Expression patterns of estrogen receptors in the central auditory system change in prepubertal and aged mice

Abstract

Estrogens are important in the development, maintenance and physiology of the CNS. Several studies have shown their effects on the processing of hearing in both males and females, and these effects, in part, are thought to result from regulation of the transcription of genes via their classical estrogen receptor (ER) pathway. In order to understand the spatiotemporal changes that occur with age, we have studied the expression of ERs in the central auditory pathway in prepubertal and aged CBA mice with immunohistochemistry. In prepubertal mice a clear dichotomy was noted between the expression of ERα and ERβ. ERβ-positive neurons were found in the metencephalon whereas the majority of ERα was found in mesencephalon, diencephalon or the telencephalon. In the aged animals a different pattern of ER expression was found in terms of location and overall intensity. These age-induced changes in the expression pattern were generally not uniform, suggesting that region-specific mechanisms regulate the ERs' age-related expression. Neither the prepubertal nor the aged animals showed sex differences in any auditory structure. Our results demonstrate different age-dependent spatial and temporal changes in the pattern of expression of ERα and ERβ, suggesting that each ER type may be involved in distinct roles across the central auditory pathway in different periods of maturation.

Fig. 1

Mean and standard deviation of ABR thresholds for the aged mice (3 females and 3 males, 26–28 mon old). Thresholds at 3 and 48 kHz were beyond the limits of the equipment. Thresholds between 6 and 32 kHz ranged from 45 to 74 dB SPL. The hearing losses varied from moderately-severe to profound with some individuals having greater hearing losses at both low and high frequencies.

Fig. 2

Representative western blot showing the specificity of the antibodies detecting ERα and ERβ expression in whole cell extracts of mouse cochlear nucleus (CN), inferior colliculus (IC) and ovary (Ov). ERα was detected in CN (lane 1), IC (lane 2) and ovary (lane 3); ERβ was also found in CN (lane 4), IC (lane 5) and ovary (lane 6). Incubation of blots with either anti-ERα or anti-ERβ IgG detected the specific bands of 66 kDa in lanes 1–3 or 51 kDa in lanes 4–6 respectively. GAPDH (at 37 kDa in lanes 1–6) was used as a loading control. The molecular weight marker is seen on the left. 20µg of protein were loaded in each lane.

Fig. 3

Estrogen receptor α (ERα) immunocytochemistry in prepubertal mice: (a) the auditory nerve nucleus (AuN) and the CN showing the dorsal cochlear nucleus (DCN), the anterior ventral cochlear nucleus (VCA), the posterior ventral cochlear nucleus (VCP). Higher magnification (20×) images showing the (b) AuN, (c) VCA and VCP, and (d) DCN with the granule cells in the periphery (scale bar=50 µm).

Fig. 4

ERα immunocytochemistry throughout the central auditory pathway of prepubertal mice: (a) nucleus of the trapezoid body (Tz), (b) lateral superior olive (LSO), (c) dorsal nucleus of lateral lemniscus (DLL), (d) ventral nucleus of lateral lemniscus (VLL), (e) medial geniculate nucleus (MGN), (f) auditory cortex (AuC), (g) dorsal nucleus of inferior colliculus (DCIC), (h) central nucleus of inferior colliculus (CIC), (i) external nucleus of inferior colliculus (ECIC) and brachium of inferior colliculus (BIC) (scale bar=50 µm).

Fig. 5

Estrogen receptor β (ERβ) immunocytochemistry in prepubertal mice: (a) the auditory nerve nucleus (AuN) and the CN showing the dorsal cochlear nucleus (DCN), the anterior ventral cochlear nucleus (VCA), the posterior ventral cochlear nucleus (VCP). Higher magnification (20×) images showing the (b) AuN, (c) VCA and VCP, and (d) DCN (scale bar=50 µm).

Fig. 6

ERβ immunocytochemistry throughout the central auditory pathway of prepubertal mice: (a) nucleus of the trapezoid body (Tz), (b) lateral superior olive (LSO), (c) dorsal nucleus of lateral lemniscus (DLL), (d) ventral nucleus of lateral lemniscus (VLL), (e) medial geniculate nucleus (MGN), (f) auditory cortex (AuC), (g) dorsal nucleus of inferior colliculus (DCIC), (h) central nucleus of inferior colliculus (CIC), (i) external nucleus of inferior colliculus (ECIC) and brachium of inferior colliculus (BIC) (scale bar=50 µm).

Fig. 7

ERα immunocytochemistry in aged mice: (a) the auditory nerve nucleus (AuN) and the CN showing the dorsal cochlear nucleus (DCN), the anterior ventral cochlear nucleus (VCA), the posterior ventral cochlear nucleus (VCP). Higher magnification (20×) images showing the (b) A, (c) VCP, (d) VCA, and (e) DCN (scale bar=50 µm).

Fig. 8

ERα immunocytochemistry throughout the central auditory pathway of aged mice: (a) nucleus of the trapezoid body (Tz), (b) lateral superior olive (LSO), (c) dorsal nucleus of lateral lemniscus (DLL), (d) ventral nucleus of lateral lemniscus (VLL), (e) medial geniculate nucleus (MGN), (f) auditory cortex (AuC), (g) dorsal nucleus of inferior colliculus (DCIC), (h) central nucleus of inferior colliculus (CIC), (i) external nucleus of inferior colliculus (ECIC) and brachium of inferior colliculus (BIC) (scale bar=50 µm).

Fig. 9

ERβ immunocytochemistry in aged mice: (a) the auditory nerve nucleus (AuN) and the CN showing the dorsal cochlear nucleus (DCN), the anterior ventral cochlear nucleus (VCA), the posterior ventral cochlear nucleus (VCP). Higher magnification (20×) images showing the (b) AuN, (c) VCP, (d) VCA, and (e) DCN (scale bar=50 µm).

Fig. 10

ERβ immunocytochemistry throughout the central auditory pathway of aged mice: (a) nucleus of the trapezoid body (Tz), (b) lateral superior olive (LSO), (c) dorsal nucleus of lateral lemniscus (DLL), (d) ventral nucleus of lateral lemniscus (VLL), (e) medial geniculate nucleus (MGN), (f) auditory cortex (AuC), (g) dorsal nucleus of inferior colliculus (DCIC), (h) central nucleus of inferior colliculus (CIC), (i) external nucleus of inferior colliculus (ECIC) and brachium of inferior colliculus (BIC) (scale bar=50 µm).