Laboratory Animal Nutrition

1. Laboratory Animal Nutrition Timo Nevalainen University of Eastern Finland

2. Nutrient requirements • Each species has specific needs for essential nutrients • e.g. vitamin C essential for guinea pigs, not for rats • National Research Council has published detailed documents for each species • Requirements are ”minimum needs” under ad lib / maximum growth conditions

3. Nutrient requirements documents for all species freely available: http://lab.nap.edu/nap-cgi/discover.cgi?term=nutrient+requirements&restric=NAP&mw=&submit.x=6&submit.y=11

4. Why to fulfill nutrient requirements? • Essential needs of specific nutrients for each species and each period in life, for maintaining good health, and • For obtaining reliable results Skin lesions in Wistar rats due to dietary Vitamin B 6 and B12 deficiency

5. Toxic levels of nutrients • Influence of methionine on atherosclerosis in apoE deficient mice • 2.2 and 4.4% dietary methionine • ->toxic effects Zhou Art.Thromb.Vasc.Biol.2001

6. Responsible concentrations • Purified AIN diet • Control 0.5% methionine (NRC level) • Highest level: 1.5% methionin • No toxic effects • Reliable results • Good welfare

7. Types of eating & digestion • Rodents practice coprophagia • Rabbit also coprophagic; and is a rear end fermentor • Dog and cat are carnivores • Sheep and goat are ruminants; and front end fermentors • Swine is a omnivore like humans

8. Types of diets used • Natural ingredient diets = made of natural ingredients, such as maize gluten, corn oil, wheat • Frequently used • Composition such that pelleting is possible • Relatively cheap Ritskes-Hoitinga et al. 1991

9. Natural-ingredient diets”Chow diets” • ”Standard” diets commercially available for each species • Special diets for Growth/maintenance and Reproduction • Composition meets the needs for each species and condition more than sufficiently

10. Diet types • Natural chows • Chemically defined • Purified diets

11. Problem with natural ingredient diets?? • Beynen 2001: • Between brand variation • Between batch variation • Batch analysis certificate strongly advised

12. Between-brand variation in 9 natural-ingredient minipig diets Catalogue values Ritskes-Hoitinga, Bollen 1999

13. Between-batch variation4 batches of minipig diet Ritskes-Hoitinga, Bollen 1999

14. Results from feeding10 brands of rodent chows Ritskes-Hoitinga et al. 1991

15. Variable composition gives variable results: 2 yr tox studies in 5 labs Haseman 1984, Roe 1994

16. Standardization requires description • Survey of scientific articles • Brand name told in 10-15% • Composition shown in 2% • Conclusion: • If diet has no effect, then OK • If diet has effect, no way to repeat

17. Solution for variation? • Stick to one batch • Difficulty: limited duration • Analysis for critical elements in the diet • Detailed description of diet in scientific articles

18. Diet forms • Full chows in use • Only diet and water available • Standard and expanded pellet • Powdered diets • Test substances can be mixed • Diet is wasted • Special equipment for feeding • Liquid and colloid diets

19. Measuring diet consumption • Diet used = eating + spillage • Metabolism cage – only reliable • In group housing get averages • Values calculated or from reports are for guidance only

20. Designing special diets • If one wishes to change the fat content of the diet, isocaloric exchange is needed, because, • Fat contains more calories per gram than carbohydrates and protein • When changing the fat concentration in the diet, the energy concentration changes • A change in fat concentration must be accompanied by an exchange with e.g. carbohydrates on the basis of calories • If this is done on the basis of weight, then the animals will ingest a different amount of ALL nutrients

21. Diet changes • Dilution effect • All additions have some • Energy rich additions are problematic • Energy rich drinking solution are no different • Maximum dilution 10-20 % • Results in different amounts of feed eaten

22. Feeding types • Ad libitum. Takes place mostly during dark • Meals • Day or night? Usually restricted feeding time • Restricted feeding • amount given is limited • Pair feeding • Same amount for study and control groups

23. Standardization in relation to biological rhythms • All processes show circadian rhythms • Feeding should be in accordance with their natural rhythms • Otherwise disturbed health, well-being and results • More research needed

24. Advantages of restricted feeding • Animals live longer • Fewer spontaneous tumors • In carcinogenicity study • test compound may decrease appetite -> carcinogenicity of compound may be underestimated • some assessment of diet consumption is necessary to find out which is the case

25. ’Work for food and enrichment’ • Hypothesis: • Rats will work ONLY for food they necessarily need, provided work intensity is properly set • Study • pellets placed in drilled holes of aspen board • 3 rats (250 g- ) in a cage, 20 week study • 3 groups • A & B = diet enrichment combination (2 ‘doses’) • C = ad libitum

26. p < 0.001

27. Diet problems • Excessive protein • Many diets contain 20-28 % protein • Maximum growth with 17% protein • Suitable protein content for breeding is on 17-20 % • In long-term studies excessive protein may lead to kidney failure

28. Rodent chows influence on nephrocalcinosis • Kidney calcification in female rats • Severity increases with higher dietary P levels • Variance in nephro-calcinosis 0/6 – 6/6 • Histological scores 0.0 – 1.3 Ritskes-Hoitinga et al. 1991

29. Diet problems • Pesticide & heavy metal residues • Most common cause for rejection of a batch (GLP) • Ref: Food and Drug Administration (FDA), 1978, Nonclinical Laboratory Studies. Good Laboratory Practice Regulations. Fed Regis, Part 2, 59986-60025.

30. Why animal breeders use high energy diets? • Scientists buy animals by weight • Ad libitum feeding with high energy diet ->rats with certain weight fast • No change visible

31. References • Quick reference guide on nutrition, www.felasa.eu, 2001 • Beynen, Principles of LAS, 2003 • Chwalibog & Ritskes, Handbook of LAS, 2004 • Ritskes and Strubbe, The welfare of lab. animals, 2004 • Ritskes, The laboratory mouse, 2004 • Dietary Restriction, ILSI Press, 1995

32. Pop quiz • Study on effects of alcohol on longevity • Rats drink 10 % ethanol in water • What is being studied: • Effect of ethanol? • Effect of decreased eating? • Effect of decreased drinking? • All of the above?