Estrogens play a salient role in the development and maintenance of both male and female nervous systems and behaviors. The plainfin midshipman (Porichthys notatus), a teleost fish, has two male reproductive morphs that follow alternative mating tactics and diverge in multiple somatic, hormonal, and neural traits, including the central control of morph-specific vocal behaviors. After we identified duplicate estrogen receptors (ERβ1 and ERβ2) in midshipman, we developed antibodies to localize protein expression in the central vocal–acoustic networks and saccule, the auditory division of the inner ear. As in other teleost species, ERβ1 and ERβ2 were robustly expressed in the telencephalon and hypothalamus in vocal–acoustic and other brain regions shown previously to exhibit strong expression of ERα and aromatase (estrogen synthetase, CYP19) in midshipman. Like aromatase, ERβ1 label colocalized with glial fibrillary acidic protein (GFAP) in telencephalic radial glial cells. Quantitative polymerase chain reaction revealed similar patterns of transcript abundance across reproductive morphs for ERβ1, ERβ2, ERα, and aromatase in the forebrain and saccule. In contrast, transcript abundance for ERs and aromatase varied significantly between morphs in and around the sexually polymorphic vocal motor nucleus (VMN). Together, the results suggest that VMN is the major estrogen target within the estrogen-sensitive hindbrain vocal network that directly determines the duration, frequency, and amplitude of morph-specific vocalizations. Comparable regional differences in steroid receptor abundances likely regulate morph-specific behaviors in males and females of other species exhibiting alternative reproductive tactics. J. Comp. Neurol. 521:2850–2869, 2013. © 2013 Wiley Periodicals, Inc.