Effects of angiotensin II microinjected into medial amygdala on male sexual behavior in rats

Research was undertaken to study the role of central angiotensin in the modulation of male sexual behavior, testing the effect of angiotensin II (Ang II) injections into the medial amygdaloid nucleus (MeA). The sexual behavior of adult male Wistar rats was evaluated, 15 min after bilateral intra-amygdaloid microinjection (0.3 microl) of saline and 5 doses of Ang II: 10; 25; 50; 100, and 150 fmol. The effects of the Ang II receptor blockade were also studied. We tested the effect of coinjection of Ang II (50 fmol) with the AT1 antagonist, losartan (20 pmol) and the AT2 antagonist, CGP 42112 (1 pmol). Ang II inhibited sexual behavior and this inhibition was prevented by the coinjection of AT1 antagonist, losartan, or the AT2 antagonist, CGP 42112. Results show that Ang II has a powerful effect on male sexual behavior, which may be mediated by both AT1 and AT2 receptors.     

Angiotensin II receptor subtypes play opposite roles in regulating phosphatidylinositol hydrolysis in rat skin slices.

Among the many functions of angiotensin II (Ang II) it now appears that Ang II is a growth factor. The concentration of Ang II in rat skin has been shown to increase during wound healing. To investigate the intracellular effect of Ang II in skin we determined the levels of total cytoplasmic inositol phosphates after incubation of skin slices with different doses of Ang II. 10(-6) M of Ang II increased significantly the phosphatidylinositol (PI) hydrolysis, and the effect was dose dependent up to 10(-4) M Ang II. The majority of inositol phosphates yielded after 1 hour incubation in the presence of lithium was InsP1, with lesser amount of InsP2. Losartan, the Ang II AT1 antagonist, at a dose of 10(-4) M blocked the effect of Ang II, while PD123319, the Ang II AT2 antagonist, had no antagonistic action; PD123319 at the higher dose of 10(-3) M, however, potentiated the effect of Ang II on PI hydrolysis. The results suggest that PI hydrolysis is a second messenger system for Ang II in rat skin. Also, the two subtypes of Ang II receptors mediate opposite effects on PI hydrolysis: Ang II binding to AT1 receptors increases inositol phosphate production, while Ang II binding to AT2 receptors decreases inositol phosphate production.     

Angiotensin II modulates the activity of Na+,K+-ATPase in cultured rat astrocytes via the AT1 receptor and protein kinase C-delta activation

In astrocytes the activity of the Na+,K(+)-ATPase pump maintains an inwardly directed electrochemical sodium gradient used by the Na+-dependent transporters and regulates the extracellular K+ concentration essential for neuronal excitability. We show here that incubation of cultured rat astrocytes with angiotensin II (Ang II) modulates Na+,K(+)-ATPase activity, in a dose- and time-dependent manner. Na+,K(+)-ATPase activation was mediated by binding of Ang II to AT1 receptors as it was completely blocked by DuP 753, a specific AT1 receptor subtype antagonist. Stimulation of Na+,K(+)-ATPase activity by Ang II was dependent on protein kinase C (PKC) activation because PKC antagonists abolished the inducing effect of Ang II and the PKC activator phorbol 12-myristate 13-acetate enhanced transporter activity. Ang II stimulated translocation of PKC-delta but not that of other PKC isoforms from the cytosol to the plasma membrane. These results indicate that the activity of Na+,K(+)-ATPase in astrocytes is increased by physiological concentrations of Ang II and that the AT1 receptor subtype mediates the Na+,K(+)-ATPase response to Ang II via PKC-delta activation.     

The synergistic induction of cyclooxygenase-2 in lung fibroblasts by angiotensin II and pro-inflammatory cytokines

Although we have demonstrated that Angiotensin II (Ang II) signaling plays a role in colon and lung tumorigenesis, the precise mechanisms by which Ang II stimulates tumorigenesis remain unclear. The aim of this study was to investigate the synergistic induction of COX-2 by Ang II and pro-inflammatory cytokines in lung fibroblasts. We also compared the efficiencies of Ang II-dependent COX-2 induction in lung epithelial cells and stromal cells. Ang II induced COX-2 expression in lung fibroblasts in a dose-dependent manner (10(-9) to 10(-7) M) through the Ang II subtype 1 receptor (AT(1)). In addition, Ang II synergistically stimulated the induction of COX-2 by pro-inflammatory cytokines, IL-1beta, or TNF-alpha. Our results indicate that the pro-tumorigenic function of Ang II is attributable, in part, to its strong stimulatory effect of COX-2 expression in lung fibroblasts in which synergistic stimulation with pro-inflammatory cytokines was evident. It is also suggested that the AT(1) receptor in lung fibroblasts may be a rational target for chemoprevention of lung cancer.     

Estrogen prevents intestinal inflammation after trauma-hemorrhage via downregulation of angiotensin II and angiotensin II subtype I receptor

Although angiotensin II (Ang II) plays a key role in development of organ ischemia-reperfusion injury, it remains unclear whether it is involved in development of intestinal injury following trauma-hemorrhage (T-H). Studies have shown that 17beta-estradiol (E2) administration following T-H improves small intestinal blood flow; however, it is unclear whether Ang II plays a role in this E2-mediated salutary effect. Male Sprague-Dawley rats underwent laparotomy and hemorrhagic shock (removal of 60% total blood volume, fluid resuscitation after 90 min). At onset of resuscitation, rats were treated with vehicle, E2, or E2 and estrogen receptor antagonist ICI 182,780 (ICI). A separate group of rats was treated with Ang II subtype I receptor (AT1R) antagonist losartan. At 24 h after T-H, plasma Ang II, IL-6, TNF-alpha, intercellular adhesion molecule (ICAM)-1, cytokine-induced neutrophil chemoattractant (CINC)-1 and CINC-3 levels, myeloperoxidase (MPO) activity, and AT1R expression were determined. T-H significantly increased plasma and intestinal Ang II, IL-6, TNF-alpha levels, intestinal ICAM-1, CINC-1, CINC-3 levels, MPO activity, and AT1R protein compared with shams. E2 treatment following T-H attenuated increased intestinal MPO activity, Ang II level, and AT1R protein expression. ICI administration abolished the salutary effects of E2. In contrast, losartan administration attenuated increased MPO activity without affecting Ang II and AT1R levels. Thus Ang II plays a role in producing small intestine inflammation following T-H, and the salutary effects of E2 on intestinal inflammation are mediated in part by Ang II and AT1R downregulation.     

Intracisternal galanin/angiotensin II interactions in central cardiovascular control

The aim of this work was to investigate the interactions between angiotensin II (Ang II) and galanin(1-29) [GAL(1-29)] or its N-terminal fragment galanin(1-15) [GAL(1-15)] on central cardiovascular control. The involvement of angiotensin type1 (AT1) receptor subtype was analyzed by the AT1 antagonist, DuP 753. Anesthesized male Sprague-Dawley rats received intracisternal microinjections of Ang II (3 nmol) with GAL(1-29) (3 nmol) or GAL(1-15) (0.1 nmol) alone or in combination. The changes in mean arterial pressure (MAP) and heart rate (HR) recorded from the femoral artery were analyzed. The injection of Ang II and GAL(1-15) alone did not produce any change in MAP. However, coinjections of both Ang II and GAL(1-15) elicited a significant vasopressor response. This response was blocked by DuP 753. Ang II and GAL(1-15) alone produced an increase in HR. The coinjections of Ang II with GAL(1-15) induced an increase in HR not significantly different from the tachycardia produced by each peptide. The presence of DuP 753 counteracted this response. GAL(1-29) alone elicited a transient vasopressor response that disappeared in the presence of Ang II. The coinjections of Ang II with GAL(1-29) and with DuP 753 restored the transient vasopressor effect produced by GAL(1-29). GAL(1-29) produced a slight but significant tachycardic effect that was not modified in the presence of Ang II. The presence of DuP 753 did not modify the tachycardic response produced by Ang II and GAL(1-29). These results give indications for the existence of a differential modulatory effect of Ang II with GAL(1-15) and GAL(1-29) on central blood pressure response that might be dependent on the activity of the angiotensin AT1 receptor subtype.     

Apoptosis induction and inhibition of cellular proliferation by angiotensin II: possible implication and perspectives

The renin-angiotensin system plays a pivotal role in the regulation of fluid, electrolyte metabolism and blood pressure. Molecular cloning and pharmacological studies have defined two major classes of Angiotensin II (Ang II) receptors, designated AT1 and AT2. Recently, it has been well recognized that Ang II, beside its classical physiological actions, is a profibrogenic peptide and displays characteristics of a growth factor. The emerging picture suggests that angiotensin receptor subtypes exert opposing features in many aspects of their biological function, most importantly in cellular growth and proliferation. Accordingly, the proliferative and/or growth-promoting effects of Ang II are thought to be mediated by AT1 receptor, whereas the AT2 receptor subtype may have growth-inhibitory properties. The novel finding that Ang II is able to induce apoptosis by AT2 receptors in diverse cell types is of great scientific interest, as recent studies revealed a role for apoptosis as a deliberate form of cell death in the pathogenesis of various cardiovascular diseases such as heart failure and vascular remodeling. Furthermore apoptotic cell death might occur during the development of progressive glomerulosclerosis. It is tempting to speculate that autocrine-paracrine vasoactive substances such as Ang II might regulate these apoptotic processes during pathogenic conditions.     

Angiotensin II bi-directionally regulates cyclooxygenase-2 expression in intestinal epithelial cells

We previously demonstrated that angiotensin II (Ang II) receptor signaling is involved in azoxymethane-induced mouse colon tumorigenesis. In order to clarify the role of Ang II in COX-2 expression in the intestinal epithelium, the receptor subtype-specific effect on COX-2 expression in a rat intestinal epithelial cell line (RIE-1) has been investigated. Ang II dose- and time-dependently increased the expression of COX-2, but not COX-1 mRNA and protein. This stimulation was completely blocked by the AT(1) receptor antagonist but not the AT(2) receptor antagonist. Ang II and lipopolysaccharide (LPS) additively induced COX-2 protein in RIE-1 cells, whereas the LPS-induced COX-2 expression was significantly attenuated by low concentrations of Ang II or the AT(2) agonistic peptide CGP-42112A only in AT(2) over-expressed cells. These data indicate that Ang II bi-directionally regulates COX-2 expression via both AT(1) and AT(2) receptors. Control of COX-2 expression through Ang II signaling may have significance in cytokine-induced COX-2 induction and colon tumorigenesis.     

Involvement of c-Src tyrosine kinase in SHP-1 phosphatase activation by Ang II AT2 receptors in rat fetal tissues

Angiotensin II (Ang II) AT(2) receptors are abundantly expressed in rat fetal tissues where they probably contribute to development. In the present study we examine the effects of Ang II type 2 receptor stimulation on SHP-1 activation. Ang II (10(-7) M) elicits a rapid and transient tyrosine phosphorylation of SHP-1, maximal at 1 min, in a dose-dependent form, blocked by the AT(2) antagonist, PD123319. SHP-1 phosphorylation is followed in time by tyrosine dephosphorylation of different proteins, suggesting a sequence of events. Ang II induces association of SHP-1 to AT(2) receptors as shown by co-immunoprecipitation, Western blot and binding assays. SHP-1 activity was determined in immunocomplexes obtained with either anti-AT(2) or anti-SHP-1 antibodies, after Ang II stimulation (1 min), in correlation with the maximal level of SHP-1 phosphorylation. Interestingly, following receptor stimulation (1 min) c-Src was associated to AT(2) or SHP-1 immunocomplexes. Preincubation with the c-Src inhibitor PP2 inhibited SHP-1 activation and c-Src association, thus confirming the participation of c-Src in this pathway. We demonstrated here for the first time the involvement of c-Src in SHP-1 activation via AT(2) receptors present in an ex vivo model expressing both receptor subtypes. In this model, AT(2) receptors are not constitutively associated to SHP-1 and SHP-1 is not constitutively activated. Thus, we clearly establish that SHP-1 activation, mediated by the AT(2) subtype, involves c-Src and precedes protein tyrosine dephosphorylation, in rat fetal membranes.     

125I[Sar(1)Ile(8)] angiotensin II has a different affinity for AT(1) and AT(2) receptor subtypes in ovine tissues

Iodinated angiotensin II (Ang II) and its analogues are often assumed to have equal affinities for AT(1) and AT(2) receptor subtypes. However, using saturation and competition binding assays in several tissues from pregnant, nonpregnant, and fetal sheep, we found the affinity of 125I[Sar(1)Ile(8)] Ang II for Ang II receptors was different (P<0.05) between tissue types. The dissociation constants (Kd) and half maximal displacements of [Sar(1)Ile(8)] Ang II (Sar IC(50)) were directly related (P<0.05) to proportions of AT(1) receptors, and inversely related (P<0.05) to proportions of AT(2) receptors in tissues from all groups combined, in tissues from individual groups (pregnant, nonpregnant or fetal), and in some individual tissues (uterine arteries and aortae). This suggests that 125I[Sar(1)Ile(8)] Ang II has a different affinity for AT(1) and AT(2) receptors in ovine tissues. The Kds of 125I[Sar(1)Ile(8)] Ang II for "pure" populations of AT(1) and AT(2) receptors were 1.2 and 0.3 nM, respectively, i.e. affinity was four-fold higher for AT(2) receptors. We corrected the measured proportions of the receptor subtypes using their fractional occupancies. In tissues which contained at least 10% of each receptor subtype, the corrected proportions were significantly altered (P<0.05), even in some tissues, to the extent of being reversed.     

Differential response of human cardiac fibroblasts to angiotensin I and angiotensin II

The vasoactive peptide angiotensin II (Ang II) has been implicated as a mediator of myocardial fibrosis. We carried out a comparative investigation of the effects of Ang II and its precursor Ang I on collagen metabolism and proliferation in cultured human cardiac fibroblasts. Cardiac fibroblasts responded to both Ang I and Ang II with concentration-dependent increases in collagen synthesis but no proliferation. The stimulatory effect of Ang II was abolished by the AT(1) receptor antagonist losartan but not the AT(2) receptor antagonist PD123319. The response to Ang I was not affected by either antagonist, nor by the angiotensin-converting enzyme (ACE) inhibitor captopril. In conclusion, Both Ang I and Ang II stimulate collagen synthesis of human cardiac fibroblasts, the effect of Ang II occurring via the AT(1) receptor whilst Ang I appears to exert a direct effect through non-Ang II-dependent mechanisms. These results suggest distinct roles for angiotensin peptides in the development of cardiac fibrosis.     

Renal nuclear angiotensin II receptors in normal and hypertensive rats

Accumulation of Angiotensin II (Ang II) in the kidneys of hypertensive rats infused chronically with Ang II occurs by AT1 receptor mediated internalization of Ang II, which may interact with intracellular targets, including nuclear binding sites. The aims of this study were to determine if kidney cell nuclei have specific Ang II binding sites and if chronic infusion of Ang II (70 ng/min; n=9) influences the nuclear Ang II binding capacity. Kidneys were harvested from control and Ang II infused rats and the renal cortexes were homogenized to obtain crude membrane preparations and nuclear fractions. Ang II binding sites were measured with a single point assay by incubating each fraction with 10 nM 125I-Sar-Ile-Ang II in the absence (total binding sites) or presence of either 2.5 M Sar-Leu-Ang II or 25 microM losartan to detect specific AT or AT1 binding sites. Both fractions exhibited specific Ang II binding sites that were displaced by both saralasin and losartan. In control rats, crude membrane preparations had 792 +/- 218 and the nuclear fraction had 543 +/- 222 fmol/mg protein AT1 receptors. AT1 receptor levels in membrane (885 +/- 170 fmol/mg protein) and nuclear fractions (610 +/- 198 fmol/mg protein) were not significantly different in Ang II infused rats. These data support the presence of nuclear Ang II receptors predominantly of the AT1 subtype in renal cells. Chronic Ang II infusion did not alter overall Ang II receptor densities.     

AT1 angiotensin II receptor mediates intracellular calcium mobilization in normal and cancerous breast cells in primary culture

Angiotensin II (Ang II) increases intracellular calcium concentration ([Ca2+]i) in both normal and cancerous human breast cells in primary culture. Maximal [Ca2+]i increase is obtained using 100nM Ang II in both cell types; in cancerous breast cells, [Ca2+]i increase (delta[Ca2+]i) is 135+/-10nM, while in normal breast cells it reaches 65+/-5 nM (P<0.0001). In both cell types, Ang II evokes a Ca2+ transient peak mediated by thapsigargin (TG) sensitive stores; neither Ca2+ entry through L-type membrane channels or capacitative Ca2+ entry are involved. Type I Ang II receptor subtype (AT1) mediates Ang II-dependent [Ca2+]i increase, since losartan, an AT1 inhibitor, blunted [Ca2+]i increase induced by Ang II in a dose-dependent manner, while CGP 4221A, an AT2 inhibitor, does not. Phospholipase C (PLC) is involved in this signaling mechanism, as U73122, a PLC inhibitor, decreases Ang II-dependent [Ca2+]i transient peak in a dose-dependent mode.Thus, the present study provides new information about Ca2+ signaling pathways mediated through AT1 in breast cells in which no data were yet available.     

Angiotensin II and its receptors in human endocardial endothelial cells: role in modulating intracellular calcium

The aims of the present study are to investigate the presence and distribution of angiotensin II (Ang II), as well as AT1 and AT2 receptors, in endocardial endothelial cells (EECs) and to determine if the effect of Ang II on intracellular calcium in these cells is mediated via the AT1 or the AT2 receptor. Immunofluorescence and 3D confocal microscopy techniques were used on 20-week-old fetal human EECs. Our results showed that Ang II and its receptors, the AT1 and the AT2 types, are present and exhibit a different distribution in human EECs. Ang II labelling is found throughout the cell with a fluorescence signal higher in the cytosol when compared with the nucleus. Like Ang II, the AT1 receptor fluorescence signal is also homogeneously distributed in human EECs but with a preferential labelling at the level of the nucleus, while the AT2 receptor labelling is solely present in the nucleus. Using fluo-3 and 3D confocal microscopy technique, superfusion of human EECs with increasing concentration of Ang II induced a dose-dependent sustained increase in free cytosolic and nuclear Ca2+ levels. This effect of Ang II on human EEC's intracellular Ca2+ ([Ca2+]) was completely prevented by losartan, an AT1 receptor antagonist. Our results suggest that Ang II, as well as AT1 and AT2 receptors, is present but differentially distributed in EECs of 20-week-old fetal human hearts, and that the AT1 receptor mediates the effects of Ang II on [Ca2+]i in these cells.     

Regulation of insulin-like growth factor-1 by the renin-angiotensin system during regression of cardiac eccentric hypertrophy through angiotensin-converting enzyme inhibitor and AT1 antagonist

Angiotensin II (Ang II) mediates its effects through its non-tyrosine-kinase G protein coupled Ang-II type 1 receptor (AT1). Growing evidence indicates that a functional insulin-like growth factor-1 (IGF-1) tyrosine kinase receptor is required for Ang-II-induced mitogenesis. Along with Ang II, we have previously shown that changes in IGF-1 receptor binding at myofibers are causative agents for cardiac eccentric hypertrophy. This study investigated the interaction of the renin-angiotensin system with the IGF-1 receptor during the development and regression of cardiac hypertrophy. Alterations in IGF-1 binding were evaluated in the CHAPS-pretreated perfused heart. Four weeks of aortocaval shunt increased relative heart mass by 76% without a major change in body mass or systolic blood pressure. Binding studies showed that IGF-1 has a higher affinity for the cardiac myofibers of shunt than sham rats. Two weeks of treatment with the angiotensin-converting enzyme (ACE) inhibitor captopril (0.5 g/L in drinking water) or the AT1-antagonist losartan (10 mg/(kg x day)) reduced cardiac hypertrophy by 54 and 42%, respectively. However, while both ACE inhibition and AT1-antagonist treatments produced equivalent regression in ventricular hypertrophy, captopril was more efficacious than losartan in the regression of atrial hypertrophy. Regression of cardiac hypertrophy in the shunt by either captopril or losartan was accompanied with a reduction or normalization of the elevated IGF-1 affinity. Thus, the induction and regression of cardiac eccentric hypertrophy seems to be largely dependent on cross talk between the renin-angiotensin system and the IGF-1 axis at the receptor level.     

Pharmacological characterization of angiotensin II binding sites in the canine pancreas.

High affinity 125I-angiotensin II (Ang II) binding sites were characterized in the canine pancreas. Total binding increased with protein concentration and equilibrium was reached within 60-90 min at 22 degrees C. Specific binding was saturable and averaged 70% of total. Scatchard analysis of binding yielded a KD of 0.48 +/- 0.18 nM with a Bmax of 32.8 +/- 6.5 fmol/mg protein (mean +/- SEM, n = 6). The addition of the reducing agent dithiothreitol increased specific binding two-fold. The rank order of displacement of 125I-Ang II binding by native angiotensin peptides was Ang II greater than or equal to Ang III greater than AngI greater than Ang(1-7) much greater than Ang(1-6). The use of the specific Ang II antagonists CGP 42112A, PD 123177, and DuP 753 revealed that the pancreas expresses two receptor subtypes. The majority of Ang II binding sites in the pancreas could be classified as type 2 (AT2), although type 1 (AT1) sites were also detected. In vitro autoradiography revealed binding sites localized over islet cells, acinar and duct cells, as well as the pancreatic vasculature. In addition, the autoradiographic studies confirmed the predominance of the AT2 receptor subtype throughout the pancreas.     

Angiotensin II mediates catecholamine and neuropeptide Y secretion in human adrenal chromaffin cells through the AT1 receptor

The aim of the present work was to study the effect of angiotensin II (Ang II) on catecholamines and neuropeptide Y (NPY) release in primary cultures of human adrenal chromaffin cells. Ang II stimulates norepinephrine (NE), epinephrine (EP) and NPY release from perifused chromaffin cells by 3-, 2- and 12-fold, respectively. The NPY release is more sustained than that of catecholamines. We found that the receptor-AT(2) agonist, T(2)-(Ang II 4-8)(2) has no effect on NE, EP and NPY release from chromaffin cells. We further showed that Ang II increases intracellular Ca(2+) concentration ([Ca(2+)](i)). The selective AT(1)-receptor antagonist Candesartan blocked [Ca(2+)](i) increase by Ang II, while T(2)-(Ang II 4-8)(2) was ineffective. These findings demonstrate that AT(1) stimulation induces catecholamine secretion from human adrenal chromaffin cells probably by raising cytosolic calcium.     

Angiotensin II induced increase in frequency of cytosolic and nuclear calcium waves of heart cells via activation of AT1 and AT2 receptors

The aim of this work is to verify if Angiotensin II (Ang II) affects the frequency of spontaneous cytosolic and nuclear Ca2+ waves in chick embryonic cardiomyocytes and if this effect is mediated via the activation of AT1 and/or AT2 receptors. Using the rapid scan technique of confocal microscopy, we observed that Ang II (10(-8)M) increases the frequency of cytosolic and nuclear Ca2+ waves. This effect was accompanied by a decrease in the amplitude of nuclear Ca2+ waves and an absence of effect on the amplitude of cytosolic Ca2+ waves. The effect of the octapeptide on both frequency and amplitude of the nuclear waves was prevented by the AT1 receptor antagonist L158809. However, blockade of the AT2 receptor using the antagonist PD123319 (10(-7)M) only prevented the effect of Ang II on the frequency of Ca2+ waves. Furthermore, the effect was prevented by both a PKC inhibitor (bisindolylmaleimide) and a PKC activator (phorbol 12,13-dibutyrate). In addition, the Ang II effect was not prevented by the blocker of the pacemaker current If. These results demonstrate that Ang II, via the activation of its receptors AT1 and AT2, affects the frequency of spontaneous Ca2+ waves and this effect seems to be mediated by the PKC pathway.     

Metabolism of angiotensin II is required for its in vivo effect on dopamine release in the striatum of the rat

The effect of angiotensin (Ang) IV, an inhibitor of insulin-regulated aminopeptidase (IRAP), on extracellular dopamine levels in the striatum of freely moving rats was examined using in vivo microdialysis. The Ang IV was administered locally in the striatum through the microdialysis probe. A concentration-dependent (10-100 microm) increase in extracellular striatal dopamine was observed. The effect of Ang II (10-100 microm), which has only a weak affinity for IRAP, was similar to that observed for Ang IV. The effects of both peptides could not be blocked by the AT1 antagonist candesartan (10 nm and 1 microm) nor by the AT2 antagonist S-(+)-1-([4-(dimethylamino)-3-methylphenyl]methyl)-5-(diphenyl-acetyl)-4,5,6,7-tetrahydro-1H-amidazo(4,5-c) pyridine-6-carboxylic acid (1 microm), suggesting that the observed effects are both AT1 and AT2 independent. The effect of Ang II could be blocked by the aminopeptidase-A inhibitor (S)-3-amino-4-mercaptobutylsulphonic acid as well as the aminopeptidase-N inhibitor 2-amino-4-methylsulphonylbutane thiol, indicating that the effect of Ang II is mediated via metabolism into Ang IV. Other IRAP inhibitors, such as Divalinal-Ang IV and LVV-haemorphin-7, had similar effects on extracellular dopamine levels as compared with Ang IV. We propose a role for IRAP as mediator for the effects of Ang IV and related peptides on extracellular dopamine levels in the striatum of the rat.