The cows were fed 5 kg of grass hay and 0.5 kg of concentrate twice daily and had free access to tap water.
One intravenous catheter (UK catheter kit, 14 G, 30 cm, Unitika Ltd, Osaka, Japan) was inserted into each jugular vein 1 d prior to the ACTH infusion to alleviate the stress associated with the infusion and blood collection.
An extension tube (TOP Ltd, Tokyo, Japan) was connected to the catheter for blood collection and fixed to the halter.