Sulpiride-induced effect on rat hormonal state.

<p>C; controls treated with normal saline; SULP: sulpiride (dopamine D₂-antagonist); B[a]P: benzo[a]pyrene; OIL: olive oil; T3: triiodothyronin expressed in ng/dl; T4: thyroxin expressed in μg/dl; TSH: thyroid-stimulating hormone expressed in ng/ml; GH: growth hormone expressed in ng/ml; PRL: prolactin expressed in ng/ml; Corticosterone expressed in mg/ml; Insulin expressed in pg/ml. Values are expressed as mean ± SE (n = 10). The asterisks indicate the significance of the differences between SULP-treated rats and controls, and between B[a]P-exposed rats with and without concomitant treatment with SULP</p><p>*P<0.05</p><p>**P<0.005</p><p>***P<0.001</p><p>Sulpiride-induced effect on rat hormonal state.</p

<i>In vivo</i> assessment of the effect of D₂-receptor blockade on the activation of GH/STAT5b signalling pathway.

<p>Western blotting showing the SULP-mediated suppression of STAT5b phosphorylation. Numbers in the western blot captures indicate the STAT5b phoshorylation level following treatment compared to the control level that was set at 1. Lanes C: control; SULP: sulpiride (selective dopamine D₂-antagonist); B[a]P: benzo[a]pyrene; OIL: olive oil.</p

The dopamine D₂-receptor-mediated control of the insulin/PI3K/AKT/FOXO1 signalling pathway activation.

<p>Dopamine stimulates D₂-ARs on pancreatic β-cells and restricts the release of insulin in response to increased plasma glucose levels [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128708#pone.0128708.ref061" target="_blank">61</a>]. In contrast, blockade of D₂-dopaminergic receptors by sulpiride, increases insulin release [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128708#pone.0128708.ref022" target="_blank">22</a>], which in turn, stimulates insulin receptors (IR) in hepatocyte plasma membranes, an effect resulting in the phosphorylation of the Insulin Receptor Substrate (IRS) at different docking sites, where the phosphatidylinositol 3-kinase (PI3K) binds. Activated PI3K converts phosphatidylinositol biphosphate to phosphatidylinositol triphosphate, which subsequently activates protein kinase B (AKT). Upon activation AKT phosphorylates the transcription factor forkhead box O1 (FOXO1), which then translocates into the cytoplasm thus terminating <i>CYP1A1</i>, <i>CYP1A2</i> and <i>CYP1B1</i> gene transcription [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128708#pone.0128708.ref022" target="_blank">22</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128708#pone.0128708.ref075" target="_blank">75</a>].</p

D₂-dopaminergic receptor-mediated effect on hepatic total P450 content and CYP1B1.

<p>(A) Assessment of sulpiride effect on total P450 content in the liver of rats treated with either normal saline or benzo[a]pyrene. (B) Assessment of sulpiride effect on hepatic CYP1B1 mRNA expression following treatment with either normal saline or benzo[a]pyrene. Bonferroni’s correction and Tukey post-hoc tests took place in the comparisons of the data presented here. (C vs SULP, C vs B[a]P and B[a]P vs (B[a]P+SULP)). C: controls treated with normal saline; SULP: sulpiride (dopamine D₂-antagonist); B[a]P: benzo[a]pyrene; OIL: olive oil; *P<0.05, **P<0.01, #P<0.005, ***P<0.001.</p

Sulpiride-induced effect on hypothalamic catecholamines, norepinephrine (NE) and dopamine (DA).

<p>The evaluation of the effect of sulpiride (SULP), a dopamine D₂-antagonist, on hypothalamic dopaminergic activity was based on the alterations observed at DA turnover ratio (HVA+DOPAC)/DA. DA, NE, DOPAC and HVA levels were expressed in nmoles/g tissue. HVA: homovanilic acid; DOPAC: dihydroxyphenylacetic acid. Values are expressed as mean ± SE (n = 10). The asterisks indicate the significance of the differences between SULP-treated rats and controls, and between benzo[a]pyrene (B[a]P)-exposed rats with and without concomitant treatment with SULP;</p><p>*P< 0.05</p><p>**P< 0.01</p><p>***P< 0.001</p><p>Sulpiride-induced effect on hypothalamic catecholamines, norepinephrine (NE) and dopamine (DA).</p

D₂-dopaminergic receptor-mediated regulation of hepatic CYP1A1.

<p>Assessments of the effects induced by the selective dopamine D₂-antagonist sulpiride (SULP) on <i>CYP1A1</i> relative mRNA expression by using quantitative PCR assays, on CYP1A1 apoprotein levels by using Western blotting, and on CYP1A1-catalyzed EROD activity by using a fluorometric assay. Bonferroni’s correction and Tukey post-hoc tests took place in the comparisons of the data presented here (C vs SULP, C vs B[a]P and B[a]P vs (B[a]P+SULP)). Numbers in the western blot captures, correspond to the lower lanes and indicate the relative CYP1A apoprotein expression following treatment compared to the control expression level that was set at 1. The antibody used against CYP1A1 is not highly specific and recognizes CYP1A2 as well. C: controls treated with normal saline; SULP: sulpiride (dopamine D₂-antagonist); B[a]P: benzo[a]pyrene; OIL: olive oil; *P<0.05, **P<0.01, ***P<0.001.</p

Dopamine D₂-Receptor Antagonists Down-Regulate CYP1A1/2 and CYP1B1 in the Rat Liver

<div><p>Dopaminergic systems regulate the release of several hormones including growth hormone (GH), thyroid hormones, insulin, glucocorticoids and prolactin (PRL) that play significant roles in the regulation of various Cytochrome P450 (CYP) enzymes. The present study investigated the role of dopamine D₂-receptor-linked pathways in the regulation of CYP1A1, CYP1A2 and CYP1B1 that belong to a battery of genes controlled by the Aryl Hydrocarbon Receptor (AhR) and play a crucial role in the metabolism and toxicity of numerous environmental toxicants. Inhibition of dopamine D₂-receptors with sulpiride (SULP) significantly repressed the constitutive and benzo[a]pyrene (B[a]P)-induced <i>CYP1A1</i>, <i>CYP1A2</i> and <i>CYP1B</i> expression in the rat liver. The expression of <i>AhR</i>, heat shock protein 90 (<i>HSP90)</i> and AhR nuclear translocator <i>(ARNT)</i> was suppressed by SULP in B[a]P-treated livers, whereas the <i>AhRR</i> expression was increased by the drug suggesting that the SULP-mediated repression of the CYP1 inducibility is due to inactivation of the AhR regulatory system. At signal transduction level, the D₂-mediated down-regulation of constitutive <i>CYP1A1/2</i> and <i>CYP1B1</i> expression appears to be mediated by activation of the insulin/PI3K/AKT pathway. PRL-linked pathways exerting a negative control on various CYPs, and inactivation of the glucocorticoid-linked pathways that positively control the AhR-regulated CYP1 <i>genes</i>, may also participate in the SULP-mediated repression of both, the constitutive and induced <i>CYP1</i> expression. The present findings indicate that drugs acting as D₂-dopamine receptor antagonists can modify several hormone systems that regulate the expression of <i>CYP1A1</i>, <i>CYP1A2</i> and <i>CYP1B1</i>, and may affect the toxicity and carcinogenicity outcome of numerous toxicants and pre-carcinogenic substances. Therefore, these drugs could be considered as a part of the strategy to reduce the risk of exposure to environmental pollutants and pre-carcinogens.</p></div

<i>In vivo</i> assessment of the effect of D₂-receptor blockade on the insulin/PI3K/AKT/FOXO1 and mTOR signalling pathway.

<p>Western blotting showing the SULP-mediated AKT phosphorylation that activated FOXO1 in the nucleus, which in turn translocated into the cytoplasm (reduced FOXO1β phosphorylation in the nucleus and increased FOXO1β phosphorylation in the cytoplasm). Numbers in the western blot captures indicate the relative FOXO1β, AKT and mTOR phosphorylation level following treatment compared to the control level that was set at 1. C: controls treated with normal saline; SULP: sulpiride (selective dopamine D₂-antagonist); B[a]P: benzo[a]pyrene; OIL: olive oil.</p