Competition between the Brain and Testes under Selenium-Compromised Conditions: Insight into Sex Differences in Selenium Metabolism and Risk of Neurodevelopmental Disease

Abstract

Selenium (Se) is essential for both brain development and male fertility. Male mice lacking two key genes involved in Se metabolism (Scly(-/-)Sepp1(-/-) mice), selenoprotein P (Sepp1) and Sec lyase (Scly), develop severe neurological dysfunction, neurodegeneration, and audiogenic seizures that manifest beginning in early adulthood. We demonstrate that prepubescent castration of Scly(-/-)Sepp1(-/-) mice prevents behavioral deficits, attenuates neurodegeneration, rescues maturation of GABAergic inhibition, and increases brain selenoprotein levels. Moreover, castration also yields similar neuroprotective benefits to Sepp1(-/-) and wild-type mice challenged with Se-deficient diets. Our data show that, under Se-compromised conditions, the brain and testes compete for Se utilization, with concomitant effects on neurodevelopment and neurodegeneration.

Significance statement: Selenium is an essential trace element that promotes male fertility and brain function. Herein, we report that prepubescent castration provides neuroprotection by increasing selenium-dependent antioxidant activity in the brain, revealing a competition between the brain and testes for selenium utilization. These findings provide novel insight into the interaction of sex and oxidative stress upon the developing brain and have potentially significant implications for the prevention of neurodevelopmental disorders characterized by aberrant excitatory/inhibitory balance, such as schizophrenia and epilepsy.

Keywords: neurodegeneration; neurodevelopment; oxidative stress; parvalbumin interneurons; selenium; sex differences.

Figure 1.

Redox imbalance and age-dependent decrease of Gad67 and PGC-1 expression in the IC of Scly^−/−Sepp1^−/− mice. A, Images of IC Gad67 immunohistochemistry from male WT, Sepp1^−/−, and Scly^−/−Sepp1^−/− mice at 4 and 8 weeks of age. B, Mean (±SEM) normalized Gad67 optical density relative to WT controls. C, Images of IC PGC-1 immunohistochemistry from male WT, Sepp1^−/−, and Scly^−/−Sepp1^−/− mice. D, Mean (±SEM) density of PGC-1 cells per mm³. E, Images of IC GFAP immunohistochemistry from male WT, Sepp1^−/−, and Scly^−/−Sepp1^−/− mice. F, Mean (±SEM) GFAP optical density (n = 3 or 4 per group per time point). *p < 0.05 compared with WT. **p < 0.01 compared with WT. ^#p < 0.05 compared with Sepp1^−/−. ^##p < 0.01 compared with Sepp1^−/−. Scale bar, 100 μm.

Figure 2.

ApoER2 is preferentially expressed on metabolically active PV interneurons that comprise major sites of selenoprotein synthesis. A, Images of ApoER2 (green), PGC-1 (red), and PV (blue) in the IC and retrosplenial cortex. B, Images of ApoER2 (green), selenophosphate synthetase 2 (SPS2; red), and PV (blue) in the IC and retrosplenial cortex. Yellow arrows indicate triple-labeled neurons in merged images. Scale bar, 100 μm.

Figure 3.

Impaired maturation of IC PV interneurons in Scly^−/−Sepp1^−/− mice. A, Immunofluorescent images of PV (red) and WFA (green) from the IC of WT, Sepp1^−/−, and Scly^−/−Sepp1^−/− mice at 4 weeks. Yellow arrows indicate WFA-labeled PV interneurons in merged images. B, Images taken at 8 weeks. C, Mean (±SEM) density of PV interneurons per mm³. D, Mean (±SEM) density of WFA-labeled PV interneurons per mm³ (n = 3 per genotype for each time point). *p < 0.05 compared with WT. ^#p < 0.05 compared with Sepp1^−/−. Scale bar, 100 μm.

Figure 4.

Castration of juvenile Scly^−/−Sepp1^−/− mice promotes survival, prevents behavioral deficits, and attenuates neurodegeneration. A, Experimental timeline. B, Survival curve of sham (n = 31) and castrated (n = 20) Scly^−/−Sepp1^−/− mice. C, Mean (±SEM) latency to fall off rotorod (sham: n = 7; castrated: n = 17). D, Mean (±SEM) distance traveled during the open field test (sham: n = 9; castrated: n = 13). E, Mean (±SEM) speed before 85 dB white noise and in the 10 s period after start of 85 dB white noise (sham: n = 5; castrated: n = 16). F, Images of cFos immunohistochemistry from sham and castrated Scly^−/−Sepp1^−/− mice. G, Mean (±SEM) density of cFos-positive cells per mm³ (n = 6 per group). H, Images of silver-stained brain sections from sham and castrated Scly^−/−Sepp1^−/− mice containing the XSCP (left) and the IC (right). I, Mean (±SEM) optical density of silver staining (n = 3 per group). CIC, Central nucleus of inferior colliculus; DG, dentate gyrus; ECIC, external cortex of inferior colliculus; MGN, medial geniculate nucleus. *p < 0.05. **p < 0.01. Scale bar, 200 μm.

Figure 5.

Castration attenuates oxidative stress and promotes maturation of GABAergic inhibition in the IC of Scly^−/−Sepp1^−/− mice. A, Images of immunohistochemistry for Gad67, PGC-1, and GFAP in the IC of sham and castrated Scly^−/−Sepp1^−/− mice. B, Mean (±SEM) normalized Gad67 optical density relative to sham Scly^−/−Sepp1^−/− mice (sham: n = 4; castrated: n = 5). C, Mean (±SEM) density of PGC-1 cells per mm³ (sham: n = 3; castrated: n = 4). D, Mean (±SEM) GFAP optical density (sham: n = 5; castrated: n = 7). E, Immunofluorescent images of PV (red) and WFA (green) from the IC of sham and castrated Scly^−/−Sepp1^−/− mice. Yellow arrows indicate WFA-labeled PV interneurons in merged images. F, Mean (±SEM) density of PV interneurons per mm³. G, Mean (±SEM) density of WFA-labeled PV interneurons per mm³ (sham: n = 4; castrated: n = 5). *p < 0.05. Scale bar, 100 μm.

Figure 6.

Castration increases brain selenoprotein and Se levels in Scly^−/−Sepp1^−/− mice. A, Western blot showing levels of GPx1, GPx4, SelM, PGC-1, and α-tubulin in brainstem and whole-brain samples of sham and castrated Scly^−/−Sepp1^−/− mice. B, Mean (±SEM) normalized protein levels in brainstem relative to sham Scly^−/−Sepp1^−/− samples (n = 4 per group). C, Mean (±SEM) normalized protein levels in whole brain (n = 4 per group). D, Mean (±SEM) normalized GPx activity in brainstem (n = 8 per group) and whole brain (n = 4 or 5 per group) samples relative to sham Scly^−/−Sepp1^−/− samples. E, Mean (±SEM) whole brain Se levels in male (n = 4), castrated (n = 5), and female (n = 4) Scly^−/−Sepp1^−/− mice. F, Mean (±SEM) whole brain Se levels in male WT (n = 4), Sepp1^−/− (n = 6), and Scly^−/−Sepp1^−/− (n = 4) mice. G, Mean (±SEM) testes Se levels in male WT (n = 4), Sepp1^−/− (n = 6), and Scly^−/−Sepp1^−/− (n = 4) mice. H, Mean (±SEM) testosterone levels in male WT (n = 5), Sepp1^−/− (n = 5), and Scly^−/−Sepp1^−/− (n = 4) mice. *p < 0.05 compared with male Scly^−/−Sepp1^−/−. ***p < 0.001 compared with male Scly^−/−Sepp1^−/−. ^%%p < 0.01 compared with castrated Scly^−/−Sepp1^−/−. ^##p < 0.01 compared with male WT. ^###p < 0.001 compared with male WT. ^$$p < 0.01 compared with male Sepp1^−/−.

Figure 7.

Castration delays neurological impairment and neurodegeneration in Sepp1^−/− mice challenged with a Se-deficient diet. A, Experimental timeline. B, Mean (±SEM) latency to fall off rotorod (n = 10 per group). C, Mean (±SEM) speed before 85 dB white noise and in the 10 s period immediately after start of 85 dB white noise (n = 10 per group). D, Images of silver-stained brain sections containing the XSCP (left) and the IC (right). E, Mean (±SEM) optical density of silver staining in sham (n = 5) and castrated (n = 4) Sepp1^−/− mice. F, Mean (±SEM) normalized GPx activity in brainstem and whole-brain samples (n = 6 per group). CIC, Central nucleus of inferior colliculus; ECIC, external cortex of inferior colliculus. *p < 0.05. **p < 0.01. Scale bar, 200 μm.

Figure 8.

Castration provides neuroprotection to wild-type mice challenged with periadolescent MK-801 exposure and a Se-deficient diet. A, Experimental timeline. B, Mean (±SEM) latency to fall off rotorod. C, Mean (±SEM) latency to locate the escape tunnel during Barnes maze training. D, Mean (±SEM) number of incorrect holes checked before locating the escape tunnel during training. E, Mean (±SEM) latency to locate the escape tunnel during trial block 3. F, Mean (±SEM) number of incorrect holes checked before locating the escape tunnel during trial block 3 (n = 10 per group). G, Mean (±SEM) normalized GPx activity in samples from cortex, hippocampus, and brainstem (n = 4 per group). H, Immunofluorescent images of PV (red) and WFA (green) from the retrosplenial cortex. Yellow arrows indicate WFA-labeled PV interneurons in merged images. I, Mean (±SEM) density of PV interneurons per mm³. J, Mean (±SEM) density of WFA-labeled PV interneurons per mm³. *p < 0.05 compared with MK-801 group. ^#p < 0.05 compared with Control group. Scale bar, 100 μm.