BDNF and Sprouty2 expression in the frontal cortex of rats treated with haloperidol or olanzapine.

<p>Adult rats were treated with haloperidol (2 mg/kg) or olanzapine (10 mg/kg) through drinking water for 45 days, and (A) protein and (B) mRNA levels of BDNF and Sprouty2 in frontal cortex were estimated after the treatment. BDNF protein levels were measured by ELISA and Sprouty2 proteins levels were estimated by Western blot analysis. mRNA levels of both BDNF and Sprouty2 were estimated by qRT-PCR analysis. Open bars represent haloperidol-treated rats whereas filled bars represent olanzapine-treated rats. Values are expressed as fold change relative to vehicle-treated rats. *p<0.05, n = 10–12 per group.</p

Effect of antipsychotic treatment on BDNF and sprouty2 mRNA expression

a<p>Kendall's rank correlation.</p>b<p>Spearman's correlation.</p><p>Lifetime antipsychotic dose in fluphenazine milligram equivalents.</p><p>p values are from ANOVA.</p

Correlations between BDNF or sprouty2 mRNA levels and confounding variables

a<p>Correlation value</p>b<p>significance, p value from ANOVA</p><p>Pearson's product moment correlation shown for age, PMI, brain pH, brain weight and refrigeration interval.</p><p>Kendall's rank correlation tau shown for gender, hemisphere, smoking status, age of onset, duration of illness, lifetime alcohol use and lifetime substance abuse.</p

List of primer sequences used for postmortem studies.

<p>List of primer sequences used for postmortem studies.</p

Correlation between BDNF and sprouty2 mRNA levels

a<p>Pearson's correlation.</p

Normalized BDNF mRNA expression in schizophrenia, bipolar and control subjects.

<p>Values are expressed as mean±SEM. The number of individuals per sample is indicated within each bar. BDNF expression is significantly low in schizophrenia and bipolar subjects as compared to control subjects. Level of significance as compared to control subjects is shown above each bar.</p

Chronic CORT-induced Flk1 regulation is mediated through calcium.

<p>(<i>A</i>) Calcium chelator BAPTA-AM blocked CORT (CORT)-induced reduction in Flk1 protein levels. BAPTA-AM (50 µM) was applied 30 min before CORT (1 µM) treatment to cultured neurons at DIV 5. Cell lysates were collected at 48 h after CORT treatment and Flk1 protein levels were determined by western blot analysis. CON means DMSO treatment. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in Flk1 protein levels as compared to CON. *<i>P</i><0.01 versus CON; #<i>P</i><0.01 versus CORT (Bonferroni's test). (<i>B</i>) Chronic CORT treatment increases NCS-1 protein levels in neurons. CORT (CORT; 1 µM) was applied to mouse primary cortical neurons at DIV 5. NCS-1 protein levels were determined by western blotting analysis at 48 h following CORT treatment. CON means DMSO treatment. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in NCS-1 protein levels as compared to CON. *<i>P</i><0.01 (Bonferroni's test). (C) Chronic CORT treatment increases NCS-1 protein levels in mouse frontal cortex. NCS-1 protein levels in frontal cortex of mice treated with CORT (5 mg/kg) or vehicle control (CON; 0.45% hydroxypropyl-β-cyclodextrin) for 7 weeks were determined by western blot analysis. Data represent mean±SE (<i>n</i> = 6) expressed as fold change in NCS-1 protein levels as compared to CON. β-actin is the loading control.*<i>P</i><0.05 (Bonferroni's test).</p

GR downregulation is involved in chronic CORT-induced downregulation of Flk1.

<p>(A) GR downregulation following chronic CORT exposure in neurons. CORT (CORT; 1 µM) was applied to mouse primary cortical neurons at DIV 5. GR protein levels were determined by western blotting analysis at 48 h following CORT treatment. CON means DMSO treatment. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in GR protein levels as compared to CON. *<i>P</i><0.05 (t test). (B) Chronic CORT treatment increases GR protein levels in mouse frontal cortex. GR protein levels in frontal cortex of mice treated with CORT (5 mg/kg) or vehicle control (CON; 0.45% hydroxypropyl-β-cyclodextrin) for 7 weeks were determined by western blot analysis. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in GR protein levels as compared to CON. β-actin is the loading control.*<i>P</i><0.05 (<i>t</i> test). (C) RU486 (RU, a GR antagonist) blocked CORT-induced reduction in GR protein levels. RU (1 µM) was applied 30 min before CORT (1 µM) treatment to cultured neurons at DIV5. Cell lysates were collected at 48 h after CORT treatment and GR protein levels were determined by western blot analysis. CON means DMSO treatment. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in GR protein levels as compared to CON. *<i>P</i><0.01 versus CON; #<i>P</i><0.01 versus CORT (Bonferroni's test). (D) RU486 (RU, a GR antagonist) blocked CORT-induced reduction in Flk1 protein levels. Data represent mean±SE (<i>n</i> = 5) expressed as fold change in Flk1 protein levels as compared to CON. *<i>P</i><0.01 versus CON; #<i>P</i><0.01 versus CORT (Bonferroni's test). (E) Western blot analysis of Flk1 protein expression after immunoprecipitation with GR antibody in lysates collected from DIV6 neurons. NoAb: no anti-GR antibody; total: 10% input from total cell lysates. (F) Western blot analysis of GR protein expression after immunoprecipitation with Flk1 antibody in lysates collected from DIV6 neurons. NoAb: no anti-Flk1 antibody. (G) Immunoprecipitation of Flk1 in cell lysates from corticosterone (CORT) or vehicle control (CON; DMSO) treated for 48 h. Western blotting was performed with anti-GR and anti-Flk1 antibodies. Data represent mean±SE (n = 4) expressed as fold change in GR protein levels (normalized to Flk1 protein levels) as compared to CON. *<i>P</i><0.05 versus CON (<i>t</i> test).</p

Long-term Continuous CORT treatment decreases Flk1 protein levels <i>in vitro</i> and <i>in vivo</i>.

<p>(<i>A</i>) CORT (CORT; I µM) was applied to mouse primary cortical neurons at DIV 5. Flk1 protein levels were determined by western blotting analysis at 48 hand 72 h following CORT treatment. CON means DMSO treatment. Data represent mean±SE. (<i>n</i> = 6) expressed as fold change in Flk1 protein levels as compared to CON. β-actin is the loading control. *<i>P</i><0.05 (Bonferroni's test). (<i>B</i>) Flk1 protein levels in frontal cortex of mice treated with CORT or vehicle control (CON; 0.45% hydroxypropyl-β-cyclodextrin) for 7 weeks. Data represent mean±SE (<i>n</i> = 6–8) expressed as fold change in Flk1 protein levels as compared to CON. *<i>P</i><0.01 (t test).</p

Demographic data for postmortem samples.

<p>Demographic data for postmortem samples.</p