Leydig cell functional capacity reflects the numbers and differentiation status of the steroidogenic Leydig cells in the testes and becomes more or less fixed in early adulthood with the final establishment of the hypothalamo-pituitary-gonadal (HPG) axis after puberty. Factors influencing Leydig cell functional capacity and its role in puberty are poorly understood. Using a bovine model of dairy bulls fed four different nutritional regimes from 1 month to 12 months, and applying circulating Insulin-like peptide 3 (INSL3) as an accurate biomarker of Leydig cell functional capacity, showed that a high plane of nutrition in the first 6 months of life, but not later, significantly increased INSL3 in young adulthood. Moreover, INSL3 concentration at 4 months indicated a marked differential in early feeding regime and correlated well (negatively) with the timing of puberty, as reflected by the age in days for the first production of an ejaculate with >50 million sperm and >10% forward motility, as well as with testis size at 18 months. Reversing the diet at 6 months was unable to rectify the trend in either parameter, unlike for other parameters such as testosterone, body weight, and scrotal circumference. This study has shown that early prepubertal nutrition is a key factor in the development of Leydig cell functional capacity in early adulthood and appears to be a key driver in the dynamic progression of puberty.