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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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pSGK1 following chronic saline and ethanol administration.Western blot analysis of pS422 SGK1 15 minutes following chronic saline (SS), acute ethanol (2g/kg) (SE), chronic ethanol (EE) or acute saline (ES) treatment. Panels show: (a) Acute and sensitized locomotor response (cm/10 min.) following saline (SS, ES) or ethanol (EE, SE) administration. * p < 0.05 versus SS and ES groups, # p < 0.05 versus SE, ES and SS groups, (b) Quantification of pS422 SGK1, * p < 0.05 versus SS and EE groups, # p < 0.05 versus SS and EE groups, c) Quantification of SGK1, and (d) Representative Western blot.
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pone-0072979-g010: pSGK1 following chronic saline and ethanol administration.Western blot analysis of pS422 SGK1 15 minutes following chronic saline (SS), acute ethanol (2g/kg) (SE), chronic ethanol (EE) or acute saline (ES) treatment. Panels show: (a) Acute and sensitized locomotor response (cm/10 min.) following saline (SS, ES) or ethanol (EE, SE) administration. * p < 0.05 versus SS and ES groups, # p < 0.05 versus SE, ES and SS groups, (b) Quantification of pS422 SGK1, * p < 0.05 versus SS and EE groups, # p < 0.05 versus SS and EE groups, c) Quantification of SGK1, and (d) Representative Western blot.

Mentions: We have shown that Sgk1, Fkbp5 and possibly other glucocorticoid responsive genes were no longer regulated following chronic ethanol administration (Figure 3a–d). However, due to the disparate responses of Sgk1 mRNA and protein levels (Figure 8a-b) with acute ethanol, we hypothesized that SGK1 protein levels or phosphorylation might show compensatory changes to the blunted HPA axis. We therefore evaluated SGK1 pS422 levels following chronic ethanol sensitization studies. There again was overall effect of treatment on locomotor activity (one-way ANOVA, F3,27= 45.87 p < 0.01) (Figure 10a). Post-hoc analysis revealed that ethanol-ethanol (EE) treated animals showed significantly greater locomotor activity compared to saline-ethanol (SE), saline-saline (SS) and ethanol-saline (ES) treated animals. In addition, SE treated animals showed greater locomotor activity than SS and ES treated animals. In evaluating SGK1 pS422 levels in these animals, there was an overall significant effect of treatment on SGK1 pS422 levels (one-way ANOVA, F3,21= 8.75, p < 0.01) (Figure 10b,d). Post-hoc analysis showed that SGK1 pS422 levels were significantly increased in animals treated acutely with ethanol compared to SS and EE treated animals. Interestingly, SGK1 pS422 levels were also significantly elevated in ES treated animals compared to SS and EE treated animals. SGK1 pS422 did not differ between SS and EE treated animals. Total SGK1 levels did not differ between SS, SE, EE and ES treated animals in any group 15 minutes following acute ethanol administration (Figure 10c-d).


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

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

pSGK1 following chronic saline and ethanol administration.Western blot analysis of pS422 SGK1 15 minutes following chronic saline (SS), acute ethanol (2g/kg) (SE), chronic ethanol (EE) or acute saline (ES) treatment. Panels show: (a) Acute and sensitized locomotor response (cm/10 min.) following saline (SS, ES) or ethanol (EE, SE) administration. * p < 0.05 versus SS and ES groups, # p < 0.05 versus SE, ES and SS groups, (b) Quantification of pS422 SGK1, * p < 0.05 versus SS and EE groups, # p < 0.05 versus SS and EE groups, c) Quantification of SGK1, and (d) Representative Western blot.
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getmorefigures.php?uid=PMC3750005&req=5

pone-0072979-g010: pSGK1 following chronic saline and ethanol administration.Western blot analysis of pS422 SGK1 15 minutes following chronic saline (SS), acute ethanol (2g/kg) (SE), chronic ethanol (EE) or acute saline (ES) treatment. Panels show: (a) Acute and sensitized locomotor response (cm/10 min.) following saline (SS, ES) or ethanol (EE, SE) administration. * p < 0.05 versus SS and ES groups, # p < 0.05 versus SE, ES and SS groups, (b) Quantification of pS422 SGK1, * p < 0.05 versus SS and EE groups, # p < 0.05 versus SS and EE groups, c) Quantification of SGK1, and (d) Representative Western blot.
Mentions: We have shown that Sgk1, Fkbp5 and possibly other glucocorticoid responsive genes were no longer regulated following chronic ethanol administration (Figure 3a–d). However, due to the disparate responses of Sgk1 mRNA and protein levels (Figure 8a-b) with acute ethanol, we hypothesized that SGK1 protein levels or phosphorylation might show compensatory changes to the blunted HPA axis. We therefore evaluated SGK1 pS422 levels following chronic ethanol sensitization studies. There again was overall effect of treatment on locomotor activity (one-way ANOVA, F3,27= 45.87 p < 0.01) (Figure 10a). Post-hoc analysis revealed that ethanol-ethanol (EE) treated animals showed significantly greater locomotor activity compared to saline-ethanol (SE), saline-saline (SS) and ethanol-saline (ES) treated animals. In addition, SE treated animals showed greater locomotor activity than SS and ES treated animals. In evaluating SGK1 pS422 levels in these animals, there was an overall significant effect of treatment on SGK1 pS422 levels (one-way ANOVA, F3,21= 8.75, p < 0.01) (Figure 10b,d). Post-hoc analysis showed that SGK1 pS422 levels were significantly increased in animals treated acutely with ethanol compared to SS and EE treated animals. Interestingly, SGK1 pS422 levels were also significantly elevated in ES treated animals compared to SS and EE treated animals. SGK1 pS422 did not differ between SS and EE treated animals. Total SGK1 levels did not differ between SS, SE, EE and ES treated animals in any group 15 minutes following acute ethanol administration (Figure 10c-d).

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