In the midge world, Belgica antarctica larvae are something akin to superheroes. Resistant to anoxia for weeks, tolerant of pH ranging from 3-12 and able to survive losing 65% of their body water, these creatures are bywords for tough. But possibly their most remarkable characteristic is their ability to survive being frozen solid. An Antarctic veteran, Richard Lee has been intrigued by the residents of this remote continent ever since his first visit as a postdoc in 1980. Puzzled by B. antarctica's stress tolerance, Lee, David Denlinger and their colleagues wondered whether the insect's larvae would acquire increased cold tolerance as a result of a previous exposure to a cold snap. Lee explains that this phenomenon, known as`rapid cold hardening', is well studied in many freezing intolerant insects,but has never been reported in creatures that miraculously survive being frozen alive. Could B. antarctica's remarkable freeze-tolerance be extended by rapid cold hardening(p. 399)?
Lee and Denlinger headed south with postdocs Joseph Rinehart and Scott Hayward. The team was also accompanied by a local high school science teacher,Luke Sandro, who related his experiences doing science in Antarctica to colleagues and students over the internet in daily web-logs. Working long hours in the austral summer, the team gathered thousands of midge larvae from the mud and penguin detritus around the US's Palmer Station.
Back in the base's lab, the team tested the larvae's freeze-tolerance by cooling them to -10°C; 75% of the larvae died. But when they precooled the larvae for an hour at -5°C before plunging them to -10°C, the insects'tolerance increased dramatically, with the death rate falling to less than 15%. The larvae had undergone rapid cold hardening.
Shipping some of the untreated larvae in icepacks back to the northern hemisphere, Lee was surprised to find that the insects had serendipitously become acclimated to winter conditions by the time they arrived in Ohio. Michael Elnitsky, Lee's graduate student, began testing the winter acclimated insects' cold tolerance, finding that they survived much lower temperatures than the summer-adapted larvae in Antarctica. But how would these winter-adapted insects respond to a dose of rapid cold hardening?
Elnitsky precooled the larvae to -5°C for 1 hour before freezing them at -15°C for 24 hours. This time, more than 85% of the larvae survived a temperature that was fatal for the summer larvae. In fact, most of the larvae survived -20°C after rapid cold hardening. Lee, was astonished; rapid cold hardening had extended the limit of freeze tolerance significantly.
But how are the insects able to cold harden so quickly? First Lee and Elnitsky tested to see whether an episode of rapid cold hardening caused the insect to produce protective antifreeze molecules, but could find no evidence. Next, they began investigating how tissues responded to the cold, with and without rapid cold hardening. The team found relatively high levels of cell death in tissues that had not been rapidly cold hardened, while the death rates were much lower in rapidly cold hardened cells. Lee believes that the physiological basis of rapid cold hardening is cellular, but finding the mechanism is going to be a tricky task.
