Changes in plasma cortisol concentrations during the first 4 hours after castration of 6-week-old lambs by one of four methods were measured. The methods were application of a rubber ring above the testes, application of a ring to shorten the scrotum below the testes, use of a castration clamp plus a ring, and use of a castration clamp alone. The efficacy of local anaesthetic in reducing or abolishing the cortisol responses to castration by all these methods except short scrotum was assessed. Local anaesthetic was injected into the neck of the scrotum, both spermatic cords, the scrotal neck plus spermatic cords, or into both testes. The combined clamp plus ring method was used to test whether or not clamp damage to afferent nerves from the testes would reduce the cortisol response to ring castration. The short scrotum method elicited a significantly lower cortisol response than that caused by ring castration. Injection of local anaesthetic into both spermatic cords marginally reduced the cortisol response to ring castration, but the response was virtually abolished by prior scrotal neck, or scrotal neck plus spermatic cords, or intra-testicular local anaesthetic injections. The clamp plus ring method, where each spermatic cord and the associated scrotal tissue were crushed separately for 1, 5 or 10 seconds with no overlap between the two crush lines, did not elicit lower cortisol responses than the ring-only method. Local anaesthetic injected into the scrotal neck virtually abolished, and spermatic cord injections markedly reduced, the cortisol responses to combined clamp plus ring castration. The IO-second clamp application caused a more protracted cortisol response than did ring-only castration, whereas the cortisol responses to 1-second clamp application and to ring-only castration were similar. Local anaesthetic injected into the scrotal neck or spermatic cords did not affect significantly the cortisol response to the IO-second clamp application. The following conclusions were drawn from this work: the testes as well as the scrotum were sources of noxious sensory input after ring application; the clamp plus ring method used here was ineffective in reducing the pain-induced distress (as indicated by cortisol concentrations) caused by ring-only castration; injection of local anaesthetic into the scrotal neck or into the testes prior to ring application blocked most noxious sensory input from both the scrotum and the testes; and on the basis of the present cortisol responses the 10-second clamp applications method could not be recommended in reference to the ring-only method, but the l-second clamp application method might be equally acceptable for castrating 6-week-old-lambs. Of the methods examined, injections of local anaesthetic into the scrotal neck or both testes were the most effective in reducing the pain-induced distress caused by ring-only castration.