Some of Berthold's previous publications were specifically linked to glands. Scientists up until the 19th century had little idea how glands affected the body. Castrati - men who were castrated before puberty - were kept in harems to ensure females remained chaste; from the middle ages boys were castrated to keep youthful-sounding singing voices due to the lack of an Adam's apple, but in adulthood they also kept a straight hairline, grew large chests and unusually long limbs. Whole-body effects were similarly clear in animals: castrated cock chickens (capons) did not develop the typical male secondary sexual characteristics, namely aggression, crowing, muscle development, sexual proclivity and most visibly the cockscomb and wattle; they were docile and developed tender flesh, which was considered a delicacy.
Upon autopsy, he found that the transplanted testes had grown a new vasculature but no other bodily connection: this finally demonstrated that whatever was controlling the secondary characteristics was transported from the testes via the bloodstream. The prevailing theory, that sexual characteristics were mediated via the nervous system, was thus disproved.
Despite this ground-breaking result, Berthold abandoned completely any further developmental work in this field and his contemporaries also showed little interest. The non-reductionist philosophy prevalent amongst physiologists at Göttingen might have coloured their thinking; Rudolph Wagner repeated the experiment without success, and perhaps this also had an effect. No major progress was made until five decades later with some dramatic successes in the treatment of thyroid-based illnesses by Victor Horsley and, soon after, his student George Murray