Recent studies have demonstrated that rather than being a collection of heterogeneously dispersed cells, the pituitary gland is wired by multiple and specific endocrine cell networks to synchronise hormone release. Prolactin (PRL) is primarily important for lactation. In response to changing physiological demands during pregnancy and lactation the pituitary has the ability to expand and contract its cell number. We have investigated changes in lactotroph morphology and cell-to-cell contacts in virgin, pregnant and lactating mice. Anterior pituitary sections from virgin, 1 week pregnant (1P), 3 week pregnant (3P) and lactating mice (n=4 per group) were immunogold labelled for PRL and examined by quantitative electron microscopy to determine lactotroph size, secretory granule characteristics and rough endoplasmic reticulum (rER) density. The identity of cell types contacting lactotrophs were also quantified. Lactotroph number was increased in lactation but cytoplasmic area was not significantly different between groups. Secretory granule density was significantly (P<0.01) increased in 1P mice but decreased in 3P (P<0.01) and the density of rER was significantly greater in lactating and 1P and 3P groups compared to virgin consistent with greater secretory activity. In virgin mice, lactotrophs made junctional contacts with all the major classes of pituitary endocrine cell types but mostly with GH, PRL and folliculo-stellate cells, and only a few ACTH, LH and TSH cells which probably reflects the relative proportion of the cell types in the pituitary. In lactation there was a significant increase (P<0.01) in the number of contacts made with other lactotrophs and a significant decrease (P<0.01) in contacts made with folliculo-stellate cells. Folliculo-stellate cells release a number of factors inhibitory to PRL release, such as annexin 1, and these data suggest that increased lactotroph connectivity coupled with a retraction of the folliculo-stellate cell network contributes to increased PRL secretion in lactation.