From channels to cats: an overview of isradipine pharmacology.
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Animals↗
Biomedical subjects
U T Rüegg
Publications and source records attributed to U T Rüegg.
Antivasoconstrictor effects of isradipine. A quantitative approach in anesthetized rats and conscious rabbits.
Calcium antagonists blunt the constrictor effects of vasoconstrictor agents. Differences in the sensitivity of several vasoconstrictors to the constrictor effect of calcium antagonists were quantified in experiments on rats and rabbits. In rats, the dose-response curves for pressor effects to angiotensin II were shifted in parallel to the right after treatment with isradipine, but not with prazosin and dihydralazine, suggesting that the antivasoconstrictor effect of isradipine was of a specific type. Blood pressure increases elicited by angiotension II or norepinephrine (alone, or with propranolol or propranolol plus prazosin) were also measured in rabbits and, again, isradipine caused parallel shifts of the dose-response curves. Both agonists thus appear to stimulate calcium entry, at least in resistance vessels, and via the same pathway, the L-type calcium channel. However, the angiotension II pressor effects were more sensitive to isradipine than those of norepinephrine in the presence of propranolol, and even more so in the presence of propranolol plus prazosin. These results suggest subtle and as yet unexplained differences, possible related to the activity of the adenylate cyclase system, in the chain of events leading to activation of the L-channels in vivo.
Anesthesia, General↗
Staurosporine, K-252 and UCN-01: potent but nonspecific inhibitors of protein kinases.
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Alkaloids↗
Differential blockade of agonist- and depolarization-induced 45Ca2+ influx in smooth muscle cells.
ATP stimulated 45Ca2+ influx in rat aortic smooth muscle cells in a concentration-dependent manner (EC50 = 3.6 +/- 0.5 X 10(-7) M). ADP and GTP were less effective than ATP in stimulating 45Ca2+ influx; AMP was weakly active and the adenosine agonist 5'-(N-ethyl-carboxamido)-adenosine (NECA) had no effect. ATP gamma S was about equieffective with ATP, whereas alpha,beta-methylene-ATP (APCPP) did not induce 45Ca2+ influx. Stimulation of 45Ca2+ influx by ATP was not abolished by the dihydropyridine Ca2+ channel antagonist darodipine (PY 108-068), which completely blocked depolarization-induced 45Ca2+ influx. Inorganic cations (La3+, Cd2+, Co2+, Ni2+, Mn2+, and Mg2+) were able to inhibit both agonist- and depolarization-induced 45Ca2+ influx. Cd2+, however, was approximately 20 times more selective in blocking K+-stimulated than agonist-stimulated 45Ca2+ influx. These data indicate that ATP-stimulated Ca2+ influx in rat aortic smooth muscle cells is resistant to darodipine but is reduced by La3+, Cd2+, and other inorganic blockers of Ca2+ channels.
Adenosine Triphosphate↗
Receptor-operated calcium-permeable channels in vascular smooth muscle.
The extracellular Ca2+ dependence of agonist stimulation of vascular smooth muscle (VSM) has been investigated in rat cultured aortic smooth muscle cells (SMCs) and isolated mesenteric resistance vessels (MRVs). Agonists such as [Arg8]vasopressin (AVP), angiotensin II (Ang II), and adenosine-5'-triphosphate (ATP) stimulated 45Ca2+ entry into the SMCs that was (a) independent of the extent to which the membranes were polarized, and (b) was not inhibited by organic Ca2+ channel antagonists. Measuring the intracellular Ca2+ concentration [( Ca2+]i) after stimulation with agonists revealed a rapid increase of [Ca2+]i, which was followed by a sustained rise that was insensitive to Ca2+ antagonists. In Ca2+-free medium, only the initial peak of [Ca2+]i was still observed, but the sustained response to the agonists disappeared completely. This observation indicates that the sustained elevation seen in Ca2+-containing medium was the consequence of agonist-induced Ca2+ entry. In MRVs, a corresponding Ca2+-antagonist-insensitive, agonist (norepinephrine and AVP)-induced tonic tension was also identified. Moreover, agonists were able to induce sustained tension in the MRVs regardless of whether the membrane was normally polarized or was previously depolarized (80 mM K+) upon their administration. The agonist-stimulated 45Ca2+ entry in the SMCs could be blocked by the multivalent cations La3+, Cd2+, Mn2+, Co2+, Ni2+, and Mg2+ (in this order of potency). Depolarization-induced 45Ca2+ influx was inhibited by these cations in the same order of potency, but was significantly more sensitive to Cd2+ and significantly less sensitive to La3+ than that stimulated by agonists. Treatment with 2-nitro-4-carboxyphenyl-N,N-diphenyl-carbamate (NCDC, a proposed inhibitor of phospholipase C) reduced both the agonist-induced 45Ca2+ influx and the sustained elevation of [Ca2+]i in the SMCs. NCDC also abolished both contraction and depolarization induced by agonists in the MRVs. The kinase C stimulator phorbol-12-myristate-13-acetate (PMA) inhibited the agonist-induced 45Ca2+ influx and sustained increase in [Ca2+]i in the SMCs, whereas the kinase C inhibitor staurosporine had no effect. In the MRVs, in contrast, PMA had no influence on agonist-induced contractions. Staurosporine (1 microM), however, completely prevented these contractions, as did NCDC, but, unlike NCDC, it did so without affecting the agonist-induced depolarization. These data support an important role of receptor-operated Ca2+-permeable channels in VSM activation by agonists and suggest that these channels may be controlled by intracellular enzymic pathways and second messenger systems.
Animals↗
Cyclosporin A augments angiotensin II-stimulated rise in intracellular free calcium in vascular smooth muscle cells.
Pretreatment of rat vascular smooth muscle cells with the immunosuppressive drug cyclosporin A caused concentration- and time-dependent increases in both the amplitude and duration of the angiotensin II-induced rise in cytosolic free calcium, as measured with quin 2. Cyclosporin A had no significant effect on basal quin 2 fluorescence. However, cyclosporin A increased the basal 45Ca2+ influx. This stimulation of 45Ca2+ influx was not blocked by nifedipine (10(-6) M). Cyclosporin A also augmented the angiotensin II-stimulated influx and efflux of 45Ca2+. These results demonstrate that cyclosporin A increases the permeability of the plasma membrane for Ca2+ and also augments the angiotensin II-induced increases in cytosolic free calcium.
Angiotensin II↗
Vasopressin increases 45Ca2+ influx in rat aortic smooth muscle cells.
[Arg8]Vasopressin (AVP)-induced 45Ca2+ influx was examined in vascular smooth muscle cells derived from rat aorta. AVP stimulated the 45Ca2+ influx in a concentration-dependent manner. The effect was abolished in the presence of La3+. The dihydropyridine calcium channel antagonist darodipine did not affect the AVP-induced influx of 45Ca2+. These data suggest that AVP stimulates in these cultured aortic smooth muscle cells a receptor-operated channel (ROC) that is permeable to Ca2+.
Animals↗
Platelet-activating factor mobilizes intracellular calcium in vascular smooth muscle cells.
The effect of platelet-activating factor (PAF) on polyphosphoinositide metabolism and 45Ca2+ efflux was examined in a vascular smooth muscle cell line (A7r5). PAF stimulated a rapid but transient production of inositol trisphosphate and inositol bisphosphate which, in the presence of lithium, resulted in an accumulation of inositol monophosphate. In addition, PAF induced a rapid efflux of 45Ca2+ from preloaded cells, an effect which was concentration-dependent. These data suggest that PAF mobilizes intracellular Ca2+ via the production of inositol trisphosphate.
Animals↗
Serotonin increases the production of inositol phosphates and mobilises calcium via the 5-HT2 receptor in A7r5 smooth muscle cells.
Serotonin (5-HT) induces inositol phosphate production and the efflux of 45Ca2+ in a smooth muscle cell line (A7r5) derived from rat aorta. These effects were pharmacologically characterised and compared to data obtained in radioligand binding studies performed with the 5-HT2 ligand [3H]ketanserin in rat brain cortex membranes. 5-HT causes in increase in the levels of inositol trisphosphate (InsP3), inositol bisphosphate (InsP2) and inositol phosphate (InsP1). InsP3 production was rapid and transient whereas InsP1 accumulated in a time and concentration dependent manner. The 5-HT stimulated InsP1 accumulation (pEC50 = 6.48) was potently and competitively inhibited by the 5-HT2 specific antagonists, pirenperone and ketanserin, whereas antagonists of other 5-HT receptors were active only at high concentrations. There was a significant correlation between inhibition of 5-HT stimulated InsP1 accumulation and 5-HT2 binding (r = 0.98, P = 0.0035). 5-HT stimulated the efflux of 45Ca2+ from preloaded cells with a pEC50 of 7.59. The rank order of potency for agonist induced Ca2+ efflux, 5-HT greater than alpha-methyl-5-HT greater than 1-methyl-5-HT greater than RU 24969 (5-methoxy-3[1,2,3,6,-tetrahydropyridin-4-yl]-1-H indole) greater than 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)-tetralin) greater than 8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)-tetralin) greater than 5-CT (5-carboxamidotryptamine) is typical for a 5-HT2 receptor mediated event. The effect of 5-HT was competitively blocked by ketanserin (pA2 = 8.22).(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Vasopressin induced production of inositol trisphosphate and calcium efflux in a smooth muscle cell line.
Phosphatidylinositol metabolism and 45Ca2+ efflux were examined in a vascular smooth muscle cell line (A7r5). [Arg 8]Vasopressin stimulated the rapid formation (measurable at 1 sec) of inositol phosphates in a concentration-dependent manner. The time course for formation of inositol phosphates was similar to that for 45Ca2+ efflux from preloaded cells. The efflux of 45Ca2+ in response to [Arg8]vasopressin could be inhibited by a vasopressin antagonist. This supports the hypothesis that inositol 1,4,5-trisphosphate plays a role in vasopressin stimulated calcium mobilisation from an intracellular source in cultured vascular smooth muscle cells.
Animals↗
A smooth muscle cell line suitable for the study of voltage sensitive calcium channels.
A cell line originating from the fetal rat aorta has been studied with respect to 45Ca2+ uptake. Kinetic experiments showed an initial rapid uptake followed by a slow linear phase; both the initial rate and the maximum uptake were increased in the presence of 55 mM potassium chloride. The calcium channel antagonists, darodipine (PY 108-068) and verapamil, inhibited both the basal and the potassium chloride stimulated uptake. Neither tetrodotoxin nor furosemide affected either basal or depolarisation induced 45Ca2+ uptake. Blockade of the Na+/K+ ATPase by ouabain and of the Ca2+ ATPase by vanadate caused a net increase in cellular 45Ca2+ accumulation.
Animals↗
Lack of evidence for voltage dependent calcium channels on platelets.
Intracellular calcium was measured in human platelets using the fluorescent calcium indicator Quin 2. A concentration dependent increase was observed with thrombin. Depolarisation induced by high KCl concentrations did not alter [Ca++]i. The calcium agonist Bay K 8644 did not affect resting levels or thrombin stimulated elevation of intracellular calcium. The calcium antagonists diltiazem, verapamil and PN 200-110 did not inhibit the thrombin stimulated elevation in [Ca++]i. Pretreatment of platelets with adenylate cyclase stimulants reduced the rate and magnitude of the maximal [Ca++]i elevation due to thrombin. In addition, thrombin stimulation of 45Ca++ influx was insensitive to Bay K 8644, verapamil, diltiazem and Pn 200-110. We conclude that functional voltage sensitive calcium channels are not present on human platelets.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Effects of calcium entry blockers on calcium fluxes in a vascular smooth muscle cell line.
A smooth muscle cell line was used for studies on the activity of the voltage-sensitive calcium channel (VSCC). Upon exposure to a depolarizing solution or to a calcium channel activator, an increase in 45Ca2+ uptake was observed. Calcium entry blockers inhibited the uptake with potencies very similar to those observed in relaxing depolarization-induced constriction of aortic rings. The two enantiomers of the dihydropyridine PN 200-110 showed about a 300-fold potency difference in blocking 45Ca2+ uptake.
Animals↗
Studies on the mechanism of action of the vasoconstrictive dihydropyridine, CGP 28392.
The effects of the vasoconstrictive dihydropyridine, CGP 29392, upon tension development and 45Ca2+ uptake by vascular smooth muscle were studied using isolated rabbit aorta. CGP 28392 did not elicit contractile responses when administered alone but lowered the threshold for the contractile response to elevated extracellular K+. CGP 28392 also increased the magnitude of the contractions elicited by submaximal concentrations of K+. This effect was competitively antagonized by PY 108-068, a Ca antagonist of the dihydropyridine class. Similarly, the relaxing effects of PY 108-068 upon KCl-constricted rabbit aorta were competitively antagonized by CGP 28392. The calcium content of unstimulated tissues was only minimally increased by CGP 28392 but the 45Ca2+ uptake stimulated by depolarizing levels of K+ was markedly augmented by this agent. Further analysis of the data indicated that CGP 28392 did not alter the relationship between tension development and 45Ca2+ uptake. In contrast, CGP 28392 was much less effective in augmenting the contractile response and 45Ca2+ uptake elicited by noradrenaline. These results thus support the hypothesis that CGP 28392 predominantly facilitates Ca2+ entry through potential-operated Ca2+ channels.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Growth of dissociated neurons in culture dishes coated with synthetic polymeric amines.
Polyornithine and polylysine, the most commonly used coating materials in the preparation of neuronal cultures, can be replaced by polyethyleneimine (PEI) and other synthetic polymeric amines. PEI supports attachment and growth of neurons from fetal rat brain equally well as polyornithine. Furthermore, differentiation of the cultured neurons, as judged by the expression of choline acetyltransferase and of binding sites for substance P, was very similar in cultures grown in dishes coated with PEI or polyornithine.
Animals↗
Correlation between substance P content of primary sensory neurones and pain sensitivity in rats exposed to antibodies to nerve growth factor.
An experimental autoimmune model was used to study effects of nerve growth factor deprivation on the peptide content of sensory neurones and on pain sensitivity. Rats exposed during development to anti-nerve growth factor antibodies showed an increased threshold towards nociceptive stimuli. These animals also showed a significant decrease in substance P levels in their dorsal root ganglia and spinal cord. These results suggest that substance P-containing primary sensory neurones are involved in certain types of nociception.
Animals↗
Inactivation and solubilization of opiate receptors by phospholipases A2.
(1) As previously shown, stereospecific binding of opiates to membrane bound receptors is inhibited by treatment with small amounts of phospholipase A2 from Vipera russelli. This effect is quantified and compared with the enzymes from the venoms of Naja Naja siamensis, Apis Mellifica and from porcine pancreas. All enzymes are equally effective. The inhibition is due to partial phospholipid hydrolysis leading to inactivation of membrane-bound receptor. (2) Bee venom phospholipase A2 together with the synergistically acting peptide, melittin, causes receptor solubilization up to 80% of preformed receptor-ligand complex can be solubilized in this manner. (3) Lysophosphatidylcholine, a product of phospholipid hydrolysis, solubilizes performed receptor-ligand complex to a similar extent. Several other detergents were tested for their ability to solubilize receptor-ligand complex. Digitonin appears to be most effective in solubilizing such a complex.
Animals↗
Purification and chemical characterization of melittin and acetylated derivatives.
Melittin, the main basic and hydrophobic peptide of bee venom, displays marked detergent-like properties. At high peptide concentration, and depending on salt and pH, it forms a tetramer. This is prevented by using urea. A purification procedure in presence of 4.0 M urea was developed to prepare melittin in its monomeric form, free of other venom constituents such as N alpha-formyl melittin, degradation products of peptides and phospholipase A2. NH2-residues on the melittin molecule were modified by reaction with acetic anhydride to alter the asymmetrical charge distribution supposed to confer detergent-like properties to the molecule. This gave rise to di- and mono acetyl derivatives which could be used, once isolated, to study further the melittin structure-activity relationship.
Animals↗