Dundee University carried out experiments on more than 50,000 animals in 2013.
Figures obtained by the British Union for the Abolition of Vivisection found that UK universities experimented on more than four million animals in 2013.
Edinburgh University carried out more than 240,000 experiments that year, more than any other institution.
Dundee University carried out 51,830 experiments on animals, which was the third highest number of any Scottish university.
The vast majority, 51,393, involved mice.
Researchers also experimented on 396 rats, 28 hamsters, 10 frogs and three rabbits.
At St Andrews University, there were 2,441 experiments on animals. Again, most of these were on mice (1,617) followed by rats (313), birds (217) frogs (60) and fish (30).
Experiments were also carried out on 201 seals “in their natural habitat”. This research into seal behaviour is to aid conservation.
A Dundee University spokesman said: “Animals are used for research only in instances where no alternatives are available. The results of these studies are of great importance in better understanding and developing treatments for serious health problems including Alzheimer’s, Parkinson’s disease, cancer, heart disease, diabetes, genetic disorders and infectious diseases.
“All our work is carried out in accordance with the 3Rs concept reduction in the number of animals used, refinement of procedures so the least possible harm is done in collecting information, and the replacement of animals with other research methods wherever possible.
“Compliance with the provisions of the law is monitored closely by the university and by Home Office inspectors who make regular and unannounced visits.”