Ring Preparations Research Articles

Direct Vasoactive Properties of Thienopyridine-Derived Nitrosothiols

Thienopyridines (ticlopidine, clopidogrel, and prasugrel) require in vivo metabolism to exhibit a critical thiol group in the active form that binds to the P2Y12 platelet receptor to inhibit platelet activation. We hypothesized that formation of thienopyridine-derived nitrosothiols (ticlopidine-SNO, clopidogrel-SNO, and prasugrel-SNO) occurs directly from the respective parent drug. Pharmaceutical-grade thienopyridine (ticlopidine, clopidogrel chloride, clopidogrel sulfate, clopidogrel besylate, or prasugrel) was added to nitrite in aqueous solution to form the respective thienopyridine-SNO (Th-SNO). An isolated aortic ring preparation was used to test vasoactivity of the Th-SNO derivatives. Increasing nitrite availability resulted in increased Th-SNO formation for all drugs (other than ticlopidine). Th-SNO induced significant endothelium-independent relaxation of preconstricted aortic rings. Clopidogrel-chloride-SNO displayed rapid-release kinetics in a chemical environment, which was reflected by immediate and transient vasorelaxation when compared with the SNO derivatives of the other thienopyridines. Accounting for differences in yield, clopidogrel-chloride-SNO exhibited the greatest propensity to immediately relax vascular tissue. Th-SNO derivatives exhibit nitrovasodilator properties by supplying NO that can directly activate vascular soluble guanylate cyclase to induce vasorelaxation. Differences in SNO yield and vasoactivity exist between thienopyridine preparations that might be important to our understanding of the direct pharmacological effectiveness of thienopyridines on vascular and platelet function.

Chronic β-adrenoceptor Antagonist Treatment Controls Cardiovascular Remodeling in Heart Failure in the Aging Spontaneously Hypertensive Rat

Selective β1-adrenoceptor antagonists are part of standard therapy to prolong survival in human heart failure. This study has measured structural, functional, and electrical changes in the cardiovascular system of aging male spontaneously hypertensive rats (SHRs) to determine whether β1-adrenoceptor antagonist treatment can prevent or reverse the development of cardiovascular remodeling and heart failure in these rats. Fifteen-month-old male Wistar-Kyoto (WKY) rats or SHRs were treated with increasing metoprolol doses (30 mg·kg·d for 4 weeks, then 50 mg·kg·d for 4 weeks, then 80 mg·kg·d for 16 weeks po). Cardiovascular structure and function were determined using organ wet weight, in vivo echocardiography, histological analysis of inflammation and collagen, isolated heart and thoracic aortic ring preparations, and single cell microelectrode measurements. From 15 months, untreated SHRs developed left ventricular dilation, hypertrophy, inflammatory cell infiltration and fibrosis, and action potential prolongation together with progressive systolic, diastolic, and endothelial dysfunction and increased cardiac stiffness. Treatment with metoprolol decreased systolic blood pressure at 21 months only but improved survival, decreased ventricular weight, prevented chamber dilation, reduced inflammation, decreased fibrosis, attenuated action potential prolongation, improved systolic and diastolic function, decreased stiffness and improved endothelium-independent vascular responses. Chronic metoprolol treatment markedly attenuated both cardiac and vascular remodeling in the aging SHRs, thus attenuating the onset of heart failure and improving survival, independent of blood pressure reduction.

Fenofibrate attenuates nicotine-induced vascular endothelial dysfunction in the rat

The study has been designed to investigate the effect of fenofibrate on nicotine-induced vascular endothelial dysfunction (VED) in rats. Nicotine (2mg/kg/day, i.p., 4weeks) was administered to produce VED in rats. The development of VED was assessed by employing isolated aortic ring preparation and estimating serum and aortic concentration of nitrite/nitrate. Further, the integrity of vascular endothelium was assessed using the scanning electron microscopy of thoracic aorta. The expression of mRNA for p22phox and eNOS was assessed by using reverse transcriptase–polymerase chain reaction. Serum thiobarbituric acid reactive substances concentration (TBARS) and aortic superoxide anion concentration were estimated to assess oxidative stress. Moreover, the serum lipid profile was assessed by estimating serum cholesterol, triglycerides and high density lipoprotein. The administration of nicotine induces VED by increased oxidative stress, altered lipid profile and impaired the integrity of vascular endothelium as assessed in terms of decrease in expression of mRNA for endothelial nitric oxide synthase (eNOS), impairing the integrity of vascular endothelium and subsequently decreasing serum and aortic nitrite/nitrate and attenuating acetylcholine-induced endothelium dependent relaxation. Further, nicotine produced oxidative stress, assessed in terms of increase in serum TBARS and aortic superoxide anion generation and increase in expression of mRNA for p22phox. Nicotine altered the lipid profile by increasing the serum cholesterol, triglycerides and decreasing the high density lipoprotein. However, treatment with fenofibrate (32mg/kg, p.o.) markedly prevented nicotine-induced VED by decreasing oxidative stress and improving integrity of vascular endothelium, normalising the altered lipid profile, increasing the concentration of serum and aortic nitrite/nitrate, enhancing the acetylcholine-induced endothelium dependent relaxation and decreasing serum TBARS and aortic superoxide anion generation. Thus, it may be concluded that fenofibrate has vascular protecting potential, by improving the integrity and function of vascular endothelium.

Effect ofMucuna pruriensSeed Extract Pretreatment on the Responses of Spontaneously Beating Rat Atria and Aortic Ring toNaja sputatrix(Javan Spitting Cobra) Venom

Mucuna pruriens Linn. (velvet bean) has been used by native Nigerians as a prophylactic for snakebite. Rats pretreated with M. pruriens seed extract (MPE) have been shown to protect against the lethal and cardiovascular depressant effects of Naja sputatrix (Javan spitting cobra) venoms, and the protective effect involved immunological neutralization of the venom toxins. To investigate further the mechanism of the protective effect of MPE pretreatment against cobra venom toxicity, the actions of Naja sputatrix venom on spontaneously beating rat atria and aortic rings isolated from both MPE pretreated and untreated rats were studied. Our results showed that the MPE pretreatment conferred protection against cobra venom-induced depression of atrial contractility and atrial rate in the isolated atrial preparations, but it had no effect on the venom-induced contractile response of aortic ring preparation. These observations suggested that the protective effect of MPE pretreatment against cobra venom toxicity involves a direct protective action of MPE on the heart function, in addition to the known immunological neutralization mechanism, and that the protective effect does not involve action on blood vessel contraction. The results also suggest that M. pruriens seed may contain novel cardioprotective agent with potential therapeutic value.

ChemInform Abstract: Oxidation of Pyrrole‐2‐carboxylates with o‐Chloranil and Its Synthetic Application.

t-Butyl 3,4-dialkyl-1H-pyrrole-2-carboxylates were oxidized with o-chloranil in the presence of MeOH to afford the corresponding 5-methoxypyrrolin-2-one derivatives. The resulting 5-methoxypyrrolin-2-one was reacted with various nucleophiles under acidic conditions to afford the functionalized pyrrolinone derivatives in good yields. Toward elucidation of the structure and function of the linear tetrapyrrole (bilin) chromophores in phytochromes, we have succeeded in synthesizing phytochromobilin (PΦB), phycocyanobilin (PCB), modified PCBs, biliverdin (BV) and its analogs including sterically locked derivatives in free acid forms by developing efficient methods for the preparation of each pyrrole ring and a new coupling reaction between them. In order to synthesize different types of locked chromophores, it was necessary to prepare various pyrrole and pyrrolinone derivatives bearing a wide variety of functional groups. In our previous syntheses, pyrrole rings with an electron withdrawing group were constructed by a modified Barton reaction starting from aldehydes, 1-nitroalkanes, and isonitrile compounds. The produced tosyland ester-substituted pyrroles were converted into Aor D-ring and Bor C-ring, respectively. It would be ideal for the synthesis of the locked bilin chromophores if the various types of pyrroles and the related pyrrolinones could be available from a common pyrrole by a simple manipulation. Recently we have developed a regioselective oxidation of the α-position of the C-4 substituents on t-butyl pyrrole-2-carboxylates with DDQ and the subsequent substitution reaction via azafulvene intermediates (Scheme 1). Herein we describe an oxidation of t-butyl pyrrole-2-carboxylates with o-chloranil and its application toward the synthesis of various types of functionalized pyrrolinone derivatives. N H CO2 tBu DDQ AcOH N H CO2 tBu R OAc R Nu base N H CO2 tBu Nu

Inhibitory effects of sertraline in rat isolated perfused kidneys and in isolated ring preparations of rat arteries

Sertraline is often prescribed to patients suffering with end stage renal disease, but its action on kidney has not been investigated. We aimed to investigate the pharmacological action of sertraline on rat kidney with emphasis on the underlying mechanisms involved in the vascular actions of the drug. The effects of sertraline were evaluated in rat isolated perfused kidneys and on ring preparations of mesenteric or segmental rat renal artery. In kidneys, sertraline prevented the effects of phenylephrine on perfusion pressure, glomerular filtration rate, urinary flow and renal vascular resistance. In mesenteric rings sertraline inhibited phenylephrine-induced contractions with potency 30-times lower than verapamil. Sertraline reversed sustained contractions induced by phenylephrine or 60mm K(+) within a similar concentration range. In segmental isolated rings, sertraline also reversed contractions induced by phenylephrine or 60mm K(+) with the same concentration range, but with higher potency compared with mesenteric preparations. Under Ca(2+) -free conditions, sertraline did not change the intracellularly-mediated phasic contractions induced by phenylephrine or caffeine. Sertraline was ineffective against contractions induced by extracellular Ca(2+) restoration after thapsigargin treatment and Ca(2+) store depletion with phenylephrine. Conversely, sertraline decreased the contractions induced by Ca(2+) addition in tissues under high K(+) solution or phenylephrine plus verapamil. In rat isolated kidneys and in rat ring preparations of mesenteric or renal vessels, sertraline had antispasmodic effects that appeared to be caused by a direct action on vascular smooth muscle cells. Its actions were ineffective against Ca(2+) -releasing intracellular pathways, but appeared to interfere with sarcolemmal Ca(2+) influx with reduced permeability of both receptor- and voltage-gated Ca(2+) channels.

Reactivity of intraosseous femoral head arteries after long term glucocorticoid treatment: An experimental study in immature pigs

In the pathogenesis of steroid induced femoral head necrosis only intra- and extravascular factors have been discussed. Vasoreactivity of intraosseous femoral head arteries has not previously been investigated. This study investigates the effect of long term glucocorticoid treatment on reactivity of intraosseous femoral head arteries in a porcine model. From 24 immature female Danish Landrace pigs from 12 litters, 12 animals received 100 mg methyl-prednisolone (orally) daily for 3 months. Their 12 sister pigs served as controls and received no corticosteroid. After sacrificing the animal, resistance arteries (diameter approximately 250 μm) were isolated from the femoral head epiphyseal cancellous bone and mounted as ring preparations on a small vessel myograph for measurement of isometric force development. The vasocontractory response to increasing doses of noradrenaline was not altered by methylprednisolone treatment. After submaximal precontraction by noradrenaline, vasorelaxation by sodium nitroprusside was not altered by methylprednisolone treatment. Only part of both experimental groups showed vasorelaxation to substance P and increasing doses of bradykinin. Increasing dose of endothelin-1 evoked vasoconstriction in both experimental groups. Sensitivity to endothelin-1 was increased after 3 months of methylprednisolone treatment. The response of isolated intraosseous femoral head arteries to endothelin-1 after long term glucocorticoid treatment in the pig is enhanced and appears to be unchanged for other physiologic vasoactive substances.

ChemInform Abstract: Highly Reactive 4‐Membered Ring Nitrogen‐Containing Heterocycles: Synthesis and Properties

AbstractReview: 59 refs.

Sympathomimetic effects of Scoparia dulcis L. and catecholamines isolated from plant extracts.

The herb Scoparia dulcis L. is used in Brazilian folk medicine to treat bronchitis, gastric disorders, haemorrhoids, insect bites and skin wounds, and in oriental medicine to treat hypertension. A previous study has shown that extracts of S. dulcis have analgesic and anti-inflammatory properties; in this work the sympathomimetic activity of an ethanolic extract of Scoparia dulcis L. has been investigated in rodent preparations in-vivo and in-vitro. Administration of the extract (0.5-2 mg kg-1, i.v.) to anaesthetized rats produced dose-related hypertension blocked by the alpha-adrenoceptor antagonist prazosin (1 mg kg-1). Partition of the extract in chloroform-water yielded an aqueous phase 20 times more potent than the extract; this produced hypertension in either reserpine-treated or pithed rats. In untreated and reserpine-treated rats the same fraction (1-3 x 10(3) micrograms mL-1) produced concentration-dependent contractions of the vas deferens musculature parallel to those obtained with noradrenaline (10(-8)-10(-4)M). Prazosin (10(-7)M) reduced the maximum contractile effect of the aqueous fraction, and shifted the concentration-response curves for noradrenaline to the right. The aqueous fraction (25 and 50 micrograms mL-1) increased the inotropism of electrically driven left atria of rats, the effect being blocked by propranolol (0.4 microgram mL-1). In preparations of guinea-pig tracheal rings the aqueous fraction (1-3 x 10(3) micrograms mL-1) relaxed the muscle contraction induced by histamine (10(-4) M) in proportion to the concentration. The effect was antagonized competitively by propranolol (1.5 microM). High-performance liquid-chromatographic analysis of the aqueous fraction revealed the presence of both noradrenaline and adrenaline in the plant extract. The results indicated that both catecholamines may account for the hypertensive and inotropic effects obtained after parenteral administration of S. dulcis extracts. This sympathomimetic activity is, however, unrelated to the previously reported analgesic and anti-inflammatory properties of the plant extract, but may explain its effectiveness upon topical application in the healing of mucosal and skin wounds.

Influence of beta-adrenoceptor-mediated relaxation on alpha-adrenoceptor-mediated contraction of rat pulmonary artery to adrenaline or noradrenaline.

Concentration-response (contraction) curves to either adrenaline or noradrenaline were obtained on isolated ring preparations of pulmonary artery from rats. In preparations from young rats the curve for adrenaline was bell-shaped, unless beta-adrenoceptors were blocked with propranolol (1 X 10(-6) M). The maximum contraction to adrenaline was less in the absence than in the presence of propranolol. In preparations from aged rats the adrenaline curve was no longer bell-shaped, even in the absence of propranolol. This reflected a decrease in the beta-adrenoceptor-mediated relaxant responses of preparations from aged rats, seen as a separation between the concentration-response (relaxation) curves to adrenaline on preparations (phenoxybenzamine-treated, KCl-contracted) from young and aged rats. In preparations from young rats the noradrenaline curve was not bell-shaped, but if the preparations were from young rats treated with thyroxine (T4), then a bell-shaped curve for noradrenaline was obtained, unless beta-adrenoceptors were blocked by propranolol. These data could be explained by an increase in beta-adrenoceptor-mediated relaxant responses of preparations from T4-treated rats, seen as a separation in the concentration-response (relaxation) curves to noradrenaline on preparations from control and T4-treated rats, respectively. Thus alpha-adrenoceptor-mediated contractile responses of rat pulmonary artery preparations, to adrenaline or noradrenaline, can be attenuated by activation of beta-adrenoceptors, mediating relaxation, and the extent of this attenuation changes under the influence of factors, such as ageing or T4-treatment, which modify beta-adrenoceptor-mediated relaxation in this blood vessel type.

Age-dependence of the effects of pinacidil on rat aorta

The effect of the K+ channel opening drug, pinacidil, has been examined on aortic ring preparations from young (2 months) and aged (greater than 24 months) rats. The potency (neg log IC50) values for pinacidil in relaxing K+ (20 mM)-contracted preparations were in the range expected for its K+ channel opening (hyperpolarizing) effects but were not significantly different between young (6.34) and aged (6.31) rats. Thus, ageing does not affect the drug's potency as a K+ channel opening drug. The more marked depression of the maximum response to noradrenaline by pinacidil (10 microM) in aged rats (85% reduction) compared with young rats (43% reduction), reflected a reduced alpha-adrenoceptor reserve for noradrenaline in preparations from aged rats. Pinacidil, in concentrations greater than 10 microM, was able to relax preparations contracted with 80 mM K+ suggesting that it may have a second mechanism which does not involve hyperpolarization. It was more potent in producing this effect on the preparations from aged rats.

Choice and concentration of contractile agent influence responses of rat aorta to vascular relaxant drugs.

Concentration-response (relaxation) curves to diltiazem, forskolin, isobutylmethylxanthine (IBMX), procaterol, isoprenaline and sodium nitrite were obtained on isolated ring preparations of rat aorta contracted either submaximally or maximally with noradrenaline or KCl. Diltiazem was more potent on KCl-contracted than on noradrenaline-contracted preparations whether the preparations were submaximally or maximally contracted. The other relaxant drugs were at least 20-fold less potent against KCl than against noradrenaline, but only on maximally contracted preparations. On submaximally-contracted preparations there was no potency difference between preparations contracted with these two contractile agents. Increasing the KCl concentration had a marked influence on the location of the concentration-response (relaxation) curves to all the drugs except diltiazem. This influence was different for drugs that act via cyclic AMP and those that act via cyclic GMP. It is concluded that both the choice of contractile agent (noradrenaline or KCl) and the concentration (especially of KCl) influence relaxant responses of rat aorta to vasodilator drugs.

Effects on tracheal smooth muscle of adenosine and methylxanthines, and their interaction

AbstractThe relaxant effects of xanthine and 15 different methylxanthines were studied in a guinea-pig isolated tracheal ring preparation. Substitution in the 3- and 1-positions of the xanthine molecule were found to be of greatest importance for improving tracheal relaxant potency. The unsubstituted xanthine and all 9-methylated xanthines were weakly active. Adenosine occasionally contracted (slightly) the tracheae but a concentration-dependent relaxation was always recorded. Endogenous adenosine did not seem to contribute to the tracheal tone. Xanthines methylated in the 1-position consistently antagonized the relaxant effect of adenosine and produced graded rightward shifts of the concentration-response line to adenosine (competitive antagonism). Xanthines with an unsubstituted 1-position or with comethylation in the 9-position were generally without antagonism. The results support our view that bronchodilating xanthine derivatives lacking universal adenosine antagonism can be produced.

Investigation of the antimuscarinic and other actions of proadifen in-vitro.

The possibility that proadifen (SKF 525A) antagonizes endothelium-dependent relaxations to acetylcholine (ACh) in isolated blood vessel preparations via a muscarinic receptor blocking action has been investigated. In phenylephrine-contracted rat isolated aortic ring preparations (with endothelium), proadifen (10-100 microM) shifts ACh relaxant curves to the right without affecting the maximal response, yet endothelium-dependent relaxations to ATP are unaffected. At lower concentrations, proadifen (1-10 microM) antagonizes negative inotropic responses to ACh and ATP in guinea-pig left atria, antagonizes contractile responses to ACh and elevated [K+] in guinea-pig ileal preparations, displaces (-)-[3H]quinuclidinyl benzilate from muscarinic binding sites in membrane homogenates of guinea-pig ileal longitudinal muscle and reduces contractile responses to elevated [K+] in rat aortic ring preparations. It is concluded that proadifen may possess complex interactions with muscarinic receptors and Ca2+ entry blocking properties in concentrations 10-100 times lower than those reported to inhibit cytochrome P450-catalysed reactions.

Effects of potassium-channel opener HOE 234 in guinea-pig airways.

The mechanism of the bronchospasmolytic effect of HOE 234 ((3S,4R)-3-hydroxy-2,2-dimethyl-4-(2-oxo-1-pyrrolidinyl) 6-phenylsulphonylchromane hemihydrate), a novel opener of ATP-sensitive potassium channels, has been studied by in-vitro testing in ring preparations of trachea and different parts of the principle bronchus of guinea-pigs, using methacholine, histamine and KCl as preconstrictors. The contribution of prostanoids was estimated in the presence and absence of indomethacin. Regional differences in the bronchodilatory effect of HOE 234 were compared with that of salbutamol. HOE 234 had a concentration-dependent relaxing effect on the smooth muscle preparations. In the absence of indomethacin no regional differences were seen for the effect of HOE 234 against the metacholine or KCl preconstriction, whereas with histamine a particularly strong relaxation was detectable in trachea. In the presence of indomethacin, distal bronchus after methacholine and trachea after KCl preconstriction were relaxed significantly more strongly than the other parts. The relaxation of the histamine-constricted trachea rings was not increased in comparison with that without indomethacin pretreatment. Thus the bronchospasmolytic effect of HOE 234 varied depending on the pretreatment, method of preconstriction and part of the airways examined. It was strongest in trachea rings with histamine preconstriction but its potency was clearly less than that of salbutamol. The latter had regionally different effects, being stronger in trachea than in the bronchi for all methods of preconstriction. The results suggest that the formation of bronchoconstricting prostanoids can attenuate parts of the relaxing effect of HOE 234. The possible advantages of HOE 234 as an anti-asthma drug might be related to an effect on bronchial hyper-reactivity and mucus secretion, because as a direct bronchodilator it is inferior to salbutamol.

ChemInform Abstract: Preparation of a Cycloheptane Ring from a 1,2‐Diketone with High Stereoselectivity.

AbstractAn efficient synthesis of cycloheptane‐1,3‐diones is developed starting from dienedione substrates.

Mechanism of attenuation of diabetes mellitus and hypercholesterolemia induced vascular endothelial dysfunction by protein tyrosine phosphatase inhibition

The study has been designed to investigate downstream mechanisms in the PTPase inhibition mediated attenuation of diabetes mellitus and hypercholesterolemia-induced vascular endothelial dysfunction. Diabetes mellitus was induced in rats using streptozotocin (55 mg/kg, i.v. once), while hypercholesterolemia was produced by feeding high cholesterol diet. After 4 weeks of streptozotocin and Cholesterol rich diet administration, vascular endothelium dysfunction was assessed, in terms of attenuation of acetylcholine-induced, endothelium-dependent relaxation (Isolated Aortic Ring Preparation), a decrease in serum nitrate/nitrite level, as well as mRNA expression of eNOS (rtPCR) and disruption of integrity of vascular endothelium (Electron microscopy). After 14 days of daily administration, sodium orthovanadate (8 mg/kg, p.o., 16 mg/kg, p.o and 24 mg/kg, p.o) and atorvastatin (30 mg/kg, p.o) (positive control) significantly improved acetylcholine-induced endothelium-dependent relaxation, serum nitrate/nitrite level, mRNA expression of eNOS and maintained integrity of vascular endothelium. However, this ameliorative effect of SOV was significantly blocked by UCN-01, (PDK inhibitor) and L-NAME (Inhibitor of eNOS). Therefore, it may be concluded that sodium orthovanadate, a specific inhibitor of PTPase, may stimulate PDK and eNOS and consequently improve vascular endothelium dysfunction. Thus, inhibition of PTPase might be a useful approach in the therapeutics of vascular endothelium dysfunction.

Gynura procumbens Merr. decreases blood pressure in rats by vasodilatation via inhibition of calcium channels

INTRODUCTION:Gynura procumbens has been shown to decrease blood pressure via inhibition of the angiotensin‐converting enzyme. However, other mechanisms that may contribute to the hypotensive effect have not been studied.OBJECTIVES:To investigate the cardiovascular effects of a butanolic fraction of Gynura procumbens in rats.METHODS:Anaesthetized rats were given intravenous bolus injections of butanolic fraction at doses of 2.5–20 mg/kg in vivo. The effect of butanolic fraction on vascular reactivity was recorded in isolated rat aortic rings in vitro.RESULTS:Intravenous administrations of butanolic fraction elicited significant (p<0.001) and dose‐dependent decreases in the mean arterial pressure. However, a significant (p<0.05) decrease in the heart rate was observed only at the higher doses (10 and 20 mg/kg). In isolated preparations of rat aortic rings, phenylephrine (1×10‐6 M)‐ or potassium chloride (8×10‐2 M)‐precontracted endothelium‐intact and ‐denuded tissue; butanolic fraction (1×10‐6–1×10‐1 g/ml) induced similar concentration‐dependent relaxation of the vessels. In the presence of 2.5×10‐3 and 5.0×10‐3 g/ml butanolic fraction, the contractions induced by phenylephrine (1×10‐9–3×10‐5 M) and potassium chloride (1×10‐2–8×10‐2 M) were significantly antagonized. The calcium‐induced vasocontractions (1×10‐4–1×10‐2 M) were antagonized by butanolic fraction concentration‐dependently in calcium‐free and high potassium (6×10‐2 M) medium, as well as in calcium‐ and potassium‐free medium containing 1×10‐6 M phenylephrine. However, the contractions induced by noradrenaline (1×10‐6 M) and caffeine (4.5×10‐2 M) were not affected by butanolic fraction.CONCLUSION:Butanolic fraction contains putative hypotensive compounds that appear to inhibit calcium influx via receptor‐operated and/or voltage‐dependent calcium channels to cause vasodilation and a consequent fall in blood pressure.

Role of calcium in contractile responses of calf cardiac vein during cooling

The effects of cooling (to 28 °C) were studied on the responses induced by carbachol (10(-9) - 3 × 10(-4) M) and serotonin (5-HT, 10(-8) - 3 × 10(-4) M) in calf cardiac vein preparations and the role of calcium ions in these effects were analyzed. Ring preparations of veins obtained from calf hearts were suspended in organ baths containing 25 mL of Krebs-Henseleit solution, maintained at 37 °C and continuously gassed with 95%O(2)-5%CO(2). After a resting period, preparations were contracted with carbachol (10(-9) - 3 times; 10(-4) M) and 5-HT (10(-8) - 3 times; 10(-4) M) at 37 °C. The same protocol was repeated at 28 °C after the preparations were allowed to equilibrate at this temperature for 60 min. In order to analyze the role of calcium ions (Ca(2+)) in the cooling-induced vascular response, concentration-response curves to carbachol and 5-HT were obtained in the presence of verapamil (10(-6) M), caffeine (3 times; 10(-4) M), and Ca(2+ )free medium in the presence of EGTA at 28 °C. During cooling to 28 °C, the EC(50) values, to carbachol and 5-HT were significantly higher than at 37 °C. Cooling to 28 °C in the presence of verapamil, caffeine or Ca(2+ )free medium in the presence of EGTA increased the EC(50) values, to both carbachol and 5-HT. These results suggest that Ca(2+) plays an essential role in the cooling-induced changes of calf cardiac vein preparations treated with carbachol and 5-HT.

Potassium-induced intermittent vasomotion in rat isolated pulmonary artery

A novel intermittent vasomotion induced by potassium in rat pulmonary artery was investigated with a view to characterize the ion channel mechanisms governing such secondary oscillatory activity. Isometric force was recorded from ring preparations of rat isolated pulmonary arteries incubated in a modified Krebs buffer containing K⁺ 15-18 mM and nitro-L-arginine methyl ester (10 µM). Tissues exhibited a stable pattern of on-off vasomotion consisting of intermittent contractile wave (ICW) activity with a periodicity of 7-8/hr and a rising phase of oscillatory ramping-up of contractile tone at 7 cycles/min. L-channel antagonists arrested (nicardipine; 3 nM) or retarded (verapamil, 30 nM) ICW activity with a concomitant wave asynchronization or decrease in amplitude. Mibefradil (30-100 nM) inhibited ICW ramping-up without affecting ICW period. Niflumic acid (1.0-3.0 µM) exerted dual actions on ICW amplitude but arrested ICW cycling at 10 µM. K⁺-channel blockers produced shortening of ICW period (4-aminopyridine, Ba²⁺ 30 µM; Cs⁺ 3.0-6.0 mM) and increase (tetraethylammonium; 1.0 mM) or decrease (Ba²⁺, 100 µM) in amplitude. Cyclopiazonic acid caused ICW asynchronization (0.3 µM) or cessation (1.0 µM) of ICW cycling. Fasudil retarded ramping-up contractile oscillations without changing ICW period. The inhibitory effects of nicardipine, niflumic acid and cyclopiazonic acid were partially surmounted by small additional increments in [K⁺](e). Our findings support the concept that a secondary vasomotive oscillator operates in rat pulmonary artery which enables the activity of the primary oscillator to be regulated in a cyclic manner via sarcolemmal L-type Ca²⁺ channels and an array of K conductances.