saralasin and candesartan

The proximal tubule (PT) is major site for the reabsorption of filtered HCO(3)(-). Previous work on the rabbit PT showed that 1) increases in basolateral (BL) CO(2) concentration ([CO(2)](BL)) raise the HCO(3)(-) reabsorption rate (J(HCO(3))), and 2) the increase that luminal angiotensin II (ANG II) produces in J(HCO(3)) is greatest at 0% [CO(2)](BL) and falls to nearly zero at 20%. Here, we investigate the role of angiotensin receptors in the [CO(2)](BL) dependence of J(HCO(3)) in isolated perfused PTs. We found that, in rabbit S2 PT segments, luminal 10(-8) M saralasin (peptide antagonist of ANG II receptors), lowers baseline J(HCO(3)) (5% CO(2)) to the value normally seen at 0% in the absence of inhibitors and eliminates the J(HCO(3)) response to changes in [CO(2)](BL). However, basolateral 10(-8) M saralasin has no effect. As with saralasin, luminal 10(-8) M candesartan (AT(1) antagonist) reduces baseline J(HCO(3)) and eliminates the [CO(2)](BL) dependence of J(HCO(3)). Luminal 10(-7) M PD 123319 (AT(2) antagonist) has no effect. Finally, we compared PTs from wild-type and AT(1A)-null mice of the same genetic background. Knocking out AT(1A) modestly lowers baseline J(HCO(3)) and, like luminal saralasin or candesartan in rabbits, eliminates the J(HCO(3)) response to changes in [CO(2)](BL). Our accumulated evidence suggests that ANG II endogenous to the PT binds to the apical AT(1A) receptor and that this interaction is critical for both baseline J(HCO(3)) and its response to changes in [CO(2)](BL). Neither apical AT(2) receptors nor basolateral ANG II receptors are involved in these processes.

Topics: Angiotensin II Type 1 Receptor Blockers; Angiotensin-Converting Enzyme Inhibitors; Animals; Benzimidazoles; Bicarbonates; Biphenyl Compounds; Carbon Dioxide; Data Interpretation, Statistical; Female; Hydrogen-Ion Concentration; In Vitro Techniques; Kidney Tubules, Proximal; Mice; Mice, Knockout; Perfusion; Rabbits; Receptor, Angiotensin, Type 1; Saralasin; Signal Transduction; Tetrazoles

We demonstrated that angiotensin II (Ang II, 10-1000 nM) induced proliferation of cultured rabbit gingival fibroblasts in a concentration-dependent manner. The Ang II-induced proliferation was inhibited by CV-11974 (AT1 antagonist; 1 microM) and saralasin (AT1/AT2 antagonist; 1 microM), but not by PD123,319 (AT2 antagonist; 1 microM), suggesting that Ang II-induced proliferation was mediated via AT1 receptors present in and/or on gingival fibroblasts. The results of Western blot analysis indicated the presence of AT1 and AT2 receptors in/on the fibroblasts. In a subsequent radioligand binding assay, the binding of [3H]Ang II to the fibroblasts was specific and saturable with both high- and low-affinity sites. Competition binding experiments indicated that Ang II completely displaced [3H]Ang II binding, and CV-11974 and PD123,319 maximally displaced up to approximately 63% and 37% of the total binding, respectively. Ang II and CV-11974 completely displaced the [3H]DuP753 binding but PD123,319 did not, indicating a single population of binding site. These findings demonstrate that gingival fibroblasts contain both AT1 and AT2 receptor subtypes for Ang II, and support that Ang II stimulation of AT1 receptors results in proliferation of the fibroblasts.

Topics: Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Angiotensin Receptor Antagonists; Animals; Benzimidazoles; Binding, Competitive; Biphenyl Compounds; Blotting, Western; Cell Division; Cells, Cultured; Dose-Response Relationship, Drug; Fibroblasts; Gingiva; Kinetics; Losartan; Rabbits; Radioligand Assay; Receptors, Angiotensin; Saralasin; Tetrazoles; Tritium

Rodents are the unique species carrying duplicated angiotensin (Ang) type 1 (AT1) receptor genes, Agtr1a and Agtr1b. After separately generating Agtr1a and Agtr1b null mutant mice by gene targeting, we produced double mutant mice homozygous for both Agtr1a and Agtr1b null mutation (Agtr1a-/-; Agtr1b-/-) by mating the single gene mutants. Agtr1a-/-, Agtr1b-/- mice are characterized by normal in utero survival but decreased ex utero survival rate. After birth they are characterized by low body weight gain, marked hypotension, and abnormal kidney morphology including delayed maturity in glomerular growth, hypoplastic papilla, and renal arterial hypertrophy. These abnormal phenotypes are quantitatively similar to those found in mutant mice homozygous for the angiotensinogen gene (Agt-/-), indicating that major biological functions of endogenous Ang elucidated by the abnormal phenotypes of Agt-/- are mediated by the AT1 receptors. Infusion of Ang II, AT1 blockers, or an AT2 blocker was without effect on blood pressure in Agtr1a-/-; Agtr1b-/- mice, indicating that AT2 receptor does not exert acute depressor effects in these mice lacking AT1 receptors. Also, unlike Agt-/- mice, some Agtr1a-/-; Agtr1b-/- mice have a large ventricular septum defect, suggesting that another receptor such as AT2 is functionally activated in Agtr1a-/-, Agtr1b-/- mice.

Topics: Adrenal Glands; Anesthetics; Angiotensin II; Angiotensinogen; Animals; Benzimidazoles; beta-Galactosidase; Biphenyl Compounds; Blood Pressure; Imidazoles; Infusions, Intravenous; Kidney; Losartan; Mice; Mice, Knockout; Myocardium; Phenotype; Pyridines; Receptor, Angiotensin, Type 1; Receptors, Angiotensin; Saralasin; Staining and Labeling; Tetrazoles; Thiopental; Zygote

The present study was undertaken to investigate the role of angiotensin II (Ang II) in ovulation and ovarian steroidogenesis and prostaglandin (PG) production via the Ang II receptors in rabbit ovaries. In in vitro perfused rabbit ovaries, PD123319, a selective nonpeptide antagonist for AT2 receptors, reduced the Ang II-induced ovulation in a dose-dependent manner, whereas CV-11974, a selective nonpeptide antagonist for AT1 receptor, did not affect the Ang II-induced ovulation. Ang II also significantly stimulated the meiotic maturation of ovulated ova and follicular oocytes in the absence of gonadotropin. The addition of PD123319 at 10 (-6) M to the perfusate significantly inhibited the Ang II-induced oocyte maturation. Ang II did not stimulate the production of progesterone by perfused rabbit ovaries but significantly stimulated the production of estradiol (E2) and PGs. When PD123319 at 10(-6) M was added to the perfusate 30 min before the onset of Ang II administration, the Ang II-stimulated production of E2 and PGs was significantly blocked. Saralasin, a peptide analog of Ang II, inhibited the specific binding of [125I] iodo-[Sar1, Ile8] Ang II to rabbit ovarian membranes in a concentration-dependent manner, yielding an inhibitory constant (IC50) value of 1.58 x 10(-9) M. PD123319 and CV-11974 also inhibited the binding of [125I]iodo-[Sar1, Ile8] Ang II; however, PD123319 and CV-11974 were 15 and 40 times less potent than saralasin, respectively. Autoradiographic study revealed that an intense localization of Ang II receptors in the rabbit ovaries was present in the granulosa cell layers and the stroma of the preovulatory follicles. AT2 receptors were predominantly located in granulosa cells, whereas AT1 receptors were more concentrated in the stroma and thecal cell layers. In summary, Ang II induced ovulation and oocyte maturation and stimulated the production of E2 and PG by perfused rabbit ovary in vitro via the AT2 receptor. Thus, locally produced Ang II may be part of a novel intraovarian paracrine or autocrine control mechanism during the ovulatory process.

Topics: 1-Sarcosine-8-Isoleucine Angiotensin II; Angiotensin II; Angiotensin Receptor Antagonists; Animals; Autoradiography; Benzimidazoles; Biphenyl Compounds; Cellular Senescence; Female; Imidazoles; Oocytes; Ovary; Ovulation Induction; Pyridines; Rabbits; Receptors, Angiotensin; Saralasin; Tetrazoles

We have previously shown that mechanical stress induces activation of protein kinases and increases in specific gene expression and protein synthesis in cardiac myocytes, all of which are similar to those evoked by humoral factors such as growth factors and hormones. Many lines of evidence have suggested that angiotensin II (Ang II) plays a vital role in cardiac hypertrophy, and it has been reported that secretion of Ang II from cultured cardiac myocytes was induced by mechanical stretch. To examine the role of Ang II in mechanical stress-induced cardiac hypertrophy, we stretched neonatal rat cardiac myocytes in the absence or presence of the Ang II receptor antagonists saralasin (an antagonist of both type 1 and type 2 receptors), CV-11974 (a type 1 receptor-specific antagonist), and PD123319 (a type 2 receptor-specific antagonist). Stretching cardiac myocytes by 20% using deformable silicone dishes rapidly increased the activities of mitogen-activated protein (MAP) kinase kinase activators and MAP kinases. Both saralasin and CV-11974 partially inhibited the stretch-induced increases in the activities of both kinases, whereas PD123319 showed no inhibitory effects. Stretching cardiac myocytes increased amino acid incorporation, which was also inhibited by approximately 70% with the pretreatment by saralasin or CV-11974. When the culture medium conditioned by stretching cardiocytes was transferred to nonstretched cardiac myocytes, the increase in MAP kinase activity was observed, and this increase was completely suppressed by saralasin or CV-11974. These results suggest that Ang II plays an important role in mechanical stress-induced cardiac hypertrophy and that there are also other (possibly nonsecretory) factors to induce hypertrophic responses.

Topics: Angiotensin II; Angiotensin Receptor Antagonists; Animals; Benzimidazoles; Biphenyl Compounds; Cardiomegaly; Cells, Cultured; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; Mitogen-Activated Protein Kinase Kinases; Myocardium; Phenylalanine; Protein Kinases; Rats; Rats, Wistar; Receptors, Angiotensin; Saralasin; Signal Transduction; Stress, Mechanical; Tetrazoles