Oestrogen regulates the secretion of gonadotropin releasing hormone (GnRH) and this could be mediated by noradrenergic systems originating in the brainstem. Whilst it is known that noradrenergic cells possess oestrogen receptors (ER), it is not known whether ER-immunoreactive (-ir) cells in the brainstem project to the regions of the hypothalamus in which GnRH neurons are found. We have used dual-label immunocytochemistry to determine the extent to which ER-alpha is found in noradrenergic cells in the brainstem of the ovariectomized (OVX) ewe. Noradrenergic/adrenergic cells were identified by immunostaining for dopamine beta-hydroxylase (DBH). Cells that stained for both DBH and ER were found in both the A1 and A2 cell groups, with the highest levels found in the most caudal regions. In the A1 group, at the most caudal extent, 73% of ER-ir cells were DBH-positive and 19% of DBH-ir cells were ER-positive. The degree of co-localization decreased in a linear manner towards the rostral brainstem. In the caudal half of A2, 9-14% of ER-ir cells were DBH-positive and 20-25% of DBH cells were ER-positive. Less than 2% of DBH-ir cells in the A5 group were dual-labelled and none of the cells in the A6 and A7 groups were ER-positive. The retrograde tracer FluoroGold was injected into the preoptic area of nine OVX ewes and labelled cells were examined in the brainstem to determine the extent of co-localization of ER. Only injections in the rostroventral part of the medial preoptic area near to the organum vasculosum of the lamina terminalis resulted in the labelling of cells in the brainstem. One ewe with very strong labelling of the brainstem was selected for detailed mapping. In the ventrolateral medulla, half the ER-ir cells in the most caudal regions were retrogradely labelled. Almost all the ER-ir cells in the mid-region of the ventrolateral medulla were retrogradely labelled but no co-localization of retrograde tracer and ER was observed rostral to obex. There were many ER-ir cells and retrogradely-labelled cells in the nucleus of the solitary tract but only a few double-labelled cells. Similarly, numerous ER-ir cells and retrogradely labelled cells were observed around the lateral edges of the caudal fourth ventricle and across to the lateral parabrachial nucleus but there were few double-labelled cells. These results suggest differential regulation of noradrenergic cells by oestrogen, with a direct action of the hormone confined to the cells in the most caudal region of the A1 and A2 cell groups. The cells of the caudal ventrolateral medulla which contain ER-ir cells that project to the preoptic area may be important in the mediation by noradrenaline of the actions of oestrogen on GnRH secretion in the ewe.