Arctic Froggies: I Bet You Freeze Good on the Dance Floor
During a cold, winter’s evening, what do you like to do? Wrap up in a thick blanket, turn the heating on and make a mug of hot chocolate? These, unfortunately, are not luxuries that the wood frog can afford. The wood frog (Rana sylvatica), also known as the Alaskan wood frog and mistakenly known as the Alaskan tree frog, is an amphibian with a wide distribution amongst some of the coldest regions of the world, occurring most frequently in North America. The wood frog has gained status in the scientific community over the last 100 years due to its almost unparalleled ability to freeze itself during harsh winters. The wood frog is also an obligate breeder, naturally occurring in ephemeral wetlands, which creates an even harsher environment for itself, adding to its standing.
Winter is coming!
Around the world, there are approximately 4,740 species of frog in total, however the only one of these with the ability to freeze is the wood frog. The wood frog typically lives on the forest floor and here it encounters freezing temperatures, sometimes reaching subzero. To deal with these temperatures, up to two thirds of the bodily water inside the wood frog freeze, making the frog hard enough to break through a window were it to be thrown at one. During this time, many bodily functions stop occurring – metabolic activity, waste production and perhaps most importantly, their hearts stop beating and this can occur for weeks at a time. The Alaskan wood frog in particular has refined this skill the best, as they can be frozen to temperatures as low as -16 degrees Celsius and live, compared to Ohioan wood frogs which can only survive -4 degrees Celsius. The term “freeze/thaw” is commonly used when referring to weathering, however the wood frog may experience many “freeze/thaw” cycles in that during one winter, it may freeze and thaw multiple times, due to the shift in temperature between night and day. Once thawed, the frog can simply hop away, as shown in the video below.
So how do they do it?
In the process of freezing, typically a cell will experience shrinkage due to ice formation pulling water out of the cells inside the body, eventually killing the cell as a cell cannot survive with no transport medium or nutrients. To combat this problem, the wood frog’s blood is full of cryoprotectants, which are equally dispersed across the entire body of the frog. The job of the cryoprotectants is to lower the freezing point of any tissue and also to reduce the quantity of ice that can form in any body part at any time. A high concentration of cryoprotectant in the blood, therefore, will reduce the amount of ice that can form, hence removing the stress put on cells and tissues during freezing. The main cryoprotectants are glucose and urea, and these compounds have been found in much higher concentrations in the Alaskan wood frog than the Ohioan, shown in the picture to the right, which explains the difference in temperature tolerance. In the process of freezing, the synthesis of glucose occurs in the liver, before it is transported around the rest of the body, which is why glucose concentration is highest in the liver in the picture on the right. However, there is a large difference between even the freezing potential of lab frogs and wild frogs. In a study conducted by the University of Alaska, scientists found that in wild frogs, cryoprotectant concentration and particularly glucose, was 13 times higher across all bodily tissue, 3 times higher in the liver and 10 times higher in heart tissue.
The one draw back to this freeze/thaw process, is that as soon as an ice crystal touches the frog, regardless of surrounding temperature, the immediate and natural bodily reaction is to begin the freezing process, sometimes unnecessarily putting osmotic stress onto the cells and tissues. The activities of 17 enzymes were followed in a separate experiment during the freezing process, and it was found that liver phosphorylase activity increased by 520% and a similar compound to glucose, the enzyme glucose-6-phosphatase, saw an increase of 140%. At these levels, the frogs create an internal environment which can become lethal and so this regulation has to be exact in order for them to not over protect themselves too much.
The freezing hero to the rescue!
The freezing methods currently available for medicinal storage and fertility treatments are not 100% reliable, however researchers believe that they can change this with the help of the wood frog. Using a method known as “Sandals“, short for “Small Angle Neutron Diffractometer for Liquids and Amorphous Samples”, the researchers have been able to recreate the conditions that the wood experiences while freezing, and have discovered that glycerol hinders ice crystal formation in water, even when cooled to temperatures as low as -35° C. The glycerol acted as a cryoprotectant, by forming a large network around water molecules present and prevented them from freezing. The results of this study means that the wide scale production of cryoprotectants could soon be a realistic prospect, boosting the fertility treatment industry by providing a reliable way of freezing sperm and egg cells, but also providing a safer way to store medicine and food.