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Ethanol regulation of serum glucocorticoid kinase 1 expression in DBA2/J mouse prefrontal cortex.

Costin BN, Dever SM, Miles MF - PLoS ONE (2013)

Bottom Line: Three separate results suggested that ethanol regulated Sgk1 via circulating glucocorticoids: acute ethanol increased glucocorticoid receptor binding to the Sgk1 promoter; adrenalectomy blocked ethanol induction of Sgk1 mRNA; and chronic ethanol exposure during locomotor sensitization down-regulated HPA axis activation and Sgk1 induction by acute ethanol.Chronic ethanol modifies both SGK1 protein and HPA-mediated induction of Sgk1 mRNA.These adaptive molecular responses of glucocorticoid-responsive gene expression and SGK1 in prefrontal cortex may contribute to mechanisms underlying behavioral responses to chronic ethanol exposure.

View Article: PubMed Central - PubMed

Affiliation: Virginia Commonwealth University Alcohol Research Center, Virginia Commonwealth University, Richmond, Virginia, USA.

ABSTRACT

Background: We previously identified a group of glucocorticoid-responsive genes, including Serum Glucocorticoid kinase 1 (Sgk1), regulated by acute ethanol in prefrontal cortex of DBA2/J mice. Acute ethanol activates the hypothalamic pituitary adrenal axis (HPA) causing release of glucocorticoids. Chronic ethanol dysregulates the HPA response in both humans and rodents, possibly contributing to important interactions between stress and alcoholism. Because Sgk1 regulates ion channels and learning and memory, we hypothesized that Sgk1 contributes to HPA-dependent acute and adaptive neuronal responses to ethanol. These studies characterized acute and chronic ethanol regulation of Sgk1 mRNA and protein and their relationship with ethanol actions on the HPA axis.

Results: Acute ethanol increased Sgk1 mRNA expression in a dose and time dependent manner. Three separate results suggested that ethanol regulated Sgk1 via circulating glucocorticoids: acute ethanol increased glucocorticoid receptor binding to the Sgk1 promoter; adrenalectomy blocked ethanol induction of Sgk1 mRNA; and chronic ethanol exposure during locomotor sensitization down-regulated HPA axis activation and Sgk1 induction by acute ethanol. SGK1 protein had complex temporal responses to acute ethanol with rapid and transient increases in Ser422 phosphorylation at 15 min. following ethanol administration. This activating phosphorylation had functional consequences, as suggested by increased phosphorylation of the known SGK1 target, N-myc downstream-regulated gene 1 (NDRG1). After repeated ethanol administration during locomotor sensitization, basal SGK1 protein phosphorylation increased despite blunting of Sgk1 mRNA induction by ethanol.

Conclusions: These results suggest that HPA axis and glucocorticoid receptor signaling mediate acute ethanol induction of Sgk1 transcription in mouse prefrontal cortex. However, acute ethanol also causes complex changes in SGK1 protein expression and activity. Chronic ethanol modifies both SGK1 protein and HPA-mediated induction of Sgk1 mRNA. These adaptive molecular responses of glucocorticoid-responsive gene expression and SGK1 in prefrontal cortex may contribute to mechanisms underlying behavioral responses to chronic ethanol exposure.

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SGK1, NDRG1, and NR3C1 basal expression patterns in neurons and oligodendrocytes in the PFC of D2 mice.NDRG1, NR3C1 and SGK1 were co-localized with the neuronal marker MAP2 and the oligodendrocyte marker CNPase. DAPI staining was used as a nuclear marker (right columns). Panels show (from left to right): (a) SGK1 staining, MAP2 staining, co-localization; (b) NDRG1 staining, MAP2 staining, co-localization; (c) NR3C1 staining, MAP2 staining, co-localization; (d) SGK1 staining, CNPASE staining, co-localization; (e) NDRG1 staining, CNPASE staining, co-localization; (f) NR3C1 staining, CNPASE staining, co-localization. Scale bar equals 50 µm.
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pone-0072979-g011: SGK1, NDRG1, and NR3C1 basal expression patterns in neurons and oligodendrocytes in the PFC of D2 mice.NDRG1, NR3C1 and SGK1 were co-localized with the neuronal marker MAP2 and the oligodendrocyte marker CNPase. DAPI staining was used as a nuclear marker (right columns). Panels show (from left to right): (a) SGK1 staining, MAP2 staining, co-localization; (b) NDRG1 staining, MAP2 staining, co-localization; (c) NR3C1 staining, MAP2 staining, co-localization; (d) SGK1 staining, CNPASE staining, co-localization; (e) NDRG1 staining, CNPASE staining, co-localization; (f) NR3C1 staining, CNPASE staining, co-localization. Scale bar equals 50 µm.

Mentions: The studies on NDRG1 phosphorylation (Figure 9a-9b) suggested that Sgk1 activation by ethanol might be occurring in oligodendrocytes since NDRG1 is known to predominantly expressed in that cell type within the central nervous system. We performed immunohistochemistry to further decipher the cell type(s) in which SGK1 may respond to ethanol and transduce downstream signaling. We performed double labeling experiments using neuronal (MAP2) and oligodendrocyte (CNPase) markers in addition to NR3C1, NDRG1 and SGK1 labeling to determine the cell types where NR3C1, SGK1 and NDRG1 are expressed basally in the PFC of D2 mice. We found that SGK1 and NR3C1 are expressed in both neurons and oligodendrocytes in the PFC of D2 mice (Figure 11a,c,d,f). Whereas, NDRG1 is only expressed in oligodendrocytes (Figure 11b,e). Neurons, cells positive for MAP2, were also positive for SGK1 (Figure 11a) and NR3C1 (Figure 11c), but not NDRG1 (Figure 11b) at least as detectable by immunofluorescence. Oligodendrocytes, cells positive for CNPase, were also positive for SGK1 (Figure 11d), NDRG1 (Figure 11e) and NR3C1 (Figure 11f). These results suggest that SGK1 could regulate acute responses to ethanol in both neurons and oligodendrocytes through glucocorticoid signaling mechanisms. Additionally, NDRG1 signaling might have a role as a downstream mediator of SGK1 responses to acute ethanol in oligodendrocytes.


Ethanol regulation of serum glucocorticoid kinase 1 expression in DBA2/J mouse prefrontal cortex.

Costin BN, Dever SM, Miles MF - PLoS ONE (2013)

SGK1, NDRG1, and NR3C1 basal expression patterns in neurons and oligodendrocytes in the PFC of D2 mice.NDRG1, NR3C1 and SGK1 were co-localized with the neuronal marker MAP2 and the oligodendrocyte marker CNPase. DAPI staining was used as a nuclear marker (right columns). Panels show (from left to right): (a) SGK1 staining, MAP2 staining, co-localization; (b) NDRG1 staining, MAP2 staining, co-localization; (c) NR3C1 staining, MAP2 staining, co-localization; (d) SGK1 staining, CNPASE staining, co-localization; (e) NDRG1 staining, CNPASE staining, co-localization; (f) NR3C1 staining, CNPASE staining, co-localization. Scale bar equals 50 µm.
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getmorefigures.php?uid=PMC3750005&req=5

pone-0072979-g011: SGK1, NDRG1, and NR3C1 basal expression patterns in neurons and oligodendrocytes in the PFC of D2 mice.NDRG1, NR3C1 and SGK1 were co-localized with the neuronal marker MAP2 and the oligodendrocyte marker CNPase. DAPI staining was used as a nuclear marker (right columns). Panels show (from left to right): (a) SGK1 staining, MAP2 staining, co-localization; (b) NDRG1 staining, MAP2 staining, co-localization; (c) NR3C1 staining, MAP2 staining, co-localization; (d) SGK1 staining, CNPASE staining, co-localization; (e) NDRG1 staining, CNPASE staining, co-localization; (f) NR3C1 staining, CNPASE staining, co-localization. Scale bar equals 50 µm.
Mentions: The studies on NDRG1 phosphorylation (Figure 9a-9b) suggested that Sgk1 activation by ethanol might be occurring in oligodendrocytes since NDRG1 is known to predominantly expressed in that cell type within the central nervous system. We performed immunohistochemistry to further decipher the cell type(s) in which SGK1 may respond to ethanol and transduce downstream signaling. We performed double labeling experiments using neuronal (MAP2) and oligodendrocyte (CNPase) markers in addition to NR3C1, NDRG1 and SGK1 labeling to determine the cell types where NR3C1, SGK1 and NDRG1 are expressed basally in the PFC of D2 mice. We found that SGK1 and NR3C1 are expressed in both neurons and oligodendrocytes in the PFC of D2 mice (Figure 11a,c,d,f). Whereas, NDRG1 is only expressed in oligodendrocytes (Figure 11b,e). Neurons, cells positive for MAP2, were also positive for SGK1 (Figure 11a) and NR3C1 (Figure 11c), but not NDRG1 (Figure 11b) at least as detectable by immunofluorescence. Oligodendrocytes, cells positive for CNPase, were also positive for SGK1 (Figure 11d), NDRG1 (Figure 11e) and NR3C1 (Figure 11f). These results suggest that SGK1 could regulate acute responses to ethanol in both neurons and oligodendrocytes through glucocorticoid signaling mechanisms. Additionally, NDRG1 signaling might have a role as a downstream mediator of SGK1 responses to acute ethanol in oligodendrocytes.

Bottom Line: Three separate results suggested that ethanol regulated Sgk1 via circulating glucocorticoids: acute ethanol increased glucocorticoid receptor binding to the Sgk1 promoter; adrenalectomy blocked ethanol induction of Sgk1 mRNA; and chronic ethanol exposure during locomotor sensitization down-regulated HPA axis activation and Sgk1 induction by acute ethanol.Chronic ethanol modifies both SGK1 protein and HPA-mediated induction of Sgk1 mRNA.These adaptive molecular responses of glucocorticoid-responsive gene expression and SGK1 in prefrontal cortex may contribute to mechanisms underlying behavioral responses to chronic ethanol exposure.

View Article: PubMed Central - PubMed

Affiliation: Virginia Commonwealth University Alcohol Research Center, Virginia Commonwealth University, Richmond, Virginia, USA.

ABSTRACT

Background: We previously identified a group of glucocorticoid-responsive genes, including Serum Glucocorticoid kinase 1 (Sgk1), regulated by acute ethanol in prefrontal cortex of DBA2/J mice. Acute ethanol activates the hypothalamic pituitary adrenal axis (HPA) causing release of glucocorticoids. Chronic ethanol dysregulates the HPA response in both humans and rodents, possibly contributing to important interactions between stress and alcoholism. Because Sgk1 regulates ion channels and learning and memory, we hypothesized that Sgk1 contributes to HPA-dependent acute and adaptive neuronal responses to ethanol. These studies characterized acute and chronic ethanol regulation of Sgk1 mRNA and protein and their relationship with ethanol actions on the HPA axis.

Results: Acute ethanol increased Sgk1 mRNA expression in a dose and time dependent manner. Three separate results suggested that ethanol regulated Sgk1 via circulating glucocorticoids: acute ethanol increased glucocorticoid receptor binding to the Sgk1 promoter; adrenalectomy blocked ethanol induction of Sgk1 mRNA; and chronic ethanol exposure during locomotor sensitization down-regulated HPA axis activation and Sgk1 induction by acute ethanol. SGK1 protein had complex temporal responses to acute ethanol with rapid and transient increases in Ser422 phosphorylation at 15 min. following ethanol administration. This activating phosphorylation had functional consequences, as suggested by increased phosphorylation of the known SGK1 target, N-myc downstream-regulated gene 1 (NDRG1). After repeated ethanol administration during locomotor sensitization, basal SGK1 protein phosphorylation increased despite blunting of Sgk1 mRNA induction by ethanol.

Conclusions: These results suggest that HPA axis and glucocorticoid receptor signaling mediate acute ethanol induction of Sgk1 transcription in mouse prefrontal cortex. However, acute ethanol also causes complex changes in SGK1 protein expression and activity. Chronic ethanol modifies both SGK1 protein and HPA-mediated induction of Sgk1 mRNA. These adaptive molecular responses of glucocorticoid-responsive gene expression and SGK1 in prefrontal cortex may contribute to mechanisms underlying behavioral responses to chronic ethanol exposure.

Show MeSH
Related in: MedlinePlus