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H Porzig

Publications and source records attributed to H Porzig.

At least 37 records · Page 2Linked to original sources
PubMed1988

Potential-dependent allosteric modulation of 1,4-dihydropyridine binding by d-(cis)-diltiazem and (+/-)-verapamil in living cardiac cells.

We have studied allosteric effects of the Ca channel blockers d-(cis)-diltiazem, (+/-)-verapamil, and (S)- and (R)-devapamil on the specific binding of the 1,4-dihydropyridine derivative (+)-[3H]PN 200-110 to intact tissue cultured rat heart cells. In polarized cells (membrane potential, -38 +/- 4 mV) d-(cis)-diltiazem (5 microM) increased the affinity of the radiolabel 2-4-fold causing 100-187% enhancement of binding at (+)-PN 200-110 concentrations below 0.5 nM. (+/-)-Verapamil (0.1-3 microM) had a similar, although smaller, effect on (+)-PN 200-110 binding. The two enantiomers of devapamil were without effect. In depolarized cells (membrane potential, 0 mV) d-(cis)-diltiazem had a small and the phenylalkylamines a strong inhibitory effect on (+)-PN 200-110 binding, mainly due to a reduction of binding affinity. At 50% receptor occupation by the radioligand, (R)-devapamil, (+/-)-verapamil, and (S)-devapamil displaced 40, 55, and 75%, respectively, of specifically bound radiolabel. Half-maximal effects were reached with 50, 20, and 4.5 nM, respectively, of the three compounds. Compared with nominally Ca-free medium (containing 3-5 microM Ca), addition of 1.25 mM CaCl2 caused an increase in the maximal binding capacity for (+)-PN 200-110 in both polarized and depolarized cells. However, Ca had only marginal effects on the allosteric interactions between (+)-PN 200-110, d-(cis)-diltiazem, and verapamil. We conclude from our results that positive cooperative interactions between Ca channel blockers prevail under conditions in which the voltage-dependent Ca channel can fluctuate between closed, open, and inactivated states. Negative cooperativity is usually observed under conditions in which all channels are inactivated (depolarized cells, fragmented membranes). Therefore, it is impossible to predict the type and the extent of allosteric interactions in vivo from studies in cell homogenates.

Allosteric Regulation↗
PubMed1987

Voltage-dependent cooperative interactions of calcium channel ligands in intact cardiac cells.

Voltage-dependent Ca channels were studied in living, tissue cultured rat heart cells using patch clamp analysis of single channels and radioligand binding studies at different membrane potentials. Ca channel activating and blocking dihydropyridines (DHP) show cooperative interaction and, therefore, bind to at least two different binding sites on the channel protein. Cooperative interactions between activating and blocking DHP, between DHP and verapamil and between DHP and diltiazem are all voltage dependent. The type of interaction in polarized cells and hence, the possible effects in vivo cannot be predicted from studies in cell homogenates.

Allosteric Site↗
PubMed1986

Studies on Ca channels in intact cardiac cells: voltage-dependent effects and cooperative interactions of dihydropyridine enantiomers.

We have investigated the effects of two oppositely acting enantiomers of the 1,4-dihydropyridine derivative 202-791 on voltage-dependent Ca channels by combining electrophysiological techniques and binding studies. The (S)-enantiomer of 202-791 promoting prolonged openings of single Ca channels, and thereby increasing transmembrane Ba currents, was classified as channel activator. The (R)-enantiomer favoring a closed state of the channel, and thereby reducing Ba currents, was classified as a channel blocker. Both compounds shifted the steady state current inactivation curve toward more negative potentials. At holding potentials positive to -20 mV, the Ca channel-activating effect of the (S)-enantiomer turned over into a blocking effect. In cells with normal resting potential the combination of the two enantiomers revealed a possible positive cooperative effect resulting in an enhancement of the open state probability of the channels. At depolarized holding potentials the activator enhanced the inhibitory effect of the blocker. Binding studies in intact cells were performed by using the radiolabeled channel-blocking dihydropyridine 3H-(+)-PN 200-110. The results showed a strong increase in binding affinity but no change in binding capacity when the cells were depolarized. Analysis of the interactions of (S)- and (R)-202-791 with this radioligand indicated stimulation of 3H-(+)-PN 200-110 binding by the (S)-enantiomer in polarized cells (membrane potential -38 +/- 4 mV). This effect could be attributed to an increase in binding affinity. The (R)-enantiomer had no such positive cooperative effect, but acted as a purely competitive ligand. Depolarization to 0 mV increased the apparent affinity of both enantiomers by factors of 38 (blocker) and 12 (activator), but abolished the cooperative effect of (S)-202-791 on the binding of the radioligand. Ca ions had little effect on the binding of 3H-(+)-PN 200-110 in polarized cells. However, in the presence of the activating (S)-enantiomer, Ca transformed the usual hyperbolic binding isotherm of the radioligand into a strongly sigmoid curve. Sigmoidicity was minimal with 3-5 microM Ca and maximal with 0.5 mM Ca. Together these data demonstrate homotropic and heterotropic cooperative interactions between channel activator and channel blocker. They indicate that at least two high affinity binding sites for dihydropyridines are associated with voltage-dependent Ca channels. Voltage dependence of both--binding affinity and cooperativity--suggests that these binding sites are located close to a structural component of the channel which is involved in the potential-sensitive gating process.

Aging↗
PubMed1986

The voltage-dependent effect of 1,4-dihydropyridine enantiomers on Ca channels in cardiac cells.

We studied voltage-dependent binding and action of 1,4-dihydropyridine enantiomers (Sandoz (+)-(S) & (-)-(R)-202-791) in intact cardiac cells. Drug action was studied by patch clamp method. (+)-enantiomer primarily prolonged open time of Ca channel, thus enhancing Ca currents, while (-)-enantiomer predominantly favored closed state of the channel, reducing the currents. Electrophysiological study also revealed that steady-state inactivation was greatly enhanced by both enantiomers. Therefore, it seems that both enantiomers have a capability of activating and blocking effects. Strongly voltage-dependent binding affinity of these enantiomers was revealed by displacement of 3H-(+)-PN200-110 with each enantiomers. Binding affinity of both enantiomers in depolarized cells was much higher than in polarized cells. These results indicate that both enantiomers have higher affinity when Ca channels are inactivated.

Animals↗
PubMed1985

Quantitative differences in the pharmacological effects of (+)- and (-)-cathinone.

The optically pure isomers of cathinone were prepared by separating synthetic cathinone racemate and used to study central and peripheral effects of these indirect sympathomimetics in rats and guinea pigs. The (-)-isomer was significantly more potent than the (+)-isomer in stimulating locomotor activity whereas no difference was observed with respect to their cardiac effects. In analogy to observations with (+)- and (-)-amphetamine such variable isomer discrimination may be due to different stereoselectivities of amine uptake mechanisms in the target tissues.

Alkaloids↗
PubMed1985

Regulation by 8-Br-cAMP of beta-adrenoceptors in cultured myocardial cells.

Primary cultures of myocardial cells from neonatal rats were exposed for up to 5 days to the cyclic AMP derivative 8-Br-cAMP. After one day of exposure to the nucleotide, an increase in specific binding capacity of the hydrophilic beta-adrenoceptor antagonist 3H-CGP 12177 was observed in intact cells. (-)-Isoprenaline displaced the radioligand from its binding site. Neither the KD of the antagonist, nor that of the agonist were significantly affected by 8-Br-cAMP. The loss of beta-adrenoceptors during desensitization by long term treatment of the cells with isoprenaline could be partially prevented by 8-Br-cAMP. Only in desensitized, but not in normal cells was isoprenaline-induced cAMP formation significantly enhanced by 8-Br-cAMP pretreatment. This may indicate that beta-adrenoceptors which appear during 8-Br-cAMP exposure are poorly coupled to the adenylate cyclase. Alternatively, the change in receptor density may be accompanied by alterations of other components in the beta-adrenergic system, e.g. an inhibition of the adenylate cyclase. We suggest that cAMP-dependent feed-back regulation of the beta-adrenergic system may play a role during postnatal myocardial differentiation.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1984

Plasma catecholamines, beta-adrenergic receptors, and isoproterenol sensitivity in endurance trained and non-endurance trained volunteers.

Six male non-endurance trained subjects (S) and six marathon runners (M) underwent graded treadmill exercise (T) and isoproterenol stimulation (I; 2 and 4 microgram X min-1). beta-adrenergic receptor density was additionally determined as the amount of 3H-Dihydroalprenolol (DHA) specifically bound on intact polymorphonuclear leucocytes. Heart rate, VO2 uptake, lactate, plasma noradrenaline, and adrenaline were estimated during T. Heart rate, stroke volume, cardiac output, as well as lactate, glucose, free fatty acids (FFA), and glycerol levels in the blood were determined during I. M showed the known training-dependent responses during T, such as lower heart rates, lactate levels, and plasma catecholamines at identical work loads, as well as higher VO2 max than S. I-induced cardiac output increase was quite similar in both groups. Stroke volume, however, increased significantly in M and stayed constant in S. Lactate decreased (S), glucose increased significantly (M), glycerol increased similarly in both groups, FFA rise was less marked in S. I-induced stroke volume response (I) may be indicative of a more economic regulation of heart work in M than S. Lactate decrease and less marked FFA increase, as observed in S, may be the result of a somewhat higher cardiac energy demand, dependent on less economic heart work. Higher DHA-binding as observed in M, as well as stroke volume response and glucose increase, may be indicators of a training-dependent rise in sensitivity to catecholamines. The unsolved question is, however, to what extent beta-receptor responses in intact blood cells are significant for receptor behavior in other organs.

Adult↗
PubMed1984

Recovery of beta-adrenoceptors and cyclic AMP response after long term treatment of intact heart cells with beta-blockers.

We have studied the recovery of receptor binding and of isoprenaline-stimulated cyclic AMP responses after chronic (2-5 days) exposure of tissue-cultured living rat heart cells to several beta-adrenoceptor antagonists. Most experiments were performed with [3H]- (+/-)-carazolol and [3H]-(+/-)-CGP 12177, as prototypes of high affinity lipophilic and hydrophilic ligands respectively. Chronic antagonist treatment did not alter the total number of receptors nor did it cause intracellular accumulation of the ligands. At the end of the treatment, radiolabelled antagonists were displaced either by 'infinite' dilution of the incubation medium or by competitive displacement with the non-labelled ligand (-)-timolol. In dilution assays dissociation of carazolol from specific sites was biphasic with t 1/2 values of 41 +/- 14 and 219 +/- 15 min. Dissociation of CGP 12177 was monophasic with t 1/2 of 102 +/- 2 min. Timolol enhanced the dissociation rates of both radioligands and suppressed the slow phase of carazolol dissociation. Isoprenaline-stimulated cyclic AMP formation did not recover in parallel with the release of the two antagonists from receptor binding sites. To reach about 80% of control values for receptor availability or cyclic AMP response required 3 h and 24 h washout periods, respectively, after carazolol (0.2 nM) treatment, or 1.5 and 12 h washout periods after CGP 12177 (4 nM) treatment. Such a 'decoupling' effect was not observed during recovery from chronic exposure to the antagonists, timolol and propranolol. We conclude that some antagonists cause a novel form of desensitization that is not linked to their partial agonistic potency. Moreover, carazolol-type drugs seem to induce an additional isomeric form of the beta-receptor that is not recognized by other antagonists. These observations could explain the well known discrepancy between long duration of action and rapid removal from the circulation of several antagonists in current therapeutic use.

Adrenergic beta-Antagonists↗
PubMed1984

Changes in beta-adrenoceptor binding properties and receptor-cyclase coupling during in vitro maturation of rat reticulocytes.

The loss of beta-adrenergic responsiveness during reticulocyte maturation was studied under tissue culture conditions in a defined cell population from rats. Initial beta-receptor density and receptor-mediated cAMP formation in culture medium (RPMI 1640) exceeded the values obtained in isotonic KC1-buffer by 56 and 120% respectively. During cell cultivation receptor density and hormonal responsiveness decreased rapidly by 40-50% of their initial values within the first 20 h. In the following 3 days the rate of loss varied between 10 and 15%/d. The same time course was observed for the maturation-dependent decrease in forskolin-stimulated cAMP formation. ATP depletion of cultivated cells caused a complete and irreversible loss of cAMP response within 90 min. Our results indicate that cell metabolism regulates the strength of the hormonal response. A defect in adenylate cyclase or in cyclase N-protein interaction seems to be rate-limiting for the functional inactivation of the beta-adrenergic system during reticulocyte maturation.

Adenosine Triphosphate↗
PubMed1983

[Beta-adrenergic receptors and plasma catecholamine behavior in trained and untrained athletes].

6 sports students (VO2 max. 54.0 +/- 2.6 ml/kg . min) and 6 marathoners (VO2 max. 65.7 +/- 2.0 ml/kg . min) performed graded treadmill tests. Free plasma catecholamines (noradrenaline and adrenaline), heart rate, lactate were assessed at rest and during exercise. The behaviour of beta-adrenergic receptors of polymorphonuclear leukocytes was estimated additionally before the treadmill test. The maximum running velocity was 14.6 +/- 0.8 km/h (sports students) and 17.3 +/- 0.6 km/h (marathoners). Noradrenaline was approximately 46% (v = 12 km/h) to 67% (v = 14 km/h) lower in the group of marathoners than in sports students; adrenaline showed no or smaller differences between both groups. No significant differences in the plasma catecholamine behaviour occurred between the groups at rest and during maximum exercise. Specific binding of 3H-Dihydroalprenolol to intact cells was higher in the marathoners than in sports students (p greater than 0.01). Scatchard analysis revealed a maximum binding of 21.1 fmol/10(7) cells (sports students) and 35.3 fmol/10(7) cells (marathoners), which indicated approximately 1,300 (sports students) and 2,150 binding sites cell (marathoners). Inverse correlations between noradrenaline (r = -0.63), VO2max. (r = -0.79) and the specific binding of 3H-DHA were observed. The reduction of plasma catecholamines (approximately 46-67%) and the increase in specific binding sites (approximately 67%) were in the same range.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1983

[Determination of beta-receptors on intact polymorphonuclear leukocytes in autologous plasma].

The binding of tritium labelled radioligand dihydroalprenolol was investigated on live polymorphonuclear leukocytes of 6 endurance trained (VO2 max. 65.7 +/- 2.0 ml/kg . min), and 9 non-endurance trained subjects (VO2 max. 52.0 +/- 4.0 ml/kg . min). The specific binding of dihydroalprenolol is seen as an indicator of the beta-receptor density. The specific binding of dihydroalprenolol is defined as the difference between the total binding and that amount of dihydroalprenolol that could not be displaced. The leukocytes were reincubated for the binding studies in their autologous plasma. The specific binding of dihydroalprenolol on live polymorphonuclear leukocytes shows a levelling off behaviour at a concentration of approximately 2 nmol/l dihydroalprenolol in trained as well as in untrained subjects. The specific binding amounts to about 85% (0.1 nmol/l dihydroalprenolol) to 51% (2.0 nmol/l dihydroalprenolol) of the total binding. Based on Scatchard analysis, Bmax was determined as 41.2 fmol/10(7) cells (trained subjects) and 21.6 fmol/10(7) cells (untrained subjects). KD is 0.44 nmol/l dihydroalprenolol (untrained subjects), and 0.49 nmol/l dihydroalprenolol (trained subjects). The beta-adrenergic binding sites are approximately 1300 (untrained subjects), and 2500 binding sites/cell (trained subjects). The specific binding of dihydroalprenolol on live polymorphonuclear leukocytes is significantly higher in trained than in untrained subjects. This training dependent change in beta-receptor density may be an indicator of an increased sensitivity to catecholamines.

Alprenolol↗
PubMed1983

In vitro maturation of the rat reticulocyte beta-adrenoceptor adenylate cyclase system.

Reticulocytes but not mature erythrocytes are equipped with a functional beta-adrenergic system characterizing cells which are under adrenergic control. We have followed the decline of beta-adrenergic functions in a defined, phenylhydrazine-induced population of rat reticulocytes during maturation under cell culture conditions in vitro. beta-Agonist or forskolin-stimulated cyclic 3',5'-AMP (cAMP) formation decreased by 35 +/- 5% within 18 h and by 75 +/- 7% within 5 days. Regulatory control, i.e. the desensitization after chronic agonist exposure was observed at least for the first 18 h of incubation. The protein components of the system (receptors, nucleotide binding protein (N-protein), cyclase) seem to survive longer than 5 days. Our results suggest that an impaired interaction between N-protein and adenylate cyclase may be the initial step in the inactivation of the beta-adrenergic system in reticulocytes.

Adenylyl Cyclases↗
PubMed1982

Beta-adrenergic actions on cardiac cell membranes.

Adrenergic beta receptors are located on the heart cell surface membrane and can be identified directly by ligand-binding studies in intact cells. The maximal specific binding capacity of a labeled beta-adrenergic antagonist, [3H]-CGP, corresponds to about 6000 receptors per cell. Binding of adrenergic agonist results in changes in the membrane ion permeability for Ca which appears to involve an increase in the cAMP content of cardiac cells. Studies employing tracer methods and electrophysiological techniques suggest that the number of functional Ca channels in myocardium is regulated by beta-adrenergic receptors through the involvement of cAMP-mediated phosphorylation.

Adenylyl Cyclases↗
PubMed1982

Competitive and non-competitive interactions between specific ligands and beta-adrenoceptors in living cardiac cells.

We have used primary cultures of hearts from newborn rats to study beta-adrenoceptor properties in living myocardial cells. Receptors were labelled with the lipophilic antagonists 3H-(+/-)-carazolol and 125I-(+/-)-cyanopindolol (CYP) or with the hydrophilic antagonist 3H-(+/-)-CGP 12177. Under equilibrium conditions all ligands bound to a saturable homogeneous class of specific sites with a maximal binding capacity of approximately 100 fmol/mg protein (corresponding to approximately 5000 sites/cell). After 90-180 min preincubation of intact cells with 3H-carazolol or 3H-CGP 12177 only 80% of these antagonists could be displaced from specific binding sites by competing ligands. In the simultaneous presence of the antagonist (-) timolol 100% of specifically bound radiolabelled ligand remained displaceable. In competitive displacement experiments the radioligands did not affect the apparent affinity of the displacing nonlabelled antagonists timolol and CGP 12177, but agonist affinity was markedly changed. The apparent KD values for (-)-isoprenaline were 1560 and 2720 nmol/l in the presence of carazolol and CYP, but only 32 nmol/l in the presence of CGP 12177. This antagonist-dependent difference in agonist KD values was observed only in intact cells but not in membrane particles prepared from heart homogenates of newborn rats, where high agonist affinity was seen during displacement of all radioligands. The KA value for isoprenaline-stimulated cAMP accumulation in living cells was 30 nmol/l in 5-day cultures. A direct proportionality existed between agonist receptor occupation and cAMP accumulation in the presence of CGP 12177 as estimated by the KA/KD ratio. In the presence of carazolol the KA/KD ratio decreased from 1 to 0.02 suggesting that low affinity receptors were not coupled functionally to adenylate cyclase. These results indicate that some lipophilic antagonists which appear to be inert competitive ligands in fragmented membranes, alter receptor binding properties in intact cells. These antagonists seem to promote the transformation of receptor sites into a new "inactivated" state where competitive interactions between different ligands are inhibited.

Animals↗
PubMed1981

Quantitative changes in beta-adrenergic responses of isolated atria from hyper- and hypothyroid rats.

Concentration-response curves for the chronotropic and inotropic effects of isoprenaline, in the absence and presence of propranolol, were obtained on heart atria isolated from normo- or dysthyroid rats. Hyperthyroidism increased the chronotropic potency and efficacy of the beta-adrenergic agonist. The results are compatible with the view that thyroid hormone increases the density of functional beta-adrenoceptors in cardiac pacemaker tissue.

Animals↗
PubMed1981

Beta-adrenergic receptors and responses in the heart.

This article summarizes some essential steps leading to one of the most prominent beta-adrenergic effects on the heart, the increase in force of contraction. Binding of beta-adrenergic agonists and antagonists, activation of the enzyme adenylate cyclase by agonist binding, increase in membrane permeability to calcium ions, possibly via cyclic AMP, and increase in force of contraction and its relation to enhanced calcium influx are briefly discussed.

Action Potentials↗