covering than the jiggly eggs of the red-eyed tree frog. So maybe warning vibes don’t travel as well through the tougher eggs, or early hatching would pose its own problems. Warkentin says she’s still musing about how to make sense of the evolutionary pattern.

Exactly how snaky shaking warns red-eyed tree embryos still holds some mysteries as well, Warkentin and her colleagues say in the Feb. 15 Journal of Experimental Biology. Earlier egg-vibration studies had established that the embryos don’t need scent or sight to tell a snake shake from a storm blast. Embryos distinguished between the two when researchers subjected eggs to vibrations only, based on leaf-movement recordings. The new work, in cooperation with Boston University’s mechanical engineering department, shows that the embryos are more likely to hatch at low-frequency vibrations but that frequency alone won’t distinguish threat from routine. Frequencies from benign and lethal causes overlap broadly, so the embryos must be combining multiple clues to make such clever choices.

Prehatch preschool

For the cleverest choices, embryos actually learn the latest about their environment’s opportunities and dangers. “This may not be how most people think of the learning they did in school,” says Alicia Mathis of Missouri State University in Springfield. She uses a broad definition: “Learning is experiencing something that causes you to change your behavior.”

In this sense, prenatal learning of smells and tastes has been a traditional line of embryo research. Exposing an embryo to a compound, sometimes by giving it to mom, sets up the baby to react to the substance after birth. After researchers added a dash of anise seed oil to food for pregnant dogs (golden retrievers, Labradors and mutts), for example, the newborn pups tended to point their noses toward an anise-scented cotton swab instead of one wetted with plain water. Pups didn’t have the same interest in vanilla swabs, showing they had learned to recognize the anise scent before they were born, researchers from Queen’s University Belfast in Northern Ireland reported in 2006.

Similar prenatal exposures or lessons work in people, rats, sheep and rabbits as well as in some birds, reptiles, amphibians and even invertebrates.

Embryos can learn visually, too. As cuttlefish develop, the outer egg membrane turns clear enough for the embryos to peek out at the big world around them. To see whether the youngsters were paying attention, Ludovic Dickel at the University of Caen Lower Normandy in France and colleagues set crabs within sight of cuttlefish eggs. After hatching, the youngsters preferred to eat crabs, Dickel and his colleagues reported last summer in Animal Behaviour. Without that embryonic view of food, newly hatched cuttlefish preferred shrimp to crab.

One lesson from cuttlefish and other reports of prenatal learning is a new caution in describing animal behavior, says Mathieu Guibé, who works on the Dickel lab’s cuttlefish project. “Now that we know embryos can learn, scientists must avoid interpreting a behavior shown at birth as something innate,” he says.

After birth, plenty of creatures can manage the fancier task of associative learning, like Pavlov’s dogs drooling for food at the mere sound of a bell. To see if embryos can learn this way too, Mathis scared wood frog eggs. Research on stressful stimuli typically focuses on negative after-effects, she says, but “negative is not always bad.” Perhaps scary experiences would teach an embryo some valuable lessons about danger.

Mathis, Douglas Chivers of the University of Saskatchewan in Canada, and colleagues let wood frog eggs get a good long taste of water dosed with a slurry of freshly ground-up wood frog tadpoles. To a little embryo on the brink of tadpole-hood itself, this could be scary, and the researchers combined it with water from tanks of fire-belly salamanders. In nature, the salamanders don’t live around wood frogs, so the scent by itself should have been ecologically irrelevant.

After the eggs hatched, though, tadpoles that had been exposed to a double-scent cocktail reacted to salamander scent alone as a threat, Mathis reported last year in Proceedings of the Royal Society B. Untraumatized tadpoles didn’t.

In a twist on that lesson, Chivers and Maud Ferrari, also of the University of Saskatchewan, exposed wood frog embryos to salamander scent, with no dead-tadpole spicing this time. Once the little ones hatched, the researchers gave them the full cocktail of ground-up tadpole plus salamander. In theory, such an alarming brew could give youngsters a long-term horror of the salamanders. Yet tadpoles that had been exposed to salamander scent as embryos did not develop a phobia after this treatment. Tadpoles without the early lesson did. An embryonic experience of harmlessness can be as important as a lesson in danger, the researchers say in the April 23 Biology Letters.

Learning when not to panic also avoids unnecessary costs. When Warkentin’s frog embryos escaped a snake attack, they faced consequences. They didn’t become eggs for breakfast, but they were undersized and underpowered as tadpoles darting away from deadly fish or shrimp. And in wood frogs, youngsters that learn fear don’t forage as freely or grow as fast. Trade-offs, trade-offs. Not a bad life lesson for smart embryos of any species.

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K.M. Warkentin.  “Oxygen, gills, and embryo behavior: mechanisms of adaptive plasticity in  hatching.” Comparative Biochemistry and Physiology 2007.

Susan Milius is the life sciences writer, covering organismal biology and evolution, and has a special passion for plants, fungi and invertebrates. She studied biology and English literature.