IACUC Animal Protocol Writing Workshop 101

1. IACUC Animal Protocol Writing Workshop 101 How to Write an Effective and Successful Animal Activity Protocol: Hands On Training

2. A Good Sources of Information • Working with the IACUC: Writing an Animal Protocol AALAS Publication: American Association for Laboratory Animal Science www.aalas.org “Bookstore” link • How to Write an Application Involving Research Animals www.niaid.nih.gov/ncn/clinical/researchanimals/tutorial/index.htm • Guidelines for the Care and Use of Mammals in Neuroscience and Behavioral Research ILAR Publication: Institute for Laboratory Animal Research www.national-academies.org/ilar

3. Protocol Form 6 sections • General information • Number of animals • Surgical and non-surgical procedures • Alternative considerations and literature search • Statement of Assurances • IACUC actions

4. 1.Principal Investigator Must be full-time UMC faculty (Instructor – Professor)

5. Other Personnel Identify all persons associated with the animal work. This information assists the LAF and IACUC in contacting knowledgeable persons to answer questions/concerns relative to the animal experimentation. For access to the animal facility, all personnel working on this protocol must be listed.

6. Laboratory Personnel Training Certification

7. Project Title • This title will be used in the IACUC database to identify the project. (may be the same as the grant title) • The title should briefly state procedures to be carried out and the animal species to be used.

8. Project Title - Examples • Tissue Harvest in Rats • Health Monitoring of Guinea Pigs • Production of Transgenic and Gene Knockout Mice • Evaluation of Drugs for Osteoporosis in Squirrel Monkeys • Breeding protocol for AXt-1a knockout mice

9. 4. Proposal is • New - designation of a de novo submission • Revision of Existing Protocol - designation of amendments/revisions to the protocol. Any “revised” information should be easily identified [highlighted, bold,italics, colored font] • 3 year Full Submission • designation of a previously approved protocol that must be re-written after three years • Needs to include Appendix K

10. 5. Outside Contracts? Will any “outside” contracts be used in this study that involve live animals (antibody production, toxicology screening, subcontracts with other facilities, etc)? No Yes (provide information on the level of involvement) Mark YES if any non-UMC entity will be involved in any animal care and use activities. Examples include housing your animals at another institution for specialized procedures, use of a contract toxicology research lab, antibody production techniques by another lab, etc.

11. 6. Funding Source This question aids the IACUC and LAF in allocation of resources. In times of lean animal housing space, the LAF will further give top priority to those extramurally-supported protocols. Generally, the IACUC does not attempt to provide scientific review of a protocol, yet is obligated to provide some level of scientific review for those departmentally-funded protocols lacking any peer review.

12. 6. Funding Source • Title:  Grant Application Title • Grant Funding Agency: NIH, NSF, American Heart, etc    • Covered Dates: 11-01-04 thru 1-31-07 • Departmental Funds  • Has this proposal received any peer review? • Yes No Pending

13. 7. Procedure Category • Acute (anesthesia without recovery or euthanasia) • Acute procedures not only refer to an anesthesia without recovery, but also those studies where no advance forms of animal manipulations have occurred. • Examples include • animal sacrifice • tissue collection • terminal surgical procedure

14. 7. Procedure Category • Survival • Survival procedures not only include survival surgical procedures but any animal manipulations that may be conducted on the animal. • Examples include • any type of dosing (oral, feeding, injections, topical, gavage, etc) • sample collection (blood, urine, CSF) • behavioral experimentation (swim test, radial arm maze, single unit recording)

15. 8. Dates of Study • Anticipated start date of study: 2-23-04  thru 1-31-07 • B. Study Duration (maximum 3 years): 2 years Response assists the LAF in planning/coordinating space for the study.

16. 9. Animal Requirements List all animals required to complete the study. Animal numbers must be calculated for a period of three years. Complete all columns of the chart. If using nonhuman primates, must complete Appendix A. If using time pregnant animals, must complete Appendix B. If using animals from an in-house breeding program, must complete Appendix B.

17. 10. Breeding Program • Will animals be involved in a breeding program at UMMC or will time-pregnant animals be used? • No • Yes (provide information in Appendix B) • Mark YES for any study proposal involving animal mating or for proposals utilizing time-pregnant animals. Must also include Appendix B.

18. 11. Potential Hazards

19. 11. Potential Hazards Specific check boxes are included for chemical, radioisotopes, and/or infectious agent exposures in research animals. Boxes should be checked where appropriate and the proposal must be reviewed by the responsible oversight group. IMPORTANTLY, any YES answers must also include specific details of any specialized animal care procedures to be used, paying particular attention to occupational safety of animal caretakers. The explanation should state type of exposures, personal protective equipment to wear, decontamination procedures, and any other relevant information.

20. 12. Animal Husbandry

21. 12. Animal Husbandry • Standard: Standard methods for UMC include • ad lib access to water • LAF feeding of commercial diets • conventional housing

22. 12. Animal Husbandry • Nonstandard: Examples include • providing limited access to water • limited quantities of water • feeding specialized diets (e.g., high fat, High/Low sodium) • feeding specialized amounts • “unconventional” housing (metabolic cage, single housing) • behavioral manipulation of environment (e.g., changes in light-dark cycle)

23. 13. Housing Mark YES for any proposal that requires an animal to be maintained in a laboratory/testing location in excess of 12 hours. Any location maintaining animals for greater than 12 hours will be considered as a housing location and must be maintained in accordance with the USDA, OLAW, and AAALAC. The IACUC will need to provide approval for such sites.

24. 14. Objectives in lay terminology

25. 14. Objectives in lay terminology

26. 15. Rationale

27. 16. Brief Outline

28. 16. Brief Outline Appropriate responses to this question will briefly outlineall animal manipulations. This question also contains a check list of related appendices (A through K). Investigators will note that many specific details of surgical procedures, sample collection, administration of test compounds, antibody production, etc will necessitate completion of the appropriate appendices. An important consideration for inclusion is a time-line of events. This time-line helps the IACUC clarify the proposed work.

29. 16. Brief Outline For example, a study seeks to feed an experimental diet to rabbits then surgically instrument each animal with telemetry devices. Following the surgery, the animals will be monitored for 4 weeks and intermittent urine/fecal samples will be collected. 2 weeks Diet 1 Surgical manipulations 6 week old rabbits 4 weeks Diet 2 2 weeks Urine/fecal sample Sacrifice

30. Example Answer • In this response, #16 would include information about the diet, especially if potential nutrient deficiencies were anticipated. General information about the surgical procedure would be included. Likewise, any information about urine/fecal collection methods would need to be a part of the response (metabolic cage, catheterization, litter pan retrieval, etc). • A probable time-line might be: • Experimental diet (4 weeks) →Telemetry implant (1 day) → Monitor (4 weeks) → behavioral test (forced swim) → euthanasia • Additionally, the following appendices must be included: • Appendix C [Surgery] • Appendix D [Biological samples] • Appendix F [Administration Test Compound (diet)]

31. Appendices Check and complete only those appendices that apply to the animal manipulations in your experimental design.

32. 17. Justification of Animal Number • The total number of animals needed to complete item #16 should be explained in this response. Likewise, this number should match the requested number provided in #9. • It is often beneficial to include a flow diagram or chart/table that defines the animal use. • Break numbers down into number of animals/group, number of groups, number of replicates, etc. • Any animals needed for pilot work or training new technical personnel should be accounted for. • Responses should have some statistical merit.

33. 18. Location and Transportation • A. Indicate room(s) where animal procedures will be conducted. • This response is noted in the IACUC database as the experimental area. • Semi-annual program reviews will include this location into subsequent inspections. • Likewise, affirmative answers to #13 (Housing) should also be listed in this response.

34. 18. Location and Transportation B. Studies involving animal transportation to locations other than the housing area must identify the animal transport device, the nature of shrouds used to cover the transport device, and describe the route of transport Numerous laboratories are required to remove their animals from the LAF environment to their respective labs. The method and route of transport should be courteous to Medical Center visitors/patients, conscious of occupational exposures, and respectful of those opposing the use of animals in research. All animal movement must occur in appropriate restraint boxes/cages and with a concealment shroud/drape. Consult LAF SOPs for descriptions of routes of appropriate animal transportation.

35. 19A. Euthanasia • At what point in the proposed experiments will animals normally be euthanized (experimental end-points)? Or at what point will any individual animal be euthanized? • This response would correlate with the experimental end points described in #16. The question seeks to clarify the IACUC’s understanding of when any particular animal will conclude the study.

36. 19B. Euthanasia B. What criteria will be used to determine if an animal is to be euthanized prior to, rather than at, the anticipated end-point of an experiment? Note: Contact LAF, ext.4-1385, for recommendations on the assessment criteria. The appropriate response will articulate potential adverse consequences or effects of the experiment. Investigators should list potential surgical complications, adverse effects of drug/diet administration, or failure to manage pain and distress. Specific, objective criteria may also be incorporated (e.g., 20% loss in body weight, hematocrit falls below 25%, creatinine levels above 2.5 mg/dl, etc) to note when an animal is removed from the study.

37. 19C. Euthanasia C. If proposing a natural death (or death due to manipulations), explain and justify. This type of scenario is described as “death as an endpoint”. These studies must firmly justify the scientific need to allow an animal to advance through a potentially painful/distressful terminal event. Unless information to the contrary is known, it is assumed that all “death as an endpoint” studies will result in some level of pain and/or distress to the animal while in the moribund state. Ideally, some definitive, objective criteria can be used to “measure” when an animal can be removed. For example, in a model of renal failure, perhaps BUN levels >100 coupled with Cr values >3.5 are indicative of end-stage kidney disease and will provide the necessary endpoints. Likewise, in an oncology study, perhaps tumor growth beyond a specific size could be used.

38. 20. Euthanasia Procedures Procedures must be consistent with the 2000 Report of the AVMA Panel on Euthanasia [copy located in the LAF Training & Procedural Manual]. Likewise, secondary methods should be proposed to ensure and/or guarantee death. Secondary methods are typically those involving exsanguination, pneumothorax, or tissue harvest.

39. Assurances • Institutional requirement for all persons to be registered and known by the health services organization. • Have all personnel received a medical evaluation from UMC Student/Employee Health? • Have all personnel become familiar with the LAF Training & Procedural Manual?

40. Assurances The LAF Training and Procedural Manual is jointly produced by the LAF and the IACUC. The manual is designed to answer many commonly asked questions and address particular areas related to animal-based research programs at UMC. Each Principal Investigator is issued a copy of the manual. The manual should be available to all research personnel in the laboratory in an effort to foster a better understanding of animal care and use programs.

41. Assurances To comply with the USDA’s Animal Welfare Act [9 CFR, part 2, Section 2.31 (d)(1)(ii) and (iii)] you must complete a written NARRATIVE of the sources you consulted to determine whether or not alternatives exist to procedures that may cause pain and distress. Likewise, this search will ensure that the proposed studies are not unnecessarily duplicative.

42. Assurances Review of the available resources and previous experiments have determined that the proposed activity is not unnecessarily duplicative of previously reported activities? The question seeks to prompt the investigator to conduct a literature review and assure the IACUC that the proposal does not duplicate existing work.

43. Appendix A: Environmental Enhancement/ Enrichment • Enrichment Techniques • Are there any enrichment forms/techniques that are included in this protocol? YES NO • Description • Describe the above techniques • No Enrichment? • Are there any forms of enrichment/enhancement that should not be used in this study? YES NO • Justification for exemption YES NO • If YES, provide complete justification for this exemption

44. Appendix B: Breeding Programs Complete Appendix B for all proposals planning on establishing a breeding colony or for those studies utilizing time-pregnant animals. Studies incorporating breeding programs or offspring from time-pregnant animals will be required to report annual production (number of offspring used) at the time of IACUC protocol annual renewal. 1. Description Provide a specific description of the type of breeding program to be utilized (harem, monogamous pair, etc). 2. Personnel Responsible Identify personnel responsible for the breeding program. 3. Records Identify personnel responsible for maintaining breeding program records.

45. Appendix B: Breeding Programs 4. Adults How many adults will be utilized in this breeding program? 5. Final Disposition of Adults What is the final disposition of these adults at the conclusion of their breeding program? 6. Offspring How many offspring are anticipated from each breeding or time-pregnancy? 7. Final Disposition of Offspring What is the final disposition of any offspring not utilized in the experimental program?

46. Appendix C Surgery & Management of Surgical Pain and Distress 1. A Complete description of surgical procedure

47. Appendix C Item 2A: Formulary

48. Appendix C: Anesthesia 2b. Anesthesia Who will conduct the anesthesia procedure(s)? List Individual(s) by name 2c. Anesthesia criteria What criteria will be used to assess anesthetic depth and how will this be monitored? toe pinch, reduced/absent corneal reflex, withdrawal reflex 3. Experience/training Describe the training/experience, including species of animals, of personnel conducting the anesthetic procedure? List individual named in #2B and indicate level of training, number of years actively involved in procedure, and what species he/she has worked

49. Appendix C: Surgical Procedures 4. Surgical procedure Who will conduct the Surgical procedure(s)? List individual(s) by name 5. Experience/training Describe the training/experience, including species of animals, of personnel conducting surgical procedures? List individual named in #4 and indicate level of training, number of years actively involved in procedure, and what species he/she has worked. 6. Location of procedures Where will the surgical procedures be conducted? Indicate Room in which dedicated space for surgical procedures is located, e.g., LAF procedure room, R-717, N-412.

50. Criteria for Assessing Level of Anesthesia • Respiration rate • Heart rate • ECG • Toe pinch • Tail pinch • Corneal reflexes • Color of mucous membranes • Muscle relaxation • Other • Pulse oximeter • respirometer