Journal of Chemical Neuroanatomy 23 (2002) 59 – 71 Distribution of aromatase immunoreactivity in the forebrain of red-sided garter snakes at the beginning of the winter dormancy Randolph W. Krohmer a, *, Gerald J. Bieganski a , Daniel D. Baleckaitis a , Nobuhiro Harada b , Jacques Balthazart c a Department of Biology, Saint Xaier Uniersity, 3700 West 103rd Street, Chicago, IL 60655, USA b Molecular genetics, School of Medicine, Fujita Health Uniersity, Toyoake, Aichi 470 -11, Japan c Center for Cellular and Molecular Neurobiology, Research Group in Behaioral Neuroendocrinology, Uniersity of Liege (Bat. L1), 17 place Delcour, B-4020 Liege, Belgium Received 17 November 2000; received in revised form 27 April 2001; accepted 9 September 2001 Abstract Until recently, it has been difficult to identify the exact location of aromatase containing cells in the brain. The development of new antibodies has provided a sensitive tool to analyze the distribution of aromatase immunoreactive (ARO-ir) material at a cellular level of resolution. In the present study we examined, for the first time, the distribution of ARO-ir cells in the brain of a reptile, the red-sided garter snake, at the beginning of the winter dormancy. ARO-ir cells were found at all rostro-caudal levels in the red-sided garter snake brain. Although weakly stained cells were distributed throughout the brain, more intensely immunoreactive cells were primarily concentrated in the preoptic area, anterior hypothalamus, septum and nucleus sphericus. Although androgens are elevated upon emergence from hibernation in the male red-sided garter snake, initiation of courtship behavior appears to be independent of direct androgen control. To date, the only known stimulus found to initiate courtship is a period of low temperature dormancy followed by exposure to warm temperatures. Circumstantial data, however, suggest an indirect role in the activation of male copulatory behavior for estrogenic metabolites of testosterone produced in the brain by aromatization during the winter dormancy. This study provides the first documentation of the distribution of ARO-ir cells in a reptilian species and demonstrates that while the aromatase enzyme occurs in most regions of the brain, the ARO-ir cells that appear to contain the highest concentration of enzyme are clustered in brain areas classically associated with the control of courtship behavior and mating in vertebrates. These data are consistent with the idea that estrogens locally produced in the brain may participate in some way to the activation of sexual behavior in this species also. This notion should now be experimentally tested by analyzing annual changes in aromatase activity and immunoreactivity and assessing the effects of pharmacological blockade of the enzyme activity at different times of the year. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Aromatase; Immunocytochemistry; Neural aromatase; Red-sided garter snake; Thamnophis sirtalis parietalis www.elsevier.com/locate/jchemneu Abbreiations: III, third ventricle; Acc, nucleus accumbens; aDVR, anterior dorsal ventricular ridge; AHPOA, anterior hypothalamus-preoptic area; AC, anterior commissure; AOB, accessory olfactory bulb; AOT, accessory olfactory tract; Bst, bed nucleus of the stria terminalis; d, dorsal cortex; dm, dorsomedial nucleus of thalamus; ep, external plexiform layer; gl, glomerular layer; Hy, hypothalamus; ig, internal granular layer; ip, internal plexiform layer; l, lateral cortex; LFB, lateral forebrain bundle; LV, lateral ventricle; m, medial cortex; ME, median eminence; mit, mitral cell layer; MOB, main olfactory bulb; nLOT, nucleus of the lateral olfactory tract; MOT, main olfactory tract; NS, nucleus sphericus; Nsa, anterior septal nucleus; Nsl, lateral septal nucleus; Nsm, medial septal nucleus; oc, optic chiasm; optr, optic tract; ov, olfactory ventricle; ot, olfactory tubercle; pDVR, posterior dorsal ventricular ridge; POA, preoptic area; ra, rostral amygdaloid nucleus; sm, stria medullaris; sr, rostral septal nucleus; str, striatum; v, ventral cortex; va, ventral amygdaloid nucleus; val, ventral amygdaloid nucleus, lateral portion; vam, ventral amygdaloid nucleus, medial portion; vm, ventromedial nucleus of the hypothalamus; vn, vomeronasal nerve. * Corresponding author. Fax: +1-773-779-9061. E-mail address: rwkrohmer@hotmail.com (R.W. Krohmer). 0891-0618/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved. PII:S0891-0618(01)00145-4