Identical mesenteric,femoral and renal vascular responses to angiotensins II and III in the dog

The effects of angiotensin II (AII) and its 1-des Asp analog (AIII) given intra-arterially (0.3–30 ng/kg) were compared in the mesenteric, femoral, and renal vascular beds in anesthetized dogs in which flow was measured with an electromagnetic flowmeter. As has been shown previously, AII and AIII produced similar changes in renal blood flow. In view of the reduced pressor activity of AIII it was surprising to find strikingly similar responses to AII and AIII in the mesenteric and femoral vascular beds. We conclude that the difference in pressor activity of these agents is attributable to something other than differences in their peripheral vascular receptor, and perhaps may be due to differences in their central actions.     

Change of the activities of angiotensins I and II on the vascular smooth muscle by crude kallikrein.

Incubation in vitro of angiotensin I (A I) with crude kallikrein induced a potentiation in the response to decapeptide of the isolated continuously superfused rabbit aorta. Crude kallikrein when incubated with angiotensin II (A II) caused a decrease in response to octapeptide of the same assay tissue. Converting enzyme inhibitor, SQ 20881, produced a competitive inhibition in the response to A I preincubated with crude kallikrein but did not alter the inhibitory effect of the enzyme on A II response. Pure kallikrein did not induce any change in the responses to both peptides when used at the same concentrations. The competitive inhibitor of A II (N,N-dimethyl) Gly1-Ile5-Ile8-angiotensin II (DMGIA II), abolished the effects of both A II- and A I-preincubated with crude kallikrein. From these results it was concluded that crude kallikrein-induced potentiation in the response to A I of the aorta is probably due to the conversion of decapeptide to octapeptide by an enzyme fraction in crude kallikrein preparation. These results also indicate that crude kallikrein (Padutin) is not a pure enzyme preparation and probably contains some other enzyme fractions which are responsible from the changes of the vascular activities of angiotensin-peptides.     

Direct effects in vivo of angiotensins I and II on the canine sinus node.

The chronotropic responses to angiotensins I and II (5 micrograms in 1 mL Tyrode's solution) injected into the sinus node artery were assessed before and after the intravenous administration of captopril (2 mg/kg) and saralasin (20 micrograms/kg) in anaesthetized dogs. The effects of angiotensin II given intravenously were also observed. The animals (n = 8) were vagotomized and pretreated with propranolol (1 mg/kg, i.v.) to prevent baroreceptor-mediated responses to increases in blood pressure. Injection of angiotensin I into the sinus node artery induced significant increases in heart rate (114 +/- 6 vs. 133 +/- 6 beats/min) and in systemic systolic (134 +/- 13 vs. 157 +/- 14 mmHg; 1 mmHg = 133.3 Pa) and diastolic (95 +/- 10 vs. 126 +/- 13 mmHg) blood pressures. Similar results were obtained when angiotensin II was injected into the sinus node artery, but intravenous injection induced changes in systolic (138 +/- 8 vs. 180 +/- 25 mmHg) and diastolic (103 +/- 8 vs. 145 +/- 20 mmHg) blood pressures only. Captopril induced a significant decrease in systolic (118 +/- 11 vs. 88 +/- 12 mmHg) and diastolic (84 +/- 9 vs. 59 +/- 9 mmHg) blood pressures without affecting the heart rate (109 +/- 6 vs. 106 +/- 6 beats/min). Saralasin produced a significant increase in systolic (109 +/- 7 vs. 126 +/- 12 mmHg) blood pressure only. Increments in heart rate and systolic and diastolic blood pressures in response to angiotensins I and II were, respectively, abolished by captopril and saralasin. It was concluded that angiotensin II has, in vivo, a direct positive chronotropic effect that can be blocked by saralasin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of vascular distending pressure on regional flows in isolated perfused dog lungs

To confirm the regional differences in vascular pressure vs. flow properties of lung regions that have been documented in zone 2 conditions [pulmonary venous pressure (Ppv) less than alveolar pressure], regional distending pressure vs. flow curves in zone 3 were generated by use of isolated blood-perfused dog lungs (3 right and 5 left lungs). Each lung was kept inflated at constant inflation pressure (approximately 50% of full inflation volume) while radioactively labeled microspheres were injected at different settings of Ppv. To achieve maximal vascular distension, Ppv was increased to approximately 30 cmH2O above alveolar pressure for the first injection. Subsequent injections were made at successively lower Ppv's. The difference between pulmonary arterial pressure and Ppv was kept constant for all injections. As was found in zone 2 conditions, there were differences in the regional distending pressure vs. flow curves among lung regions. To document the regional variability in the curves, the distribution of flow at a regional Ppv of 30 cmH2O above alveolar pressure was analyzed. There was a statistically significant linear gradient in this flow distribution from dorsal to ventral regions of the lungs but no consistent gradient in the caudad to cephalad direction. These results indicate that, even in near-maximally distended vessels, the dorsal regions of isolated perfused dog lungs have lower intrinsic vascular resistance compared with ventral regions.     

Excitatory action of angiotensins II and IV on hippocampal neuronal activity in urethane anesthetized rats

The renin–angiotensin system of the mammalian brain seems to interfere with the process of cognition and has been associated with the hippocampal function in relation to mechanisms of learning and memory. In our investigation, the effects of angiotensin II (Ang II) and angiotensin IV (Ang II) on neuronal activity have been studied in the hippocampus of adult rats anesthetized with urethane. Excitatory effects of both angiotensins predominated over inhibitory effects. Angiotensins also induced an enhancement of burst discharges. These angiotensin-induced effects were blocked by the specific angiotensin antagonists. Our findings showed that the different effects of Ang II and Ang IV in behavioral studies are not similarly reflected in a different change of the discharge rate and/or pattern of hippocampal neurons after microiontophoretic administration of both substances.     

Effect of angiotensin II on vascular resistance in whole-body perfused dogfish

• 1.1. The effect of angiotensin II (AII), norepinephrine (NE), epinephrine (E) and isoproterenol (ISO) was observed on the branchial and systemic circulations in a whole-body-pump perfused dogfish preparation.
• 2.2. NE and E increased systemic blood flow resistance, but decreased branchial resistance.
• 3.3. ISO decreased both systematic and branchial blood flow resistance.
• 4.4. AII had no significant effect on either branchial or systemic resistance.

     

Action of brain cathepsin B,cathepsin D,and high-molecular-weight aspartic proteinase on angiotensins I and II

The action of three previously isolated electrophoretically homogeneous brain proteinases—cathepsin B (EC 3.4.22.1), cathepsin D (EC 3.4.23.5), and high-molecular-weight aspartic proteinase (M_r=90K; EC 3.4.23.−)—on human angiotensins I and II has been investigated. The products of enzymatic hydrolysis have been identified by thin-layer chromatography on Silufol plates using authentic standards and by N-terminal amino acid residue analysis using a dansyl chloride method. Cathepsin D and high-molecular-weight aspartic proteinase did not split angiotensin I or angiotensin II. Cathepsin B hydrolyzed angiotensin I via a dipeptidyl carboxypeptidase mechanism removing His-Leu to form angiotensin II, and it degraded angiotensin II as an endopeptidase at the Val³-Tyr⁴ bond. Cathepsin B did not split off His-Leu from Z-Phe-His-Leu. Brain cathepsin B may have a role in the generation and degradation of angiotensin II in physiological conditions. Special Issue dedicated to Dr. Eugene Kreps.     

Analysis of angiotensins I,II, III,and iodinated derivatives by high-performance liquid chromatography

Angiotensins I, II, and III were separated by reversed-phase high-performance liquid chromatography on an octadecylsilyl column. The peptides were isocratically eluted with 50 mm NaH₂PO₄-25% ($\text{v}\text{v}$) acetonitrile, pH 6.0. The retention times were 3.3, 6.0, and 9.6 min for angiotensin II, III, and I, respectively. ¹²⁵I-Angiotensins II, III, and I eluted with retention times of 5.4, 16.8, and 19.9 min, respectively, under the same chromatographic conditions used for the unlabeled angiotensins. The effect of iodination of the tyrosine residue on the retention time was also demonstrated by chromatographic comparison of tyrosine and diiodotyrosine. Saralasin (Sar¹, Ala⁸-angiotensin II), a partial agonist of angiotensin II, and des-Asp¹, Ile⁸-angiotensin II, an inhibitor of angiotensin III, eluted with retention times of 2.5 and 3.9 min, respectively.     

In vitro renin inhibition to prevent generation of angiotensins during determination of angiotensin I and II

To prevent in vitro generation of angiotensins, the renin inhibitor CGP 29287 (CGP) was added to blood sampling tubes. Plasma immunoreactive angiotensin (ir-ANG) I and II were simultaneously measured by radioimmunoassay after rapid and quantitative extraction from a single plasma sample on phenylsilylsilica (Bondelut PH). True plasma ANG-(1-8)octapeptide was determined after additional separation of the different angiotensins by high performance liquid chromatography. Ir-ANG II/CGP showed the known linear relationship with ANG-(1-8)octapeptide (r = 0.87, n = 23), but - in contrast to studies without addition of CGP - the y-axis intercept which presumably represents cross-reacting angiotensins other than ANG II was very small. Ir-ANG II/CGP concentrations fell below 1 fmol/ml after converting enzyme inhibition. The results suggest that CGP 29287 prevents in vitro generation of ANG I and ANG II as well as the ANG-metabolites. Ir-ANG I/CGP measured after Bondelut PH extraction of the plasma was strongly correlated with ir-ANG I obtained after blood ethanol extraction (r = 0.97, n = 23). Thus, it is now possible to measure reliably both ANG I and ANG II within the same plasma extract after a simple extraction procedure.     

Influence of angiotensins (I,II & III) on prostaglandin production by minced rat renal medulla

Minced rat renal medulla was incubated for 30 min at 37 °C in the presence of angiotensin, I, II or III (100 ng/ml) to determine the existence of a direct stimulating effect on prostaglandin (PG) production. PGE₂, PGF_2α, 6-keto PGF_1α and Thromboxane B₂ (TXB₂)_were determined by radioimmunoassay.For analysis of data variance, the results were separated according to whether the net output of PGE₂ was above or below 1.5 ng PGE₂ equivalent/mg tissue/30 min. Under low-output conditions, angiotensin I, II or III stimulated PGE₂ production significantly (p<0.02) and tended to augment PGF_2α production, while under high-output conditions no effect on PGE₂ or PGF_2α production was observed.Under either output condition, angiotensin I, II or III had no effect on 6-keto PGF_1α and TXB₂.     

Enzymatic formation of angiotensins II and III in the hindlimb circulation of dogs

Experiments were performed in 14 anesthetized dogs to (1) to determine if the reductions in hindlimb blood flow produced by [des-Asp1] angiotensin I were due to its local enzymatic (kininase II) conversion to angiotensin III and (2) to quantitate the extent of conversion of angiotensin I to angiotensin II and of [des-Asp1] angiotensin I to angiotensin III in the hindlimb circulation. Graded doses of these peptides were administered as bolus injections directly into the left external iliac artery while measuring flow in this artery electromagnetically. Dose-response relationships were determined before and during the inhibition of kininase II activity with captopril or antagonism of angiotensin receptor sites with [Ile7] angiotensin III. Captopril inhibited the vasoconstrictor responses to angiotensin I and [des-Asp1] angiotensin I, but did not affect the responses to angiotensins II or III, or norepinephrine. [Ile7] angiotensin III inhibited the vasoconstrictor responses to all four angiotensin peptides but did not alter the responses to norepinephrine. These findings indicate that the hindlimb vasoconstrictor responses to [des-Asp1] angiotensin I were due to the local formation of angiotensin III. The extent of conversion of [des-Asp1] angiotensin I to angiotensin III that occurred in one transit through the hindlimb arterial circulation was estimated to be 36.7%, which was not different from the estimated 36.4% conversion of angiotensin I to angiotensin II. We conclude that angiotensin I and [des-Asp1] angiotensin I are converted to their respective vasoactive forms (angiotensins II and III) to a similar extent in the hindlimb circulation via the action of kininase II.     

Influence of angiotensins (I,II, & III), bradykinin and arachidonic acid on renomedullary PGE production in vitro

Renomedullary tissue from rabbit or rat was incubated with angiotensin I, II, III, arachidonic acid, bradykinin, indomethacin and meclofenamate to study their effect on PGE₂ production.Arachidonic acid and bradykinin enhanced PGE₂ production significantly. Indomethacin and meclofenamate inhibited PGE₂ production by more than 70%. Angiotensin I, II and III did not influence PGE₂ production. These results suggest that bradykinin and arachidonic acid stimulate PGE₂ production by a direct cellular action whereas the angiotensins do not.     

In vitro studies on the subcellular location of glucosidase I and glucosidase II in dog pancreas

When programmed with yeast prepro--factor mRNA, the heterologous reticulocyte/dog pancreas translation system synthesizes two pheromone related polypeptides, a cytosolically located primary translation product (pp--F_cyt, 21 kDa) and a membrane-specific and multiply glycosylated e-factor precursor (pp--F₃, 27.5 kDa). Glycosylation of the membrane specific pp--F₃ species is competitively inhibited by synthetic peptides containing the consensus sequence Asn-Xaa-Thr as indicated by a shift of its molecular mass from 27.5 kDa to about 19.5 kDa (pp--F₀) , whereas the primary translation product pp--Fcyt is not affected. Likewise, only the glycosylated pp--F₃ structure is digested by Endo H yielding a polypeptide with a molecular mass between PP--F₀ and pp--Fcyt. These observations strongly suggest that the primary translation product is proteolytically processed during/on its translocation into the lumen of the microsomal vesicles. We believe that this proteolytic processing is due to the cleavage of a signal sequence from the pp--Fcyt species, although this interpretation contradicts previous data from other groups. The distinct effect exerted by various glycosidase inhibitors (e.g. 1-deoxynojirimycin, N-methyl-dNM, 1-deoxymannojirimycin) on the electrophoretic mobility of the pp--F₃ polypeptide indicates that its oligosaccharide chains are processed to presumbly Man₉-GlcNAc₂ structures under thein vitro conditions of translation. This oligosaccharide processing is most likely to involve the action of glucosidase I and glucosidase II as follows from the specificity of the glycosidase inhibitors applied and the differences of the molecular mass observed in their presence. In addition, several arguments suggest that both trimming enzymes are located in the lumen of the microsomal vesicles derived from endoplasmic reticulum membranes.Abbreviations dNM 1-deoxynojirimycin - N-Me-dNM N-methyl-dNM - dMM 1-deoxymannojirimycin - CCCP carbonylcyanide m-chlorophenyl hydrazone