Annexin 1 (ANXA1), a 37K protein, is a member of the super-family of Ca²⁺- and phospholipid-binding annexin proteins. In the anterior pituitary ANXA1 is expressed mainly by folliculo-stellate (FS) cells and mediates the early delayed feedback inhibition exerted by glucocorticoids on the release of ACTH and other pituitary hormones. It has been previously demonstrated that TtT/GF cells (a FS cell line) express and externalize ANXA1 in response to glucocorticoid treatment. However, ANXA1 lacks a cleavable signal sequence and externalization is not affected by inhibitors of the secretory pathway. We have previously shown that glyburide, an ATP-binding cassette (ABC) transporter inhibitor, inhibits the externalization of ANXA1 from TtT/GF cells and pituitary tissue. Here we investigated if ABCA1 is involved in ANXA1 externalization. The use of the ABCA1-transporter inhibitors geranyl-geranyl pyrophosphate and sulfobromophthalein significantly inhibited ANXA1 externalization. Partial silencing of ABCA1 expression in TtT/GF cells by siRNA also significantly decreased the amount of cell surface ANXA1. However, anterior pituitary tissue from ABCA1-null mice was found to externalize ANXA1 normally. Since compensation by other ABC family members may occur in vivo, ANXA1 externalization was studied in two transfection models: Xenopus oocytes injected with ABCA1 mRNA and AtT20 D1 corticoctroph cells co-transfected with ABCA1-GFP and ANXA1. ABCA1-expressing oocytes, but not water-injected controls, were found to externalize ANXA1. Expression of ABCA1 in AtT20 D1 cells significantly increased the amount of cell surface ANXA1 compared with mock-transfected and ANXA1-only transfected controls. Together these data provide evidence for a role of ABCA1 in ANXA1 export.