Winter ecology is like all life - it begins with chemical interactions and finally with energy expenditures. Here are some background details to use in exploring this aspect.

1. Chemical and Solar
Adaptations
to Winter

2. The density of pure water ice at 0°C is 0.9168
g/ml, nearly 9% lighter than liquid water at
0°C, which has a density of 0.99987 g/ml. It is
enough to keep ice floating on top of water
and allows aquatic organisms to survive the
winter. This drop in density occurs because
the hydrogen bonds in water create an open
hexagonal lattice, leaving space between the
molecules.

3. Ice Crystal Snow

4. Colligative Properties
• Some of the properties of solutions do notSome of the properties of solutions do not
depend on the amount and type of solutedepend on the amount and type of solute
present in solution.present in solution.
– Ie food coloring doesn’t affect the boiling pointIe food coloring doesn’t affect the boiling point
of water (much)of water (much)
• Properties that depend on theProperties that depend on the
concentration of solute particles but not onconcentration of solute particles but not on
their identity are called Colligativetheir identity are called Colligative
properties.properties.

5. Electrolyte and non Electrolyte
• Electrolytes are substances that dissolve
in water to give a solution that conducts an
electric current
– Sports drinks and salt water
– Ionic compounds are usually strong
electrolytes because they separate
completely in water
– Covalent compounds can be strong, weak or
non electroyltes

6. • Non-electrolytesNon-electrolytes: a liquid or solid: a liquid or solid
substance that does not allow the flowsubstance that does not allow the flow
of an electric current, either in solutionof an electric current, either in solution
or in its pure state, such as water oror in its pure state, such as water or
sucrose.sucrose.
• Nonvolatile substance is one that hasNonvolatile substance is one that has
little tendency to become a gas underlittle tendency to become a gas under
existing conditionsexisting conditions

7. What about Electrolytes?
• Electrolytes break apart into ions. Each
ion has an effect on boiling point and
freezing points. If a solution has more or
less ions it will change the boiling points
and melting points even more.

8. Plants and Animals in Winter
• Plants and Animals must adapt to the
coming of winter by:
– Behavioral Adaptations
• Bird Migration
• Bear Hibernation
– Physical Adaptations
• Deciduous trees drop leaves in the fall
• Wood frogs and Cope’s grey tree frogs survive by
freezing solid

9. Physical Adaptations
• With most physical adaptations to winter
there is an important chemical adaptation
– This is in response to:
1) Water freezing which, causes
2) Ice crystals to form damaging cells
• Both plants and animals have a variety of
chemical adaptations

11. Productivity
• Energy pyramid is skewed in winter
– Primary producers shut down
– Movement and travel require more energy
– Exposure cause energy drain.
– Food production low
– Need for more food is high

12. Plant sources of heat and loss

13. The complex system of acclimation
carbonic anhydrase II
Catalizes hydration
Of Carbon Dioxide
Kinases enzyme
catalyzes phosphorous
cold response and
improves freezing
tolerance of the
transgenic plants
regulator
CBF – blood flow or fluid
Protein binding gene
Radical Death
gene Melatonin induced
Histomine usually in plant
seed coating

14. Deciduous Trees, Abscisic
Acid, Fructose, Glucose and
Sucrose

15. Sugar Maple
• Taking a sugar
substance from a tree
can be used to create
maple syrup
• The maple syrup
aqueous solution boils
at 219 degree F
• That is 7 degree higher
than water.
• It also depresses the
freezing point

16. Deciduous Trees
• Deciduous trees start producing abscisic
acid
– Due to reduction in photoperiod (seasonal)
– In response to reduction of water
– Because of drop in nutrients for the tree

17. Abscisic Acid
• Is released in response stress
• The main cause of abscission (leaf loss) in
deciduous trees
• Inhibits cell division in the cambian (why tree
rings in winter are narrower)
• Hardens cell membranes to help protect against
ice crystals

18. Growth inhibiting substance as Abscisic acid or Abscissin II, which was once
called as ‘Dormin’. Besides ABA, plants also contain other natural growth
inhibitors such as Coumerin, Ferulic acid, Para ascorbic acid, phaseic acid,
violoxanthin, etc. In addition, plant chemists have identified some synthetic
growth inhibitors. Ex. 2,3,5 tri iodo-benzoic acid, morphactins, caproic acid,
phenyl propionic acid, Malic hydrazide, etc.

19. ABA RESPONSE CHART

20. Sucrose
• Inhibits the formation of ice crystals, it gels as it
freezes
• Commonly known as table sugar
• Maple tree sap is 2% sucrose, the rest is mainly
water

21. Fructose and Glucose
• Fructose (left) is a simple sugar
• Fructose and Glucose are monosaccharides
• Glucose is synthesized from glycerol
• Fructose and glucose are the two main
constituents of sap from birch trees
• Give birch syrup a very distinct flavor

22. Frogs and Glycerol

23. Frogs
• Both the Gray Tree Frog and the Wood
Frog are able to safely freeze almost solid
• There livers produces high amounts of
glycerol
– Through osmosis, gylcerol is exchanged for
water in the cells
• Once thawed in the spring the glycerol is
synthesized into glucose (energy boost)

24. The world of freezing Herps
• About a dozen species of amphibians and reptiles are known to
tolerate the freezing of their tissues under thermal and temporal
conditions that mimic frost exposure in nature (i.e., slow cooling to
relatively high subzero temperatures).
• Some species survive freezing at temperatures as low as -6°C and
endure freezing episodes lasting more than a month. Fully-frozen
animals, in which up to 65-70% of the body fluid has become ice,
appear dead - muscle contraction, heartbeat, and breathing have
completely ceased.
• The frozen tissues become depleted of oxygen and the cellular
energy status declines sharply. Remarkably, these animals arouse
after thawing and can soon resume normal physiological and
behavioral functions.
• Freeze tolerance is promoted by special physiological adaptations,
including an accumulation of certain cryoprotective compounds, a
redistribution of bulk water within the body, and an innate tolerance
of cells to dehydration.

25. Distribution of Rana sylvatica
Map obtained from:
http://www.exploratorium.edu/frogs/woodfrog/woodfrog_3.html
Wood frogs have adapted to live in
the North of the US, Canada and
Alaska by freezing solid during
winter and thawing early in spring.
Distribution of wood frogs (red)

26. What is freezing?
• Life is a complex set of
electrochemical reactions. The
rate at which chemical
reactions take place depends
on temperature, and usually
the lower the temperature the
lower the rate. At absolute
zero Kelvin (-460°F), the rate
is zero. Therefore, lowering the
temperature of biological
materials such as cells,
organs, or entire organisms to
absolute zero causes life to
stop indefinitely.

27. • Because we humans are mostly water, however,
any journey into the supercool is physically
traumatic.
• At about 31°F, the water in our bodies begins to
freeze.
• It starts at this temperature (rather than 32°F)
because biological water is in the form of a
solution, mainly of ions or charged atoms.
• The survival of any cells during freezing
depends at minimum on the rate the
temperature changes.
• For most organisms, even the slowest cooling
results in an assault on their cells that is just too
great.

28. How the freezing begins
• Several mechanisms ensure that wood frogs freeze without
supercooling extensively.
• First, owing to the highly permeable nature of amphibian skin, ice
surrounding the frog can instantly trigger the freezing of the body
fluids.
• Also, the frog’s winter refuge hosts an abundance of ice nucleating
agents, such as various mineral particulates, organic acids, and
certain microbes, that may cause the frog to freeze. Laboratory
experiments suggest that ingestion of these agents promote ice
formation in freeze-tolerant frogs.
• In fact, several strains of bacteria expressing potent ice nucleating
activity have been cultured from the intestines of winter-collected
wood frogs, indicating that such bacteria are retained throughout
hibernation (Lee et al. 1995).
• Inoculation by ice or ice-nucleating agents in the winter environment
probably is the primary mechanism initiating freezing in amphibians;
there is no need for ice-nucleation proteins or other endogenous ice
nuclei, as are found in some invertebrates (Costanzo et al. 1999).

29. Stresses
• Extensive freezing solidifies tissues,
arrests vascular circulation, and
deprives cells of oxygen.
• Because ice forms only in
extracellular spaces, water inside cells
is osmotically drawn externally where
it joins the growing ice lattice.
• During this process cells may shrink
substantially, potentially with damage
to membranes and structural support
systems.
• Macromolecules and solutes become
crowded in a diminishing solvent
volume, perhaps with adverse
consequences.
• Ice formation within body fluids also
poses the threat of mechanical injury
by the growing ice lattice, particularly
in compact and highly structured
tissues and organs.
• Ice fronts may shear and separate
tissues, disrupting intercellular
communication systems.
• Upon thawing, large pools of dilute
fluid form in extracellular spaces.
Cell volume, hydroosmotic balance,
and energy status must be restored.

30. • Resulting cell damage is related to the dissolved
substances or solutes in biological water, to the
cell membrane properties, and to the fact that
ice has a very tight crystallographic structure
and cannot contain solutes.
• When biological materials freeze, the solution
between cells usually freezes first.
• Solutes found in the original solution are ejected
and concentrated in the unfrozen space between
the ice crystals.
• Cells usually remain unfrozen though
supercooled.

31. Danger of Freezing
Rupturing of cell membranes by intracellular ice crystals
Normal tissue Ice formation inside cells
Images taken from: http://www.oncura.com/German/prostate-cryotherapy.html
Intracellular ice crystals
When ice forms inside cells, the crystals break the membranes damaging the tissue.
This is the most common way of freezing, however wood frogs freeze in a different
way…

32. Freezing of Wood
Frogs
• When the frog’s body becomes covered
by ice the liver is stimulated to produce
glucose.
• Their hear rate doubles, aiding in the
fast transfer of glucose to all organs.
• The frog’s large cavities are the first to
freeze avoiding the formation of
intracellular ice crystals which can be
deathly.
Images obtained from:
www.zoldmagazin.com/belso/fagvottbeka.htmal
http://seattletimes.nwsource.com/html/nationworld/2002118796_frogs14.html

33. Bacteria and dust particles found in the
frog’s large cavities act as nucleators for ice
crystal formation. Ice that is growing in these
extracellular spaces attracts water out of
cells by osmosis.
Images obtained from:
www.gsfc.nasa.gov/feature/2004/0116dust.html
www.aquat1.ifas.ufl.edu/guide/bacecoli.jpg
http://coslabindia.com/fibremodel2.htm
Bacteria Dust Particle
Ice Crystals

34. • In order to balance out the resulting difference in
potential energy between the inside and outside
of the cell, water leaves the cell through the cell
membrane.
• Inside the cell, this loss of water causes an
increase in the ionic concentration and leads to
chemical damage. Interestingly, ions, not ice
crystals, trigger cell injury during freezing.
• Theoretically, an infinitely fast rate to absolute
zero would eliminate this harm. This is not
possible, of course, and at higher cooling rates
the supercooling of water in cells causes ice to
form within those cells, which also brings about
damage.

35. Alternative crystallization process
Ice crystals in wood frogs form outside the cell
Extracellular ice formation Dehydration
Images taken from: http://www.oncura.com/German/prostate-cryotherapy.html
Normal tissue
The extracellular crystals cause dehydration of the cells through osmosis, but the
excess of glucose inside the cells protects them from freezing.

36. Wood Frog Freezing
• Cells in freeze-tolerant wood frogs experience
the same mechanism of freezing injury as any
other creatures' cells.
• The frogs freeze very slowly to a temperature
often several degrees below freezing. This
should destroy the frog's cells, yet those cells
and the frog as a whole survive.
• By lowering the amount of water that leaves the
cell during freezing, the glucose offers protection
against the rise in ionic concentration and
excessive cell shrinkage, thereby reducing
chemical harm.

37. Glucose C6 H12 06
• Glucose is a monosaccharide.
It has 6 carbons (blue) 12
hydrogens (yellow) and 6
oxygens (white).
• It can work as antifreeze,
preventing the cells from
collapsing or freezing solid.
http://personal.tmlp.com/Jimr57/textbook/chapter2/cs.htm

38. • The accumulated glucose apparently enhances
the survival of cells, tissues, and organs
because experimentally administering additional
glucose to the frog increases its tolerance to
freezing (Costanzo et al. 1993).
• One of the primary functions of glucose is to
raise the osmotic pressure of the body fluids,
which in turn reduces the amount of ice that
forms at any given temperature.
• Glucose transported into cells acts as an
osmolyte, decreasing the degree of cell
shrinkage during freezing, and also serves as a
fermentable fuel that can be metabolized in the
absence of oxygen.

39. Ice crystals and blood flow
The science of cryogenic surgery
This may look like a set of steak
knives, but it's actually ice crystals
that form in a physiological saline
solution.
These images show red blood cells
between ice crystals, at different
temperatures below freezing. (A) is the
lowest subfreezing temperature, (D) the
highest. Note how the cells shrink as they
become exposed to lower temperatures.

40. Magnetic Resonance Image of a wood
frog in the process of freezing
The dark region is where ice
crystals have formed.
The liver is the last organ to
freeze; it produces glucose
saturating all vital organs
and therefore lowering the
frog's freezing point.
Image obtained from: http://www.exploratorium.edu/frogs/woodfrog/woodfrog_4.html

41. Evolutionary limits
• The frogs have evolved to
produce just the right
composition of cryoprotectants
and gross tissue properties
that allow them to survive
freezing at the temperatures
they experience in nature.
• They cannot survive freezing
at lower temperatures.
• This is the key attribute of
evolution: it solves only the
challenge an organism
encounters and nothing else.
This is the first-ever ultrasound
image of a frozen lesion in a liver.
The arrows point to the margin of
the frozen lesion, which appears
dark because it reflects the
ultrasound pressure waves.

42. Production of
Fibrinogen
Another adaptation of wood frogs
is the production of “fibrinogen”, a
blood-clotting enzyme which helps
stop any bleeding caused by ice.

43. Recovery
• Recovery is remarkably rapid, with basic physiological and
behavioral functions usually returning within several hours of
thawing.
• In collaboration with Jack R. Layne, Jr. (Slippery Rock University),
our work has shown that recovery dynamics are characterized by
sequential restoration of fundamental to progressively more
complex functions.
• For example, the heart resumes beating even before ice in the body
has completely melted, and pulmonary respiration and blood
circulation are restored soon thereafter.
• Contractility in hindlimb muscles returns 1-2 h after thawing,
whereas function of the innervating sciatic nerve is restored within
approximately 5 h.
• Hindlimb retraction and righting reflexes return several hours later
and the frogs usually exhibit normal body postures and coordinated
motor functions within 14-24 h.
• Higher order behaviors, such as mating drive and courting behavior,
are not restored until at least several days later (Costanzo et al.
1997).

44. MRI’s showing the thawing process of wood frogs:
Dark areas are frozen
Light areas have thawed
• Wood frogs thaw out evenly; if the exterior unfroze before
the heart, liver and brain, the limbs would die due to lack
of oxygen.
Images obtained from:
http://www.exploratorium.edu/frogs/woodfrog/woodfrog_5.html

45. Eastern Box Turtle
• This species hibernates in shallow
excavations in deciduous forests
throughout the eastern United States.
Insulated from winter's cold only by a thin
blanket of leaf litter and snow (when
present), these turtles encounter frost in
their hibernacula, yet survive the freezing
of their tissues

46. • Our data, not yet fully analyzed, showed that turtles
occasionally freeze during winter, with body
temperatures falling several degrees below zero.
Survival of these freezing episodes was good. However,
in January of one winter, turtles were lulled out of their
hibernacula by unseasonably high temperatures only to
be caught abroad and immobilized by a rapidly
approaching Alberta Clipper. Two turtles were killed
outright by heavy frost (core body temperatures near –
5°C), and another succumbed some months afterwards.
Overall, the ability to survive most frost exposures
seems to be an important adaptation permitting winter
survival in the northern portion of the species’ range.

47. Painted turtles
• Painted turtles (Chrysemys picta)
inhabit freshwater habitats from
coast to coast in the northern United
States and southern Canada. These
turtles generally overwinter
underwater, except that the young,
which hatch in late summer,
commonly hibernate inside the natal
nest, only ~10 cm beneath the
ground surface. The cold hardiness
of painted turtle hatchlings is
remarkable, as many emerge from
their nests in spring after being
exposed to temperatures that may
fall to -11°C or below.

48. • The biochemical and physiological
adaptations promoting the extreme cold
hardiness seen in some turtles are still
incompletely understood. With respect to
freeze tolerance, there is no involvement of
thermal hysteresis (= antifreeze) proteins or
special ice-nucleating proteins; however, the
question of cryoprotectants is unresolved.
• Freezing turtles accumulate small quantities
of glucose, lactate, and certain amino acids,
though it seems doubtful that the
concentrations ultimately achieved, which are
much lower than those found in frozen frogs,
could substantially reduce the body ice
content.
• An innate anoxia tolerance likely helps frozen
turtles cope with ischemia, though our studies
strongly suggest that the cause of freezing
mortality is unrelated to oxygen deprivation.
Supercooled turtles also accumulate glucose
and lactate, but whether this response
improves survival is unknown.
• One thing is clear, however: cold
acclimatization is crucial to the development
of supercooling capacity, inoculation
resistance, and freeze tolerance (Costanzo et
al. 2000).

49. Supercooling
• The supercooling capacity of hatchling painted turtles is
probably the best of any vertebrate animal, as these
turtles - if carefully isolated from the ice nucleating
agents naturally found in their nest - may cool to -20°C
before spontaneously freezing. Although they may
remain unfrozen, hatchlings nevertheless perish at
temperatures below -12°C. Still, supercooling provides
protection over a remarkably broad range of
environmental temperatures, and clearly allows turtles to
survive at temperatures much lower than could be
tolerated in the frozen state.

50. The Manitoba red garter snake
• Not only does this little 20
inch snake survive harsh
minus forty (- 40) degree
cold winter Canadian
weather but also travel as
much as 50 miles to do
so.
• Narcisse, in the Manitoba
Interlake region lies
between Lake Winnipeg
and Lake Manitoba.
• Here you will see
more snakes than
anywhere else in the
world.
• The limestone
provides winter dens
below the frost line in
this special
“hibernacula”..

51. Snake sex
• First out in the springtime,
are the male red sided
garter snakes.
• Hundreds and even it
may seem thousands
may emerge all once.
The females appear soon
after, but singly or in
small numbers. The
emergence of the female
garter snakes is even
spread over several
weeks.
• As each female garter
snake appears at the
surface she will be
“mobbed” by male
suitors. “Mating balls “
are formed ,
consisting of a single
female garter snake
intertwined with as
many as 100 males

52. Hibernaculum Frenzy
• These dens are sinkholes in the
local limestone rock produced
simply when underground caverns
have collapsed. The resulting
fissures and crevices in the
limestone bedrock give the snakes
access to depths below the frost
line

53. Manitoba Mania

54. Conifers, Alcohols and
Terpenes

55. Coniferous Trees
• Conifer sap contains alcohols and
terpenes (i.e. alpha- and beta-pinene)
• Very little is understood about the specific
chemicals employed by conifers to
prevent freezing
– It is believed that the alcohols and terpenes
help them survive winter
• Spruce will employ alcohols and terpenes
to force the water out of their cells
– This prevents cell damage from ice crystals

56. Alpha- and Beta-Pinene
• Very low melting points (i.e. freezing
temperature) down to -60ºC
• Main constituent in conifer sap
• In the terpene family of compounds
– Hydrophobic alkenes

57. Alcohols
• Alcohols depress the freezing point of water
• A variety of alcohols are employed by conifers to
survive the winter
• Are characterized by an -OH group attached to
alkanes

58. Mosses and Arachidonic Acid

59. Mosses
• Many mosses continue photosynthesizing
throughout winter
• Some animals (lemmings and reindeer for
example) will eat mosses to help them
with the low temperatures of winter
(behavioral adaptation)
– This allows animals to gain the benefit of the
mosses’ natural anitfreeze -arachidonic acid

60. Arachidonic Acid
• Helps cells keep moving in low temps
• Acts to toughen cell membranes to protect
against ice crystals
• Is a second messenger to relay signals within a
cell
• Makes up 35% of the fatty acids present in moss

62. Animal heat response

63. lower critical temperature (LCT) is the
turning point at which more heat is lost
to the environment than is normally
metabolically produced.
The lower lethal temperature (LLT) is
the extreme cold temperature where an
animal can no longer produce enough
heat and dies of hypothermia.
zone of metabolic regulation

66. Solar collection
• Solar radiation plays a large part in
determining not only ambient and body
temperatures but animal behavior as
well. A wide variety of species have
developed methods to reduce the cost
of thermoregulation by behaving
certain ways: seeking shade,
burrowing, panting, gular fluttering,
wing flapping when exposed to
temperatures above the UCT, entering
short bouts of torpor or longer bouts of
hibernation, increasing insulation, or
migrating.

67. Most organisms attempt to remain within a favorable
range of temperatures. For homeotherms, this is known
as the thermal neutral zone (TNZ). This optimal range of
ambient temperatures typically lies between 30-42°C
Solar Radiation homeothermic or euthermic organisms
(Speakman, 2004). Within this range, metabolic rate is
minimal. Outside the TNZ, metabolic energy is required
to maintain TB within the optimal range. Metabolic
activity involving temperature-dependent enzyme
activity will not function properly if the animal becomes
hypo- or hyperthermic. Thus, thermoregulation is an
integral part of an organism’s energy balance.

68. Emissivity is defined relative to what is known
as a black body, a perfect emitter with an
emissivity of 1.0. Most animals have an
emissivity value within 0.90-0.98, often
dependent on surface properties such as fur or
skin color. Surfaces either reflect or absorb light
to varying degrees contingent on pigment levels
and texture that in turn affect emissivity. Dark
colors absorb energy within the infrared
spectrum, increasing the absorptivity of
inanimate objects or organisms compared to
light colors which reflect visual wavelengths of
solar energy.

69. Heat Loss
An animal can either increase its heat production or
reduce its heat loss to maintain homeothermy.

70. In a countercurrent heat exchanger, the hot fluid
becomes cold, and the cold fluid becomes hot.

72. A homeothermic animal is in a continual state of dynamic equilibrium between heat
production and heat loss. The continual adjustment of physiological and behavioral
responses to the changing energy flux in the environment results in short-term
temperature changes in the animal