Semen Collection Semen collection was carried out once a week for a year. Animals were trained to deliver semen on presentation of a discriminative stimulus in operant conditioning, combined with positive reinforcement. Some modifications to the training techniques and collection procedure were made on the basis of those described in previous reports (Schroeder and Keller, 1989; Robeck and O'Brien, 2004). Collection took place at poolside; the animal cooperatively floated with its ventrum above water level. A hand signal was used to instruct the animal to extrude the penis. Upon full extrusion, the tip of the penis was guided into a 500-mL collection bottle (polypropylene; Nalgene Labware; Nalge Nunc International, Rochester, New York) with an opening of 5.3 cm in diameter. The penis was squeezed gently and ejaculation occurred within seconds. After ejaculation was complete, a lid was immediately placed over the bottle and the subject was released from the position. A collection session entailed repeated presentations of a subject, under voluntary behavior, for ejaculation, and the semen collected on each presentation was considered an ejaculate. An absence of semen in spite of correct presentation and effort in response to stimulation was considered a failure to ejaculate and the occurrence of this, or micturition, signified the end of a collection session. Ejaculate samples were incubated at 37°C (Plentikool; Igloo Product Corp, Katy, Texas, or Minitub; Abfullund Labortechnik GmbH & Co, KG, Tiefenbach, Germany) and transported to a laboratory for evaluation within an hour of collection.

http://onlinelibrary.wiley.com/doi/10.2164/jandrol.108.006692/full