Preferences for nesting material as environmental enrichment for laboratory mice

Preferences for nesting material asenvironmental enrichment for laboratory mice

H. A. Van de Weerd1, P. L. P.Van Loo1, L. F. M. Van Zutphen1,J. M. Koolhaas2 & V. Baumans''Department of laboratory Animal Science, Utrecht University,The Netherlands and 2Department ofAnimal Physiology, University of Groningen, Haren, The Netherlands .

SummaryBehavioural and psychological needs of laboratory animals generally cannot adequately bemet in standard laboratory cages. Environmental enrichment, which provides a morestructured environment can enhance the well-being of laboratory animals. They may performmore of their species-specific behaviour and may control their environment in a better way.An easily applicable form of enrichment for laboratory mice is nesting material. Six differenttypes of nesting materials were evaluated in a preference test with male and female animals oftwo strains (C57BL/6Tor BALB/c, n=48). No significant differences in preference were foundbetween the strains or between the sexes.

All mice showed a clear preference for cages with tissues or towels as compared to paperstrips or no nesting material, and for cages with cotton string or wood-wool as compared towood shavings or no nesting material. Paper-derived materials were preferred over wood-derived materials, although the results also suggest that the nature (paper or wood) of thenesting material is less important than its structure, which determines the nestability of thematerial. Nesting material may be a relatively simple method to contribute to the well-beingof laboratory mice.

Keywords Preference tests; environmental enrichment; nesting material; nestbuilding; mice

Standard laboratory cages are designed tofulfil the most essential needs in a laboratoryanimal's life, such as provision of food, waterand a substrate, e.g. bedding or a grid floor, toavoid contact with their excreta. Animalshowever, also have behavioural and psycho-logical needs, most of which cannot be metin these cages. The animals are able toperform only a part of their complete species-specific behavioural repertoire (Van de Weerdet al. 1994), which can result in abnormalbehaviour such as stereotypies (Wemelsfelder1990). Furthermore, animals maintained in

Correspondence to: H. A. Van de Weerd, Department ofLaboratory Animal Science, PO Box 80166, 3508 TDUtrecht, The Netherlands. Tel: ++31-30-2532033, Fax: ++31-30-2537997, Email: [email protected]

Accepted 12 December 1996

unresponsive environments and highly un-natural groupings are less adequate modelsfor extrapolating experimental results tohumans (Markowitz & Gavazzi 1995),Environmental enrichment may enhance thewell-being of laboratory animals by providinga more structured environment whichenables them to perform more of theirspecies-specific behaviour and which givesthem more control over their environment(Scharmann 1991, Chamove 1989). As aconsequence, animals from enriched envir-onments may be more physiologically andpsychologically stable and better representa-tives of the species and thus ensure betterdata collection and scientific results (Benn1995, Scharmann 1991, Markowitz &Gavazzi 1995). The introduced enrichment

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should be interesting for the animals bymeeting their behavioural requirements but,from the human point of view, it should beeasy to provide, remove and clean (Van deWeerd & Baumans 1995).

Nesting material is an easily applicableform of enrichment for laboratory mice. Bothmales and females will build a nest whenoffered nesting materials (Lisk et a1. 1969,Lee 1972, 1973). Females build nests duringpregnancy to bear and raise their young (Lisket a1. 1969, Broida & Svare 1982). Hormonessuch as progesterone play an important rolein this maternal nestbuilding (Lisk et a1.1969). Between strains both qualitative andquantitative differences in maternal nest-building exist (Broida & Svare 19821.Inlaboratory mice, the use of nesting materialreduces the preweaning mortality of pups andenhances the number of litters (Porter &Lane-Petter 1965). Similar results have beenreported for rats (Nolen & Alexander 19661.Norris and Adams (19761found that the typeof nesting material used markedly affects thepreweaning survival rates in the rat.

Nesting material is also used, both byfemales and males, as a source of protectione.g. against extreme environmental tempera-tures (Lisk et a1. 1969). Behavioural adapta-tions, such as nestbuilding, must be used fortemperature regulation, when the physiolo-gical systems alone are inadequate to main-tain body temperature (Lynch & Hegmann19721.Lee and Wong (1970) showed that theamount of cotton used for nestbuildingincreased with decreasing temperatures,although significant differences were foundbetween strains of mice. Nests also offer anopportunity to hide from predators and in thelaboratory to avoid aggressive conspecifics, orto provide a shelter from overexposure tolight.

When applying enrichment, it is necessaryto evaluate the suitability of the enrichmentprogramme, as various species or strains mayrespond differently to the methods ofenrichment (Beaver 1989). Preference testscan be used to determine some generalprinciples about species relevant propertiesof enrichment devices (Mench 1994). Choicetests have been used to assess the relativepreference for or avoidance of several housing

Van de Weerd et a/.

conditions in laboratory animals (Blom et a1.1992). Blom et a1. (1996) and Mulder (19751offered mice different types of beddingmaterial. Ottoni and Ades (1991) allowedhamsters to choose between nestboxes inrelation to food and nesting material andgerbils were offered a choice between par-tially darkened or transparent cages (Van denBroek et a1. 1995). Pregnant mice and rats,chose wood-derived bedding over beddingswith other origins (e.g. clay, com cob) tobuild nests before parturition (Mulder 1974,1975). In Blom's study (1996), mice showed apreference for shredded filter paper in com-parison with smaller particled bedding ma-terial.

Nesting material has been studied mainlyin relation with pregnancy or cold exposure.The aim of the present study was toinvestigate whether the nesting materialstested may serve as enrichment and to detectpossible differences in preference for thesematerials between or within strains of mice.For this purpose male and non-pregnantfemale mice of two inbred strains were testedin a preference test and their choices fordifferent types of nesting material (wood orpaper derived) added to otherwise standardenvironmental conditions were evaluated.

Animals, materials and methods

AnimalsFemale and male mice of two strains (C57BL/6JIcoU and BALB/cAnCrRyCpbRivU, n=48)were used. They were bred and raised with-out any nesting material. At the start of theexperiment they were 8-10 weeks of age. Theexperiment was conducted in two cohorts,the first experiment (male mice) lasted sevenweeks, the second (female mice I lasted fourweeks. The animals were housed (per strainand sex) in groups of six animals in a housingsystem, consisting of two Macrolon type IIcages (375 mm2, UNO Roestvaststaal, Zeve-naar, The Netherlands), connected with apassage tube, similar to the tubes used in thepreference test system. Both cages weresupplied with food pellets ad libitum (RMH-B, Hope Farms, Woerden, The Netherlands),tap water ad libitum and sawdust bedding



Preferences for nesting material

(Lignocel 3/4, Rettenmaier and Sohne, Ell-wangen Holzmuhle, Germany). The animalswere kept in conventional rooms withcontrolled photo period (12:12 L:D, lights onat 07:00 h, approx. 200 lux at 1 m above thefloor), temperature (20-22 0c), relative hu-midity (50-60%) and ventilation (15 airchanges per hour). Environmental conditionsin the experimental rooms were similar,except for the light intensity which wasapproximately 300 lux at 1 m above the floor,in order to approach light intensities instandard animal rooms.

Preference test systemThe preference test system used in this studyhas been validated and described in detail byBlom et a1. (1992). In short, a multiplehousing system was used consisting of eithertwo or four test cages (Macrolon type II)connected by non-transparent tubes (PVC,inner dimensions: 2.6 x 2.6 x 25 cml to acentral cage (15x 15x 18 cm, transparentperspex). When testing with a two-cagesystem the central cage was divided diagon-ally by a PVC sheet (19x 17 cm). A total ofsix multiple housing systems were useddivided over two four-tiered constructions intwo similar experimental rooms. Each con-struction was turned gently during testing toprevent bias due to external influences in theexperimental room which could interferewith the choice behaviour of the mouse.

The test cages were supplied with 50 g ofsawdust bedding (LignoceI3/41, a food hopperwith equal amounts of food pellets (100 g,RMH-B)and tap water in bottles. The centralcage had no food, water or bedding. Themovements of the mice between the testcages were detected automatically by meansof photo-electrical devices in the passagetubes. The signals were sent to a computerwhich calculated dwelling times per cage(software: Gate-Watch, Metris System En-gineering, Leiden, The Netherlands).

Behavioural observationsOne.of the six multiple housing systems wasequipped with a video camera system. Eachtest cage, including the central one, wasprovided with a video camera (Panasonic

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WV-151O).The cameras were connected withthe photo-electrical devices, so that move-ments of the mouse could be followed in thetest system. The signals from the videocameras were sent to a time-lapse videorecorder (Panasonic AG-6700) which couldrecord 24 h of testing (recording: 1/9 ofnormal speed). During the night the experi-mental room with the video equipment hadred lights (approx. 5 lux at 1 m) to enablevideo recordings.

Procedure; nesting materialsMice were introduced into the test systembetween 15:00 and 17:00 h and tested in-dividually during 48 h. A group of six mice(of one sex and one strain) were testedsimultaneously. The behaviour of one animalwas recorded for 12 h during day time(second day of the test) and for 12 h duringnight time (second night of the test). Foodand water of each test cage were weighedbefore and after the experiment.

Three test series were performed to test sixdifferent types of nesting material. In each ofthe first two series (series 1: paper, series 2:wood), three nesting materials were com-pared (the fourth cage in each series con-tained sawdust bedding without nestingmaterial). In the third series, per strain andsex the nesting materials which most ani-mals had chosen in the first two series werecompared in a two-cage system. Figure 1shows the nesting materials and Table 1describes the materials and gives theamounts provided per series (approximatelyequal volumes).

Statistical analysisThe dwelling data were analysed by distin-guishing three time frames: the total ofdwelling times during the 48 h of theexperiment, the dwelling times during 12 hof day light (second day of the testl and thedwelling times of 12 h of night time (secondnight of the test). These two latter periodssynchronized the periods of collected beha-vioural data (video tape recordings).

The method of statistical analysis used hasbeen described by Blom et a1. (1995).Briefly,per test series the dwelling time data (in



136 Van de Weerd et al.

dwelling times. Statistical significance waspreset at P<O.OS.

Overall significant differences betweenchoice cages in dwelling times and amount offood and water consumed were furtheranalysed using a paired t-test to indicatewhich of the cages were preferred or avoided.As multiple comparisons were made, thelevel of statistical significance was preset atP<0.0083 (Bonferroni's adaptation).

Material Trade name Appearance (size) Amount

Table 1 Nesting materials

lCeltona, Cuijk,The Netherlands, 2Kimberly-Clark Corpora-tion)', EEC,3BMI, Helmond,The Netherlands, 4VNK, Haar-lem,The Netherlands

Fig1 The six different nesting materials tested inthe preference test. Top row (paper series): papertowel (left), tissues (middle), folded paper strips(right). Bottom row (wood series): cotton string(left), wood-wool (middle), wood shavings (right)

Behavioural dataThe behavioural data on video tape wereviewed and analysed using a behaviouralobservation software package (The Observerv 2.0, Noldus Info Tech B.V., The Nether-lands). The tapes were viewed at normalspeed, thus behaviour was seen nine timesfaster than the original behaviour. Every 5 sthe behaviour was scored, which correspondsto one sample every 45 s in reality. Thefollowing ethogram was used to classify thebehaviour (based upon Blom et a1. 1992):

Sleeping (sl)=movements are absent while the animalis in a sitting or lying position. Very shortor minor movements during a longresting period (e.g. turning) are not con-sidered as an interruption.

Manipulation (man)=manipulation of the nesting material,includes shredding, fraying, dragging andnestbuilding behaviour.

Grooming in the nest (gr-il=while sitting or standing in its nest, themouse is shaking, scratching, wiping orlicking its fur, snout, ears, tail or genitals.

Grooming outside the nest (gr-ol=same as gr-i, but outside the nest.

Ingestive behaviour (ing)=includes eating and drinking behaviour.Eating: gnawing on food particles fromthe food hopper or from the sawdust,coprophagy is included as well. Drinking:licking the nipple of the drinking bottle.

Locomotion (loco)=. all other movements (e.g. walking, run-ning, jumping).

Climbing (eli)=climbing on or hanging from the bars ofthe wire cage lid or food hopper, or

5 g

1piece2 pieces

5 g5 g

5 9

Folded strips(11 x 0.3 cm)Cotton string

(variable)Wood-wool3 Strips (variable)Gold Wood shavings

shavings3 (variable)

Paper towel' Sheet (25 x 31 cm)KleenexR Sheet (20 x 21 cm)

tissues2

Enviro-dri3

Sharpy4Wood

Paper

seconds) were logarithmically transformed asthey were not always normally distributed,and to increase the independency of the data.For the same reason, central cage dwellingtimes were not included in the analysis. Dataon food and water intake were not trans-formed, because they were normally distrib-uted.

The data were analysed using multivariaterepeated measures analysis (Wilk's lambda)to evaluate the influence of type of nestingmaterial and interactions on choice beha-viour and to detect possible differencesbetween the strains, or sexes of a strain inchoice behaviour. Food and water intakewere analysed in a similar way as the




standing on the passage tube or drinkingnipple. While climbing or hanging thehind legs or tail may touch the cage walls.

Rearing (rear)=standing position with the forepaws nottouching the cage floor. The animal isstanding on its hind feet or toes, usuallysupporting itself with the tail. Thefore-paws may lean against the passagetube, cage wall or food hopper.

Digging (dig)=bedding material is pushed forwards orbackwards with nose, fore paws or hindlegs. Mouse moves around or is sitting inone place.

Descriptive statistics were used to analysethe behavioural data, because only twoanimals from each sex and strain group(n=12) were observed per test series. Theresults were used to describe the behaviour ofthe mice in the different test cages during atest series. The distribution of behaviour ineach test cage was analysed for the night andday time period separately.

Results

Figure 2 illustrates the mean relative dwell-ing times per cage for the paper series.Figures 3 and 4 show the same for the woodseries and the paper vs wood series, respec-tively. Figure 5 gives two examples of thedistribution of behaviour for the four cages inthe paper and wood series.

Cage choiceNo significant differences in cage preferences(dwelling times) were found between strainsin the paper series and the wood series,except for the total data of the paper series(MANOVA, P<O.OS). The cause for thisdifference is not very clear. Also, no sig-nificant differences between the sexes of astrain were found, in all three test series andduring all three time periods. Therefore, perstrain both sexes were analysed together inthe paired t-test analysis. Mice of both strainschose significantly for a cage during all threetime periods of both the paper and woodseries (MANOVA, for all: P<O.OOl).Figure 2illustrates that in the paper series the cages

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with the tissues and paper towels werepreferred, whereas from Fig. 3 it can beconcluded that in the wood series the cageswith the cotton string and wood-wool werepreferred. In the paper series the biggestcontrasts between cages were found to existbetween the cages with tissues and towel onthe one hand and the cage with paper stripsand the cage without nesting material on theother. This was most clear for mice of theBALB/cstrain (paired t-test, all P <0.001). Forthe CS7BLmice it was less obvious. Thedifferences between the cages with eithertissues or towel and the cage without nestingmaterial were significant (paired t-test, allP < 0.01), just as the contrast between thecages with the towel and paper strips (pairedt-test, all P<O.OOSI.

In the wood series mice of both strainsspent significantly more time in the cageswith cotton string and wood-wool in com-parison to the cage with wood shavings andthe cage without nesting material (bothstrains: paired t-test, all P< 0.0051.Onlyduring night periods the contrast between thecage with the wood-wool and the cages withwood shavings or without nesting materialwere not significant for the CS7BL.For theCS7BLstrain significant differences betweenthe cages with the cotton string and wood-wool were found (paired t-test, all P<O.OOSI.

In the paper vs wood series (Fig.4) thematerials which most mice had chosen in thepaper and in the wood series were compared,being the paper towels vs cotton string for theCS7BLmales, and the tissues vs wood-woolfor the BALB/cmales. Females of both strainspreferred the tissues and cotton string. Datawere analysed per strain. The CS7BLmicechose significantly for the cage with thepaper nesting material (males: towels, fe-males: tissues), but only during the total andday period (MANOVA, both: P<O.OOSI.BALB/cmice did not make significant cagechoices during all three time periods of thissenes.

Food and water intake

Significant differences between the strains infood and water intake were food in the woodseries only (MANOVA, both: P<O.OS).



138 Van de Weerd et a/.

NESTING MATERIAL: PAPER SERIESMale C57BL Mice Male BALB/c Mice

~ 80"W

~ 70"

'"" 80"~c 60"~Q; 40%~"tl

30%".::•• 20"~c: 10"••" 0%::;;

TISSUES TOWEL STRIPS CENTRAL

i' SO~W~ 70%

'"~ 60%.;:;Q) 50%

:§Q) 40%~"tl

cD 30%>

'; 20%

~c: 100/.a!

~ 0%Tissues TOWEL STRIPS CENTRAL

Female C57BL Mice Female BALBlc Mice

TISSUES TOWEL STRIPS

~ 80%W

~ 70%

'"" 80%.§c 60%.!:a; 40%~"tl

30%

".::•• 20%~c: 10%a!

":2 0%TISSUES TOWEL STRIPS

c::::J DAY _ NIGHT

CENTRAL

~ TOTAL

i' 80%W~. 70%

'"~ 80%

C'l 50%

~Qj 40%~"tl(J) 30%

.::i 20%

~c: 10%al

"::< 0%

c::::J DAY _ NIGHT

CENTAAL

~ TOTAL

Fig 2 Results of the preference test with three types of paper-derived nesting material. Mean relativedwelling times (and SEM) per cage for day (=12 h), night (=12 h) and total (=48 h) period, for mice of twostrains (n=48)

BALB/c mice consumed overall more foodthan the CS7BL mice. In the paper and woodseries no significant differences between thesexes of the strains were found in cage choicefor food or water intake.

In the paper series mice of both strainsmade significant cage choices for waterconsumption, but not for food consumption.CS7BL mice drank most in the cages withtissues and no nesting material (MANOVA,P<O.OOl); BALB/c mice in the cages withtissues and towels lMANOVA, P<O.OOSj.Significant contrasts were found between thecage with paper strips and the cage withoutnesting material (CS7BL: paired t-test,p<O.On and the cages with paper strips andtissues (paired t-test, CS7BL: P<O.Ol; BALB/

c: P <0.00 11.In the wood series significantcage choices for food intake were made by theBALB/c mice; they ate most in the cages withwood shavings and no nesting material(MANOVA, P<O.OOSj. The main contrastswere found between the cages with woodshavings and either wood-wool or cottonstring (BALB/c: paired t-test, P<O.Ol).Significant cage choices for water consump-tion were made by the CS7BL mice, theydrank most in the cages with cotton stringand no nesting material (MANOVA, P<O.OS).

In the paper vs wood series, food and waterintake of the BALB/c mice did not differsignificantly between the cages. The C57BLmice did not make a significant cage choicefor water consumption, but the male mice




ate significantly more in the cage with thepaper towel than in the other cage (MANO-VA, P<O.OS), whereas the females did notmake a significant cage choice.

Behavioural dataA striking behaviour performed by approxi-mately half the number of animals, is thecombining of nesting materials by draggingmaterial from one cage to another. In thepaper series the animals that dragged,alwaysmade a combination of tissues and towel. Inthe wood series the cotton string wascombined with wood-wool and sometimes afew wood shavings were added. In general,the preferred materials were combined.

Figure 5 illustrates the distribution of

139

behaviour during daytime and during thenight for the BALB/cmice (paper series) andthe CS7BLmice (wood series). During day-time the mice mostly slept in their preferredcage, where they also showed manipulationof the nesting material and grooming in thenest. During the night the mice were moreactive and they performed active behaviours(ingestive behaviour, locomotion, climbing,rearing, diggingl in all cages of the testsystem, although a fair amount of sleepingwas performed in the preferred cage.

DiscussionThe results indicate that mice preferred cageswith nesting material and that they discri-minate between different nesting materials

NESTING MATERIAL: WOOD SERIESMale C57BL Mice Male BALB/c Mice

i 90%

W

~ 80%..70%Q)

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Q) 30%.2:;;

20%~c: 10%

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STRING WOOL SHAVINGS CENTRAL

i" 90%

W

~ 80%..70%Q)

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60%Q)~ 40%"tl

Q) 30%>';20%~

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STRING WOOL SHAVINGS CENTRAL

Female C57BL Mice Female BALB/c Mice

STRING WOOL SHAVINGS

i 90%

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~ 80%..70%Q)

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~ TOTAL

i" 90%W~ 80%

~ 70%E

'Z 60%Clc:

60%a;;; 40%"tl

~ 30%

:; 20%

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Q)

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Iz:22l TOTAL

Fig 3 Results of the preference test with three types of wood-derived nesting material. Mean relativedwelling times (and SEM) per cage for day (=12 h), night (=12 h) and total (=48 h) period, for mice of twostrains (n=48)



140 Van deWeerd et at.

NESTING MATERIAL: PAPER vs WOODMale Mice Female Mice

i 90% ~ 90%

W C57BL BALBlc w C57BL BALB/c

f!! 80% f!!- 80%

'" '" 70%" 70% "E 60% E 60%0 '"~ 50% :§ 50%

~ ~~ 40% ~ 40%"0 "0

" 30% " 30%.2: .2:iii iii 20%~ 20% ~c: 10% c: 10%co coQ) Q)

0%::E 0% ::ETOWEL STRING CENTRAL TISSUE~, WOOL CENTRAL TISSUES STRING CENTRAL TISSUES STRING CENTRAL

c::J DAY _ NIGHT ~ TOTAL c::::J OAY _ NIGHT ~ TOTAL

Fig 4 Results of the preference test with the two most preferred nesting materials from the paper andwood series (as indicated). Mean relative dwelling times (and SEM) per cage for day (=12 h), night (=12 h)and total (=48 h) period, for micE! of two strains (n=48)

BEHAVIOUR

65%J45% I

::;; 25% I [UJ 15%~c:.2E::J 10%

"0

">'E~ 5%

c:.•Ql::;

0%

Paper Series: BALB(c Mice

DAY 1 65% ~_ 45%

~ 25%

~ 15%

c:.2'§

10%:l"0

Ql.:::iii~ 5%

c:.•":2

0%

Wood Series: C57BL Mice

DAY 1

sl man gr-I gr-o ing loco eli rear dig sl man gr-i gr-o ing loco eli rear dig

65%}i 45%W 25% -~ 15%

c:o

~:= 10%"0

".:::iii~ 5%c:co"::;;

0%

NIGHT 1 65% E~:~~~n~ 15%

c:.2~::I 10%"0

Ql>~~ 5%

c:coQ)

::E0%

NIGHT 1

sl man gr-i gr-o ing loco eli rear dig

c=J Ti•• u _ Towel ~ Slrip. ~~ - c:=J Centr

sl man gr-; gr-o Ing loco eli rear dig

c:J String _ Wool ~ Shavi !!!II - E::J Cenlr

Fig 5 Results of behavioural observations from preference tests for nesting material with mice of twostrains (n=8). Mean relative time (and SEM) spent on indicated behaviour during day (=12 h) and night(=12 h) period (seeAnimal.s, Materials and Methods for explanation of abbreviations)




and make consistent choices when sub-mitted to a preference test. Cages withtissues or towels are preferred over cageswith paper strips, whereas cages with cottonstring or wood-wool are preferred over woodshavings as nesting material. C57BLmicepreferred paper-derived nesting materialsover wood-derived materials. BALB/cmicedid not make a significant choice betweenthe two materials offered, but in most casescombined them. In the third series threemale BALB/cspent most time in the centralcage, where they combined all nestingmaterials. An explanation for this behaviourmight be that when testing with only twocages, a piece of PVC is dividing the centralcage in two parts, making this area smallerand relatively dark.

The fact that some animals combinenesting material might suggest that there isnot a clear preference for the nature ofnesting material (e.g.paper or wood) but thatother features of the nesting material such asthe structure (e.g.shredded or as a sheet), alsoplaya role. In a choice test with mice,Mulder (1975)found a significant preferencefor bedding materials from a wood origin(aspen and cedar), but he did not test paperproducts. Both cellulose wadding andshredded paper as well as wood chips yieldedlow to normal preweaning mortality inmouse litters in the study by Porter andLane-Petter (1965).In the study by Blom eta1. (1996),both C57BLand BALB/cmiceshowed a preference for shredded paper bed-ding instead of sawdust or wood chips.Behavioural observations indicated that ma-nipulation of the bedding and resting in nestswere performed mostly on this type ofbedding. In the study by Nolen and Alex-ander (1996)best breeding results were foundwhen providing the rats with shredded paperas nesting material, whereas in the study byNorris and Adams (1976)better breedingresults were obtained with wood-wool in-stead of paper tissues. These results alsosuggest that the nature of the nestingmaterial might be less important than thestructure (e.g. shredded or as a sheet). Thestructure may be important because itdetermines the nestability of the material. Inthe present study, the characteristic feature

141

which the preferred nesting materials have incommon is that the mice can manipulatethem to build nests, and by doing this, theyare able to structure their environment.Towel and tissues were shredded to buildnests and the wood-wool and cotton stringwere shaped into the desired form. Whennesting material is put into the home cages ofmice they start building nests within min-utes after introduction (Watson 1993,Schneider & Chenoweth 1970).With cagecleaning, these nests can be transportedcompletely to the clean cage. In the presentstudy several animals, especially of theBALB/cstrain also combined the two pre-ferred nesting materials to make morecomplicated nests. Pennycuik (1973)alsoobserved that mice moved nesting material(wood-wooll to the nestbox selected asnestsite. The behavioural observations in thepresent study showed that 10-20% of thetime budget was spent on manipulation ofthe nesting material during day or night.

Another aspect of the nesting materialwhich could be an important criterion forselection by the mice is the degree of lightabsorption. Mice are nocturnal animals whooften prefer to hide and sleep at dark placesduring daytime. Exposure to light can causedamage to the eyes (Clough 1987).However,most of the nesting materials preferred in thepresent study allowed some penetration oflight [e.g.the tissues and cotton string). Onlythe paper towels could provide a shelter forlight, but only if the mice were completelycovered by the materials of their nest, whichwas mostly not the case.

In general, there were no differences incage choice between the day and nightperiods. During darkness the mice spentmost of their time in the same cage as duringlight periods, which was illustrated by thebehavioural observations. However, duringthe night period the mice also visited theother cages of the test system. In general, theresults showed that the mice visited all thecages of the housing system with approxi-mately the same frequency, however thepreference is based on the duration of theirstay. Since sleeping requires most time, thecage selected for resting is the preferred cageby definition IBlom et a1. 1992).Figure 5



142

illustrates the increased activity of the miceduring the night. In contrast with the presentresults, experiments with rats showed thatdifferent cages are preferred during day andnight, suggesting that various behaviouralactivities require different cage floor covering(Van de Weerd et a1. 1996).

The study by Blom et a1. (1992) showedthat from the recorded behaviour, 65% of thetime is spent on sleeping, grooming anddigging behaviour. In the present study, thesame amount of time is spent on sleeping,grooming in the nest and manipulation of thenesting material. In the study by Blom et a1.mice did not have nesting material, so itseemed that they performed digging as a kindof redirected behaviour for nestbuildingactivities. In the present study nestbuildingbehaviour could be performed with thenesting material and digging was less fre-quently observed.

Mice of both strains consumed equalamounts of food in all four cages of the paperseries. However, most water was consumedin the cage provided with tissues. This wasalso the preferred cage for all mice, exceptmale C57BL. In the wood series the BALB/cmice ate most in the cage without nestingmaterial (males) and the cage with the woodshavings (females) and the C57BL mice drankmostly in the cage with the cotton string. Inthe third series only male CS 7BL mice had apreference for the cage with the paper towelto consume food. These results are not veryconsistent and are only partly in concordancewith the results of Blom et 01. (1996), whofound that eating and drinking behaviour ofthe mice was similar for the four test cageswith different types of floor covering. Ham-sters, on the contrary, consumed most foodclose to their nests, because they preferrednestboxes nearest to the food source (Ottoni&. Ades 1991). Food intake differed signifi-cantly between the strains, with BALB/cmice eating more than C57BL mice. This canbe explained by a difference in weight; BALB/c mice are in general heavier than C57BLmice. Water intake did not differ between thestrains. Although no differences between thestrains were observed regarding the choice ofnesting material, there was strain-specificbehaviour towards the nesting material.

Van de Weerd et a/.

Fig 6 Dome-shaped nest made of shreddedtissues by a male BALB/cmouse

Tissues and towels were shredded morethoroughly by BALB/c mice than by C57BLmice. Also the shape of the nests differed.The BALB/c mice build dome-shaped nests(Fig. 6), whereas the C57BL mice made bowl-shaped nests. This has also been described byLee and Wong (1970). Other studies have alsoreported differences in the shapes of sleepingnests between mice of different strains (Lee1972, 1973, Lynch &. Hegmann 1972, 1973).These differences in nestbuilding betweenstrains are probably genetically determined(Lynch &. Hegmann 1972).

The results of this study show thatlaboratory mice prefer nesting materialwhich they can use for nestbuilding. Byproviding them with nesting material theanimals are able to use an active strategy tomanipulate and control more aspects of theirenvironment, which is important for theeffectiveness of enrichment (Mench 1994,Sluyter et a1. 1995). Nesting material is easilyapplicable in standard cages and thus may bea relatively simple method to contribute tothe well-being of laboratory mice.

Acknowledgments The authors are grateful to BMIIHelmond) and Broekman Instituut (Someren) forsupplying the nesting materials.

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Preferences for nesting material as environmental enrichment for laboratory mice

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