The 4 th International Academic Conference on Environmental and Occupational Medicine

1. The 4th International Academic Conference on Environmental and Occupational Medicine Kunming, Yunnan, PR China Oct. 16-19, 2006 Assessing deficits of central nervous system functions in the detection of eventual environmental and occupational health hazards Michael Bornhausen, MD Institute of Animal Physiology, Faculty of Veterinary Medicine, University of Munich, Germany

2. Former affiliation Institute of Toxicology German National Research Centre of Environmental and Health Sciences (GSF-FZ) München-Neuherberg, Germany Research topics: - radiobiology - spectral analysis of EEG (FFT) - single nerve cell activity, spike train analysis - operant-behavior - learning, memory - design and evaluation of alternative methods in toxicology

3. General aim Assessment of the dynamics of changes of CNS functions Specific aim Detection of cognitive effects eventually induced by chronic exposure to GSM or UMTS electromagnetic fields in three consecutive generations of rats

4. Acknowledgement Part of this work was supported by the German Federal Office of Radiation Protection (Bundesamt für Strahlenschutz, BfS) and done in cooperation with: • Dr. H. Scheingraber, Max-Planck-Institute of Extra-terrestrial Physics (MPE), Garching, • Dipl.-Ing. C. Kögler, ck-3d IT solutions, Emmering, Germany • Prof. Ping LIU, Prof. Xinan WU, Department of Public Health, Kunming Medical College, Kunming, Yunnan, PR China

5. Key words • health hazards • electromagnetic fields (EMFs), cell-phone • Central Nervous System (CNS) • cognition (learning, memory) • operant-behavior • microstructures (inter-response intervals, IRIs) • dynamics of serial properties in operant-behavior tests • rats

6. Literature • M. Bornhausen and H. Scheingraber: Prenatal exposure to 900 MHz, cell-phone electromagnetic fields had no effect on operant-behavior performances of adult rats. Bioelectromagnetics 21:566-574, 2000 • C.B. Ferster and B.F. Skinner: Schedules of Reinforcement. Appleton-Century-Crofts, New York 1957 • R.P.Jensh: Behavioral testing procedures: a review. In: E.M. Johnson, D.M. Kochhar (eds.): Handbook of experimental pharmacology, vol. 65. Teratogenesis and reproductive toxicology. Berlin, Heidelberg, New York: Springer Verlag, 1983 • M.H. Repacholi: Low-level exposure to radiofrequency electromagnetic fields. Health effects and research needs. Bioelectromagnetics 19:1-19, 1998 • B. Weiss, J.M. Ziriax, M. Christopher Newland: Serial properties of behavior and their chemical modification. Animal Learning & Behavior 17(1):83-93, 1989

8. China U.S.A. Germany • area 9 598 029 km2 9 629 047 km2 357 046 km2 • population 1 288 700 000 291 500 000 82 600 000 • population/km2 134 30 231 • CO2-emission/subject 2.5 tons 19.8 tons 10.0 tons • energy consumpt./subj. 880 kgoe 7 960 kgoe 4 230 kgoe • smokers 35.6 % 23.6 % 35 % • meat consumpt./subj. 47 kg 122 kg 87 kg • use of paper/subject 33 kg 331 kg 233 kg • no. of persons/room 1.1 0.5 0.5 • water consumpt./subj. 1 203.3 l 5 024.7 l 1 586.3 l /day (agriculture, industry, households) • TV-sets/1000 subj. 292 844 586 • cars/1000 subjects 16 774 565 (Courtesy of National Geographic, March 2004; sources UN, WHO; oe = oil equivalent)

9. American life style "... If everybody on Earth would live like the US-Americans we would need three planets to keep our living standard." Peter RAVEN, botanist and environmentalist (Courtesy of National Geographic, March 2004)

10. Persistent organic pollutants (POPs) Long-term exposure to even small doses of • dioxines • furanes • polychlorinated biphenyls (PCB) • endosulfanes • polybrominated diphenylethers (PBDE) • DDT • etc. may have serious CNS consequences in man.

11. Rationale for experimental studies of cognition after chronic exposure to environmental hazards (e.g. GSM- or UMTS-electromagnetic fields [EMFs]) • changes of function are precursors of structural changes • functions of the Central Nervous System (CNS) are more sensitive than functions of other organs • CNS functions are most sensitive during prenatal development

12. No effects after prenatal exposure M. Bornhausen and H. Scheingraber: Prenatal exposure to 900 MHz, cell-phone electromagnetic fields had no effect on operant-behavior performances of adult rats. Bioelectromagnetics 21:566-574, 2000

13. Reasons for the choice of automated operant-behavior testsNegative example: Appearance of “developmental landmarks” (various physical signs and neuromuscular reflexes) in rat pups must be carefully and repeatedly checked by experienced personnel postnatal day 0 day of birth 2 surface righting 4 pinna detachment 8 cliff avoidance 9 negative geotaxis 11 forelimb grasp 15 bar holding 16 eye opening, forelimb hanging, air righting 17 development of fur 19 ear unfolding 25-30 descent of testis, opening of vagina

14. Operant-behavior • Operant-behavior tests are run in a battery of 10 standardized test chambers (“Skinner boxes“). • Subjects are required to press a lever for food reinforcement (pellets of 45 mg) . • 15h-nocturnal test sessions (16:00-07:00 CET) are subdivided by alternating 30 min ON- and 60 min OFF-cycles. • Final tests require the subjects to respect a blocking interval of 16 sec after a reinforcement.

15. Operant-behavior test chambers(“Skinner boxes“)

16. Operant-behavior schedules • Differential Reinforcement of Zero Rate (DR0) • Differential Reinforcement of High Rate (DRH) • Differential Reinforcement of Low Rate (DRL)

17. Actual sequence of operant-behavior tests • DR0 5 min (ss. are automatically trained to press a lever for food reinforcement) • DRH 2/1 (ss. are required to press the lever 2 times in 1 sec for a reinforcement) • DRH 4/2 (ss. are required to press 4 times the lever in 2 sec for a reinforcement) • DRL 1/16 (ss. are required to respect a blocking interval of 16 sec after a reinforcement; a precocious bar press resets the interval to its start. The interval is signaled by a green cue light.)

18. Actual BfS-sponsored research project Detection of cognitive effects – if any – after chronic exposure to GSM- or UMTS- electromagnetic fields (EMFs) in three consecutive generations (F0, F1, F2) of rats

19. Acceptance problem • Cellular phones are welcome; base stations, however, are being disputed. • Advantages of individual cellular phones in emergencies and accidents are recognized. • But people living in the vicinity of base stations and having visual contact to the antenna, very often say NO. • Controversial discussions concerning an alleged health risk of base stations are often very emotional and can be characterized by a syndrome well known to psychologists: • ignorance of scientific facts • unsufficient transfer of knowledge • and “ANGST” (fear of something you don’t understand)

20. Focus of public concern • digital pulses of cell-phone signals are thought to be particularly dangerous • continuous exposure by base stations • high risk of sensitive people • definition of exposure limits relative to thermal effects only • health hazards by non-thermal effects • unsufficient scientific data, especially concerning eventual consequences of long term exposure (5 % of the general population in Germany, only, believe that mobile communication does present health risks.)

21. Microstructures • Microstructures of operant-behavior test sessions are shown by pattern of inter-response intervals (IRIs, i.e. time between consecutive lever presses with a resolution of 1 msec). • IRI-pattern differentiate between “learners“ and “non-learners“.

22. Superimposed scatter plots of the 10 ON-cycles of a DRL session

23. Assessing dynamic vs. static changes Static results at the end of a test session (f.i. the ratio of correct responses to all responses) do not reflect the dynamics of test requirement acquisition.

26. Dynamic signal When looking at the sequence of 10 consecutive ON-cycles of a 15h-nocturnal test session - a signal becomes apparent that is proportional to the ability of the subject to acquire the test requirement (i.e. to "learn").

27. Susceptibility of the "learning" process • We are not only interested in the fact whether a subject has finally acquired the test requirement (i.e. has "learnt" the task). • We go further and measure the intrinsic properties of the "learning" process itself.

28. The phenomenon of learning - can be appropriately described by an S-shaped curve parameters: • start time • base line • rise time • saturation • turning point

29. Measuring serial properties of operant-behavior Inter-response intervals (IRIs) between consecutive lever presses are recorded (in multiples of 1 msec), weighed and normalized relative to an empirically determined standard IRI-distribution, and expressed as "points". The sequence of points, then, is fitted to an S-shaped curve by an appropriate algorhythm.

30. Mathematics of S-shaped curve fit Minimize the sum of squares of a set of equations. x=arg min(sum(func(y)2)) Description: Return the point which minimizes the sum of squares of M (non-linear) equations in N unknowns given a starting estimate, x0, using a modification of the Levenberg-Marquardt algorhythm.

31. Examples of actual research The following graphs (scatter plots and corresponding S-curve fits) are examples of our actual and ongoing research project focusing on the question: Are there any deficits of cognition (learning ability, memory) after long-term exposure to cell-phone EMFs in rats ?

36. Changes of CNS functions Obviously, the ability of an animal subject to acquire an operant-behavior test requirement (i.e. to "learn") can be mathematically assessed (in terms ofpoints, rise time, turning point, saturation, start time, etc.) and used to quantify subtle deficits of CNS functions.

37. Conclusions • Operant-behavior contingencies are useful in the assessment of potential health risks, in toxicology, environmental protection, etc. • The analysis of microstructures of operant-behavior test performance is a powerful tool to demonstrate changes of CNS functions. • The dynamics of test acquisition (i.e. "learning”) can be measured by S-curve fits and used to enhance test sensitivity of CNS functions in situations of potential health hazards.

38. Thank you for your attention !