Chronic corticosterone administration down-regulates metabotropic glutamate receptor 5 protein expression in the rat hippocampus

Abstract

Several lines of evidence suggest a dysfunctional glutamate system in major depressive disorder (MDD). Recently, we reported reduced levels of metabotropic glutamate receptor subtype 5 (mGluR5) in postmortem brains in MDD, however the neurobiological mechanisms that induce these abnormalities are unclear. In the present study, we examined the effect of chronic corticosterone (CORT) administration on the expression of mGluR5 protein and mRNA in the rat frontal cortex and hippocampus. Rats were injected with CORT (40 mg/kg s.c.) or vehicled once daily for 21 days. The expression of mGluR5 protein and mRNA was assessed by Western blotting and quantitative real-time PCR (qPCR). In addition, mGluR1 protein was measured in the same animals. The results revealed that while there was a significant reduction (-27%, P=0.0006) in mGluR5 protein expression in the hippocampus from CORT treated rats, mRNA levels were unchanged. Also unchanged were mGluR5 mRNA and protein levels in the frontal cortex and mGluR1 protein levels in both brain regions. Our findings provide the first evidence that chronic CORT exposure regulates the expression of mGluR5 and are in line with previous postmortem and imaging studies showing reduced mGluR5 in MDD. Our findings suggest that elevated levels of glucocorticoids may contribute to impairments in glutamate neurotransmission in MDD.

Figure 1

Effects of repeated corticosterone injection (40 mg/kg for 21 days) on adrenal (A) and body weights (B). Adrenal to body weight ratio (C) was significantly lower in the CORT group (−51%) relative to control group after 21 days. These well-known effects of glucocorticoids confirm the effectiveness of the CORT dose administered. Values (circles) and mean values (horizontal lines) are presented. CORT, rats treated with corticosterone.

Figure 2

mGluR5 antibody specificity test using mGluR5 blocking peptide. Lanes 1–8 were loaded with 10 μg of total hippocampal protein. Lanes 1–4 show mGluR5 immunoreactivity (monomer at ~150 kDa and presumably dimer at ~300 kDa) detected using anti-mGluR5 antibody. In lanes 5–8 mGluR5 immunoreactivity was completely abolished after incubation with anti-mGluR5 antibody in the presence of mGluR5 blocking peptide. Lower panel shows immunoreactive actin (lanes 1–8) probed with anti-actin antibody on the same blots as a control for protein loading and transfer. For molecular weight visualization 10ul of MagicMark XP Western Protein Standard (Invitrogen; cat. no. LC5602) was loaded into lane 0.

Figure 3

Effects of repeated corticosterone injection on mGluR5 protein in the hippocampus (A) and frontal cortex (B). Representative immunoblots of mGluR5 and actin from six experimental animals (upper panels) and scatter plot of mGluR5 protein levels normalized to actin (lower panels) are shown. In the hippocampus the average mGluR5/actin ratio from rats treated with CORT (n=12) was significantly lower (−27%) than that of controls (n=12). In the frontal cortex the average mGluR5/actin ratio from rats treated with CORT (n=12) was unchanged as compared to control group (n=12). Normalized values (circles) and mean values (horizontal lines) are presented. CORT, rats treated with corticosterone.

Figure 4

Effects of repeated corticosterone injection on mGluR1 protein in the hippocampus (A) and frontal cortex (B). Representative immunoblots of mGluR1 and actin from six experimental animals (upper panels) and scatter plot of mGluR1 protein levels normalized to actin (lower panels) are shown. In both brain regions the average mGluR1/actin ratios from subjects treated with CORT (n=12) were unchanged as compared to controls (n=12). Normalized values (circles) and mean values (horizontal lines) are presented. CORT, rats treated with corticosterone.

Figure 5

Effects of repeated corticosterone on mGluR5 mRNA expression in the hippocampus (A) and frontal cortex (B). mGluR5 mRNA levels in both the hippocampus and frontal cortex were unchanged in CORT treated rats (n=12) as compared to controls (n=12). Normalized values for the individual rats (circles) and mean values (horizontal lines) are presented. CORT, rats treated with corticosterone.