NOV 2 62003 · n:r.li8pOl1 is requ;red by law (7 USC 2143). Failul'8 to rwport aca:ni'ing to the ~Ialions e:atl s.o a_farm for

... result In In order to cease and deSist and to be subject 10 penalties al provtded for in Seelon 21! additional infometion.

UNITED STATES OEPARlMENT OF AGRICULTURE I. CERTIFICATE NUMBER: 14-F-0009 ANIMAL AND PlANT HEALT1i INSPECTION SERVICE

CUSTOMER NUMBER: 463

U.S.D.A.lHuman Nutrition Res. etr. At Tufts U ANNUAL REPORT OF RESEARCH FACILITY 711 Washington Street

( TYPE OR PRINT) Boston, MA 02111

Telephone: (617) ·556·3200

J. REPORTING FACIUTY (usr aJlloc:ation. where animlls were houaed or used in adual research. Iestlng, or experimenta1ion. or held for these purposes. AtIad1 addItional Sheets if ncessary)

FACILITY LOCAnONS ( so.. ) • See A1achod Uoting

I REPORT OF ANIMALS USED BY OR UNDER CONTROL OF RESEARCH FACILITY rAllach addlllonal.h_1f_IV or .... APHIS Form 7023A )

A.

An.Imals Covered By1lle_,

wellaro Regulotlona

B. NUl'T'Cer of C. Nurrbetof lnimall being anImalsupan btecl, condlUoned, whId1 teaching, or held tor use in rooearch. teaching. testing, el(pellments, or_monts. ,....~

research, Dr condUded surgery but not YE invol"ling no pain, usecl for such distress. Druse 01 purposes. palrwellBVing

drugs.

D. NulTlOer of anlmels upon MtId1 expet1mentl. tAchlng• .-r<:h. surgery. or tes.ts were conducted IIWDI"Iing 8ICCXlI1l)Bnying pain or diau.. to the animal. an forwhld\approprillle anesthetic. anIIlgestc, or trv.nqulllZlng dnJQI were usad.

E. Nu..-ofanl..... upon ...cIl ...chlng, _"me.... ,I F. l'WI!!IlIrch, surgery or tests were c:onducUId involving a_nying palnordlllnss.toltlotmimaisandlorlMl

l

TOTALNUMllER

=-":'~=":...t~~h' . OFANIt.\AlS results, or intlll'p'8tatiOn of the teaching. .-earth,

(COLUMNS_menls, surgery. or _. (An expIonatlon of .... pn:x:edures prodUdng pain 01" dtslress in Ihese anlrnala a C+D+E)the Il!8IQI"IS lIUch drugs were nor used I'TLIII: be ertaehed t11Is.-pcl1 }.

4. Dogs

5. Cala

8. Guinea Pig.

7••Ha_ 37 492 529

8. R8bblta 2 2

O.NoII-human PrlrnafBs

10. Sheep

11. Pigs

12. Olhar Fa"" Animals

13. OlborAnimals

Mice 3,363 3,199 1,479 279 4,957 Rats 662 183 398 581

Ferrets 70 121 191

I· ASSURANCE STATEMENTS I11 _lyOCClll"a1>Ie__......... _ ... ct.lrtng.ondlal-.;ocIuIIond ....Oi/.,_.IncIudIno~_of--'Mlc,_IIld_Ilzing_._

-.In;. 1lIotIng, IIUIVO'Y. or_'"willica.-..101_ bytlis _1ocI11ty. . ­2)' Eech principol_goIOrhu _ ol_topoinfUl~

3) Thllllocliity 10_10.... _ ond~"- it..M, ond Olla _rwl_-....to.._ ond_boopodlled ond ~ by .... prIrdpoI-.-..ond~ ~ AnilMJ Care."d U.. CammIa.. (IACUC). A IUmmIIY of alll1lGh uClPdOnl" II atlilchecl to lNa_nrual npOrt. In 8ddiIIan to IcIentifring 1M ~~ this UII'MJ)' WIt bri.....,.-of....-....••W11I1 ..... opecIoo ond_ar__

"l ""_lng_onf"'!"IlI__ __of_-...ry,,"",ondto_"'_of__af_,,"",ond_ityllu~ouVaiIy"'_"'

CERTIFICATION BY HEADQUARTERS RESEARCH FACILITY OFFICIAL ( Chief Executlva OIIICII" or Legally Reoponlllble InatIIu1IClI1lIf 0ftIc:iaI )

N------- -- --------- ----- --------- ----- ----------------------- -------------- ----- ---------- ------ ESIGNED --------- -- ------------- ------------ ----------- --- 10':'0

3 (OCT 88). wIIlch is _otl.)

I

· .. ,. Jean Mayer f

United States Department of Agriculture Human Nutrition Research Center on Aging . iI

At Tufts University ,/

I~O -l7

Nutritional Immunology Laboratory October 15, 2001

To: ---- -------- ---------- -------- --------- ------ ----- ----- Committee, HNRCA

From: ------- ------------ -------- ------ i7 .f\1'rN'-­Re: Category E animals in Amendment to Protocol MS-31

The major limiting factor in conducting our study is the large number of animals needed to collect sufficient number ofmacrophages for our experiments. This inherent difficulty can be overcome by intraperitoneal injection of thioglycollate (TG) which elicits recruitment ofmacrophages to peritoneal cavity. TG is a widely used stimulatory agent which induces non-infectious acute peritoneal inflammation in mice and rats. Administration ofTG has been shown to increase the total number ofmacrophages up to four-fold, which will reduce the· number of animals necessary for addressing our specific aims.

A number of recent studies have successfully demonstrated that TG-elicited macrophages can be used in the study of some gene expression and signal transduction. However, the feasibility ofasing TG-elicited macrophages to study COX-2 gene expression is not mown.

To test this, we need to inject TG intraperitoneally to mice three days before they are euthanized by CO2 asphyxiation for macrophage collection. Peritoneal injection will cause discomfort and moderate pain in mice, which unfortunately can not be alleviated. Thus we have classified the animals under category E.

711 Washington Street Boston, Massachusetts 02111 FAX: (617) 556-3344

....

To: Animal Care and Use Committee

From: --------------- ------- PI of WA-} Protocol

RE: Justification of Category E in WA-} Protocol: Effects of Combined Chemopreventive Agents (9-cis retinoic acid, celecoxib, and 1,25(OH)z vitamin DJ ) Against NNK-induced Lung Carcinogenesis in AJ Mice

Protocol WA-l will include USDA Category E research in which some experimental animal groups will experience pain and/or distress without alleviation. This letter will verify a lack of alternative methods and assure the committee that the proposed research does not . unnecessarily duplicate previous experiments.

We propose to conduct an in vivo intervention study to investigate the effectiveness of9­cis retinoic acid, 1,25(OH)~ vitamin D3, and a COX-2 inhibitor drug alone and in combination as anti-carcinogenic agents in the AI mouse model oflung cancer. Lung tumors in strain AlJ mice resemble human lung adenocarcinoma and have become the preferred test system to study this form of cancer. The target of chemoprevention is premalignant lung disease, making animal models essential for evaluating the efficacy of compounds and interactions in the suppression of tumor progression. Because symptoms rarely occur in the early stages of human lung cancer and many of these early cancers go undiagnosed, mice genetically predisposed to this form of cancer allow us to study lung cancer chemoprevention over the course of months and with fewer animals than similar studies with human subjects. The induction of lung tumors in AJ mice progresses through several distinct stages similar to the stages of human lung cancer. In both mice and humans, adenocarcinomas progress to adenomas and ultimately carcinomas. Further, tumor initiation by a tobacco-derived carcinogen. 4-(methylnitrosamino)-I-(3-pyridyl)-1­butanone (NNK), in AJ mice is characterized by premalignant legions containing a gene alteration that is also present in some human cancers. This makes the AJ mouse an ideal model in which to study lung cancer chemopreventative agents that may be of benetit to the human population. Although \ve cannot alleviate tumor formation in the N?\K-injected control group, the treatment group using combined chemopreventive agents should alleviate tumor formation/distress/animal pain.

While mechanistic hypotheses and data from cellular studies suggest that combinations of vitamins and anti-inflammatory drugs may be effective in lung cancer chemoprevention, there is a clear lack of in vim \vork in this area. This will be the first study to examine vitamin A and vitamin D interactions in an animal model of lung cancer and the tirst study to combine these vitamins with a COX-2 inhibitor to examine synergistic effects. If successful, this study could lead to new approaches in cancer chemoprevention, utilizing combinations of chemopreventive vitamins and drugs in smaller and less toxic doses, thereby avoiding the side effects commonly seen in early clinical trials testing single agents. This research cannot be done using cell models as results cannot be applied to in vivo tumorigenesis.

.,.'

TO: FROM:

The HNRC Animal Care and Use Committee ---------- ----------- ------ NEPS Laboratory

RE: Justification of Category E in protocol RO-t7, ''Roles of TNF and interleukin-l in stress-induced cachexia: Effects of age in transgenic mice"

Our protocol RO-17 addresses the question ofwhether the cytokines involved in cachexia are the same as sarcopenia (namely TNF, IL-I, and IL-6). This line of research pertains to the mission of the NEPS laboratory, ie, the understanding and alleviation of physiological or pathological processes leading to sarcopenia, wasting and cachexia.

In RO-17, turpentine will be delivered subcutaneously in one of the hind limbs of wild type and IGF-I transgenic mice. Unfortunately, turpentine injection, although not lethal, results in a sterile abscess that cause pain. This pain is comparable to that felt by humans with a thigh abscess. We anticipate the abscess to be maximal 16 days after injection, and to gradually shrink thereafter. Unfortunately, the pain will not be alleviated by pain killers, as these drugs may induce changes in the levels of muscle cytokines, one of the major endpoints of this study. Because sub-clinical inflammation is a recognized feature ofhuman aging, the proposed experiments are germane to the issue of age-related changes in protein catabolism, inflammation, and immune responses.

~----- ----- ­

NOV 2 6211D3~<ol~l?:.--i'{..

B Jean Mayer

~ ~ ::; "" United States Department ofAgriculturea ~ Human Nutrition Research Center on Aging~ ~ At Tufts University .o--i.yET\.~

November 10, 2003

Elizabeth Goldentyer, D.V.M. Regional Director - Animal Care APHIS, Eastern Regional Office 920 Main Campus Drive, Suite 200 Raleigh, NC 27606-5213 Reference: USDA Annual Report (Registration No.: 14-F-0009)

Dear Dr. Goldentyer:

The enclosed documents represent the U.S.D.A. Human Nutrition Research Center on Aging at Tufts University's (HNRCA) "Annual Report of Research Facilities" for the Federal fiscal year, October 1, 2002 through September 30, 2003. Aspects of this report that reqUire comment are:

1) Animals reported under Category E:

a) Mild non-infectious peritoneal inflammation was induced in sixty-one (61) mice by the intraperitoneal injection of thioglycollate to increase the total number of peritoneal macrophages available (which reduced the number of animals used) for peritoneal macrophage harvest. The letter of justification for category E research was submitted with the IACUC animal protocol and is attached.

b) Lung tumors were induced in one hundred one (101) mice to examine the combined synergistic effects of vitamin A, vitamin D and COX-2 inhibitors to evaluate their role in lung cancer chemoprevention. The letter of justification for category E research was submitted with the IACUC animal protocol and is attached.

c) Sarcopenia was induced in one hundred fifteen (115) mice by the subcutaneous injection of sterile turpentine into the hind limbs of the mice to evaluate if thecytokines involved in cachexia are the same as those of sarcopenia (namely TNF, IL-1 and IL-6) in an effort to understand and potentially alleviate the physiological or pathological processes leading to sarcopenia, wasting and cachexia. The letter of justification for category E research was submitted with the IACUC animal protocol and is attached.

---- ~#ve;1estions ------------- ----- ---- ort, please do not hesitate to contact me.

------ - --- --- ---------- -- --------- -------- --------------------------------- ---------- ------- ------------- ------ ------- --------------

---- --- ---------- -- -------------

711 Washi~it'ti:CSC?r~et Boston, Massachusetts 02111 FAX: (617) 556-3344

This r!pcu"t is required' Dy law (? USC 21 43) r~re to repOrt "ac:.::ordin; lome re.gulationsc:an result In an order to "UZSI and desist and 10 b~ ,ut;;ed.1o penaIl~ as pr'O'Yidecl for in Sedion 2150

UNIi:;:O .$TATa$ O:PA.E:;'iMENlO:E AGFtlCUL.'i'l.Jq£ . ANtMAl AND Pl..ANi" H:Al.."ir;: INS!=IECTION S.e;:rvJCS'

ANNUAL REPORT OF RESEARCH FACILITY (TYPE OR PRINT)

2. f'oe,l..OQUAfO"!'RS i\!S5J.RC)O\ F~OUjf (t-."3me and Addreac. as r.giet.,..d wl1h uSO. inc;ud. 2i;:t Cad.) " _

USDA,APHIS,WS,NWRC 4101 LAPORTE AVE. FT. COLLINS, CO 80521

f R,EPOF('T' OF ANIMALS USSO BY OA UNDER CO!"lTROL OF FtEsEAJ='CCH F....OU'i'Y (Allen addiliOnal sbMts It t\8:aS8ry OC'UM APHIS FORM. ~023A) ... B.Numbel"of

C"~'af O~ N\IfTIber of8ll"lim.....·upon IGo ............. ., 0";""''' upon ""'i.., '0­ IF.anifT'lllla beirte' an~upc-n -tidl ecperirN=ntS. e~tItS......eareh. surpcy. or1eSU were A ....I~ea...c:-e= b"'''­ 'Whic:::'t\.A.=k~. ,.~~.~~. ~ducled ~"9 ..ceornp.~g"'"" or distr­__

By:'rtl8Anima eoncDtic",.c;f.cr .......... nJlQt'Y. Of \eSl& w":"l 10 1ht: an,",_ls and la:"whieh 1he .....,. d .pprgprie~ "iO'i'Al..l'.IO

W.''''re RegulafiQnl: he~ for us-. in ~rneM&.or tor-dueled ift'wQMng .............dc. ~ or tranQUl1i2.ing dn.Jpa '""'OUki O!EAHI~

••=hinD. t.sting. tes:s ~... ac:::cmP*il"l)"mg p.in Ol" ~... • ~.t1~ ..... ~oeduru. resultz. or ~ntit. conduc..d 6istf"eS:S'lOthe ..nirnaI. ""erpretalign of 'D'lc'teaChing. raaearch" ,......rct\.0l"

""""'""" no _nd for wh~ .pj:lI~pr"te experimel"lta. surgery. or 1eIn&. fl',n ~NIIion d . ceo"'" c­autpery bUl: not pain. diatre.c. ClIo ancAhellc. .,..Ige.e. or :lheptOCzadl.Jlt'lN p~ p.in Qr't:£..,... in I~ D.;. E)

~tusPlclr'DIdt wa'cdpei";" ~mzil\gdlU~ -et'8 a""'.1s .nd rite N-.ons av=h d~...,.. /'lOt ~d ........ relie"'''O drug-. ..... rnuat be .a.~tt1.0 thi. nlpOl11 ,

4. Docs I I 5. Cats / I 6. Guinea ?i~s I I 7~ Hamsters I 8. Rabbits 24 34 1­ 0 I 0 34 9. Non-human Primates ·1 I ! 10. Shee;) I. ,,~ Pigs

12. Other Fann Animals

Goats 21 23 I 0 ~o. -23 13 Other A..,imals I Coyotes 28 77 I 16 35 I 128 Norwa'y Rats I 0 7 0 0 7

·Deer Mice I 18 24 I 0 - .0 I 24rMSUr=tANOe ~AT:"'$NTS

1) PrcfeSSlQNIIlyacceptable ctaI\CJ.TCS go'Yefnmp In. c.rlt. tr-..\lTtenl, and ute Dr an..".11 inelUc:&r-act appl'Ctp.........e of ~h.tic. an.lpe-.e. and .,..nquAi;cing d2"UQ'S. Df"ICI" 10. Chr.if"G and folio....1'Ig adUal roesurd\. leaching. lUting,. -"'V.')'. or ~fiol\-"- fon_d by 1h1. ,...••""" fa:iiity

~) Tnw. ~eIIt:Y ilk adherlOU 1CI11"Ia a:\Ilnd.rdI; and replMations: undtJ'\".lhaAet, .nd iI. has I"'eQrJred Ihlll: ~iont' 'to ......n.di.tds and "'~'60nl .b• .pedf'.c ..nd ~ine4by Ihe principal inveaC",a1Or and' .~C' by the. lrIafituliOnlll Animal e-re. and U.. Committee CJ.'I'oCUC). A al,l1Tlrn.ryof an aveh 4I:lIrcep'tiDft.. is .!i.ehed too 'hia .",,..'lMII ....p~_'"

addlUon ~ id_n1lfying tt'wI I.a.cuc..p~d~ tmc "","",,_ry indud_. bcri.r expl• .,..tign q the e~ns:••c \I04R •• the aP'i=ies: ."d number flanitTtala atteeted'.

"") The .uncfinp 'Wterinaro:..1""IfOr lhis r-eal'd1 f.clrJty .... appro;r..,. aV'hOn'Y 10 ~nt::ll'e thoe pr::lI'ori.~ of .dequ_. vt\e:ftnary care and 10 avet"lIlee lhe adeClU.ey of other ellP'l!'=t' or anir:"lal care and use..

'C:RTIFICATJON SY HEADQUART=RS RES=ARCH FACILI7Y OFFICIAL

(Chief Executive Officer or Legally Respo"sible Institutior>af Official) Ie.i'tify 1h.llh••boY. is 1M. tornel, .nd complete (7lJ.S.C. S.cfion 2143~

SIQ'NA,T"Jl=l!: 0;: CEQ Ol'lt INSi1T1JTiQNAL OFi='ICLA.l NAME &. "r1'i"1S 0;: CSO OR IHSTli"VilONAL. O:!='FIClAl..(i~or PrirltJ ~"ii: SIGNSO

~~ ,.~~ - ---------- --- ------------ ----------

/~Z-fjo~ APHIS FORM 7023 (~:>taee.. V$ J=OlltM 1l--2~(OCT 88).....r.'liCh is oo.o\e:\e

(AUG .'1

NOV 4

Se; reVe."'se sid!. fOiTn15 re?ortts requ.ireci by law (i USC 2'1'3) Failure to report 2ccoraln, to ~he r~g1.f.E'tions can addftbnal inf'onr.-ion re'v~ iD an ortierto cease and duW and Ie ba zub;ecllo penalties 2S prcMded tor in Ssction ),150

UNli'E) SiA"-:S D~AATM:5N'io!t' A~RICUL."iU~

ANI"""AL AND P'l,A..... .HEALTl; INS~ECTIONS;~C=:

. CONTINUATION SHEET FOR ANNUAL REPORI

OF RESEARCH FACILITY (TYPE OR PRINT)

1.. R:GIS'i'iU.i10N NO. FORM A=-PRO\f50 OM2 NO. 057'P.Q03684-F-0001 I

2.,MSAOQ~~~Si=t.!::S:AKCH 'FAC1LJi'Y rwllme and Add~ss.;as register!!d with USDA. m=lude LIp Code) •

-

USDA,APHIS,WS,NWRC 410'1 LAPORTE AVE. top; FT. COLLINS, CO 80521 4r/QA'~O;P Yr;.

IRS?'OR,T OF AN1MAL.$ USEO BY OR UNDE~ CONTFt01.. OF RESEARCH FACJUi'Y (Altach ac::ldltion8l sheets it ft8CfISSa'Y Of 1:Se A~HIS FOAM 7023A,) .'/'f/ If,,-(,Ill A. a.NUlm~erof C. Nurnbtlrof C. Nur;,bet o1anl1nam upon £.. N~b8rd animalS u;:aon which te..ding .

anirrr8ila being animal.. upon which i::xp.et1ments• e:.perlments. r..ear:f'\. .-urge')'. Of' len _ .... ....nim.is' Covered ....., which leaching. t"cNng........rch. condu=-d invoMng aceomparl)'i'n; pain 01' d"'cavesa

Sy The Anima c:onditiOl"leCI.or research" surpery, or 1esv ""'~ '0 the animat& .nd fcr'whJ::h the u.e fI£ appro;:....'e Weltare ~e.pul.'ions 1\o6'd 1orua. in experiments. or COftct:ucMWl invoMng aneslhetic. an.lpesic, Dr vanQuili..-mg dru~. woula

t.aching. testing, testa: were .CICOrhpanying! pain or n.". .~ anee:s.d th. procedures. ,....ub. or experiments. conduCled oartrau \Cl1t'W animab

interp...1.erion Of 'Ihc \caching, re:searcll.,

---------------- ~r=n.or inYcMng no and for which a~e e.xperinw=nts. Altgery. or....1L. f4.n ""n.fian of .surgery but not pain. dl8tru:s.. CJl .nealheflC, aft8llpu:i=. ar It» p~dl.Jie.p~P4'in or~.. ift rho';

,:2..&/OFl 13.0"'..- )'!l!lu-CSfarlNl:b v•• of~""" t....nQUIi:I:inG o~ were .nirn" .nd".. nt.Ulna AIC1'l drug:. ....r. not u..i:I (U$lby~) ......... ,.,r-evinll dru;:s.. u.... mU$l bf: .,..-cJu)d fO tIW ,..JXirtJ

bund Sauirrels I 0 () I~O n Squirrels I O' 13 I 0 I 0

unks 0 33 I 0 0 ccoons 0 67 0 I 0

rita in Beaver 1 9 32 I 0 5 earn Beaver I 19 22 0 I 0

ite Tail Deer 13 15 I 0 0

C;~ Tail Deer 90 57 0 I 0

es 60 0 0 0

cupine I 3 0 I 0 I 0

ket Gophers 5 0 0 0 LQOOSe 0 6 I 0 0

I I I .1

I I

I I j

.. I I t ,\Ssu;::tAHCE S1AT:J.eNTS

"IfF.

TOTAL.NO

OF ANIMALS

(Cola. C­0+ E)

J

Gr t ~O

FoX 13 Sk 33 Ra 67 Mou 37 Str 22 Wh 15

Bla 57

Vol ;>-' ..'.

0

Por U

Poc 0 Mon I 6

I I

r

') Pro"S'S~N1l1yaccaptabl.aancarda D'O"I'tming lhe c.r&, lra.tmen~.ndI:h of .nirn.b inc:Iudin,g .P;JFO~rlal-e use of ane&thelic:••n8I~sic••nc: tr8nQlIIlI%ing drops. Pricr1O. during _nc1 folbwinp actual rasean::b, leachino, te:slins;. surgery. or experirnen:ation ~e fotlowad by lhi.. ret:.a~h :.c"lly.

3) "Thi$ facili't)- is adh8ring to the sg;nd_T"l:ts and ~puI.,lons under the; Ac\" and it has required 1'-\ exceprions: lo the- .-ndards anod regulations: be apec:ir-ec:!' .nd e:Jq)1_n.d by the prtn:ipal i~i;alorand approvad by the mstiluftOnal ArUmaI Care .nd UM Commlh.. (lACUC). A summary of .11 such exca-ptiona ;05 aftached" 10 'l~. annual ,..or:.1n ..ddroon IQ lO....ury;ng th. lAClJC.a~e.-eeplicw1s. this surnt'NIry ~a • ~rioa~ e~l.n.fioo of 1he eEC"e:J1icms, •• ......n as ,..,. ~=s and nvmber Of anlmats a1feCiad.

£) T'he enending veleuinarian fo:"1h:c resurc:h fec:rnty I'roes appro;tr.,e .lJthoril¥ lC erm.r. Ihe pr'O'l'i8ion of.dequa1e veterinary =are _nd lc ove..... the adIequ.cy or ott-.. ~ "" animal care ahd ....e.

C:=RTIFICATION BY HEADQUARI :=RS R:=Si::ARCH FACJLlTY OFFICIAL. (Chief Executive Officer or Legally Responsible Institutional Official)

'cor1liy thol lilt Ib..... is lruo, c.noct, Ind complote f7 U.S-C. S.::iian 21~3~

e.-.:.=.SIGN~SIGN"'-ru~e OF C:~ OR InlSTiTlJilOPllAL O~CfAl; ~ .. ilTL.~ OF eso OP\ tNSTiiViIONAl ~FICL.&."L.(T"Joo'Peor 1=',.;,,1j

---------- --- ------------ ---------- Vrlljdy---- ~ ~ ---- APHIS FORoM 7023A

(AUG 9')

NOV -4 2004

Column E Explanation

t. Registration Number:

2. Number of animals used in this study: 35

3. Species (common name) of animals used in study: Canis latrans (coyote)

4. Explain procedure producing pain and/or distress:

Animals were fed or gavaged with suspensions containing mixtures of caffeine and theobromine to eva"luate the pot~ntial of these subst~nces as selective predacides. Dose vs. Response (percent mortality) curves for three mixtures (13:1 (theo:caf), 5: 1 (theo:caf), 100% theo) are being constructed from the toxicity testing data.

5. Prqvide justification why pain or distress could not be relieved. State method or means used to determine that pain and/or distress relief would interfere with test results,

QA-1064 "Development of a Natural, Safe and Effective Plant Based Predator Toxicant" was designed to evaluate the potential of methylxanthines (theobromine, caffeine) as a selective predator toxicant. With experimental toxicants, it is difficult to predict pain or distress experienced by the animals dosed. Administration of other substances (analgesics, etc.) prior to symptoms of intoxication might confound the pharmacological action of the methyxanthine test substances and lead to erroneous conclusions and ideally would be avoided until necessary. Although listed as a Category E study, the protocol permitted the attending veterinarian to administer analgesics, anesthetics and/or euthanasia in instances where the animals were determined to be in pain or distress.

6. What, if any federal regulations require this procedure?

!,gency: none CFR: none

Column E explanation for QA 1118

Registration Number: 84·F·0001 Number of animals used: 5

Species (common name): Mountain Beaver

Several attempts have been made to reduce damage by decreasing mountain beaver (Aplodontia rufa) populations through the use of conibear-traps. This type of lethal control is becoming politically less popular, as indicated by the passage of Initiative 713 in 2000 (RCW 77.15 section 3) which banned the use of all body gripping traps in the state of Washington. Therefore, alternative 'tools to conibear traps for reducing mountain beaver populations may be desirable. At present there are no toxicant registered for use to control mountain beaver. A previous study (QA- 885) showed that chlorophacinone was the only underground bait that was 100% effective and readily consumed by mountain beaver. LiphaTech currently holds a chlorophacinone label in the form of paraffmized pellets. These pellets are delivered in bags to prevent weather damage. Another recent study (QA 1072) conducted in the Olympia Field Station habitat pens, showed that mountain beaver cached bags. Using LiphaTech's delivery system might reduce primary hazards as mountain beaver can cache baits inside their nests.

Ten mountain beaver served as subjects. Animals were given a minimum of 2-4 weeks to ad8.pt to pen and burrow system. After adaptation period was over, the bait was placed in a trash can (761) in each of the pens. Each container has a 10cm diameter hole at the bottom to allow access and to mimic the rodent's natural burrow system. Five of the animals received a 12 oz bag of 0.005% chlorophacinone, while the other 5 animals were used as control and given no bait only bags with plain oats. Bait formulation for each treatment was presented as paraffmized pellets. The status of animals was monitored at 2-hour intervals for the first 6 hours, then again every 24 hour for the next 28 days. As administering sedatives or analgesics could affect the toxicity of the chlorophacinone baits, no drugs were administered until acute toxicity was imminent. Animals were frequently monitored to minimize any potential suffering until the completion of the study. Any animal that demonstrated severe symptoms (e.g., convulsions, comatose) ofpoisoning were euthanatized immediately.

This is a preliminary study to determine ifthe LiphaTech product might be applicable to a field application. EPA regulations require that for fmal registration of a product that 70% efficacy with death as an end point be demonstrated. Since this data will support registration we therefore followed EPA regulations with a few modifications to account for a different species (------ --- --------- Standard rat anticoagulant place pack dry bait laboratory methods, Guideline #1.217).

October 25,2004 NWRC, FY 2004 Aghis Form 7023, attch #1

Locations where animals in this report were used and/or housed:

USDA, APHIS, WS, NWRC 4101 LaPorte Avenue Fort Collins, CO 80521

USDA, APHIS, WS, NWRC Olympia Field Station 9730-B Lathrop Industrial Drive SW Olympia, WA 98512

USDA, APHIS, WS, NWRC Logan Field Station 4200 S 600 E Cache County Road Millville, UT 84326

USDA, APHIS, WS, NWRC Hawaii Field Station PO Box 10880 Hilo, HI 96721

United States Department of Agriculture Animal and Plant Health Inspection Service

Safeguarding American Agriculture NDV­

'{h~.reporll. re'lulled by law 17 USC 2143, Failure 10 repor' acclIldinll'o Ihe reoulal~~~i .i See reverse side lor Inleragency ReI>OI' ~ ,ol;ull In an ordof'r lQ ('fa.;\t;p Ami 11"~KI AlII'i In bp. subject 10 pen..111ei as prOvided.or in Section 2150

allditiunal inlollnalloo. 0180·00A·AN

UNITED STATES OEPARTMENT OF AGRICULTURE t. REGISTRATION NO.ANIMAL ANO PLANT HEALTH INSPECTION SERVICE FORM APPROVED OMB NO 0579-003682-V-001 I

Z. HEADOUARTERS RESEARCH FACIUTY (N8lfle and Address, ., leg/"eled ..ilh USDA, include Zip C_}

Department of Veterans Affairs ANNUAL REPORT OF RESEARCH FACILITY VACO 810 Vermont Avenue NW Washington, DC 20420

(TYPE OR PRINT)

, 3. REPOIlnNG FACILITY Il••• ullrocal,ons where alllma', were hOllS8<l or USll<Iln aclual'll$tWrch IlIS"II9 leaching 1If expell...en'alion or held lor lhese purpose•. Allach addItionalsheets If necessa,'t.) .' •• I

FACILI1Y I.OCATIONS (SlIe.)

531 VA Medical Center 500 West Fort Street Boise, ID 83702

LREPORT OF ANIMALS USED BY OR UNDER CONTROL OF RESEARCH FACILITY (Attach allill/llnll'" streets ., n""S"a'll or use APHIS FORM lO'l3A.) A.

Animals Covered By The Animal

Wellare Regulations

4. DOllS

5. Cats

6. Guinea Pigs

7. Hamsters

R. Rabbits

9. Non-human Primates

10. Sheep , 11. Pins

12, Other Farm Ammals

13. Otoor Animals

B. Numbero! animalS being bred. condilioned, 01 held 101' use in leaching, lesling. experiments, research, or surgery bill noI ye. used lor such purposes.

C Number 01 animals upon whIch leachIng, r_arch, e.periments. or 'es's were conducted Involving no pain. dis'ress. 01

II$O! 01 pain­relieVing drugs.

D. Nulnbe, uS animals upun which expe:rime,,'&.. 'eilChing. research, surgery, or lests were conduc'ed lII"""'i"g accompanying pain Or distress 10 the animals and lor which appropriate .nesltlelic, analgesic, or Irallquilizing d"'lls ....re used

E. Number 01 animals upon which leaching. experiments, research, surgery Of tesls were conducled illvolving accompanYing pain or dislress 10 lhe anilnals and 'or which 'he use 01 appropriate anllSlhelic. analgesIC. or 'ranquilizing drugs would have acMl<sely .Ileeled lhe procedures, resu"•• or interplelalion of the '88Chlngt research. experiment•• SUlll"'Y, or ,esls. (/In sxplallal;on of Ihe procedUf8IJ producing pam Of di.'ress ill I/le.s animals and fhe mason.s .uch drugs wer. not uslld dIU" be fllfllCl!ed to this ,..".,rn.

F.

TOTAL NO. OF ANIMAls

(COla. C .. o + El

2k 12k;

I ASSURANCE STATEMENTS

I' P."O't:5sh31IaUy acceptable ~t~nd.Jrdc governing the car". heillfn'i!''''. O1n41 u!li~.nl ammills. mcludmO appronale use 01 aoesltlehc. analgesIC. and t.anqulllLlng drugs. prior to. dunng. arllJ lullowing aClua' research. leachirwJ. lesting, surgery. or el'perirnentaliOiI were toflowed by Ihis research f.acility

21 Each fJUlICtpat lnvestigator has cOIlSldered allernahves to pailliul procedures

3). Thi$ l&.Ieilily iii oiJdhering 10 Ihe slandiJrds arwJ regullUlions under the Ad. aud it has reqUired that eKCepllollS 10 the standards and tegulations be ,pee,tied and explained by 'he prir1c.:lp.a' ilivesllgulur alld approved by the Instllillionol AnilnCi'1 Care and Usc Co.nmitlee flACUCJ A summary 01 an such ••cept6ons ia atlac;hed to thIS annual report. '" ad<Jilion ,u idenlltying Ihe IACUC·al'proved exceplio"., 'hi•••""nary includes u briel eXj1lanalion ollhe excep'ion5. as well as ,he SpeclllS and numlll!r 01 animals allecled

41 n)(~ iltlulldill9 vetcl;lIafian tor this rcStldrch lactlity h~5 aPf>Jop.wle aUlbuuly 10 enSUre Ihe VruvlSkUl u1 adequa1e veterinary care and to oversea the adet:,uu,cy ul oUter ,aSPllt:IS uf animal care Dud use.

CEK'I'JIo'ICATION 8Y IlEAIlQU AKn:S RESEARCH Jt'ACII.l1'Y ()fo'Jt'ICIAL <Chief Executive Officer or Legally ltelipllnsible Institutional Official)

I ".r"ly Ih..1 Ihe at>O"e i. 11118, """nel, alld Clx"p1&lu \1 USC Sec:lio" 21431

SIGNATURE OF C.E.O. OR INSTITUTIONAl. OFF'CIAl. DATE SIGNED

10/27/O~

APHIS FORM 7023 (Replac.es 'liS FORM 18-23 (OCT 881. whIch IS obsoJele I (AUG 91)

P4RT 1 _ I-lI=AfVlIIARTI=R~

, ThIS ,eporl is required by law (7 USC 21~3) Failure \0 re~rf ~cC"rding 10 the reg~lalion:eP.,E C .0 7200~see 'everse side 'or ------ --- -- -------- ----- Inlerageoey Rep/", Conl'o' No. (esuliin an order 10 cease and deslsi and '0 be SUbjecl 10 penalties as prOvided for in Secrton 2.150 addilional informal/on. alSO-DCA-ANI. UNITED STATES OEPARTMENT OF AGRICULTURE

ANIMAL AND PLANT HEALTH INSPECTION SI:RVICE

ANNUAL REPORT OF RESEARCH FACILITY (TYPE OR PRINT)

1. REGISTRATION NO. FORM APPROVED

82- V-DOl OMB NO. 0579~036

2. HEADQUARTERS RESEARCH FACILITY (Name and Address. as registered with USOA, inclw. Zip Code)

Dp.partm~nt. of Veterans Af~airs V,i),ca 810 V~rmn~t Av~nue NW W~sh;0gtons DC Z0420

, 3. REPORTING FACILITY (lisl all locations where al\lmals were housed OJ IJsed in actual research, resling, teaching, or experimenralion, or held lor these purposes. Allach addilional , sheels it necessary.~

FACIUTY LOCATIONS (Sites)

A.

4.

5.·

6.

7.

8.

9: 10.

1,.

12.

13.

,. .... - ~. :\..jJl :j.' ;"':ed'jca 1 Center !1nn :.:~r-:'<: + Fnrr ~tV\~::1f

~ ..... ...,..,-..,8~, i"~.e ~ ID O •..l!U;;,.

I REPORT Of ANIMALS U5ED BY OR UNDER CONTROL OF RESEARCH FACILITY (Attach a.IWilinnal sh.'I!'s H ,leces...", or USf> APHtS !'ORAl 7023A./

Animals Covered By The Animal

Welfare Regulalions

Doos

Cats

Guinea Pills

Hamsters

Rabbits

Non-human Primates

a Numberol animals being bred, condit ioned. or held for use in '.aching, lesling. experimenls. research, Or surgery but no' yet used tor Such purposn.

C Number 01 animaJs. upon whIch leaching, research, experimenlS. or lests were conducted involving no pain, distress, or use ot pail\. reieving drugs.

17

1P.

o Number oj arumals upon' . whtCh experimen,s,

teaching, research, surgery, Or les's were cooduc'ed invo&ving accompanying pain or disness 10 the animals and tor which appropreate anesthetic, ana~esic. or Iranqumzing drugs were used.

35

78

Eo

,~

23

Sheep

Pim:

Olher Farm Animals

Other Animals

!J;::t.~~

~\'; 1r..Q

.,. J .'"

")1') L.. .:~

Number 01 animals upon which teaching. f.experiments. research. surgery or tests were conducled inVolving accompanyino pain or distress 10 Ihe anomals alld lor which lhe use 01 appropriale TOTAL NO.. aneslhetic, analgesic, or ,ranquilizing drugs would OF ANIMALs have adversely aflecled the procedures. results. or inierprelation 01 the teaching. research, eJlperimenls. surgery, or tests. (An explanation 01 ICols. C .. the plocedutes ploducing pain Ot distress in these 0 .. Elanimals and the le.son8 such drugs "-Ble not used muslbe at/ached 10 Ihis teport).

181

35

96 ~~ QQ '.~·v/'0

\

I ASSUIlANCE STATEMENTS ..

I) ProlesSlOnally acceptable slandards governmg Ihe care. trealrnenl. and use 01 animals. Includmg appronale use ot anes!heUc. analgesiC, and tranquIlizing drugs, pnor 10, durango and lollowing actual research, tucning. ,esling. surgery, or experimenlation were lanowed by Ihis research 'aciUly.

2). Ea.ch princ.pal in~estiga.or has considered cMlernatives 10 paintul procedur~s.

3). Thh. lacUity is Iitdhering 10 Ihe Siandards and regulations under 1he ACI. and it has required Ihal exceplions 10 fhe s1andards and regula lions be specified attd eKplained by Ihe princlpat invesligalor and apPfoved by the InslillJliolJiJl Animal Care and Use COImnillee (IACUC). A summary of aU such exceptions is attached to this annual report, In addilion 10 idenlifying lhe IACUe-approved eKceplions, lnis summitry inctudes a briet explanation 01 the exceptions I as well as the species and number 01 animals aUected.

4) ihe ullendlng velerincuiill\ lor Ihis research lacilily has iJppropriiJle iJuthorily 10 ensure the prOVision 01 adequale veterinary COlre and 10 oversee Ihe adequacy .01 olher aspec15, or allimal care and use.

APHIS FORM 7023

CER1'JIt'ICATION BY IIEADQUARTJo~S RES£ARCH ~'ACILl1'Y Ot't'ICIAL (Chief E"eculive Officer or Legally Responsible Insliluiional Official)

_.• I certlly thaI ihe above is Iru~, correcl. iJl.d COIl.lplEne (7 USC $eclion 2143).-. SI--------------- ----- --------- ----- -------------------- -------------

~_- .

NAME & TITLE Of C.E.O. OR INSTITUTIONAL OFFICIAL (Type or PT/nl) DATE SIGNED

C\L\~~ ---------- --- --------- ---------- -------- ---------- 11/10/04

--- --- - -- '" (Replaces VS FO~8'Z3 (OCT 88), whK'.h IS obsolete)

(AUG 91) _..- r

0 .cl.. 3 2004 PART 2 - SECTOR OFFICE

Investigator: ---- ----------- --------- --------

Protocol #: OLS-00148

Justification of Category E Animals

The category E rabbits were injected with anthracydines chronically and they may develop congestive·heart failure over a 10-week period. Use of anxiolytic agents or stress reducing drugs such as barbiturates or benzodiazepenes may induce or inhibit microsomal liver metabolism or displace anthracydines from plasma protein stores thereby altering the pharmacokinetics of anthracyclines. Such an effect may interfere with the experimental outcome. The drugs may have direct effects on the proteins of the heart being studied and interfere with the results of the study .

DEC 1 3 2004

Harvel ItfCKelIJw ' This reporl is required by law (7 USC 2143). Failure 10 reporl according 10 Ihe regulalions can See reverse side tor Inleragency Report Cq,fuOI No .. J resull rl~an order (0 cease and deSist and fa be subject 10 penalties as provided for in section 2150. additional ~nformalion.IJ......@t!10~~A.AN .'C", . ­

UNITED STATES DEPARTMENT OF AGRICULTURE 1. REGISTRATION NO. Co ..."T'~ I ANIMAL AND PLANT HEALTH INSPECTION SERVICE FORM APPROVED "a. 'I .../ . /.7 . . S?- if-001 /3 ,'+ .-M. OMB NO. 0579-0036n/ ~... 0-' /Kr...,.u.....,.~/ . 2. HEADOUARTERS RESEARCH FACILIlY (Name and Add'ess. as 'egisle,ea wi/h USDA,

./.... I' V T C.J-/' I S () )- include Zip Code) .

ANNUAL REPO T OF RESEARCH FACILI~ Dp.:Jar-tMer:t of Veterans Affa i rS'jl>V-L ........ -lY'-''',-<.... (TYPE OR PRINT) VACO <;:;/'''; ~'? Y jp:::::, ( i

GI0 V~~"m0:1t J\v{~riue f~~·l ?t.'j~~~?:~=_~.Lf~ "iashir1r;to:~. DC 20420:_:;:..~~-:-j;.-·:"'/-'::;-~-/~\i"\(~'{"''':::'''''''

::;r- ..... /,/ F. /' r ~_~'--(Fv-t,. ")

3. REPORTING FACILITY (liSI aillocalions where ammals were hqused or used in aClual research, lesllflg, leaching, or experimenlalion. Of held for lhese purposes. Atlach addilIDnil.!

1 sheels il necessary.) . . ;?i:9 'I~~- 1/ DTJ ·/~

--;::-;;-::_-;-;-~:-;---;-:_"'7--;:-_:-;- F_A_Cl_LIlY LOCA_rTl_ON_S.:..:..:.(s........ :;p:.z...:~='t:-r_!....-...:)?:.:.....~::.::-::;;.=];j~I:.::~.::=::!.d-<-:::'::.;··::::::,.__ ,'eS)=-­

531 '}A" '::d i co. 1 C",;; :'>:t' ~/'--'

I REPORT OF ANIMALS USED BY OR UNDER CONTROL OF RESEARCH FACILllY (Attach all;,liliollalsheels i/ne~essary 0' use APHIS FORM 7023A)

A. B. Number of C Number at D. Nu.nber 01 annnals upon E. Number 01 animals upon which leaching. F. animals being animals upon which experimenls, I experiments, research, surgery Or teslS were

Animals Covered bred. which leaching. leaching. research. condUCled involving accompanying. pain or dislress By The Animal condilioned, or research, surgery, or tesls were 10 the animals and for whieh Ihe use 01_ appropriate TOTAL NO.

Welfare Regulalions held lor use in experimenls. or conducled involving anesthe1ic. analgesic, or tranquilizing drugs would OF ANIMALs leaching. lesling, lests were accompanying pain or have adversely affecled the procedures. results, or experiments. conducted dislress to the animals interpretalion of·the leaching, research, research,· or involving no and lor whach appropricue experiments. surgery. or tesls. (An explanation of (CO'Il. C ... surgery but not pain, dislress, or aneslhelie, analgesic, or the procedures producing pain 01 distress in these o ... E) yet used tor such use ot pain- tranquilizing drugs were animals and the reasons such drugs were not used purposes. relieving drugs. used. must be attached to this report). ­

4. DOQS 1:1 • ..,V ~nR YOUR 5. Cats INFORMATION 6. Guinea Pios

7. Hamsters

."} ...: Inf_o,)8. Rabbits 17 1

9. Non-human Primates

3510. SheeD 35

11. Pins

12. Other Farm Animals

13. Other Animals

05

76 13

I ASSURANCE STATEMENTS

1} Professionally acceptable standards governing the care, treatment, and use 01 aOlmals, IIlcludmg appronate lise of anesthetiC. analgesIC, and tranquilizing drugs, pnor to, dUling, and lollowing actual research, teaching, lesting. suryery. Or eKperimentation were lollowed by lhls research lacility.

2). Each principal invesligalor has considered alternalives 10 pamlul procedures.

3), This lacillty is adhering to the standards and regula lions under Ihe ACI, and iI has reqUired Ihal exceptions 10 lhe slanda,ds and regulations be specified and explained by the prin(:lpal investigator and approved by the Inslilutional Anilnal Care and Use Commillee (IACue). A summary of all such exceptions is attached to this ~nnual report. In addilion 10 identilying Ihe IACUe·approved exceptions, this summary includes a brief exphmalion ollhe excepliolls. as well as the species and number of animals attected.

4) The attending velerinilrian lor lhis resei)rch lacllily has i)pplOprtUle i:lulhorily to ensure the prOVision 01 i1dequilte velet'i~ary care and 10 oversee Ihe adequacy ul other aspects of i)nimal care and use.

CERTU'ICATION BY IIEADQUARTES RESEARCH to'ACILITY ()J<· ..·ICIAL (ChierExecutive Officer or Legally Responsible Institutional Orticial)

I ce.rllly ahat Ihe i1bove is Irue. corrf!C1. and cOlnplele (7 U S.C Sel:lion 2143) DEC - 1 2004 SIGNATURE OF C.E.O. OR INSTITUTIONAL OFFICIAL NAME & TITLE OF C.E.O. OR INSTITUTIONAL OFFICIAL (Type or PrrnV

~vayne --- --------- ---------- -------- ----------

DATE SIGNED

11/10/04

(Aeplaces VS FOAM 18-23 (OCT 881. which IS obsolete I APHIS FORM 7023 (AUG 91)

~-Ft' I~I'I

~~. 'frf

&d-V-(~><J 1,.

Investigator: ---- ----------- --------- ---- ---

Protocol #: OLS-OOI48

Justification of Category E Animals

The category E rabbits were injected with anthracyclines chronically and they may develop congestive heart failure over a IO-week period. Use of anxiolytic agents or streSs reducing drugs such as barbiturates or benzodiazepenes may

. induce or inhibit microsomal liver metabolism or displace anthracydines from. plasma protein stores thereby altering the phar~kineticsof anthracydines. Such an effect may interfere with the experimental outcome. The drugs may have direct effects on the proteins of the heart being studied and interfere with the results of the study .

(,OJ:..b ;+5

DEC -1 ID04

Additional information requested for Block E of APHIS Form 7023, dated February 3, 2005

The rabbit involved in this study was injected with anthracyclines or their analogs twice a week for eight weeks. Heart function was assessed by echocardiography.The rabbit was euthanized when fractional shortening was reduced to less than 25% or twenty weeks after the first anthracycline injection. The heart was then removed for further study.

FEB 1 4 2005

;~~~,;~~u~~ ~~r~e~~~~e~::e'~~J711~S~~::,~3:o ~i~~lr'~~lre'OpopeJlaC"curdon9 10 Ihde r"!l,ulallons cali' ~ ~. <.' 41J\lll'ee reverse side 'or 'nlcriJgency Report Cnnlrol N,t)-- t1 i / ....... na 'es as prov, ed Of 1/. Secliofl 2150 additionilt informa1fon 01 aO·OQA·...N ( .:fI..:.--·I

t~·

rREPORT OF ANIMALS USED BV OR UNDER CONTROL OF RESEARCH FACILITY (Attach ..,li<1illOnal shL'f>ls ,1 necessary or lise APHIS FORM lO23A)

A. B. Numbero. E. Number of animals upon which leaching.C Number of D. Number 01 DUlinals upon F.animals being experimen.ts, research, surgery or tests were..nimal~ upon which experhuenls,bred."'nimals Covered conducted involVing accompanying pain or distresswh'ch leaching. reaching, research.By The Animal condilioned. or research, 10 lhe animals and for which 'he use of appropriale TOT... l NO surgerv. or tes's wereheld lor use In Weltare Regulations anesthetic. analgesJc. or tranquiliZing drugs wouklexperimenl5. Or OF ANIMAlsCOoduCled involving leaching, lesling, have advcr~cly :Jilteclcd tho procedures, results. ortesl3 were accompanying pam orexperiments, inlerpt'etation 01 the leaching. research.condueted distre~"S 10 the animalsresearch, or experimenls, surgery. or lesls. (An explanation of;nvolVing no and 'or whIch appropriare (Co's. C + surgery bUI nol the procedures"producing pain or distress in thesepam. distress, or 0+ E)anesthetic;, analgesic, Ofyel used for such animals and the reasons such drugs were nof useduse of pain- tranquilizing drugs werepurposes. must be attached to this report).relieving drugs. used

4. DOQS

5. Cats

6. Guinea Pigs _.---.-- ..'--.

7. Hamsters

8. Rabbits 9494

9. Non-human Pnmates

10. Sheeo

11. Pios

12. Other Farm Animals

13. Other Animals

2229

Mice

Rats 62 552688 927

5301127 468 24

I ASSURANCE STATEMENTS

UNITED STATES DEPARTMENT OF AGRICULTURE ANIMAL AND PLANT HEALTH INSPECTION SERVICE

(cl \ ,I

~. / ~ / .

I ANNUAL REPORT OF RESEARCH FACILITY I (TYPE OR PRINT)

1. REGISTRATION NO. FORM APPROVED

93-V-007 OMS NO 0579·0036

'.---'

2. HEAOOUARTERS RESEARCH FACILITY (Name and Add,ess. as reglslered w,lh USDA. include ZjIJ Codel

Department of Veterans 810 Vermont Ave, N.W. Washington, DC 20420

Affairs

13, RhEPORTlNG FACILITY (llsl all localiolls where arumals were housed or used in aclual research. leslmg. leaching or experornenlalion ur held lor 'hese pu'poses Andch add,'",nal!Ii eets" necessary.) . .

FACILITY LOCATIONS (SlIes)

tl600 - VA Long Beach Healthcare System 5901 E 7th Street Long Beach, CA 90822

1) Prolesslonally acceptable standards govermng the care. Irealrnent. imu use of aUllniJls. including upprorlille lise 01 anesthellc. analgeSIC. and lranqulhLmg drugs, prior 10, durIng, and following actual research. leaching, tesling, sltrUE~ry, or experimentaaion were tollowed by Ihl:i research tacillty

2}. EiJch prlllClpal investigator hilS considered allernJUves to pc:unhJl proCt!dures.

31. This fiJclllly i~ adhering 10 the standards alld regulallolls under the Act. and i1 hilS required that cJ(ceplions to Ihe standards and regulations be specified ,and explained by Ihe prnu;lp,,' rnvesliyalor "nd approved by Ihe lnslillillorwi Anillw' Care ..nd Use Commillee (IAGUG) A summary 01 all such exceptions is allached 10 Ihis annual repor1. In addllion 10 IdentifYing the IACUC-approved excev11ol):!:l., Ihl~ summary includes Col braef explanation of the exceptions, as well as rhe Species and number of rmiUldls dflecled.

4) the -..UcrnjlUg velerillarwn tor this rt!~c~rch 1&l<.:II,ty h,us iLlppropnale aUlhurlly 10 ensUre Ihe rnuvlSioli 01 adequate velermary cure LlIUJ lu oversee Ihe ad~quil<':Y ul other iJspects u1 eJllHnal cure and use.

Clo:R'I'U'ICA'I'ION tty 1I ..;AUQUARTt;S RF.SEARCII "'ACIUl'Y O.....·ICIAI. (Chief Executive Officer --- --------- ----------- - -----------------

I "erltly Ihill Ihe ..b----- --- --- ---- --- --- -- --- ------- ----------

SIGNATURE OF C.E.O. OR INSTITUTIONAL OFFICIAL DATE SIGNED-- --- -------

l()(~(/}

APHIS FORM 7023 (ReplaCes VS FORM (AUG 91)

PAI=lT 1

Explanation of Category E:

Rabbits

PI: Haake Protocols: 00II -178

04044-02 Number of animals under Category E: 28 Procedures involving rabbits fall into Category E because of immunization with adjuvant, which is required for adequate B cell production. Rabbits are sedated with acepromazine 0.25-0.5 mglkg SC or 1M given 10-15 minutes prior to immunization.

PI: Ohning Protocol: 01002-03 Number of animals under Category E: 4 The reason that procedures involving rabbits fall into category E is because of immunization with adjuvant, which is required for an adequate antibody response. Rabbits are sedated with acepromazine 0.25-0.5 mg/kg SC or 1M given 10-15 minutes prior to immunization.

PI: Mayer Protocols: 99064-04 (VA project # 0041)

11091-02 (VA project # 0055) Number of animals under Category E: 443 The rats listed under USDA Category E were subjected to Water Avoidance stress and Colorectal Distension as described in protocols entitled "Modulation of the Pain Response to Repetitive Colorectal Distention" (VA project # 0041), "Influence of choronic water avoidance stress on visceral sensitivity in rats" (VA project # 0055). Water Avoidance causes no hann to the animals, but does cause them discomfort (psychological stress), which is a necessary part of the protocol. Colorectal distension does cause brief pain, however the stimulus is of a very short duration (20 seconds) and is also used in humans. This response cannot be elicited in anesthetized animals. Because the investigator is studying the analgesic effect of different drugs, the use of other analgesics would confound the results and interpretation of the study. None of the rats used during this time interval exhibited signs of excessive or prolonged pain (agitation, vocalization or bleeding during or following testing), which would have necessitated immediate euthanasia.

PI: SattinlPekary Number of animals under Category E: 25 25 rats were categorized under Category E for the VA merit review study because the forced swim test is considered psychologically painful. These rats cannot be anesthetized because they would drown during the forced swim test.

PI: Tache Protocol: 06069-02 Number of animals under Category E: 30 A colorectal distention model is used in rats. The study addresses the role of stress and CRF in the genesis andlor maintenance of lower gut motor function alteration and visceral hypersensitivity/pain symptoms. The distention process is unavoidable because the visceral pain response to distention needs to be compared to the response observed after the test substance administration. Because anesthesia will block the abdominal contraction response to distention and because the effect of potential analgesics is studied, the use of anesthesia or other analgesics will confound the data and their interpretation. Thus the pain response to colorectal distention is studied without prior analgesia.

PI: Wasterlain VA Merit Review # 9711-041 NIHlUCLA grant # 12105-02 Number of animals under Category E: 423 The rats were placed in Category E because they undergo a period of seizures which are not treated dUring that time. While humans who have had similar seizures do not report pain there may be some distress. The rats are not treated during the seizure period because it is the mechanism of brain damage due to untreated seizures which is being studied in order to develop better treatment.

PI: Yang VA protocol # 06068-02, UCLA ARC # 2002-032-02 Number of animals under Category E: 10 The rats were used to study the effect of energy deficiency on brain neuronal activation. The rats were fasted for 48 hours and then were euthanized.

PI: Haake Protocol #: 04044-02 Number of animals under Category E: 24 Procedures involving 24 mice fall into Category E because of immunization with adjuvant, which is required for adequate B cell production.

PI: Mayer Protocol: 03030-02 (VA project # 0052) Number of animals under Category E: 13 The mice listed under Category E were subjected to chemical stimulation of the colon as described in protocol entitiled "Modulation of stress-induced pain response in mice" (VA project # 0052). The end point of the chemical stimulation of the colon is significant pain

and discomfort that cannot be alleviated by analgesic since the aim of the study is to investigate pain behavior.

PI: Pandol Protocols: 0111-42

01008-02 Number of animals under Category E: 40 40 mice are listed under Category E for Protocols # 0111-042 and # 01 008-02. In the mouse model of experimental pancreatitis (#0111-042), high doses of cerulein (an analogue of CCK), given as intraperitoneal oudy injections, cause acute pancreatitis. In Protocol # 01008-02 the animals receive either intraperitoneal injections of cerulein or vehicle every hour for 7 hours.