u-0126 and Hypertension

Spontaneously hypertensive rats (SHRs) have increased daily or induced sodium intake compared to normotensive rats. In normotensive rats, angiotensin II (ANG II)-induced sodium intake is blocked by the inactivation of p42/44 mitogen-activated protein kinase, also known as extracellular signal-regulated protein kinase1/2 (ERK1/2). Here we investigated if inhibition of ERK1/2 pathway centrally would change sodium appetite and intracerebroventricular (icv) ANG II-induced pressor response in SHRs. SHRs (280-330 g, n = 07-14/group) with stainless steel cannulas implanted in the lateral ventricle (LV) were used. Water and 0.3 M NaCl intake was induced by the treatment with the diuretic furosemide + captopril (angiotensin converting enzyme blocker) subcutaneously or 24 h of water deprivation (WD) followed by 2 h of partial rehydration with only water (PR). The blockade of ERK1/2 activation with icv injections of U0126 (MEK1/2 inhibitor, 2 mM; 2 μl) reduced 0.3 M NaCl intake induced by furosemide + captopril (5.0 ± 1.0, vs. vehicle: 7.3 ± 0.7 mL/120 min) or WD-PR (4.6 ± 1.3, vs. vehicle: 10.3 ± 1.4 mL/120 min). PEP7 (selective inhibitor of AT1 receptor-mediated ERK1/2 activation, 2 nmol/2 μL) icv also reduced WD-PR-induced 0.3 M NaCl (2.8 ± 0.7, vs. vehicle: 6.8 ± 1.4 mL/120 min). WD-PR-induced water intake was also reduced by U0126 or PEP7. In addition, U0126 or PEP7 icv reduced the pressor response to icv ANG II. Therefore, the present results suggest that central AT1 receptor-mediated ERK1/2 activation is part of the mechanisms involved in sodium appetite and ANG II-induced pressor response in SHRs.

Topics: Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Animals; Appetite; Butadienes; Captopril; Disease Models, Animal; Furosemide; Humans; Hypertension; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nitriles; Rats; Rats, Inbred SHR; Receptor, Angiotensin, Type 1; Sodium

Arteries from hypertensive animals and humans have increased spontaneous tone. Increased superoxide anion (superoxide) contributes to elevated blood pressure (BP) and spontaneous tone in hypertension. The association between the extracellular signaling-regulated kinase 1/2 (ERK1/2)-mitogen-activated protein kinase (MAPK) signaling pathway and generation of superoxide and spontaneous tone in isolated aorta was studied in angiotensin II (ANG II)-infused hypertensive (HT) rats. Systolic BP, phosphorylation of ERK, aortic superoxide formation, and aortic spontaneous tone were compared in sham normotensive and HT rats. Infusion of ANG II (0.5 mg x kg(-1) x day(-1) for 6 days) significantly elevated the systolic BP (P<0.01). The phosphorylation of ERK1/2 vs. total ERK1/2 in thoracic aorta was enhanced, and superoxide was increased in the HT vs. the sham group (P<0.01). Spontaneous tone developed in the HT group, but not in the normotensive group. MAPK/ERK1/2 (MEK1/2)-ERK1/2 signaling pathway inhibitors, PD-98059 (10 micromol/l), and U-0126 (10 micromol/l), significantly reduced the phosphorylation of ERK1/2, superoxide generation (P<0.01), and spontaneous tone (P<0.01) in HT. These findings suggest that ANG II infusion induces the production of superoxide and spontaneous tone and that both are dependent on ERK-MAPK activation. In endothelium-denuded aorta, however, MEK1/2 inhibitors did not inhibit the spontaneous tone, even though they significantly reduced superoxide generation similar to endothelium-intact aorta. These data suggest that inhibition of ERK1/2 signaling pathway, via PD-98059 or U-0126, may regulate spontaneous tone in an endothelium-dependent manner. In conclusion, these findings support the importance of the ERK1/2 signaling pathway in modulating vascular oxidative stress and subsequently mediating spontaneous tone in HT.

Topics: Angiotensin II; Animals; Aorta, Thoracic; Blood Pressure; Butadienes; Disease Models, Animal; Endothelium, Vascular; Enzyme Activation; Flavonoids; Hypertension; Male; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Nitriles; Oxidative Stress; Phosphorylation; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Superoxides; Vasoconstriction

The epithelial sodium channel (ENaC) is expressed in a variety of tissues, including the renal collecting duct, where it constitutes the rate-limiting step for sodium reabsorption. Liddle's syndrome is caused by gain-of-function mutations in the beta and gamma subunits of ENaC, resulting in enhanced Na reabsorption and hypertension. Epidermal growth factor (EGF) causes acute inhibition of Na absorption in collecting duct principal cells via an extracellular signal-regulated kinase (ERK)-dependent mechanism. In experiments with primary cultures of collecting duct cells derived from a mouse model of Liddle's disease (beta-ENaC truncation), it was found that EGF inhibited short-circuit current (Isc) by 24 +/- 5% in wild-type cells but only by 6 +/- 3% in homozygous mutant cells. In order to elucidate the role of specific regions of the beta-ENaC C terminus, Madin-Darby canine kidney (MDCK) cell lines that express beta-ENaC with mutation of the PY motif (P616L), the ERK phosphorylation site (T613A), and C terminus truncation (R564stop) were created using the Phoenix retroviral system. All three mutants exhibited significant attenuation of the EGF-induced inhibition of sodium current. In MDCK cells with wild-type beta-ENaC, EGF-induced inhibition of Isc (<30 min) was fully reversed by exposure to an ERK kinase inhibitor and occurred with no change in ENaC surface expression, indicative of an effect on channel open probability (P(o)). At later times (>30 min), EGF-induced inhibition of Isc was not reversed by an ERK kinase inhibitor and was accompanied by a decrease in ENaC surface expression. Our results are consistent with an ERK-mediated decrease in ENaC open probability and enhanced retrieval of sodium channels from the apical membrane.

Topics: Amiloride; Amino Acid Motifs; Animals; Butadienes; Cell Line; Disease Models, Animal; Dogs; Down-Regulation; Epidermal Growth Factor; Epithelial Sodium Channel Blockers; Epithelial Sodium Channels; Hypertension; Kidney; Membrane Potentials; Mice; Mice, Transgenic; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mutation; Nitriles; Phosphorylation; Protein Kinase Inhibitors; Protein Structure, Tertiary; Protein Transport; Sodium; Sodium Channel Blockers; Syndrome; Transfection

Angiotensin II (Ang II) has been reported to inhibit insulin signaling at multiple levels in vascular smooth muscle cells (VSMC) in vitro. We have demonstrated that VSMC from spontaneously hypertensive rats (SHR) produce Ang II in a homogeneous culture.. In the current study, we investigated influences of endogenous Ang II on insulin signaling in VSMC from SHR.. Phosphatidylinositol 3-kinase (PI3-kinase) activity, insulin receptor substrate-1 (IRS-1) associated tyrosine phosphorylation, and p85 subunit of PI3-kinase were measured in VSMC from SHR and normotensive Wistar-Kyoto (WKY) rats in the absence and presence of Ang II type 1 receptor antagonist RNH6270 and mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) inhibitor U0126.. Insulin treatment increased PI3-kinase activity in VSMC from WKY rats in a dose-dependent manner. In contrast, insulin treatment of VSMC from SHR did not affect PI3-kinase activity. However, co-treatment of VSMC from SHR with RNH6270 and insulin, increased PI3-kinase activity. PI3-kinase activity, IRS-1-associated tyrosine phosphorylation and p85 subunit of PI3-kinase in VSMC from WKY rats decreased in response to treatment with Ang II and returned to control levels upon co-treatment with U0126. Basal levels of PI3-kinase activity, IRS-1-associated tyrosine phosphorylation, and p85 subunit of PI3-kinase were significantly lower in VSMC from SHR than in cells from WKY rats. U0126 treatment of VSMC from SHR significantly increased levels of PI3-kinase activity, IRS-1-associated tyrosine phosphorylation, and p85 subunit of PI3-kinase.. These results indicate that endogenous Ang II suppresses insulin signaling in VSMC from SHR by activating extracellular signal-regulated kinase. These findings suggest that tissue Ang II may play a role in insulin resistance in hypertension.

Topics: Angiotensin II; Animals; Butadienes; Dose-Response Relationship, Drug; Enzyme Inhibitors; Hypertension; Insulin; Male; Mitogen-Activated Protein Kinases; Muscle, Smooth, Vascular; Nitriles; Phosphatidylinositol 3-Kinases; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Signal Transduction