A few weeks ago Jane Viti, one of my teaching colleagues, asked me what was going to happen to the two frogs that had been living in her small, backyard pond all summer. As we talked about her frogs’ appearance, behavior, and songs I decided that they must be Northern green frogs (Lithobates clamitans melanota), and since her pond is quite shallow and expected to freeze solid over the winter, and also because it is surrounded by a very dense growth of myrtle, I speculated that the green frogs (which can either hibernate underwater or underground) would leave the soon-to-be-solid pond and dig a hibernaculum in the soil under the protective cover of the myrtle. But where these frogs sit out the winter was just the beginning of this story. Frogs are ectothermic (they rely on the heat of their environment for their body heat) and would seem quite vulnerable to freezing solid in the very cold winters of Western Pennsylvania even if they were underground or underwater. How could they survive this extreme thermal trauma?
In my Cell Biology class I talk about changes that can be seen in cell membranes in both amphibians (like the green frog) and reptiles (like turtles and snakes) as seasonal temperatures begin to fall. An enzyme is stimulated that begins to add double bonds to the fatty acids of the cell membrane phospholipids. This “desaturase” enzyme makes the altered fatty acids more crooked and thus less able to stick together. This reduces the freezing point (which usually referred to as the “melting point” for some reason) of the cellular membrane and keeps the membrane “fluid” and functional at lower and lower temperatures. Also, amphibians add cholesterol to their cell membranes, and these steroids further keep the fatty acids from clumping together even at decreasing temperatures.
These changes help to keep our frog active at temperatures that are lower than optimal, but eventually temperatures start to approach the freezing point of water, and the frog is at risk of cell and tissue damage from the freezing of the water in its blood and cytoplasm.
Let’s take a second and think about people. When skin is exposed too long to freezing temperatures cells are destroyed and “frostbite” occurs. Why do the cells die? First, the blood flow into the cold body part is curtailed in order to prevent excessive body heat loss (humans are endothermic organisms who use the heat from their metabolic activities to generate their body heat and there is only so much heat energy to go around!). The lack of blood flow into the tissue means that oxygen is no longer being delivered and cell death from lack of oxygen may occur. Also, and maybe of a more immediate concern, the lack of warm blood entering the tissue means that the fluids in the tissue and in its cells may start to freeze. Usually the interstitial fluid around the cells freezes first and these ice crystals actually start pulling water out of the inside of the cells. For a while this dehydration event may actually hold off cytoplasmic freezing, but eventually the cell will be irreversibly damaged (i.e. “killed”) by either excessive dehydration or by the inevitable freezing of its cytoplasm.
But, let’s get back to our frogs: before the frogs are exposed to freezing temperatures they undergo many physiological changes in addition to the cell membrane changes I listed above. Their livers start synthesizing and releasing large quantities of glucose (“sugar”) into their blood streams. These sugars are absorbed by the cells of the body causing the cytoplasm to become thick and syrupy and increasingly hypertonic to the surrouding interstitial fluids. Also the frog releases special proteins called Protein Ice Nucleases (or “PIN’s”) into their blood stream. These proteins will stimulate freezing of the water in the blood stream which will then inhibit the potentially lethal freezing of the water of the cytoplasm inside the cells! When the frogs are finally exposed to truly freezing temperatures the skin and then the rest of the body freezes solid (they are like little rock statues of frogs!), but the freezing is primarily confined to the blood and to fluids around the cells! The forming ice crystals in the interstitial fluid draw water out of the cells (just like in human frostbite) but the high levels of sugar inside the cells not only act as a natural antifreeze for the cell but also hang onto enough water so that the cells don’t dehydrate to the point of death!
Terrestrial frogs (like the American toad (Bufo americanus), the wood frog (Rana sylvatica), the spring peeper (Hyla crucifer), the gray tree frog (Hyla versicolor) and the northern green frog when it decides to hibernate on land) basically let themselves freeze solid in their soil hideouts. Wood frogs and tree frogs don’t even go down into the soil but just bury themselves in piles of leaves and ride out the months of freezing temperatures. During warm spells these terrestrial hibernators may even thaw out and move around, but they will typically then re-freeze and settle back into their winter slumbers. I noted in several spring and summer blogs this year the very large number of gray tree frogs in the trees around my field and yard. I wonder if all of the leaves that I have been letting pile up under my trees (because of my selective leaf-raking policies) provided these great creatures with sufficient winter habitat to favor the growth of their population?
Aquatic frogs (like the leopard frog (Rana pipens) and the American bullfrog (Lithobates catesbeianus) and the northern green frog when it decides to overwinter in a body of water) spend the winter if not frozen then nearly so in the still liquid environment of their ponds or pools. They do not bury themselves in the muds of these systems because they must continue to pick up oxygen from the surrounding water through their skin. Sometimes they sink to the bottom of their pools or ponds (they are quite solid and have no air in their lungs) but they must keep contact with the oxygen-rich water in order to survive. They also may swim about a bit when they warm up during lulls in the winter cold. Aquatic frogs can survive freezing solid in ice, but I don’t know how long they can live that way. The lack of oxygen would surely be fatal if the ice-encasement persisted for too many weeks.
So, on the next cold Fall night as you sit in your warm house wrapped in an afghan or a sweater, give a thought to the little frozen frogs outside who are waiting for their personal Spring thaws to come.