Directly supporting the overall validity of a PcG-dependent repressive mechanism holding in check the initiation of puberty is the pubertal delay observed when the decline in hypothalamic Eed expression that occurs during normal puberty is prevented via targeted lentivirus-mediated gene delivery. Over-expression of Eed in the ARC of the hypothalamus, which contains the KNDy neurons required for pulsatile GnRH release 34 reduced the number of neurons expressing detectable levels of immunoreactive kisspeptin, the content of immunoreactive kisspeptin per cell, and the abundance of Kiss1 mRNA in the ARC. Importantly, it reduced pulsatile GnRH release, delayed puberty, and disrupted estrous cyclicity. Although the animals receiving lentiviral particles carrying the EED gene were still able to ovulate, the estrus cycle profiles displayed by these animals suggested that they were ovulating sporadically. This inference is supported by the finding that these animals exposed to a fertile male delivered an average of 2 pups as compared with 12 pups delivered by rats receiving either a control virus or a virus expressing EED but targeted outside the ARC.
