1. biological dynamics  Dynamics = how systems change/evolve with time  Why are dynamics important to biological systems?  Temporal behavior of proteins, cells, organisms  metabolism, cell growth, development, protein production, aging, death, species evolution … all are time dependant processes  Inherent complexity in biological systems both in time and space  temporal patterns are related to structural ones

2. traditional biological framework  tend to think in terms of single point equilibrium processes  a stable and constant steady state time ---->

3. biological dynamics can be complex  as time goes to infinity … response doesn’t have to go to a single constant value could for example have oscillations that would go on forever unless perturbed time ---> can even have aperiodic behavior that goes on forever but never repeats (chaotic)!

4. Oscillations / Rhythms Occur in Nature At all time scales Predator Prey Population Cycles (years) Circadian Rhythms (24 hours)  sleep wake cycles Biochemical Oscillations (1 – 20 min)  metabolites oscillate Cardiac Rhythms (1 s) Neuronal Oscillations (ms – s) Hormonal Oscillations (10 min - 24 hour) Communication in Animal and Cell Populations  fireflies can synchronize their flashing  bacteria can synchronize in a population

5. Biological Clocks  Why do you think that you sleep at night and are awake during the day? External or Internal Cues?  What do you think would happen if you were in a cave, in complete darkness?  If you were in a cave.. Could you track the days you were there by the number of times you fell asleep and woke up? i.e. 1 wake – sleep = 1 day = 24 hours?

6.  Humans w/o Input or External Cues diurnal (active at day constant darkness  ~25 hour clock (>24 hours) wake up about 1 hour later each day constant light shortens the period  Rodents nocturnal (active at night) constant darkness  ~23 hour clock period (<24 hours) wake up a little earlier each day constant light lengthens the period Internal Clock Day time of day

7. Circadian Rhythms Everywhere!  actually more rare for a biological factor to not change through-out the 24 hour day  temperature  cognition  learning  memory  motor performance  perception all cycle through-out the day

8. The Circadian Clock  1. Period of ~24 hours  2. synchronized by the environment  3. temperature independent  4. self-sustained (--- therefore inherent) Defined By:

9. Clock Definitions  Period ( T )– time for the rhythm to repeat T

10. Clock Definitions - Phase Shifts  Phase Advance Phase Delay Period (T) remains the same External cues can shift the phase

11. Basis of the Clock?  Self-sustained rhythm  Inherent Period must be some inherent mechanism cells … proteins … genes??  Light Dark Pattern modulates the Phase “sets” the clock (and period slightly) What is the cellular mechanism? How does light interact?

12. Entrainment  Entrainment: Causing a gradual phase shift so that the oscillation becomes synchronized with the entraining rhythm or signal  Zeitgeber: entrainment signal German for “time” “giver”  Light Zeitgeber”: Sleep / Wake Circadian Rhythm entrains primarily to light  You know this phase shift - adjust to traveling overseas Direction and time of day you fly makes a difference light therapy strategies for jet lag

13. Zeitgeber All light inputs in mammals come in through the photosystem

14. Genes?  Drosophila (fruit flies) convenient for genetics mutation – behavior genetic screens  Screened for flies with altered circadian rhythms some too short some too long some arrhythmic (no repeating pattern at all)  What genes are mutated?

15. Seymour Benzer Purdue Physicist. PhD from Purdue in 1947 Important role in the invention of the transistor Cal Tech. became interested in genes and behavior highly original experiments 1.mutating drosophila 2.Sort for specific behavior changes or deficits 3.Search for the underlying gene mutation became the father of neurogenetics Discovered genes underlying circadian rhythms

16. Time, Love & Memory By Jonathan Weiner

17. Genes Involved?  one of them period (per) * found point mutations in period some delayed, some advanced, some arrhythmic found that Period (PER) levels oscillates in single cells with period of 24 hours! Clocks in single cells?? Are the oscillations inherent to Period? Or does another gene / protein interact to create the oscillations?

18. Mathematical Model  Based on biochemical, cellular, and gene data  Can Period support its own oscillations? hypothesis: negative feedback knew this existed from experimental data question?: can this system alone oscillate? answered this with modeling yes

19. Feedback Mechanism make mRNA make protein mRNA transport into cytosol protein phosphorylation (2Xs) phosphorylated protein travels into nucleus phosphorylated protein inhibits mRNA production This model can in fact support 24 hours oscillation of Per protein levels But no mechanism for entrainment!! How does light interact? Model … incomplete??? Goldbeter NATURE VOL 420 14 NOVEMBER 2002

20. Other genes?  timeless (tim) timeless mutations were arhythmic Timeless affected Period  Per location  Per level  Per protein oscillations  Per phosphorylation How does Period and Timeless interact? How does light interact?

21. Drosophila PER TIM Model fully phosphorylated PER and TIM form a complex that inhibits expression of both Ahhh!! Light induces TIM degradation Can this model simulate the experimental observations of effects of light? Answer … most of them!! … but still a few things missing … other genes etc. Leloup & Goldbeter, J. theor. Biol. (1999) 198, 445–459 Goldbeter NATURE VOL 420 14 NOVEMBER 2002

22. Leloup & Goldbeter, J. theor. Biol. (1999) 198, 445–459

23. example of simulation output protein oscillations mRNA oscillations PER TIM complex oscillations Leloup & Goldbeter, J. theor. Biol. (1999) 198, 445–459

24. Entrainment Light Destroys TIM When tim RNA is high … it delays the clock (phase delay) Because mRNA is ready to quickly replace the destroyed TIM When tim RNA is low … it advances the clock (phase advance) Because mRNA is not ready to quickly replace the destoyed TIM So effect of light depends on the tim mRNA levels protein oscillations mRNA oscillations PER TIM complex oscillations

25. More complete known Per / Tim Feedback Loop in Drosophila

26. Humans - Free Running Clock (FRP)  Actually variation in FRPs in humans  Can be longer or shorter than 24 hrs  Entrainment depends on the FRP of the clock and the light cycle  People who like to go to bed early and get up early often have FRP <24 and vice versa

27. Suprachiasmatic Nucleus (SCN) Wang et al. BMC Developmental Biology 2001 1:9 A symmetric pair of nuclei In the hypothalamus Just behind the nose Near the crossing of the optic nerves Is the “master circadian clock” in the brains of humans

29. Circadian rhythm of firing activity of SCN neurons

30. Dissociated neurons, however, are not in phase

31. Block the cells for days And the firing rhythm emerges again with the same phase The circadian rhythm firing is an inherent property of individual neurons

32. Clock Genes Vs. Clock Controlled Genes Clock genes Clock controlled genes Such as: Temperature Blood pressure Cognitive Hormones

33. Output Rhythms Secondary Oscillations in Body Isolated organs and cell from other part of body also display circadian oscillations in gene expression Phased slightly later than the SCN “pacemaker” Will generally dampen out (not sustain) after isolation Physiological Outputs Blood pressure (lowest just after midnight) Cognitive Performance (best in mid afternoon) Hormones Cortisol (highest in morning) Melatonin (highest at night)

34. Clock Phathologies  A type of dynamic disorder  Appears to live 25 hour day on average despite light dark cues  Not properly entrained  Has a weak zeitgeber response  Man suffers from severe depression

35. Human Pathologies  Advanced Sleep Phase Syndrome  Delayed Sleep Phase Syndrome  Light Entrainment Impacts some blind individuals