Apparently 32 non-human animals (not including microbes or fruit flies) were on the final Space Shuttle trip back from the Space Station. Air & Space Magazine posts about NASA's animal research in space. Above is an astrofrog on Spacelab in 1992.
NASA's animal research policies have come a long way since its days of shooting monkeys into space on brand new, failure-prone launch vehicles. Back then the agency mostly relied on federal law, such as the 1966 Federal Animal Welfare Act, to govern its animal care, until it started to develop more specialized policies in the 1980s. Finally in 1996, it codified a formal rule called the NASA Principles for Ethical Care and Use of Animals. Even more recently, Alex Dunlap, NASA's chief veterinarian, and others drew up an agreement for use on the station; the International Animal Welfare Agreement for Space Borne Research establishes "a base line level of animal care and use in space that all countries could agree on."
NASA's use of animal astronauts has changed along with the culture, according to Dunlap. "We've become more compassionate with time, more aware of seeing and making sure that animals get humanely treated." The agency is unlikely ever to return to the days of flying monkeys and chimpanzees. Other than humans, mice are the highest-order animal currently being sent into space. They provide the best balance of sample size (more tissue and bone structure to study) and cage logistics: their small cages are easier to store in a cramped cabin and to provide with ample air circulation.
