Rodent Breeding: Rats

Boston University is committed to observing Federal policies and regulations and AAALAC International standards and guidelines for the humane care and use of animals.

Rats have short gestation times, with litter sizes varying from 8–16 pups. Cages may become overcrowded quickly if the individual responsible for managing the breeding colony and separating animals at the proper times does not do so in a timely fashion.

If the breeding colonies are managed improperly, the animals become uncomfortable and stressed. Pups can die from being trampled. In static micro barrier cages the air quality quickly deteriorates with a high density of animals and may predispose them to respiratory disease.

In addition, rats grow throughout their lives, and may reach a body weight of 500 g or more. Rats also grow quickly: Regular monitoring is essential to ensure cages do not become too crowded. Overcrowding of cages is an animal welfare concern and can have a deleterious effect on research.

This policy provides guidelines for rat breeding colonies and defines standards and responsibilities for rat housing and rat breeding cages. The policy assigns responsibility for action when rat breeding cages become overcrowded.

Responsibilities

Investigators

Investigators are responsible for:

1. Coordinating with BU ASC for space allocation for their rat breeding colonies.
2. Managing their own breeding colonies unless arrangements for technical support have been made with BU ASC.
3. Designating a Colony Manager (someone who has received specific training on managing rat breeding colonies, and who will be the primary contact person for the lab). More than one Colony Manager may be designated by the PI, but it is preferable that this number be minimized to facilitate communication between BU ASC and the rat users.

Colony Manager

Designated Colony Manager (research staff or BU ASC staff) is responsible for separating animals according to allowed cage space as described in the Breeding Schemes, Weaning Pups, and Number of Rats Per Cage sections, below.

ASC staff and investigators

BU ASC staff and investigators managing their own rat breeding colonies must abide by the procedures outlined in this document.

Breeding Schemes Procedures

Two different breeding schemes are acceptable. In either case, the Designated Colony Manager (research staff or BU ASC staff) is responsible for carefully monitoring pregnancies.

A. Monogamous pairs

Postpartum estrus occurs within 24 hours of parturition. Thus, if a male is left in the cage, the female is likely to become pregnant again while lactating and nursing the new litter.

• Male is removed AFTER the litter is born: One male (ONLY ONE MALE PER CAGE IS ALLOWED) and one female are housed together for mating. Nesting material is provided in the cage. The rats are not separated when the female becomes pregnant or delivers the pups. To provide more space for the pups, the male is removed at first cage change after parturition. This model takes advantage of postpartum estrus and allows the female to become pregnant and nurse at the same time. Litters are born approximately 21 days apart. The 3-week old litter must be weaned prior to the birth of the new litter.

• Male is removed BEFORE the litter is born: One male (ONLY ONE MALE PER CAGE IS ALLOWED) and one female are housed together for mating. Nesting material is provided in the cage. When the female is noticeably pregnant the male is removed from the cage.

B. Harem mating

This method houses two (2) females in a cage with one male (ONLY ONE MALE AND TWO FEMALES PER CAGE IS PERMITTED).

During routine health/breeding checks, each noticeably pregnant female is removed and placed in her own cage. When the pregnant female is separated from the harem cage, she is given nesting material in her delivery cage to make a nest for her pups. Female delivers her pups and nurses them for 21 (or up to 28 days, with BU ASC approval, following the procedure below in the Weaning Pups section). Only one nursing female and litter is allowed per cage. After the pups are weaned, the female may be returned to a harem cage.

C. Checking Pregnancy and Birth

The Designated Colony Manager has primary responsibility for checking for pregnancy and birth and for recording these events on the cage card(s). When the litter is born, the cage is flagged with a new litter card and the Date of Birth (DOB) and projected weaning date are documented. However, if BU ASC staff finds births of litters when checking and changing cages, they will place a new litter card and DOB.

D. After Birth

After pups are born, the cage is left undisturbed for at least three (3) days except for replenishing of food and water as needed. In case the bedding gets very dirty or wet and the cage must be changed sooner, the following procedure will be followed:

• The female is transferred first
• Then the litter, plus a small amount of the dirty bedding (so the smell in the new cage will be familiar) is scooped up altogether with a gloved hand and transferred to the new cage.
• The same procedure is followed until the pups start moving around the entire cage.

Required Documentation and Cage Cards

A. Cage Card Required Information

• Principal Investigator (name)
• Account # (when required)
• IACUC protocol (ID#)
• User name/Colony Manager
• Contact information
• Species
• Strain or stock and specific GEM or mutant nomenclature
• Vendor (“in-house” for animals bred in-house)
  • Number of animals and sex
  • Date of birth (DOB)
  • Date of arrival*

*Not always applicable

B. Breeding Card Required Information

• Principal Investigator (name)
• IACUC protocol (ID#)
• User
• Contact information
• Litters born and DOB

C. New Litter Card

This card is placed vertically behind the cage card, so that it is visible.

• Date of Birth (DOB)
• Projected weaning date of new litter

Weaning Pups

A. Age of Pups at Weaning

Weaning age for rat pups is routinely 21 days of age. In the case of some inbred, genetically modified or mutant strains, it may be advantageous to allow the pups to remain with the female for 28 days.

1. To extend nursing time past the 21 day standard, notify BU ASC by placing a note on the New Litter Card. The Colony Manager must communicate via email to BUASC@bu.edu or LACF-list@bu.edu when all pups in a given breeding line will be routinely weaned at 28 days.
2. User writes “Extended Weaning,” “Date to wean,” and “Dam (Mother) not pregnant” on the cage card.
3. Three-week-old litters cannot remain in the same cage as a lactating female with a newborn litter.

B. Monogamous Pairs

Assuming the lactating mother is pregnant, pups are weaned at 20-21 days of age, just before the new litter is born. This will prevent trampling of newborn pups by the weanling pups, and prevent the cage from being overcrowded.

C. Harem-Mated Females

If a singly housed lactating female is alone in a cage with her litter, weaning is less urgent than with monogamous pairs. However, rat pups are routinely weaned at 21 days of age unless an exception has been approved by the IACUC and/or BU ASC.

D. Separation of Sexes at Weaning

• Male and female pups are separated at the time of weaning, placing rats of each sex in separate cages.
• If a litter contains only one pup of a given sex, provisions must be made to house this pup with others of the same sex. Newly weaned pups must not be housed singly. Possible housing options include:
  • A single female pup may be housed with the mother.
  • A single male pup may be placed with other male pups from a different litter of the same age.
• If the parents are a monogamous pair, a single male pup may be housed with the father, both being separated out into a new cage.
• A single male pup may be housed with female sibs up to six weeks of age (i.e., adulthood).
• More than one male pup may NOT be housed with female sibs.
• It is recommended that sexing of the pups be verified one week later.

E. Feeding Weaned Pups

At the time of weaning, a small amount of chow (only a few pellets so they don’t get moldy) must be provided on the cage floor for the next seven (7) days.

Number of Rats Per Cage

Rats are social animals and because male rats rarely fight—with the notable exception of retired breeders —every effort must be made to group-house rats whenever it does not interfere with the experimental design. Post-operative rats may or may not be group-housed, again depending on their postoperative needs and the experimental design.

Cage sizes may vary in different facilities. The Colony Manager is encouraged to ask BU ASC staff for information on the size of cages in which their rats are housed.

Static rat cages with filter tops OR individually ventilated cages

A. Static rat cages with filter tops and an inside floor area of 170 square inches OR Individually ventilated cages (IVC) with filter tops and an inside floor area of 160 square inches.

• One adult female rat with one nursing litter are allowed per cage
• One retired male breeder per cage
• Eight (8) young rats up to 100 gm per cage.
• Six (6) rats weighing up to 200 gm each per cage.
• Four (4) rats weighing up to 300 g each per cage.
• Three (3) rats weighing up to 400 g each per cage.
• Two (2) rats weighing up to 500 g each per cage.
• Rats over 500 g may be singly housed, or may be pair-housed, as determined by the PI and approved by the veterinarian and IACUC.

Defining Responsibility for Separating and Weaning Rats

Designated colony manager

The Designated Colony Manager (research staff or BU ASC staff) is responsible for cage card documentation and for separating and weaning according to the above guidelines.

PI/research staff

The PI or research staff separates non-breeding experimental rats, as needed, unless technical support has been arranged with BU ASC in advance.

BU ASC Actions in Cases of Overcrowding

1. The BU ASC Staff checks for overcrowding (O/C) and pregnancy when performing daily health checks and when changing cages. Any cages that are overcrowded according to the standards defined above are marked with a Problem Notification – O/C card, dated and initialed.
2. When overcrowding is noted, the responsible individual, the Designated Colony Manager, is contacted via email and is given 48 hours to correct the problem, depending on the severity of the overcrowding
   • Note: weekends and holidays count as days and are not exempt.
3. If overcrowding is not addressed within the allotted time, BU ASC Staff separates the rats and charges the PI.
4. When two litters, one newborn and one previous litter, are in one cage, separation is performed as soon as possible. In such a case, the BU ASC Staff separates the older pups into a separate cage and provides food on the cage floor. Female and new pups are left in the breeding cage. Cage is marked with a Problem Notification – O/C card (and any other information necessary to identify the rodents), dated and initialed and Colony Manager is notified via email.
5. Any time a cage is significantly overcrowded and the welfare of the animals is at stake (Emergency O/C), the animals are promptly separated into acceptable group sizes and Colony Manager is notified via email.
6. Adding/updating of number of cages are documented on the census sheet by the Colony Manager or person who separates the rats.
7. When the overcrowding is corrected, the O/C card is removed.

References

1. The Guide for the Care and Use of Laboratory Animals. 2011. NRC ILAR. P.27. Table 2.1. Recommended Space for Commonly Used Group-Housed Laboratory Rodents.
2. BU ASC Standard Operating Procedures (SOPs).
3. UMDNJ New Jersey Medical School Comparative Medicine Resources Rodent Breeding Policy and Standard Operating Procedures (SOPs).

BU IACUC Approved October 2010, Revised January 2014, Reviewed June 2019