Biomedical subjects

D J Triggle

Publications and source records attributed to D J Triggle.

At least 127 records · Page 7Linked to original sources

PubMed1985

Functional interactions between organic calcium channel antagonists in smooth muscle.

The Ca2+ channel antagonists D-600, diltiazem, and nifedipine are competitive antagonists of Ca2+ responses in K+-depolarized guinea pig taenia coli and rat mesenteric artery preparations. pA2 values for D-600, diltiazem, and nifedipine in taenia coli were 8.28, 7.44, and 9.27, respectively and in mesenteric artery, 9.6, 7.83, and 10.4, respectively. The combination of nifedipine plus diltiazem gave in both tissues antagonism greater than that calculated on the basis of additivity. This suggests, consistent with published 3H-labelled radioligand binding data, that diltiazem and nifedipine interact at distinct sites. However, the combination nifedipine plus D-600 yielded antagonism consistent with additivity of response.

Animals↗

PubMed1985

Ca2+ channel antagonist actions in bladder smooth muscle: comparative pharmacologic and [3H]nitrendipine binding studies.

The actions of a series of 15 Ca2+ channel antagonists including D-600, nifedipine, and diltiazem were examined against K+ depolarization and muscarinic receptor induced responses in guinea pig bladder smooth muscle. Responses of bladder are very dependent upon extracellular Ca2+ and sensitive to the Ca2+ channel antagonists, the tonic component more than the phasic component of response. Regardless of stimulant, K+ or methylfurmethide (MF), or component of response, the same rank order of antagonist activities is expressed, suggestive of a single structure-activity relationship and the existence of a single category of binding site which may, however, exist in several affinity states. High affinity binding of [3H]nitrendipine (KD = 1.1 X 10(-10) M) occurs in bladder membranes, and similar high affinity binding was found in microsomal preparations from other smooth muscles including guinea pig and rat lung, rat vas deferens, uterus, and stomach. [3H]nitrendipine binding in the bladder was sensitive to displacement by other 1,4-dihydropyridines, paralleling their pharmacologic activities and showing excellent agreement with binding data previously obtained for guinea pig ileal smooth muscle. Comparison of pharmacologic data for inhibition of K+- and MF-induced responses by a common series of Ca2+ channel antagonists in bladder and ileum revealed excellent correlations. Neither pharmacologic nor binding studies suggest significant differences in Ca2+ channel antagonist properties in smooth muscle from bladder and intestine.

Animals↗

PubMed1985

Conformational features of calcium channel agonist and antagonist analogs of nifedipine.

The crystal and molecular structures of methyl 2,6-dimethyl-5-nitro-4-(2-trifluoromethylphenyl)-1,4- dihydropyridine-3-carboxylate and ethyl 4-(2-difluoromethoxyphenyl)-1,4,5,7,-tetrahydro-2-methyl-5-oxof uro[3,4-b]pyridine-3-carboxylate, which are analogs of the calcium channel antagonist nifedipine reported to have agonist activity, have been determined. The conformations of these two agonists are compared with the conformational features shown by nifedipine and related 1,4-dihydropyridine calcium channel antagonists. Common conformational features shown by these agonists and antagonists allow both to bind to the same plasma membrane receptor while subtle differences in hydrogen-bonding activity of the amine group, and ester group orientation and hydrophobic fit, may control the availability of channel open and closed states.

Calcium↗

PubMed1985

Subcellular distribution of dihydropyridine isothiocyanate binding in guinea-pig ileal smooth muscle.

The subcellular distribution of the 3H--2,6-dimethyl-3,5-dicarbomethoxy-4(2-isothiocyano) phenyl-1,4-dihydropyridine (DPSCN) binding to guinea-pig ileal smooth muscle was studied by subcellular fractionation. Initial experiments on subcellular fractionation of 3H-DPSCN-labelled tissues by differential centrifugation showed that there was an excellent correlation between the levels of the label present in a fraction and the plasma membrane marker phosphodiesterase I (r = 0.98) but not between the label and the putative endoplasmic reticulum marker NADPH: cytochrome-c-reductase (r = 0.56) or the inner mitochondrial marker cytochrome-c-oxidase (r = 0.36). Centrifugation of the microsomes on a continuous sucrose density gradient showed an excellent correlation of the migration of the label with phosphodiesterase I activity (r = 0.93) but not with the activities of NADPH: cytochrome-c-reductase (r = 0.66) or cytochrome-c-oxidase (r = 0.44). Treatment of microsomes with digitonin (1 mg/ml) followed by centrifugation on continuous sucrose density gradients increased the weighted mean densities of the phosphodiesterase activity (plasma membrane marker) and the labelling by similar magnitudes (0.04 to 0.06 g/ml). The weighted mean densities of NADPH: cytochrome-c-reductase and the cytochrome-c-oxidase were not altered significantly. It is concluded that in the guinea-pig ileal smooth muscle, DPSCN labels the plasma membrane specifically.

Animals↗

PubMed1984

Interaction of phenoxybenzamine with muscarinic receptors and calcium channels.

Phenoxybenzamine (POB, 10(-6) - 10(-4) M) inhibited the responses of guinea pig ileal longitudinal smooth muscle to both muscarinic agonists and K+-depolarization but was more effective against the agonist-induced responses. POB inhibited binding of both the muscarinic antagonist [3H]quinuclidinyl benzilate (QNB) and the Ca2+ channel antagonist [3H]nitrendipine and was, paralleling its effects on mechanical responses, more effective against [3H]QNB binding. POB reduced specific [3H]QNB binding by a reduction in Bmax with no change in KD, but inhibited [3H]nitrendipine binding by reducing KD with no effect on Bmax. It is suggested that the activity of POB against Ca2+ channels may underlie the ability of POB, and other 2-halogenoethylamines, to inhibit a wide variety of apparently discrete pharmacological events.

Animals↗

PubMed1984

Comparison of high affinity binding of calcium channel blocking drugs to vascular smooth muscle and cardiac sarcolemmal membranes.

The binding of the 1,4-dihydropyridine calcium channel blocker [3H]nitrendipine to canine cardiac sarcolemmal and bovine aortic membranes was found to be rapid, specific, saturable, and reversible. Dissociation constants (Kd) determined by Scatchard analysis were 0.14 and 0.16 nM and the maximal numbers of binding sites (Bmax) were 0.96 +/- 0.2 and 0.08 +/- 0.01 pmole/mg protein for cardiac and aortic membranes respectively. Values of Kd calculated from kinetic data were approximately 0.10 nM for both membrane preparations. Competition assays with the enantiomers of a nisoldipine derivative indicated that [3H]nitrendipine binds stereoselectively. The order of potency of several nifedipine analogs for inhibition of binding of [3H]nitrendipine to cardiac and aortic membranes paralleled their relative potencies for inhibition of contraction in smooth muscle. It is concluded that the high affinity binding sites for nitrendipine in bovine aortic smooth muscle membranes are similar to those of canine ventricular sarcolemma.

Animals↗

PubMed1984

Nitrendipine and isoproterenol induce phosphorylation of a 42,000 dalton protein that co-migrates with the affinity labeled calcium channel regulatory subunit.

Slow inward calcium channels in canine cardiac membranes were affinity labeled with the calcium channel analogue, [3H]o-NCS [2,6 dimethyl-3,5-dicarbomethoxy-4-(2- isothiocyanatophenyl )-1, 4-dihydropyridine], in the presence and absence of cold o-NCS or nicardipine. A major specifically labeled peak was identified with Mr 42,000 on NaDodSO4 polyacrylamide gels. In parallel experiments the effects of the calcium channel antagonist, nitrendipine and a variety of other chemical mediators were tested for their ability to stimulate protein phosphorylation in cardiac membranes. These data demonstrate that both nitrendipine and isoproterenol induce the phosphorylation of a 42,000 dalton protein via a kinase endogenous to the cardiac membranes and that the effects of isoproterenol are attenuated by carbachol.

Affinity Labels↗

PubMed1984

Specific binding of a calcium channel activator, [3H]BAY k 8644, to membranes from cardiac muscle and brain.

BAY k 8644 is a member of a new class of drugs that directly activates Ca2+ channels. This 1,4-dihydropyridine was found to bind to both high and low affinity sites on rabbit ventricular microsomes and guinea pig brain synaptosomes. The dissociation constant obtained from Scatchard analysis with [3H]BAY k 8644 was 2 to 3 nM for the high affinity binding site, and the estimated maximal number of binding sites was 0.8 and 0.4 pmol/mg protein for heart and brain membranes, respectively, at 15 degrees C. Competition between nitrendipine and [3H]BAY k 8644 indicated a common high affinity binding site for Ca2+ channel activators and antagonists. The results suggest that the 1,4-dihydropyridine Ca2+ channel antagonists do not act as simple channel plugs.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1984

Subcellular distribution of [3H]nitrendipine binding in smooth muscle.

Distribution of specific binding sites for [3H]nitrendipine was studied in subcellular fractions isolated from rat gastric fundus smooth muscle and from rat myometrium. There was an excellent correlation between the distribution of [3H]nitrendipine binding determined at the nitrendipine concentrations of 0.138 and 1.38 nM, and the distribution of the plasma membrane markers K+-activated ouabain-sensitive p-nitrophenylphosphatase, 5'-nucleotidase, phosphodiesterase I, and Mg-ATPase, but not between the mitochondrial markers cytochrome c, oxidase, succinate-dependent cytochrome c reductase, or rotenone-insensitive NADH-dependent cytochrome c reductase or the putative endoplasmic reticulum marker NADPH-dependent cytochrome c reductase. The binding occurred with high affinity and with a similar (0.097-0.146 nM) equilibrium dissociation constant to all the fractions, even though the density of binding sites varied and was highest in the plasma membrane marker-enriched fractions. The maximal binding in the plasma membrane-enriched fraction from the rat gastric fundus smooth muscle was 0.43 +/- 0.04 pmol/mg, and in that from rat myometrium was 0.72 +/- 0.09 pmol/mg. Thus in the two smooth muscles studied the plasma membrane is the locus of the high affinity nitrendipine binding.

Animals↗

PubMed1984

BAY K 8644, a 1,4-dihydropyridine Ca2+ channel activator: dissociation of binding and functional effects in brain synaptosomes.

K+-stimulated 45Ca2+ uptake into rat brain and guinea pig cerebral cortex synaptosomes was measured at 10 s and 90 s at K+ concentrations of 5-75 mM. Net increases in 45Ca2+ uptake were observed in rat and guinea pig brain synaptosomes. 45Ca2+ uptake under resting or depolarizing conditions was not increased by the 1,4-dihydropyridine BAY K 8644, which has been shown to activate Ca2+ channels in smooth and cardiac muscle. High-affinity [3H]nitrendipine binding in guinea pig synaptosomes (KD = 1.2 X 10(-10) M, Bmax = 0.56 pmol mg-1 protein) was competitively displaced with high affinity (IC50 2.3 X 10(-9) M) by BAY K 8644. Thus high-affinity Ca2+ channel antagonist and activator binding sites exist in synaptosome preparations, but their relationship to functional Ca2+ channels is not clear.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1984

Specific and non-specific desensitization of guinea-pig ileal smooth muscle.

The effects of cis-2-methyl-4-dimethylaminomethyl-1-3-dioxolane methiodide (CD), a muscarinic agonist, histamine, substance P and K+-stimulation on the mechanical responses, Ca2+-dependence and desensitization in guinea-pig ileal longitudinal smooth muscle have been studied. The mechanical responses to all four stimulants are highly dependent upon extracellular Ca2+(Ca2+EXT) and are blocked by the Ca2+ channel antagonist nicardipine. The tonic (slow) components of response are more dependent on Ca2+EXT and are more sensitive to nicardipine (IC50 values 5.0 X 10(-8) - 2.5 X 10(-9)M) than are the phasic (fast) components of response. Tissue exposure to CD (5 X 10(-7)M, 10 min) or histamine (3 X 10(-6)M and 3 X 10(-4)M, 10 min) produces short term nonspecific desensitization but substance P (5 X 10(-8)M, 10 min) produces only specific desensitization. K+-induced responses neither desensitize nor are desensitized. Desensitization is concentration- and time-dependent for both specific and nonspecific processes. Nonspecific desensitization is protected by elevation of K+ concentration (5.36mM) in the incubating medium, by dithiothreitol and by inhibitors (mepacrine,p-bromophenacyl bromide and phenylgloxal) of phospholipase A2 and is potentiated by mellitin, an activator of phospholipase A2. Desensitization produced by the muscarinic agonist CS is protected by Gpp(NH)p (10(-4)M), but histamine-induced desensitization is unaffected. There is no loss of muscarinic receptors, measured by [3H]QNB binding following tissue exposure to low concentrations of CD (5.0 X 10(-7)M) for up to 72 h. However, an apparent loss of receptors (20-30%) is measured following 10-90 min exposure of tissue to 10(-3)M CD. It is suggested that contractions of guinea-pig ileal longitudinal smooth muscle elicited by CD, histamine, substance P or K+ mobilize a common pool of Ca2+ through a Ca2+ channel antagonist (nicardipine) sensitive pathway. However, the existence of short term nonspecific desensitization (CD and histamine), specific desensitization (substance P) or no desensitization (K+ stimulation) indicates that significant differences exist in the pathways linking initial stimulus to mechanical response. The ability of elevated K+ to protect against nonspecific desensitization suggest that post stimulus membrane hyperpolarization may represent one contributing component to nonspecific desensitization. Products of phospholipid degradation may also contribute to desensitization since inhibitors or activators of phospholipase A2 prevented or potentiated respectively, nonspecific desensitization.

Animals↗

PubMed1984

Calcium-dependence and antagonism of responses to alpha 1- and alpha 2-adrenoceptor agonists in vascular tissues from hypertensive and normotensive rats.

Inhibition by D600 (methoxyverapamil) of responses to an alpha 2-adrenoceptor selective agonist, B-HT 920, (6-allyl-2-amino-5, 6, 7, 8-tetrahydro-4H-thiazolo [4, 5-d] azepin dihydrochloride), an alpha 1-adrenoceptor selective agonist, phenylephrine (PE), and a nonselective agonist, noradrenaline (NA), was studied in isolated preparations of the aortae and carotid arteries obtained from young (5-7 weeks) and old (15-17 weeks) hypertensive (SHR) and normotensive (WKY) rats. Maximum responses of WKY tissues to B-HT 920 were the most sensitive, PE-induced responses the least sensitive and maximum responses to NA were intermediate in their sensitivity to inhibition by D600. Sub-maximal responses to NA and PE were not different in their sensitivity to inhibition by D600, but were less sensitive than the responses to B-HT 20. Sub-maximal responses to PE were significantly more sensitive to D600 inhibition than were the maximal responses to this agonist. NA-induced responses of tissues from older SHR were less sensitive to inhibition by D600 when compared to responses in WKY rats. Responses to B-HT 920, in tissues suspended in calcium-free solutions, showed the largest decline compared to NA- and PE-induced responses. We conclude that responses to B-HT 920 largely utilize extracellular calcium. PE- and NA- induced responses mobilize extracellular calcium to varying degrees depending upon the concentration of the agonist employed to elicit the response.

Adrenergic alpha-Agonists↗

PubMed1984

Calcium reactivity and antagonism in portal veins from spontaneously hypertensive and normotensive rats.

Reactivities of portal veins from spontaneously hypertensive rats (SHR) and normotensive controls (Wistar Kyoto, WKY) at 5-7 and 15-17 weeks of age were compared. Systolic blood pressures were not different at 5-7 weeks but those of SHR were significantly elevated (177 +/- 4 mmHg) (1 mmHg = 133.322 Pa) at 15-17 weeks. Spontaneous activity, frequency, and tension were greater in SHR for both age groups. Young SHR were more sensitive to K+ at 5-7 weeks but less sensitive at 15-17 weeks than age-matched WKY rats. Sensitivity to Ca2+ in a K+-depolarizing medium was higher in SHR than in WKY for both age groups. Maximum tension responses to K+ or Ca2+ were greater in SHR. The Ca2+ channel antagonists nifedipine, nitrendipine, and nisoldipine were potent inhibitors of both noradrenaline- and K+-induced responses but did not show differences in inhibitory activity between WKY and SHR.

Age Factors↗

PubMed1984

Calcium channel activation in vascular smooth muscle by BAY K 8644.

BAY K 8644 (methyl-1,4-dihydro-2,6-dimethyl-3-nitro-4-(2-trifluoromethylphenyl) pyridine-5-carboxylate) and CGP 28 392 (ethyl-4(2-difluoromethoxyphenyl)-1,4,5,7-tetrahydro-2-methyl-5-++ +oxofuro- [3,4-b]pyridine-3-carboxylate) are closely related in structure to nifedipine and other 1,4-dihydropyridine Ca2+ channel antagonists. However, both BAY K 8644 and CGP 28 392 serve as activators of Ca2+ channels. In the rat tail artery, responses to BAY K 8644 are dependent upon Ca2+ext and prior stimulation by K+ or by the alpha-adrenoceptor agonists, phenylephrine and BHT 920 (6-allyl-2-amino-5,6,7,8,-tetrahydro-4H-thiazolo[4,5-d]azepin dihydrochloride). Responses are blocked noncompetitively by the Ca2+ channel antagonists D-600 [-)-D-600 greater than (+)-D-600) and diltiazem, but competitively by nifedipine (pA2 = 8.27). This suggests that activator and inhibitor 1,4-dihydropyridines interact at the same site. BAY K 8644 potentiates K+ responses and Ca2+ responses in K+-depolarizing media. The leftward shift of the K+ dose--response curve produced by BAY K 8644 suggests that this ligand facilitates the voltage-dependent activation of the Ca2+ channel. The pA2 value for nifedipine antagonism of BAY K 8644 responses is significantly lower than that for nifedipine antagonism of Ca2+ responses in K+ (25-80 mM) depolarizing media (9.4-9.6), suggesting that the state of the channel may differ according to the activating stimulus.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1984

Cellular action of calcium channel blocking drugs.

The calcium antagonists are an exciting "class" of drugs that are already making an important impact in the treatment of coronary artery disease. All three drugs in use in the US, verapamil, nifedipine, and diltiazem, are effective coronary vasodilators. In addition, verapamil and diltiazem have antiarrhythmic activity against certain supraventricular abnormalities. Since the calcium antagonists dilate peripheral resistance vessels, they may also be useful in hypertension. There are reports also that they may be effective in migraine headache, probably because of their "antispastic" activity in cerebral vessels. Since calcium is ultimately involved in all processes that require stimulation, either electrical or chemical, it is not surprising that drugs whose primary effect appears to be an inhibition of the entry of calcium across or through cell membranes should exhibit a wide spectrum of therapeutic activity. The stereospecific binding of dihydropyridine derivatives to membrane preparations, and the effects of those and other calcium antagonists on the binding process, should yield important information on the mechanisms of calcium antagonist action.

Binding Sites↗

PubMed1984

Cellular calcium metabolism: activation and antagonism.

The role of Ca2+ as a mediator of many cellular responses, including stimulus-secretion and excitation-contraction coupling, is reviewed. Pathways of Ca2+ mobilization are discussed in terms of both intracellular and plasmalemmal processes. The extent to which specific antagonists exist, active at these several sites, is noted, with particular emphasis on the clinically available Ca2+ channel antagonists, including verapamil, nifedipine, and diltiazem.

Allosteric Regulation↗

PubMed1984

The 1,4-dihydropyridine receptor: a regulatory component of the Ca2+ channel.

Evidence that nitrendipine (2,6-dimethyl-3-carbomethoxy-5-carbomethoxy-4-(3-nitrophenyl)1,4- dihydropyridine) and other 1,4-dihydropyridines interact at a specific site to antagonize Ca2+ channel function is reviewed. This evidence derives from electrophysiologic and pharmacologic studies and is confirmed by [3H]radioligand binding to excitable tissues. The availability of 1,4-dihydropyridine activator ligands including BAY K 8644 (2,6-dimethyl-3-carbomethyl-5-nitro-4-(2-trifluoromethyl)1,4- dihydropyridine) suggests that this site is intimately concerned with Ca2+ channel regulation. The actions of the 1,4-dihydropyridine ligands are discussed with particular reference to their tissue selectivity and the functional relationships of binding sites to Ca2+ channels.

Animals↗

PubMed1984

Characteristics of the binding of [3H]nitrendipine to rabbit ventricular membranes: modification by other Ca++ channel antagonists and by the Ca++ channel agonist Bay K 8644.

This study was carried out to characterize [3H]nitrendipine binding to cardiac membranes and to test the hypothesis that high affinity binding of Ca++ channel antagonists and agonists is to Ca++ channels. Binding was specific, rapid, reversible and stereoselective. The relative order of potency of nifedipine analogs for inhibition of binding was the same as that for inhibition of smooth and cardiac muscle contraction. Results with diltiazem, verapamil and lidoflazine were consistent with the hypothesis that nondihydropyridine Ca++ channel antagonists act at one or more sites allosterically linked to the 1,4-dihydropyridine site in cardiac cells. The Ca++ channel agonist Bay K 8644 [methyl-1,4-dihydro-2,6-dimethyl-3-nitro-4-(2-trifluoromethylphenyl)-pyr idine- 5-carboxylate] displaced specifically bound [3H]nitrendipine in an apparently competitive manner with an IC50 value of 5 nM. The results suggest that organic antagonists do not act by physically blocking the Ca++ channel. The data also support the hypothesis that the high affinity binding sites for [3H]nitrendipine in isolated cardiac membranes are associated with Ca++ channels that are inactivated or are otherwise unavailable for opening.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗