Region-specific mechanisms for testosterone-induced Fos in hamster brain

Abstract

Hamsters self-administer androgens. Previously, we determined that testosterone (T) activates select steroid- and opiate-sensitive brain regions. Is T-stimulated neuronal activation androgenic? Thirty-five castrated males with physiologic T replacement (n=7/group) were pre-treated with the androgen antagonist flutamide (15 mg/kg sc) or ethanol (0.25 ml) and infused into the lateral ventricle (ICV) for 4 h with 40 microg T (TF and TE, respectively) or 40 microl vehicle (VF and VE). To determine if androgens and opiates activate overlapping brain areas, 7 additional males received 20 mug morphine sulfate ICV following ethanol injection (ME). Immediately after ICV infusion, animals were perfused. Sixty-micrometer coronal brain slices were stained for Fos. Fos-positive neurons were counted in a 0.3-mm(2) area from 5 regions previously shown to express T-induced Fos: the posteromedial bed nucleus of the stria terminalis (BSTPM), posteromedial amygdala (MeP), lateral habenula (LHb), ventral tegmental area, and lateral pontine nucleus. T induced Fos in all areas reported previously (TE vs. VE, p<0.05), except LHb (p>0.05). Morphine induced Fos in all 5 brain regions (ME vs. VE, p<0.05), indicating that androgens and opiates activate overlapping brain regions. Flutamide alone did not induce Fos (VF vs. VE, p>0.05). Moreover, flutamide treatment blocked T-induced Fos expression only in the steroid-sensitive BSTPM, suggesting that androgens mediate neuronal activation in this area (mean+/-SEM: TF: 68.4+/-13.2 vs. TE: 137.9+/-17.6, p<0.05). The absence of flutamide effects on T-induced Fos in the steroid-sensitive MeP (TE: 210.6+/-50.0 vs. TF: 215.3+/-28.2, p>0.05) suggests that distinct mechanisms activate Fos in individual androgen-responsive nuclei.

Figure 1

Photomicrographs of the posterial medial amygdala showing Fos protein expression in representative male hamsters after 4h of 40 μg ICV testosterone (1.0 μg/μl, top) or 40 μl vehicle (bottom) with pre-treatment of ethanol (left) or flutamide (right). Abbreviations: TE: Testosterone+Ethanol; TF: Testosterone+Flutamide; VE: Vehicle+Ethanol; VF: Vehicle+Flutamide. Scale bar =200 um.

Figure 2

Photomicrographs of the bed nucleus of stria terminalis showing Fos protein expression in representative male hamsters after 4h of 40 μg ICV testosterone (1.0 μg/μl, top) or 40 μl vehicle (bottom) with pre-treatment of ethanol (left) or flutamide (right). Abbreviations: TE: Testosterone+Ethanol; TF: Testosterone+Flutamide; VE: Vehicle+Ethanol; VF: Vehicle+Flutamide. Scale bar =200 um.

Figure 3

Mean±SEM Fos-immunoreactive neurons after 4h ICV testosterone (black) or vehicle (white) infusion in male hamsters (n=7/group). 30 min prior to infusions, males received either ethanol or flutamide (shaded). Abbreviations: BSTPM: posteromedial bed nucleus of the stria terminalis; LHb: lateral habenula; MeP: posterior medial amygdala; Pn: lateral pontine nucleus; TE: Testosterone+Ethanol; TF: Testosterone+Flutamide; VTA: ventral tegmental area; VE: Vehicle+Ethanol; VF: Vehicle+Flutamide. Bars with common letter subscripts are not significantly different.

Figure 4

Photomicrograph of the posterial medial amygdala showing Fos protein expression in a representative male hamster after 4 h of 20 μg ICV morphine sulfate (0.5.0 μg/μl). Males received a subcutaneous ethanol injection 30 min prior to morphine infusion. Scale bar =200 um.