Relapse susceptibility in women with substance use disorders has been linked to the estrogen, 17β-estradiol (E2). Our previous findings in female rats suggest that the influence of E2 on cocaine seeking can be localized to the prelimbic prefrontal cortex (PrL-PFC). Here we investigated the receptor mechanisms through which E2 regulates the reinstatement of extinguished cocaine seeking. Sexually mature female rats underwent intravenous cocaine self-administration (0.5 mg/inf; 14 x 2 hrs daily) and extinction, and then were ovariectomized prior to reinstatement testing. E2 (10 µg/kg, ip) alone did not reinstate cocaine seeking, but it potentiated reinstatement when combined with an otherwise subthreshold priming dose of cocaine. A similar effect was observed following intra-PrL-PFC micro-infusions of E2 and by systemic or intra-PrL-PFC administration of the estrogen receptor-β (ERβ) agonist, DPN, but not agonists at ERα or the G-protein coupled estrogen receptor (GPER1). By contrast, E2-potentiated reinstate...