Biomedical subjects

M de Gasparo

Publications and source records attributed to M de Gasparo.

At least 73 records · Page 4Linked to original sources

PubMed1992

The angiotensin AT2 receptor modulates T-type calcium current in non-differentiated NG108-15 cells.

We report here that angiotensin II (AII) and the AT2 receptor-selective ligand, CGP 42112, modulate the T-type calcium current in non-differentiated NG108-15 cells, which express only AT2 receptors. Both peptides decrease the T-type calcium current at membrane potentials above -40 mV and shift the current-voltage curve at lower potentials with maximal effect between 5 and 10 min after application. These data describe a new cellular response to AII and suggest that the AT2 receptor mediates certain neurophysiological actions of this hormone.

Action Potentials↗

PubMed1992

The angiotensin AT2 receptor stimulates protein tyrosine phosphatase activity and mediates inhibition of particulate guanylate cyclase.

The signalling mechanism and cellular targets of the AT2 receptor are still unknown. We report that angiotensin II (Ang II) inhibits basal and atrial natriuretic peptide stimulated particulate guanylate cyclase (pGC) activity through AT2 receptors in rat adrenal glomerulosa and PC12W cells. This inhibition is blocked by the phosphotyrosine phosphatase (PTPase) inhibitor orthovanadate but not by the Ser/Thr phosphatase inhibitor okadaic acid, suggesting the involvement of a PTPase in this process. Moreover, Ang II induces a rapid, transient and orthovanadate sensitive dephosphorylation of phosphotyrosine containing proteins in PC12W cells. Our findings suggest that AT2 receptors signal through stimulation of a PTPase and that this mechanism is implicated in the regulation of pGC activity. This observation is also the first example of hormonal inhibition of basal pGC activity.

Angiotensin II↗

PubMed1992

Circhoral fluctuations of serum total renin, inhibin and related hormones around the mid-cycle in normal human females.

Total renin and inhibin are secreted by the ovary. Although luteinizing hormone (LH) and/or follicle stimulating hormone (FSH) may stimulate their secretion, the close relationship between fluctuations of gonadotrophins, oestradiol, progesterone, renin and inhibin during the cycle is still conjectural. To investigate the temporal relationship between the short-term fluctuations in the circulating concentrations of LH and FSH and the ovarian hormones (oestradiol, progesterone, renin and inhibin), blood samples were collected at 15-min intervals for 6 h from 15 normal women in the late follicular (n = 4), early luteal (n = 5) or luteal (n = 6) phases of the menstrual cycle. LH levels showed the well-known pulsatile secretion with decreasing frequency and increasing relative amplitude from the late follicular to the luteal phase. Progesterone and oestradiol serum levels were pulsatile, 25% and 35-50% of which were linked to LH pulses, with time lags of 30 and 12-15 min respectively. Renin levels presented significant pulses, 26% of which were related to LH pulses with a time lag of less than 10 min; no coincidence was found between renin and oestradiol pulses. Inhibin levels presented only scattered pulses of small amplitude, which were unrelated to LH or FSH. These results show that, besides the LH-related pulses, pulsatile secretion of some ovarian hormones (oestradiol, progesterone and renin) may also occur independently of LH pulses and may be unrelated to one another. Moreover, contrary to the other ovarian hormones, inhibin seems to follow a tonic, not a pulsatile type of secretion around the mid-cycle.

Adult↗

PubMed1992

Could the pharmacological differences observed between angiotensin II antagonists and inhibitors of angiotensin converting enzyme be clinically beneficial?

Over the past several years, angiotensin I converting enzyme (ACE) inhibitors, compounds that block the formation of angiotensin II (ANG II), have become widely used in the treatment of cardiovascular disease. Recently, a new class of orally active, non-peptide inhibitors of the renin-angiotensin system, the ANG II receptor antagonists have also become available. Since both classes of compounds block the renin-angiotensin system, although at different sites, it remains to be determined whether blockade of ANG II receptors will have any specific advantage over inhibition of ACE. The following review assesses the actions of ANG II antagonists and suggests ways in which blockade of ANG II receptors may differ both pharmacologically and clinically from inhibition of ACE.

Angiotensin II↗

PubMed1992

[Identification of angiotensin II receptor and determination of its subtypes in rabbit and guinea pig fetal tissue].

Membrane angiotensin II receptors were measured in trophoblastic tissues using a 2-step procedure. The first step consisted of the relative measurement performed at a fixed 125I[Sar1 Ile8]AII concentration of 0.15 nM in order to determine which tissues had a sufficient number of binding sites for studying the competition curves. The second consisted of determining the maximal binding (Bmax) and the dissociation constant (Kd) for [Sar1 Ile8] AII and the receptor subtypes in these tissues. The relative binding measurement revealed a significant number of occupied sites in rabbit fetal placenta and chorion (159 +/- 17 and 51 +/- 10 fmol/mg proteins) and in guinea pig chorion (132 +/- 12). The mean values of the other trophoblastic tissues were 3-10-fold lower in the 2 species. The competition curves obtained from tissues with high angiotensin II binding receptors showed the predominance of the AT2 subtype in rabbit fetal placenta (AT1/AT2 = 25/75) and of the AT1 receptor in guinea pig chorion (97/3) and in rabbit chorion (90/10). The [SAR1 Ile8] AII affinity (Kd) obtained from Scatchard plot analysis was 1.2 +/- 0.2 nM (n = 5) in fetal placenta and 1.2 (n = 1) in rabbit chorion and 0.5 +/- 0.1 (n = 3) in guinea pig chorion. In these tissues, the respective Bmax values were 1,281 +/- 115 (n = 5), 263 (n = 1) and 1,188 +/- 134 fmol/mg proteins (n = 3). These findings indicate that rabbit fetal placenta and chorion and guinea pig chorion are the most important sites of action for the renin-angiotensin system present in trophoblastic tissues.

1-Sarcosine-8-Isoleucine Angiotensin II↗

PubMed1991

Radioiodinated CGP 42112A: a novel high affinity and highly selective ligand for the characterization of angiotensin AT2 receptors.

CGP 42112A, a potent angiotensin AT2 receptor selective ligand, was radio-iodinated and its binding characteristics compared with those of [125I]angiotensin II. In human myometrium (only AT2 expressed), binding was saturable (Kd 1.03 x 10(-10) M; Bmax 807 fmol/mg) and reversible (K+1 1.89 x 10(8) M-1.min-1; K-1 3.77 x 10(-3) min-1). The order of potency of a number of peptides and non-peptides was the same as when [125I] angiotensin II was used as tracer. No specific binding could be detected on membranes from vascular smooth muscle cells (only AT1 expressed). In rat adrenal glomerulosa membranes (mixed AT1/AT2), [125I]CGP 42112A bound only to AT2. [125I]CGP 42112A can therefore be used as a specific probe for AT2 receptors and will be especially useful in tissues where other subtypes are also present.

Angiotensin II↗

PubMed1991

Distribution of angiotensin II receptor subtypes in rat brain nuclei.

Angiotensin II (ANG II) receptor subtypes in rat brain were characterized and quantified by competitive radioligand binding using [125I]Sar1 Ile8 angiotensin II ([125I]sarilesin) as a tracer and ANG II, sarilesin and the subtype selective ligands DuP 753 (AT1) and CGP 42112A (AT2) as competitors. The distribution of AT1 and AT2 receptors was determined in midbrain, brainstem, hypothalamus as well as in individual hypothalamic and periventricular nuclei. Whereas in midbrain and brainstem the AT1: AT2 ratio was 40%: 60% and 70%: 30% respectively, the AT1 receptors were by far predominant in hypothalamus and in the nuclei investigated. Interestingly, we found that approximately 25% of the ANG II receptors in hypothalamus did not bind DuP 753 even at 0.1 mM. These sites which bind CGP 42112A, ANG II and sarilesin may represent a third ANG II receptor subtype.

Angiotensin II↗

PubMed1991

Angiotensin II AT2 receptors do not interact with guanine nucleotide binding proteins.

We have studied the effect of GTP gamma S on the affinity and binding kinetics of angiotensin II in plasma membrane particulate prepared from tissues expressing either only AT1 (human renal artery smooth muscle cells), only AT2 (human myometrium and bovine cerebellar cortex) or both angiotensin II receptor subtypes (rat adrenal glomerulosa). We also examined the ability of angiotensin II to stimulate GTP gamma[35S] incorporation in these membrane preparations. In contrast to its effects on angiotensin II binding to the AT1 receptor, GTP gamma S does not affect binding parameters to the AT2 receptor. Moreover, in tissues expressing solely AT2 receptors, angiotensin II was unable to induce GTP gamma[35S] incorporation. These findings indicate that AT2 receptors do not interact with G-proteins and that angiotensin II must therefore mediate some of its effects through G-protein-independent mechanisms.

Angiotensin II↗

PubMed1991

Nonpeptidic angiotensin II antagonists: synthesis and in vitro activity of a series of novel naphthalene and tetrahydronaphthalene derivatives.

Starting from the structure of the novel nonpeptidic angiotensin II antagonist DuP 753, a series of more rigid analogues was prepared by replacing the biphenyl part of DuP 753 with a naphthalene ring. Five different regioisomers (compounds 6a-e) were synthesized, and receptor binding in rat smooth muscle cell preparations as well as inhibition of angiotensin II induced contraction of rabbit aortic rings was measured and the order of potency was compared with predictions made on the basis of a molecular modeling study. In good agreement with the predictions, the 2,6-substituted regioisomer 6d and its analogue 7 (isomeric at the imidazole substituent) were found to be most potent, but were still weaker than DuP 753. Tetrahydronaphthalene derivatives with and without an additional methyl group in the alpha-position to the acidic function and with this same 2,6-substitution pattern (compounds listed in Table III) were then prepared with the expectation of getting a further increase in potency. Whereas the carboxylic acid derivatives 13a,b showed activity in the expected potency range, surprisingly no further potency increase was observed after replacement of the carboxylic acid function by a tetrazole (compounds 18a,b). These results may indicate that the compounds do not bind to the AT1 receptor in the same way as DuP 753.

Angiotensin II↗

PubMed1991

Effect of covalent dimer conjugates of angiotensin II on receptor affinity and activity in vitro.

Angiotensin II [1-8 or 2-8] analogues and [4-8] fragments were dimerized through the amino- or carboxy-terminal groups in order to try to increase their potency as reported for other hormones. The binding affinity to the angiotensin II receptor subtypes A (A IIA) and B (A IIB) was tested and compared to the potency in rabbit aortic ring. The [2-8] dimers coupled through the N-terminus show no significant change in potency in aortic ring. The [4-8] fragments coupled through the N-terminus are inactive in the ring. They have however a significantly increased affinity for the A IIA receptor, the specific function of which has not yet been reported. When angiotensin II analogues or fragments are coupled through the C-terminus, there was a significant drop in affinity and potency, confirming the importance of the free carboxyl group in position 8 for binding and activity. It is concluded that binding to the A IIB receptor correlates well with the effectiveness in aortic ring. However, in contrast to the beneficial effect reported for a large number of other hormones, dimerization of angiotensin II or its fragments is not accompanied by an increased biological activity in aortic ring.

Amino Acid Sequence↗

PubMed1991

[Angiotensin II receptors in the human placenta are type AT1].

Membrane angiotensin II receptors were measured in human placenta by means of 125I [Sar1 Ile8] All (angiotensin II antagonist) and characterized by using 2 other antagonists of angiotensin II: Dup 753 and CGP 42112A. These are specific and selective ligands which enable identification of AT1 and AT2 receptor subtypes respectively. The [Sar1 Ile8] All affinity is similar (Kd approximately 1 nmol.l-1) in the 3 different placental structures examined. However, the Bmax of villous tissues is approximately 9 times higher than that observed in chorionic plate but remains near that found in basal plate. In the central area of the placenta, mean values of 125I [Sar1 Ile8] All binding observed at a single concentration of 0.15 nmol.l-1 are 242 +/- 31 fmol/mg proteins in basal plate, 300 +/- 35 in villous tissues and 36 +/- 8 in chorionic plate. The umbilical vein and arteries respectively have 8.8 +/- 4.8 and 4.0 +/- 1.7 fmol/mg protein. The subtype analysis shows that only AT1 receptor is present in placental tissues. The Bmax values as well as those obtained by the relative measurement performed at a fixed 125I [Sar1 Ile8] All concentration of 0.15 nmol.l-1 indicate that the highest concentrations of angiotensin II receptors are found in placental villous tissues.

1-Sarcosine-8-Isoleucine Angiotensin II↗

PubMed1990

Identification and characterization of angiotensin II receptor subtypes in rabbit ventricular myocardium.

CGP 42 112 A and DuP 753 block [125I]-angiotensin II binding in rabbit ventricular myocardial membranes in a clearly biphasic manner, indicating the existence of high- and low-affinity sites for these highly selective agents. Assays using concentrations of either agent large enough to prevent high-affinity binding show that their respective high-affinity sites are distinct, and each corresponds to the low-affinity site of the other. The two receptor subsets, present in nearly equal proportions, are also distinguishable by their different sensitivities to dithiothreitol. These findings afford strong evidence for the existence of two distinct angiotensin II receptors in rabbit myocardium, corresponding to the A and B subtypes recently described in adrenals.

Angiotensin II↗

PubMed1990

Sarmesin is a partial agonist of angiotensin-II receptors in rabbit, but not in rat, aortic rings.

Sarmesin, [Sar1, Tyr(Me)4]angiotensinII], has been reported to be a competitive angiotensin II (AII) receptor antagonist in rat smooth muscle preparations (Scanlon et al., (1984), Life Science 34, 317-321). In the present study, sarmesin displaced AII from its binding sites in rat aortic smooth muscle cells and in a rabbit aorta membrane preparation (IC50 5 and 6 nM resp.; Ki 4.1 and 5.3 resp.) In rabbit aortic rings, sarmesin (0.003-3 microM) produced concentration-dependent contractions (ED50 89 nM) and this effect was inhibited by saralasin. No contraction was observed in the rat aorta up to 100 microM. In rabbit aortic rings, sarmesin, at the same concentrations that produced contraction, inhibited contractions induced by AII in a competitive manner (pA2 7, 26). These results indicate that, in rabbit aortic rings sarmesin is a partial agonist of AII receptors.

Angiotensin II↗

PubMed1990

Potentiation of angiotensin II-stimulated phosphoinositide hydrolysis, calcium mobilization and contraction of renal mesangial cells upon down-regulation of protein kinase C.

Long-term pretreatment of rat mesangial cells with 12-O-tetradecanoylphorbol 13-acetate (TPA) down-regulated protein kinase C activity and potentiated the angiotensin II-induced inositol trisphosphate (InsP3) formation. This increased response to angiotensin II occurred without a significant change in the receptor number or Kd value of angiotensin II binding to the cells. The biologically inactive phorbol ester 4 alpha-phorbol 12,13-didecanoate was without effect on angiotensin II-stimulated InsP3 generation. Long-term pretreatment with TPA also increased the angiotensin II-induced mobilization of Ca2+ and the subsequent contraction of mesangial cells.

Angiotensin II↗

PubMed1990

Biochemical characterization of two angiotensin II receptor subtypes in the rat.

Two angiotensin II receptor subtypes, A and B, have been described by means of specific and selective ligands (Biochem Biophys Res Commun 1989; 163:284-91). The present report describes the binding characteristics and the distribution of these subtypes in various rat tissues. Adrenal and uterus expressed both subtypes but in different proportions. Subtype B predominated in the adrenal glomerulosa (60%), whereas there was a greater proportion of subtype A in the medulla (70%). In uterus, subtype B was preferentially expressed (70%), and there was no difference in receptor distribution between muscle layers and serosa. Liver, kidney, and cultured aortic smooth muscle cells expressed only subtype B. In all of the tissues tested, the Ki values of various competing ligands were similar for each subtype expressed. It is proposed that subtype B is the vascular receptor. The function of subtype A, however, is still to be determined.

Adrenal Glands↗

PubMed1990

Binding characteristics and vascular effects of various angiotensin II antagonists.

Subtypes of angiotensin II (Ang II) receptors have been recently identified using specific ligands (see Whitebread et al. Biochem Biophys Res Comm 1989; 163:284-291). The present study compares the binding characteristics of different structural classes of Ang II receptor ligands in rat aortic smooth muscle cells (Ang IIB subtype) and in human uterus (Ang IIA subtype) and their effects on the constrictor response to Ang II in isolated rabbit aortic rings. Saralasin and [Sar1 Ile8]-Ang II displayed similar affinity for the two subtypes. In contrast, CGP 42112A bound with high affinity to the uterus (Ki 0.24 nM), but showed a low affinity for the aortic receptor (Ki 1,760 nM). Compound 89 displayed affinity for the aortic receptor only (Ki 26 nM) whereas Ex 169 recognized specifically the uterus receptor (Ki 310 nM). In rabbit aortic rings, saralasin, [Sar1 Ile8]Ang II, CGP42112A and compound 89 inhibited Ang II-induced contractions at concentrations similar to those required to bind to the Ang IIB receptor subtype. IC50s were 3, 0.7, 1,850, and 23 nM respectively. Ex 169 was ineffective in concentrations up to 100 microM. There was a highly significant correlation between inhibition of Ang II-induced contraction in aortic rings and binding to smooth muscle cells. This correlation does not exist with human uterus. Our results indicate that the Ang IIB receptor subtype is responsible for vascular contractions. Antagonists of this vascular receptor subtype are potential antihypertensive agents.

Angiotensin II↗

PubMed1990

Evaluation of a potent inhibitor of subprimate and primate renins.

Our attempts to synthesize a potent inhibitor of rat renin have resulted in the discovery of CGP 44 099 A, a potent inhibitor of plasma renin from all subprimate species tested so far [IC50 (in nM): dog, 0.007; rabbit, 0.033; guinea pig, 0.34; mouse, 0.4; cat, 0.57; and rat, 1.3]. This compound is also a potent inhibitor of primate renins [IC50 (in nM): human, 0.3; and marmoset, 1.4]. It is less potent against other aspartic proteinases [IC50 (in nM): porcine pepsin, 26; and bovine cathepsin D, 230). CGP 44 099 A exhibited a competitive mode of inhibition against human renin (Ki, 0.12 nM). During i.v. infusion of CGP 44 099 A in sodium-depleted normotensive rats (0.1 mg/kg/min) blood pressure (BP) was lowered by about 25 mm Hg. Plasma renin activity, angiotensin I and angiotensin II were almost completely suppressed. The converting enzyme inhibitor enalaprilat (1 mg/kg i.v.) also lowered BP by about 25 mm Hg. No further fall in BP occurred when CGP 44 099 A (0.1 mg/kg/min) was infused after pretreatment with enalaprilat (1 mg/kg i.v.) and CGP 44 099 A did not lower BP when infused in bilaterally nephrectomized rats. These results indicate that the hypotensive response induced by CGP 44 099 A in sodium-depleted rats is specifically due to the renin inhibition. Compounds such as CGP 44 099 may therefore be useful for comparing the effects of renin inhibition in different species and for studying the role of renin in various models of cardiovascular disease in nonprimate species.

Amino Acid Sequence↗

PubMed1989

Preliminary biochemical characterization of two angiotensin II receptor subtypes.

Two angiotensin II receptor subtypes (A and B) are described from rat and human tissues. They have been characterised using specific peptidic and non-peptidic ligands with affinities differing by 1000 fold or more. These subtypes are present in adrenal glomerulosa of both species. Human uterus contains only subtype A, whereas both subtypes are found in rat uterus. Vascular smooth muscle cells in culture express only subtype B. Dithio-threitol totally inhibits binding to subtype B, but enhances the affinity to subtype A. There is a good correlation between the affinities of the selected agonists and antagonists for the two subtypes in the various tissues tested which is a usual requirement for receptor classification.

Adrenal Glands↗