Effects of calcium entry blocking agents on 5-hydroxytryptamine- and noradrenaline-induced contractions of rat isolated jugular vein and aorta

Summary We calculated the contribution of the intracellular releasable calcium pool to the contractile responses induced by 5-hydroxytryptamine (5-HT) and noradrenaline (NA) by constructing time-response curves to the agonists in Ca²⁺-deficient medium in the isolated rat jugular vein and aorta. Biexponential curves were obtained compatible with a two compartment model. In the aorta the intracellular calcium pools are likely to be different for both 5-HT and NA. Moreover, we investigated the effect of maximally effective concentrations of calcium entry blocking agents (CEB's) on K⁺, 5-HT- and NA-induced contractions in Ca²⁺-containing medium. Only a moderate inhibiting effect of nifedipine, diltiazem, flunarizine and gallopamil on 5-HT and NA-induced Ca²⁺ influx could be observed; in contrast, K⁺-induced Ca ²⁺ influx could be antagonized completely. The calculated contribution of intracellular Ca ²⁺ to 5-HT-and NA-induced contractions, obtained from the experiments in Ca²⁺-free medium was much lower than that obtained after pretreatment with CEB's, leading to the conclusion that after CEB-pretreatment a Ca²⁺ influx component persists. This hypothesis was supported by the observation that contractions in Ca²⁺-free medium consist of a monophasic, fast response only, whereas after CEB-pretreatment a response similar to the control, including a slow, sustained component, was obtained. The Ca²⁺ influx component not affected by maximally effective concentrations of CEB's seems to represent an inflow of extracellular Ca²⁺ directly into the cytosol and not into an intracellular calcium store. Send offprint requests to M. A. M. Gouw at the above address     

Pharmacological properties of voltage-dependent calcium channels in functional microvessels isolated from rat brain

Summary Voltage-operated calcium channels were studied in rat intracerebral microvessels. The contractile reactivity to KCI-depolarization was assessed by the measurement of internal diameter of superfused microvessels. Dihydropyridine receptor sites associated with calcium channels were identified and characterized using ³H(+)PN 200-110 [isopropyl-4-(2,1,3-benzodiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5-methoxycarbonyl-pyridine-3-carboxylate].Depolarization induced by high-KCI solution produced a marked reduction of the internal diameter of cerebral microvessels which was associated with the appearance of rhythmic activity. The vessel contraction was reversible and abolished by nimodipine. Binding studies with ³H(+)PN 200-110 revealed the existence of a single class of specific, stereoselective and voltage-dependent binding sites which bound (+)PN 200-110 with a K _D of 88 ± 6.6 pmol l^–1 at 37°C in microvessels incubated in NaCl medium. When microvessels were incubated in KCI-medium, the apparent K _D value was reduced to 35 ± 2 pmol l^–1. B _max was not significantly changed. The effect of KCI was not related to concomitant changes in the Na concentration. The potency of various dihydropyridine derivatives in inhibiting ³H(+)PN 200-110 binding was in agreement with their pharmacological potency in smooth muscle preparations. The effect of PN 200-110 and of nimodipine was stereoselective. K _i values of PN 200-110 and of nimodipine were increased in depolarized preparations, while nifedipine's potency was unchanged. Verapamil was only a partial inhibitor of ³H(+)PN 200-110 binding. The effect of diltiazem was stereoselective: the (+)-cis isomer enhanced the binding and the (–)-cis isomer of diltiazem poorly inhibited the binding of PN 200-110.Results showed that isolated cerebral microvessels possess functional voltage-operated calcium channels, which contain potential-modulated receptors for dihydropyridine calcium entry blockers with characteristics similar to those described in other tissues.Send offprint requests to: T. Godfraind at the above address     

Effect of papaverine on cyclic nucleotide levels in the isolated rat aorta.

An increase in cAMP but no significant modification in cGMP content could be demonstrated in rat aorta strips after applying papaverine in concentrations which reduced contractile responses. Accumulation of cAMP was induced in noradrenaline-stimulated on K+-depolarized strips, under omission of external Ca2+. Thus the elevation of cAMP level preceded the reduction of contraction subsequently elicited by readdition of Ca2+. The effects could not be dissociated under the experimental conditions used here.     

Two stable cell lines for screening of calcium channel blockers

Stable cell lines are potentially excellent tools for large-scale screening of new compounds. Two carboxyterminal-deleted constructs of the two splice variants a and b of the calcium channel class C _l subunit were expressed stably in HEK 293 cells. Each cell line produced regular L-type calcium currents. The opening and closing of the calcium channel elicited by potassium depolarization was followed by Fura-2 transients. These transients were blocked by the calcium channel blocker mibefradil with a concentration for 50% inhibition of 1.7 M. The cell lines expressing the truncated cardiac _1C–a or smooth muscle _1C–b calcium channel were both blocked by nisoldipine under patch clamp conditions. Nisoldipme interacted with higher affinity with the _1C–b channel than with the _1C–a channel. These results indicate that the two cell lines retain the differential dihydropyridine sensitivity of smooth muscle and cardiac calcium channels and may be potential tools for the screening of L-type calcium channel blockers.     

Palytoxin-induced endothelium-dependent relaxation in the isolated rat aorta

Summary The effects of a potent marine toxin, palytoxin (PTX), were investigated on the contractile responses in the isolated rat aorta with or without endothelium. PTX in the concentrations of 10^–13–10^–11 mol/l showed little effect on the resting tension of the vessel with or without endothelium. PTX, 10^–10 mol/l, induced a small contraction in the aorta without endothelium but not in the aorta with endothelium. When added during the sustained contraction induced by 10^–7 mol/l norepinephrine, 10^–12 mol/l PTX sometimes (6 out of 43 strips) augmented the norepinephrine-induced contraction whereas 10 ^–11–10^–10 mol/l PTX induced a biphasic response which was composed of a transient augmentation followed by a relaxation. These effects of PTX were not observed in the aorta without endothelium. Influencesof atropine (10^–6 mol/l), indomethacin (2.5 × 10^–5 mol/l), methylene blue (5 × 10^–6 mol/l), hydroquinone (10^–4 mol/l), phenidone (5 × 10^–5 mol/l), hemoglobin (10^–6 mol/l) and p-bromophenacyl bromide (5 × 10^–5 mol/l) on the PTX (10^–10 mol/l) induced responses were examined. Methylene blue, hydroquinone, phenidone, hemoglobin and p-bromophenacyl bromide inhibited both the PTX-induced augmentation and relaxation of the norepinephrine-induced contraction. The endothelium-dependent relaxation due to 3 × 10^–7 mol/l carbachol was inhibited by atropine, methylene blue, hydroquinone, phenidone, hemoglobin and p-bromophenacyl bromide. These results suggest that PTX acts on the endothelium, modifies the synthesis or release of endothelium-derived relaxing factor and thus changes the contractile response to norepinephrine in rat aorta. Send offprint requests to H. Nagase at the above address     

钙通道阻滞剂抗肝缺血—再灌注损伤作用机制的实验研究

目的 探讨钙通道阻滞剂（ＣＣＥＢ）对肝缺血一再灌注损伤（ＨＩＲＩ）防治作用的机制。方法制备家兔ＨＩＲＩ模型,动态观察维拉帕米（ＶＰ）和地尔硫卓（ＤＴ）对肝组织及血中黄嘌呤氧化酶（ＸＯ）、超氧化物歧化酶（ＳＯＤ）活性及脂质过氧化物（ＬＰＯ）浓度的影响。结果 ＶＰ组和ＤＴ组ＯＸ活性及ＭＤＡ含量分别显著低于对照组（均Ｐ＜０．０１）,而ＳＯＤ活性与对照组比较均无显著性差异（均 Ｐ＞ ０．０５）。结论 ＣＣＥＢ抗 ＨＩＲＩ的机制与其降低 ＸＯ活性、抑制脂质过氧化反应密切相关。     

Activation mechanisms of human renal artery: effects of KCl, norepinephrine and nitrendipine upon tension development and 45Ca influx

The activation of human vascular smooth muscle by KCl-induced depolarization or norepinephrine and the inhibition produced by nitrendipine were studied in the isolated human renal artery. The contractile response of arterial rings to 80 mM KCl was abolished when extracellular Ca2+ was removed, and was inhibited by nitrendipine (IC50 = 10(-8) M). In contrast, a residual, transient contractile response to norepinephrine remained when extracellular Ca2+ was removed and the norepinephrine-induced contractions obtained in the presence of extracellular Ca2+ were not blocked by nitrendipine. KCl caused a stimulation of 45Ca influx which was completely prevented by 10(-6) M nitrendipine. Norepinephrine also caused a stimulation of 45Ca influx; however, the norepinephrine-induced 45Ca influx was not prevented by 10(-6) M nitrendipine. These findings are consistent with the concept that depolarization-induced activation of the human renal artery is primarily dependent upon a stimulation of Ca2+ influx; whereas activation by norepinephrine involves the release of intracellular Ca2+ in addition to the activation of a separate, receptor-operated Ca2+ influx pathway.     

Neuronal uptake inhibitors,nisoxetine and fluoxetine on rat vascular contractions

The importance of neuronal uptake processes in rat arteries and veins was compared using nisoxetine and fluoxetine, selective inhibitors for neuronal uptake of norepinephrine and serotonin, respectively. Nisoxetine (10^?7–10^?5 M) increased the sensitivity to exogenous norepinephrine in the portal and mesenteric veins (10-fold) and in the mesenteric artery (2.5-fold). Responses to field stimulation were also increased after nisoxetine in all three vessels. After nisoxetine, aortic responses to norepinephrine were unaltered and in all tissues, serotonin-induced contractions were reduced. Fluoxetine did not potentiate responses to norepinephrine or to field stimulation except in the mesenteric vein where sensitivity to norepinephrine was increased and the relaxation rate after field stimulation was prolonged. Although serotonin has been detected in blood vessels, fluoxetine did not increase vascular responses to serotonin. These studies suggest that rat mesenteric veins have a highly sensitive neuronal uptake mechanism for norepinephrine. Furthermore, these data provide indirect evidence against a functionally important serotonergic uptake process in rat blood vessels and suggest that neuronal uptake of serotonin does not exert a major role in terminating the vascular action of this biogenic amine.     

Effects of calcium entry blockers on tension development and calcium influx in rat uterus.

Spontaneous and potassium chloride (KCl)-induced tension development of strips of whole uterus from the day-22 pregnant rat was reduced when the tissues were incubated in a calcium ion (Ca2+)-free medium. Strips of whole uterus, in an initially Ca2+-free medium, responded to the cumulative addition of Ca2+ with graded phasic tension development and associated rapid electrical discharges. The spasms were inhibited by gallopamil (100 nM) and diltiazem (1 microM). Strips of whole uterus in a depolarizing (40 mM K+) medium, which was initially Ca2+-free, responded to the cumulative addition of Ca2+ with graded tonic tension development without associated electrical discharges. These spasms were inhibited by calcium entry blockers with a rank order of potency of nifedipine = gallopamil greater than diltiazem greater than cinnarizine. KCl-induced tension development in endometrium-free uterine strips was antagonized by calcium entry blockers with a rank order of potency of nifedipine greater than gallopamil greater than diltiazem greater than cinnarizine. Ca2+ influx into endometrium-free uterine strips was assessed by means of the 'lanthanum method'. KCl induced a concentration-dependent increase in 45Ca2+ influx which was suppressed or abolished by nifedipine (2.5 nM), gallopamil (100 nM), diltiazem (500 nM) or cinnarizine (5 microM). It is concluded that spontaneous and KCl-induced tension development of rat uterus involves Ca2+ influx from the extracellular medium into the myometrial cell. These results support the hypothesis that nifedipine, gallopamil, diltiazem and cinnarizine inhibit Ca2+- and KCl-induced tension development of rat uterus by reduction of Ca2+ influx.     

Effects of calcium channel blockers on the development of early rat postimplantation embryos in culture

Rat embryos (9.5-day-old) were cultured for 48 h in the presence of nifedipine (NIF), nimodipine (NIM), nitrendipine (NIT), gallopamil HCl (GAL), verapamil HCl (VER) and diltiazem HCl (DIL). The effects on growth and morphogenetic differentiation in vitro were monitored. Dose-response relationships were evaluated, including an assessment of the no-observed-effect-level (NOEL) or the lowest-observed-effect-level (LOEL), and the lowest concentration tested inducing abnormalities in 100% of the embryos (100% EL). The morphological alterations observed at the highest concentrations were very similar for all six drugs. The abnormalities concerned yolk sac circulation and morphology, as well as heartbeat, the morphology of the heart, head, neural tube, or fore-limbs, and the shape of the embryo. The abnormal embryos were also growth retarded (decrease in protein content and crown-rump length). Interference with calcium channel functions seems to represent an interesting model for studying a special kind of abnormal prenatal development, especially the differentiation of certain mesenchymal structures. The concentration ranges between NOELs and 100% ELs were found to be: NIM=0.1–1 g/ml; NIT and VER=1–10 g/ml; DIL=1–30 g/ml, and LOELs-100%ELs were: GAL=1–10 g/ml; NIF=10–30 g/ml.On leave of absence from: Industrial Toxicology Research Centre, Lucknow, India     

Papaverine, cyclic AMP and the dependence of the rat aorta on extracellular calcium.

The spasmolytic effects of papaverine and dibutyryl cyclic AMP (db-cAMP) were compared on isometric contractile responses induced by addition of increasing amounts of external calcium to K+-depolarized or noradrenaline-stimulated rat aorta strips. Papaverine at a concentration active on depolarized strips (3 times 10(-5) moles/1) reduced the maximal contraction (Emax) elicited by Ca2+ in these preparations, while db-cAMP did not. Contrary to what was observed on depolarized aortae, the degree of inhibition of noradrenaline-stimulated strips did not decrease with increasing extracellular calcium concentration (Ca)e. Both db-cAMP and papaverine at a concentration which did not depress Emax (5 times 10(-6) moles/1) potentiated the relaxing effect of high (Ca)e on contractions elicited by noradrenaline. In conclusion, cyclic AMP is probably implicated in the mode of action of papaverine on the noradrenaline-stimulated rat aorta. At a concentration active on depolarized strips, papaverine is also able to impair contractility directly.     

Agonist-evoked calcium entry in vascular smooth muscle cells requires IP3 receptor-mediated activation of TRPC1

Transient receptor potential canonical (TRPC) proteins have been proposed to function as plasma membrane Ca2+ channels activated by store depletion and/or by receptor stimulation. However, their role in the increase in cytosolic Ca2+ activated by contractile agonists in vascular smooth muscle is not yet elucidated. The present study was designed to investigate the functional and molecular properties of the Ca2+ entry pathway activated by endothelin-1 in primary cultured aortic smooth muscle cells. Measurement of the Ca2+ signal in fura-2-loaded cells allowed to characterize endothelin-1-evoked Ca2+ entry, which was resistant to dihydropyridine, and was blocked by 2-aminoethoxydiphenylborate (2-APB) and micromolar concentration of Gd3+. It was not activated by store depletion, but was inhibited by the endothelin ETA receptor antagonist BQ-123, and by heparin. On the opposite, thapsigargin-induced store depletion activated a Ca2+ entry pathway that was not affected by 2-APB, BQ-123 or heparin, and was less sensitive to Gd3+ than was endothelin-1-evoked Ca2+ entry. Investigation of the gene expression of TRPC isoforms by real-time RT-PCR revealed that TRPC1 was the most abundant. In cells transfected with TRPC1 small interfering RNA sequence, TRPC1 mRNA and protein expression were decreased by 72+/-3% and 86+/-2%, respectively, while TRPC6 expression was unaffected. In TRPC1 knockdown cells, both endothelin-1-evoked Ca2+ entry and store-operated Ca2+ entry evoked by thapsigargin were blunted. These results indicate that in aortic smooth muscle cells, TRPC1 is not only involved in Ca2+ entry activated by store depletion but also in receptor-operated Ca2+ entry, which requires inositol (1,4,5) triphosphate receptor activation.     

Age-related decrease in the effect of parathyroid hormone-related protein on cytosolic free calcium level and tension in rat aortic smooth muscle

Age-related changes in the effect of parathyroid hormone-related protein (PTHrP) on the cytosolic free calcium level ([Ca⁺]_i) and on the tension in rat aortic smooth muscle were investigated. The possible involvement of cAMP, a second messenger of PTHrP, in such changes was also investigated. Spiral aortic strip preparations without endothelium from 8-weeks, 6-months, and 24-months old rats were treated with fura 2/AM, and the fluorescence ratio R340/380, an index of [Ca⁺]_i was measured. Simultaneously, the tension of the preparations was measured. PTHrP-(1–34) and dibutyryl cAMP produced concentration-dependent decreases in the tension and in R340/380 in aortas precontracted with phenylephrine (10^–7 M). These effects were significantly lower in the aortas of 24-months old rats than in the vessels of 8-weeks and 6-months old rats. The effects were similar in the aortas of 8-weeks and 6-months old rats. PTHrP-(1–34) concentration-dependently increased cAMP production in vascular smooth muscle cells (VSMCs) from 8-weeks old rats. However, the increase in cAMP production was significantly lower in cultured VSMCs from 6-months and 24-months old rats than in cells from 8-weeks old rats.These results suggest that the reduced cAMP production stimulated by PTHrP and the reduced effects of cAMP with aging might contribute to the age-related changes in the decreases in tension and [Ca⁺]_i in response to PTHrP in the rat aorta.     

Cooling-induced supersensitivity to acetylcholine in the isolated airway smooth muscle of the rat

Summary Isolated tracheal and bronchial strip-chain preparations of the rat were used to study the effect of temperature on electrically or acetylcholine-induced contraction. The preparations were suspended in the organ bath containing Krebs bicarbonate solution for isometric tension recording. A decrease of bath temperature from 37°C to 20°C (cooling) had no effect on basal tone but augmented the contractile responses of the trachea and bronchus caused by stimulation of intramural cholinergic nerves (0.5–5 Hz) or acetylcholine (3 mol/1–0.3 mmol/l). Cooling-induced augmentation of the contractile response to acetylcholine was not affected by pretreatment of the tissue with physostigmine (0.1 mol/l) or tetrodotoxin (0.3 mol/l). The affinity of acetylcholine for the tracheal muscarinic receptors at 20°C, determined from its dissociation constant (K _A), was not significantly different from that at 37°C. On the other hand, acetylcholine-induced contraction of trachea which was incubated with isosmotic K⁺-rich Krebs solution and with Ca-free, EGTA (0.1 mmol/l) containing Krebs solution were both augmented at 20°C. Caffeine or vanadate, each at a lower concentration than the threshold for causing contraction by itself, augmented the contractile responses of the trachea to acetylcholine (1 mol/l–0.3 mmol/l). These potentiating effects of caffeine and vanadate were greater at 20°C then 37°C. From these observations, it is concluded that increased responsiveness of the rat airway smooth muscle to acetylcholine with lowered temperature may involve the acceleration of Ca release from intracellular storage sites, inhibition of Ca extrusion from the cell and or the inhibition of Ca reuptake by intracellular storage sites.This work was supported in part by Scientific Research Fund 58770177, from the Ministry of Education, Science and Culture, Japan     

Effects of calcium entry blockers on Ca2+-induced contraction of depolarized and noradrenaline-exposed rat resistance vessels

The effects of four calcium entry blockers (CEBs), diltiazem (D), verapamil (V), nifedipine (NF) and nicardipine (NC), were investigated on Ca2+ concentration-effect curves of rat depolarized (K+, 40 mM) or noradrenaline (NA, 3 microM)-exposed mesenteric resistance vessels. Under control conditions, NA-exposed vessels were more sensitive to Ca2+ (pD2: 4.12 +/- 0.11) than depolarized vessels (pD2: 3.16 +/- 0.02, P less than 0.01) whereas the maximal active wall tensions were not significantly different (2.86 +/- 0.11 mN/mm and 2.11 +/- 0.34 mN/mm respectively). In depolarized vessels, D, V, NF and NC induced a concentration-dependent shift to the right and a depression of the maximal effect of the Ca2+ curves, which suggested a non-competitive antagonism. The IC50 (concentration of CEB producing a 50% inhibition of the maximal contractile response from control curve) values were: D: 3 X 10(-7), V: 1.3 X 10(-7), NF: 4.5 X 10(-9), NC: 3 X 10(-9) M. In NA-exposed vessels, the CEBs produced a concentration-dependent shift to the right of the Ca2+ curves before depressing their maximal effect. This suggested that the antagonism was different from that observed in depolarized arteries. In this case, the IC50 values were: D: 4.5 X 10(-7), V: 2 X 10(-7), NF: 9 X 10(-9), NC: 7 X 10(-9) M. Although the gating mechanisms activated in this study were differently affected by CEBs, since there were marked qualitative differences in their antagonistic effects on Ca2+ concentration-effect curves, depolarization and NA promoted the entry of Ca2+ into smooth muscle cells of rat resistance vessels by mechanisms with the same sensitivity to CEBs as expressed by IC50 values.     

Role of sodium-calcium exchange and effects of calcium entry blockers on endothelial-mediated responses in rat isolated aorta

Acetylcholine relaxes rat aorta and increases aortic cyclic GMP levels by a mechanism (or mechanisms) dependent on the endothelium and on extracellular calcium. Therefore, the effects of representatives of different subclasses of calcium entry blockers, verapamil, nifedipine, diltiazem, and bepridil, on maximal acetylcholine (1 microM)-induced increases in cyclic GMP levels were investigated in rat isolated aorta. None of these compounds, at a concentration (3 microM) sufficient to maximally inhibit agonist-stimulated Ca2+ influx into vascular smooth muscle cells, significantly affected either the basal or the acetylcholine-stimulated tissue cyclic GMP levels. On replacing all but 20 mM Na+ by choline, a condition that might be expected to limit or even abolish Na+-Ca2+ exchange, or in the presence of amiloride (1 mM), an inhibitor of Na+-Ca2+ exchange, acetylcholine-stimulated increases in tissue cyclic GMP levels were abolished or inhibited by about 80%, respectively. In choline containing solution acetylcholine relaxant responses were abolished. The presence of amiloride, or the replacement of Na+ by choline, had no effect on increases in cyclic GMP levels evoked by sodium nitroprusside (0.3 microM), an agent that stimulates cyclic GMP formation in smooth muscle without intervention of the endothelium. Replacement of Na+ by Li+ but not the other treatments depressed basal tissue cyclic GMP levels by about 45% but did not abolish either acetylcholine- or sodium nitroprusside-induced relaxant responses. However, the time course of relaxant responses elicited by both these relaxant agonists in precontracted rat aortic rings with endothelium was altered by Li+ replacement; the half-time to relaxation to acetylcholine was increased by about 70-fold. It is concluded that calcium channels, as characterized in smooth muscle and cardiac tissue, are not involved in the stimulated liberation of an endothelial-derived relaxant factor by acetylcholine, but that an Na+-Ca2+ exchange process may be of importance.     

Differences between negative inotropic and vasodilator effects of calcium antagonists acting on extra- and intracellular calcium movements in rat and guinea-pig cardiac preparations

Summary In order to get more insight into the utilization of calcium in the mammalian heart and the influence of calcium antagonists on this process we have evaluated the negative inotropic and vasodilator effect of nifedipine, diltiazem, verapamil, bepridil and lidoflazine as well as of the intracellularly acting calcium antagonists ryanodine and TMB-8 in the presence of 0.9 and 1.8 mmol/l calcium in rat Langendorff hearts. The effect of ryanodine was also studied in guinea-pig Langendorff hearts. In addition, in rat and guinea-pig papillary muscles the effect of these drugs on the force of contraction was examined.With the exception of ryanodine and TMB-8 all calcium antagonists induced a pronounced coronary vasodilator effect. The rank order of potency for this effect was: nifedipine > verapamil = diltiazem = bepridil = lidoflazine in the presence of 0.9 mmol/l calcium. At a calcium concentration of 1.8 mmol/l nifedipine and verapamil proved more potent, whereas diltiazem was less active. All calcium antagonists completely suppressed the development of the left ventricular pressure. At a calcium concentration of 0.9 mmol/l the potency order for this effect was: ryanodine > nifedipine = verapamil > diltiazem = bepridil = lidoflazine > TMB-8. In the presence of 1.8 mmol/l calcium the concentration-response curves for reduction of the left ventricular pressure by nifedipine, verapamil and diltiazem slightly shifted to the right. In contrast to all calcium antagonists investigated, in guinea-pig Langendorff hearts ryanodine only partially decreased the left ventricular pressure.In rat papillary muscles ryanodine nearly completely reduced the force of contraction, whereas nifedipine only partially inhibited the force of contraction. Lidoflazine only slightly affected the force of contraction. In guinea-pig papillary muscles all drugs partially decreased the force of contraction. Lidoflazine, however, was more effective than in rat papillary muscles. In rat and guinea-pig papillary muscles the force of contraction was fully suppressed by a combination of nifedipine and ryanodine.The results of the present study suggest that in the rat heart extracellular calcium plays a major role in the maintenance of the coronary vascular tone. The development of contractile force relies on the release of activator calcium from the sarcoplasmic reticulum but also requires the influx of calcium through dihydropyridine-sensitive channels. In the guinea-pig heart activator calcium may also be released from a source complementary to the sarcoplasmic reticulum. Lidoflazine displays certain selectivity towards this particular calcium pool.Preliminary results have been presented at the Spring Meeting of the German Pharmacological and Toxicological Society, 14–17 March, 1989, Mainz (Hugtenburg et al. 1989a)     

Mobilization of stored calcium for phasic contraction induced by norepinephrine in rabbit aorta.

In Ca-free solution norepinephrine (NE) produced only a phasic contraction in the media-intima layer of rabbit aorta. The second application of NE was almost ineffective. Incubation of the muscle with Ca for a short period (Ca loading) restored the ability to produce a phasic contraction in Ca-free solution. Various ions and compounds were applied to the muscle prior to the Ca loading or prior to NE application and it was found that these treatments variously affected the NE-induced phasic contraction. The data suggest that the NE-induced phasic contraction in Ca-free solution results from Ca release from cellular sites and that a hyperbolic relationship exists between the amount of Ca at these sites and the magnitude of the contraction. These sites take up extracellular Ca by a process sensitive to La3+ but not to D600; the Ca influx stimulated by high K solution, which is sensitive to D600 and also contributes to refilling of these sites; NE releases this stored Ca by a process sensitive to procaine, caffeine and theophylline and, in this manner, elicits a phasic contraction.     

Stimulation of calcium uptake into aortic microsomes by cyclic AMP and cyclic AMP-dependent protein kinase

Summary Enhancement of calcium uptake into rabbit aortic microsomes was seen at a cyclic AMP concentration of 10^–6M in the presence of cyclic AMP-dependent protein kinase (ATP: protein phosphotransferase, EC 2.7.1.37). Protein kinase alone also increased calcium uptake, but cyclic AMP alone was without effect. The results suggest that stimulation of calcium sequestration may be the mechanism of cyclic AMP involvement in vascular smooth muscle relaxation.