Isolated Rabbit Aortic Rings Research Articles

Oxidative stress increases glyceraldehyde-3-phosphate dehydrogenase mRNA levels in isolated rabbit aorta.

We have recently shown that inhibition of endogenous Cu,Zn superoxide dismutase (SOD) by diethyldithiocarbamate (DDC) increased superoxide anion levels in isolated rabbit aortic rings, describing a useful experimental model to examine the effects of oxidative stress on the vessel wall. The present study examined the effects of oxidative stress on the steady-state mRNA levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.12). Aortic rings were incubated in physiological salt solution at 37 degrees C for up to 6 h. DDC (2 mM) decreased total SOD activity to < 5% of control levels and increased superoxide anion level ninefold. Steady-state mRNA levels of GAPDH were increased under comparable conditions. Although decreased biological activity of endothelium-derived nitric oxide was indicated by lower basal guanosine 3',5'-cyclic monophosphate levels in aortic rings treated with DDC compared with those in control rings (1.2 +/- 0.1 vs. 1.9 +/- 0.3 fmol/microgram protein, P < 0.05), neither endothelium denudation nor NG-nitro-L-arginine methyl ester had any effects on the steady-state mRNA levels of GAPDH. The cell.permeable iron chelator 1,10-phenanthroline completely prevented the increases in GAPDH mRNA levels induced by DDC. These results suggest that oxidative stress resulting from inhibition of endogenous Cu,Zn SOD causes induction of GAPDH gene expression and that the hydroxyl radical, produced through the iron-catalyzed Haber-Weiss reaction, is the intracellular reactive oxygen species responsible for the DDC-stimulated increase in GAPDH mRNA.

Angiographic contrast media relax isolated rabbit aorta through an endothelium-independent mechanism that may not depend on the presence of the iodine atom

Systemically administered iodinated angiographic contrast media evoke vasodilatation through mechanisms that are at present poorly understood. In the current investigation we have evaluated the role of the vascular endothelium in responses to an iso-osmolar formation of the non-ionic dimer iodixanol and a hyperosmolar formulation of the non-ionic monomer iopromide. Isolated rabbit aortic ring preparations with endothelium intact or removed by gentle abrasion were mounted in organ baths containing oxygenated Holman's solution, and cumulative concentration-response curves for relaxation to the contrast media were constructed after pre-constriction by phenylephrine (300 nM) in the presence of indomethacin to inhibit prostaglandin synthesis. Endothelial denudation did not influence the ability of either iodixanol or iopromide to relax the aortic ring preparations. Iopromide was significantly more potent than iodixanol when expressed in terms of iodine concentration (mg I ml-1), but both agents were equipotent when expressed in terms of molarity (mM). We conclude that relaxation of isolated rabbit aortic rings to iodixanol and iopromide under conditions where there is no fluid flow is endothelium-independent, and therefore not mediated by release of the potent endogeneous nitrovasodilator endothelium-derived relaxing factor (EDRF). Furthermore, their relaxant activity under the in vitro experimental conditions employed is attributable to a direct action on vascular smooth muscle by factors in addition to osmolality, and may depend on features that are not specifically associated with the presence of the iodine atom.

Recombinant Human Hemoglobin Inhibits Both Constitutive and Cytokine-Induced Nitric Oxide-Mediated Relaxation of Rabbit Isolated Aortic Rings

A genetically engineered recombinant human hemoglobin (rHb1.1) was recently developed for use as a blood substitute (Nature 1992;356:258-60). Like other mammalian hemoglobin (Hb) molecules, it might bind and antagonize the actions of nitric oxide (NO). We used an isolated rabbit aortic ring preparation to examine the ability of rHb1.1 to inhibit acetylcholine (ACh)- and interleukin-1 beta (IL-1 beta)-induced reductions of vasoconstrictor responses to the alpha-adrenoceptor agonist phenylephrine (PE). rHb1.1 (0.04-4.4 microM) rapidly and reversibly inhibited, in a concentration-dependent manner, both ACh- and IL-1 beta-induced decreases in PE contractile responses. These inhibitory effects of rHb1.1 were non-competitive and were equipotent to those of purified, cell-free human Hb (p.hHb). These two forms of soluble Hb were at least 10 times more potent than Hb in erythrocytes (red blood cells: RBC-Hb). Both NG-nitro-L-arginine (10 microM) a NO synthase inhibitor, and LY-83583 (10 microM), a guanylyl cyclase inhibitor, mimicked the effects of rHb1.1. The inhibitory effects of rHb1.1 were not shared by either human serum albumin (HSA 44 microM), which combines with but does not deactivate NO, or cytochrome C (44 microM), a heme-containing protein that does not bind NO; neither were they reversed by L-arginine (L-ARG) (1 mM), the presumed NO precursor. These and other results suggest that the chemical antagonism of NO is likely to be the mechanism by which rHb1.1 and other Hbs inhibit ACh- and IL-1 beta-induced decreases in the response to PE in rabbit aortic rings.

Aminopeptidase modulation of the pharmacological responses to synthetic thrombin receptor agonists

Thrombin is a contractile stimulus of isolated rabbit aortic rings and apparently produces its effects through the recently characterized cleavable receptor. A synthetic hexapeptide, NAT 6-NH 2 (new amino terminus), was found to be the minimal active structure for full activation of this receptor. The N-terminal Ser residue of NAT 6-NH 2 is crucial for biological activity. In this study we examined the metabolism of NAT 6-NH 2 in rabbit plasma, where it was rapidly degraded by aminopeptidase M. In the presence of the aminopeptidase inhibitor amastatin, no metabolism was observed. On this basis a metabolically resistant analogue, [Sar 1]NAT 6-NH 2, was designed. We compared the biological activity of thrombin, NAT 6-NH 2 and [Sar 1]NAT 6-NH 2 in the rabbit aorta and found that [Sar 1]NAT 6-NH 2 was more potent than NAT 6-NH 2; however, in the presence of amastatin the concentration-effect curve for NAT 6-NH 2 was shifted to the left of that for [Sar 1]NAT 6-NH 2. The effects of [Sar 1]NAT 6-NH 2 and of thrombin were not modified by the presence of the aminopeptidase inhibitor. We also studied the effect of amastatin on the in vivo hypotensive response to NAT 6-NH 2 and found that it was also influenced by aminopeptidase M inhibition. Our results show that aminopeptidase protection is important when evaluating responses to synthetic agonists of the thrombin cleavable receptor and that an in vivo model, the anesthesized and heparinized rabbit, may be useful for the development of agonists and antagonists of this receptor.

Conversion of Nitroxyl (HNO) to Nitric Oxide (NO) in Biological Systems: The Role of Physiological Oxidants and Relevance to the Biological Activity of HNO

Nitroxyl (HNO) and nitric oxide (NO) are chemically related compounds in that NO is the one-electron oxidation product of HNO. Previous studies from this laboratory indicated that HNO elicits pharmacological effects that are similar to those elicited by NO, namely, vascular smooth muscle relaxation and stimulation of cyclic GMP formation. The objective of the present study was to determine whether HNO could be converted to NO under physiological conditions and thereby account for the pharmacological actions of HNO. Utilizing the method of chemiluminescence detection, HNO was found to he readily converted to NO by a variety of ubiquitous biological oxidants including oxygen, superoxide dismutase, methemoglohin and flavins. The potency of HNO as a vasorelaxant using isolated rabbit aortic rings was markedly increased 30-fold by superoxide dismutase, whemas the potency of the NO-donor compound, S-nitroso-N-acetylpenicillamine (SNAP), was increased only 2-fold. These data indicate that the ready conversion of HNO to NO may account for the biological activity of HNO. Thus, HNO and HNO-donor compounds represent good sources of NO.

Inhibition of EGF-Induced Vasoconstriction in Isolated Rabbit Aortic Rings with the Tyrosine Kinase Inhibitor RG50864

The tyrosine kinase inhibitor RG50864 attenuated EGF-induced tension development dose-dependently. Similarly, contractions to a phorbolester were affected by 10 μM RG50864. In comparison, vasoconstrictor activities of norepinephrine, endothelin and Bay K 8644 remained unaltered. Western blots using antiphosphotyrosine antibodies, revealed a time-dependent increase in EGF-induced EGF-receptor autophosphorylation as well as tyrosine phosphorylation of a 55 kDalton cytosolic protein. While the extent of EGF-receptor autophosphorylation remained unaltered in the presence of 10 μM RG50864, phosphorylation of the 55 kD hand was decreased two-fold. In summary, in rabbit aorta RG50864 is an inhibitor of EGF-induced vasoconstriction; this inhibitory effect does not appear to he mediated through inhibition of EGF-receptor autophosphorylation but may involve a 55 kD cytosolic protein substrate.

Role of superoxide anion and endothelium in vasoconstrictor action of prostaglandin endoperoxide.

The vasoconstrictor actions of prostaglandin (PG) endoperoxide, PGH2, were examined in isolated rabbit aortic rings suspended for measurement of isometric tension. In aortic rings with an intact endothelium, PGH2 caused concentration-dependent contractions which were blocked by SQ 29548 (a PGH2-thromboxane A2 receptor blocker) or superoxide dismutase (a superoxide anion scavenger) but not by carbethoxyhexyl imidazole (a thromboxane A2 synthase inhibitor) or catalase (a hydrogen peroxide scavenger). In contrast U 46619, a thromboxane A2 mimic, caused contractions, which were blocked by SQ 29548 but not by superoxide dismutase. PGH2 caused significantly greater contractions in aortic rings without endothelium or in those with intact endothelium treated with NG-nitro-L-arginine, a nitric oxide inhibitor; these contractions were inhibited by SQ 29548 but not by superoxide dismutase. In aortic rings with endothelium contracted with phenylephrine, a subthreshold concentration of PGH2, but not U 46619, impaired relaxations to acetylcholine; the inhibition was prevented by treatment with SQ 29548 or superoxide dismutase, indicating that the abnormality of endothelial cell function was specific for PGH2. These observations indicate that PGH2 causes contractions and inhibits endothelium-dependent relaxation by a mechanism involving formation of superoxide anion, which interacts with endothelium-derived nitric oxide.

Effects of vanadate on vascular contractility and membrane potential in the rabbit aorta.

Isolated rabbit aortic ring with intact endothelial cell preparations precontracted with NE (10(-7) M) were relaxed by vanadate in a dose dependent manner (from 0.2 to 2 mM). Application of vanadate and ACh during the tonic phase of high K+(100 mM)-induced contraction showed a slight relaxation in contrast to that in NE-induced contraction, but sodium nitroprusside (10 microM) more effectively relaxed the aortic ring preparations in high K+ contraction than that of vanadate. Vanadate-induced relaxation in NE-contracted aortic rings was reversed by application of BaCl2 (50 microM) or glibenclamide (10 microM). Furthermore, Vanadate hyperpolarized membrane potential of smooth muscle cells in endothelium-intact aortic strips and this effect was abolished by application of glibenclamide. The above results suggest that vanadate release EDHF (Endothelium-Derived Hyperpolarizing Factor), in addition to EDRF (Endothelium-Derived Relaxing Factor) from endothelial cell. This EDHF hyperpolarize the smooth muscle cell membrane potential via opening of the ATP-sensitive K+ channel and close a voltage dependent Ca++ channel. So it is suggested that the vanadate-induced relaxation of rabbit thoracic aortic rings may be due to the combined effects of EDRF and EDHF.

EFFECT OF TOLERANCE TO GLYCERYL TRINITRATE ON VASCULAR RESPONSES IN CONSCIOUS RABBITS

1. The effect of tolerance to glyceryl trinitrate (GTN) on vasodilator and vasoconstrictor responses was examined in conscious rabbits and isolated rabbit aortic rings. 2. In conscious rabbits, depressor responses to 5 min infusions of GTN (10-40 micrograms/kg per min intravenously (i.v.)), sodium nitroprusside (SNP, 5-20 micrograms/kg per min i.v.) and acetylcholine (ACh, 3-12 micrograms/kg per min i.v.) were examined before and after transdermal treatment with GTN (20 mg/48 h). GTN pretreatment significantly attenuated GTN-induced depressor responses, indicating the development of tolerance, but did not affect the reductions in arterial pressure induced by SNP or ACh. 3. Similarly, aortic rings taken from GTN pretreated rabbits exhibited tolerance to GTN but the relaxant responses to SNP or the calcium ionophore A23187 were not affected. In the aortic rings from GTN-tolerant rabbits contractile responses to serotonin or the thromboxane-mimetic U46619 were significantly attenuated, in contrast to the responses to the alpha 1-adrenoceptor agonist phenylephrine (PE) which were significantly enhanced. 4. Similarly, in conscious rabbits, PE-induced increases in arterial pressure and hindlimb vascular resistance were significantly enhanced by GTN pretreatment but the responses to the alpha 2-adrenoceptor agonist BHT 920 were unaffected. 5. In conclusion, tolerance to GTN does not affect endothelium-dependent vasodilatation but does cause a selective enhancement of alpha 1- but not alpha 2-adrenoceptor-mediated vasoconstriction.

A novel serine protease with vasoconstrictor activity coded by the kallikrein gene S3.

A fraction separated from rat submandibular gland homogenates was found to contain a potent vasoconstrictor when tested on isolated rabbit aortic rings. The vasoconstrictor was purified by a series of chromatographic steps. The purified compound (2.77 x 10(-9) M) induced 40% of the maximum contractile response to 60 mM KCl. Constriction was slow in onset, long-lasting, rinse-resistant, and unchanged by de-endothelialization; in addition, it was dose-related and inhibited by both EGTA and verapamil, but it was not affected by DUP753, an angiotensin II receptor antagonist. The compound was found to be a protein having a pI of 7.36 and a molecular weight of approximately 29,000 and exhibiting partial immunologic identity to rat glandular kallikrein and rat tonin. After 2-mercaptoethanol treatment, it separated into heavy (approximately 19,900) and light (approximately 10,700) chains having amino-terminal sequences of AY(X)HNNDLMLL and VVGGYN(X)ETNSQ, respectively. We found that they correspond to the amino-terminal and internal sequence of a previously unidentified kallikrein-like serine protease whose mRNA, named S3, has been found in the rat submandibular gland and prostate. The vasoconstrictor is able to hydrolyze t-butoxycarbonyl-valine-proline-arginine-methylcoumarin amide (a thrombin substrate), although its Kcat/Km was only 0.02% that of rat thrombin. Both vasoconstrictor and enzymatic activity on t-butoxycarbonyl-valine-proline-arginine-methylcoumarin amide were completely suppressed by amidinophenylmethylsulfonyl fluoride and soybean trypsin inhibitor; however, they were unaffected by hirudin, a thrombin inhibitor. At pH 6.5, it released angiotensin II when incubated with sheep angiotensinogen, although it had approximately one-tenth the activity of tonin. The submandibular enzymatic vasoconstrictor is a kallikrein-like enzyme, having some properties of both tonin and thrombin. It directly contracts vascular smooth muscle, acting via a mechanism that requires intact enzymatic activity.

Central and peripheral actions of a nonpeptidic angiotensin II receptor antagonist.

Nonpeptidic imidazole derivatives were recently reported to be angiotensin II receptor antagonists with acute blood pressure-lowering activity. In the present study, we characterized the angiotensin II receptor antagonist properties of one such derivative, 4'-([2-butyl-4-chloro-5-(hydroxymethyl)-1H-imidazol-1-yl]methyl)- [1,1'-biphenyl]-2-carboxylic acid (IMI). In receptor binding studies, IMI displaced bound [125I]angiotensin II from rat uterine membranes with an IC50 of 0.17 microM. In isolated rabbit aortic rings, IMI shifted the angiotensin II concentration-response curve to the right in a parallel and concentration-dependent manner. A Schild plot of these data indicated a pA2 of 7.13 +/- 0.16 and a slope of 0.94 +/- 0.06. In rat kidney slices, IMI shifted the concentration-response curve for angiotensin II-induced inhibition of renin release to the right. Antagonism of the angiotensin II pressor response by IMI was dose dependent and reversible in ganglion-blocked, anesthetized rats. The water intake and pressor responses to intracerebroventricular angiotensin II (100 pmol) were inhibited by intracerebroventricular IMI (25 or 50 nmol) in conscious Sprague-Dawley rats. Similarly, the drinking and pressor responses to intravenous angiotensin II were blocked by intravenous IMI in conscious rats. IMI alone had no effects on mean arterial pressure or drinking when administered either intravenously or intracerebroventricularly. IMI decreased mean arterial pressure throughout 5 days of infusion in spontaneously hypertensive rats. In summary, IMI was a full competitive antagonist without partial agonist activity in peripheral tissues and the central nervous system. Moreover, the chronic administration of this angiotensin II receptor antagonist was antihypertensive in spontaneously hypertensive rats.

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.

Biosynthesis and modulation of endothelin from bovine pulmonary arterial endothelial cells

The biosynthesis and modulation of the vasoconstrictor peptide endothelin was studied in the conditioned medium from cultured bovine pulmonary artery endothelial (BPAE) cells. Conditioned medium from cultured BPAE cells produced contraction of isolated rabbit aortic rings. Incubation of BPAE cells with the protease inhibitors TPCK or isatoic anhydride attenuated the extent of conditioned medium-induced contractions. Incubation of BPAE cells with thrombin produced an enhancement of conditioned medium-induced contraction by approximately 25%. Endothelin levels in conditioned medium were measured by RIA and incubation of BPAE cells with TPCK or isatoic anhydride significantly reduced endothelin levels, whereas incubation with thrombin or transforming growth factor beta-1 stimulated the levels of endothelin in the conditioned medium. These data indicate that endothelin may be modulated by certain protease inhibitors and by platelet and immune cell mediators and suggest a potential new mode of vascular tone regulation.

Endothelin-1-Induced Vascular Contractions

Recently, a potent vasoconstrictor peptide, endothelin-1 (ET-1), was isolated from vascular endothelial cells. We investigated the inhibition of ET-1-induced contractions on isolated porcine coronary artery and rabbit aorta by calcium antagonists of the 1,4-dihydropyridine type. In addition, the vasoconstriction induced by ET-1 was compared with that induced by the calcium agonist (-)Bay K 8644. The calcium antagonists nitrendipine and nicardipine partially antagonize the vasoconstrictive effects of ET-1 in a dose-dependent manner, but this antagonism is only functional and noncompetitive. In the isolated rabbit aortic ring, ET-1 induces a half-maximal contraction. In contrast, the calcium agonist (-)Bay K 8644 has no effect on this preparation. However, when the vessels are partly depolarized by adding 15 mM K+, (-)Bay K 8644 and ET-1 express their full activity. Again here, the concentration-response curve of ET-1 is depressed by increasing concentrations of nitrendipine, but only functionally, noncompetitively. These results suggest that ET-1 acts through specific receptors different from the 1,4-dihydropyridine selective site.

Effects of Calcium Channel Activators on the Response of Rabbit Aortic Rings to Alpha&lt;sub&gt;1&lt;/sub&gt;-Receptor Agonists: Evidence for Two Modes of Action

The interaction between activators of the calcium channel and the alpha 1-adrenoceptor has been studied in isolated rabbit aortic rings. The data obtained in these investigations are not consistent with the idea of a homogenous population of alpha 1-receptors which trigger contraction via a single pathway. Our results suggest that calcium channel activators reveal two modes of spasmogenic action for alpha 1-adrenoceptor agonists. Furthermore, we have shown that there may be multiple drug recognition sites on the alpha 1-receptor.

Endothelin action on vascular smooth muscle involves inositol trisphosphate and calcium mobilization

Cultured endothelial cells release a potent vasoconstrictor peptide, endothelin. Cumulative addition of synthetic endothelin to isolated rabbit aortic rings elicited a concentration-dependent increase in contractile tension which was endothelium-independent. In cultured rabbit vascular smooth muscle cells loaded with the fluorescent dye fura 2, endothelin induced a concentration-dependent increase in [Ca2+]i over the range of 0.01 to 100 nM. Moreover, in the absence of extracellular Ca2+, endothelin could still induce an increase in [Ca2+]i. In addition, endothelin stimulated 45Ca2+ efflux from preloaded vascular smooth muscle cells in the presence and absence of extracellular Ca2+, as well as stimulating 45Ca2+ influx in a concentration-dependent manner. Measurement of inositol phosphates in [3H]-myoinositol-labelled vascular vascular trisphosphate. Unlabelled endothelin inhibited (125I)-endothelin binding to cultured rabbit vascular smooth muscle cells in a concentration-dependent manner. Binding was not inhibited by other vasoactive hormones or calcium channel ligands, suggesting cell surface receptors specific for endothelin. We conclude that one of the initial membrane events in the action of endothelin is to induce phospholipase C-stimulated PIP2 hydrolysis and that this signalling mechanism is initiated by endothelin/receptor interaction at the plasma membrane.

Endothelium-dependent relaxation of rabbit aorta by acetylcholine requires ethylenediaminetetraacetic acid.

Experiments were conducted to determine if ethylenediaminetetraacetic acid (EDTA) was essential for the acetylcholine (ACh)-induced relaxation of blood vessels. Isolated rabbit aortic rings were prepared for recording isometric tension. They were maintained in Krebs bicarbonate solution with various concentrations of EDTA. With EDTA concentrations of 0 or 0.003 mM, no ACh-induced relaxation was observed; only the contractile effect of ACh was seen. With 0.03 and 0.30 mM EDTA, ACh induced relaxation with EC50 values of 0.11 and 0.098 microM, respectively. Under the experimental conditions used, EDTA was essential for demonstration of ACh-induced relaxation.

Influence of endothelium on drug-induced relaxation of the rabbit aorta

The influence of endothelium on relaxation induced by glyceryl trinitrate (GTN) and isoprenaline (I) has been studied in isolated rabbit aortic ring preparations, in the resting state and after constriction with 5-hydroxytryptamine (5HT). In resting preparations relaxant responses to GTN and I were smaller in the presence of endothelium. In preparations constricted with 5HT relaxant responses to GTN and I were conversely greater in the presence of endothelium. These paradoxical findings are discussed in relation to basal release of endothelium derived relaxant factor.

Stereoselectivity at the Calcium Channel

The stereoisomers of the new dihydropyridine derivative 202-791 [isopropyl 4-(2,1,3-benzoxadiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5-nitro-3 -pyridinecarboxylate] were synthesized separately and tested on isolated rabbit aortic rings for effects on depolarization-induced contraction and depolarization-stimulated uptake of 45Ca2+. The racemic mixture enhanced contraction of rabbit aortic rings at low levels of depolarization but inhibited contraction and 45Ca2+ uptake at high levels of depolarization. The IC50 values were 2.0 X 10(-7) and 1.7 X 10(-7) M, respectively. The R enantiomer inhibited contraction and 45Ca2+ uptake with IC50 values of 3.2 X 10(-8) and 4.3 X 10(-8) M, respectively. This compound showed no stimulant activity. By contrast, the S enantiomer of 202-791 shifted the concentration-response curve for depolarization-induced contraction in an almost parallel fashion to the left, thus enhancing contraction. The EC50 value for this effect at a KCl concentration of 16 mM was 1.8 X 10(-7) M. This compound enhanced 45Ca2+ uptake concentration dependently at all levels of depolarization tested. Thus, the stereoisomers of a dihydropyridine derivative may behave as a calcium entry blocker or a calcium entry enhancer on vascular smooth muscle, depending only on the stereochemistry. If asymmetric compounds elicit effects suggesting a dualistic action at the calcium channel, then the stereoisomers should be prepared.

Some Recent Pharmacological Findings with Nitrendipine

The available evidence indicates that nitrendipine and other dihydropyridines with a similar pharmacological action exert their therapeutic effects by inhibiting Ca2+ channels. In our recent experiments, nitrendipine was shown to block K+-stimulated 45Ca2+ uptake and K+-induced contractions of isolated rabbit aortic rings. Its IC50 were 4.7 and 8.9 nM for inhibition of Ca2+ uptake and of contractions, respectively. There was no statistically significant difference between the two values. At higher concentrations, nitrendipine also reduced norepinephrine-induced 45Ca2+ uptake and norepinephrine-induced contractions of rabbit aortic strips. The norepinephrine-induced contractions were only slightly (21%) reduced by nitrendipine at 10 microM. Nitrendipine at 10 nM and higher concentrations inhibited K+- or angiotensin-II-(AII) induced release of aldosterone from isolated bovine adrenal glomerulosa cells. The drug was more potent and more effective in inhibiting K+- than AII-induced aldosterone release. Dantrolene, 25 microM, enhanced the inhibitory activity of nitrendipine on AII-stimulated aldosterone release. Acute renal failure produced by either glycerol or gentamicin in rats was antagonized by nitrendipine at oral doses of 15-25 mg/kg/day. Our studies confirmed previously reported observations that the usefulness of nitrendipine in the treatment of hypertension may be determined not only by its vasodilator action. We demonstrated that nitrendipine has a direct inhibitory effect on the release of aldosterone from adrenal glomerular cells. In addition to a previously described diuretic action, nitrendipine was shown to have renal cytoprotective activity.