Bradykinin action in the rat duodenum: receptor binding and influence on the cyclic AMP system.

Bradykinin binds to a single class of binding sites at rat duodenum plasma membranes. In the presence of endogenous calcium and at low bradykinin concentrations the receptor activation is followed by a stimulation of adenylate cyclase activity and the elevation of the cAMP level. In the absence of calcium and at high peptide concentrations the cAMP level is lowered via both inhibition of adenylate cyclase and stimulation of cAMP-phosphodiesterase. These different changes in the cAMP level might be correlated with the biphasic pharmacological action of bradykinin in the rat duodenum. The results suggest that one type of bradykinin (B2) receptor is able to initiate several effectuation mechanisms.

Opiate receptor binding affinities of some D-amino acid substituted beta-casomorphin analogs.

beta-Casomorphin-(5) and some analogs modified by the introduction of some D-amino acids and D-pipecolic acid as well as by C-terminal amidation were tested for their affinities to mu- and delta-binding sites in rat brain membranes. The binding affinities of these compounds are compared with the known activities in the guinea pig ileum (GPI) and mouse vas deferens (MVD) test and their antinociceptive potencies in rats. The substitution of D-proline for proline in position 4 in beta-casomorphin-(5) and beta-casomorphin-(4)amide (morphiceptin) results in derivatives with very high mu-binding affinity and mu-selectivity. These affinities correspond to the respective analgesic potencies. Both binding to mu-receptors and analgesic potency are also enhanced by the introduction of D-Phe in position 3. Testing D-Ala2 substituted derivatives with respect to their ability to compete for 3H-naloxone, we observed apparent differences between the pentapeptide amides (biphasic displacement curves) and the tetrapeptide amides (monophasic displacement curves). The substitution of L-Pro2 by D-pipecolic acid yields an analog with preferential delta-receptor affinity in the organ preparations (MVD) but preferential mu-receptor affinity in brain membranes. This finding suggests a possible difference between peripheral and central mu-binding sites.

[Synthesis and effect of affinity-labeled analogs and partial sequences of the bradykinin potentiating nonapeptide BPP9 alpha (teprotide)]. / Synthese und Wirkung affinitätsmarkierter Analoga und Partialsequenzen des Bradykinin potenzierenden Nonapeptides BPP9 alpha (Teprotide).

Affinity labeled analogues and partial sequences of the bradykinin potentiating nonapeptide BPP9 alpha inhibit the BPP9 alpha induced potentiation of the bradykinin action on the isolated guinea pig ileum. The labeled nonapeptides are more active than the labeled partial sequences. The inhibition of the potentiating action of BPP9 alpha demonstrates, that the influence on bradykinin action is not only a result of the inhibition of peptidyl dipeptide hydrolase.

[On the mode of action of bradykinin on smooth muscle (author's transl)]. / Zum Wirkungsmechanismus von Bradykinin an glatten Muskeln.

At extremely low concentrations, in the picomole and the nanomole range, bradykinin produces contraction and relaxation of smooth muscle in the gastrointestinal and the urogenital tract. At the target organ, bradykinin interacts with discriminator proteins of the plasma membranes and triggers, via changes in certain membrane functions, its biological response:--The binding to the discriminator makes specific conformative and constitutional demands on the nonapeptide. The binding results from an angular conformation which exists in the solution. The complete sequence is responsible for this specific conformation. Consequently, the biological activity of partial sequences is low. The conformational analysis of analogues used in studies on the mechanism of action showed but slight differences from bradykinin. The interaction of these analogues with the discriminator protein is disturbed to a varying extent by modifications at positions 1, 5, 8 and 9 in the side chains. The affinity for the discriminator is affected, dependently on the respective configuration, by substitution on the beta-C atom in the two phenylalanine residues.--Bradykinin is not only bound to, but also degraded at, the plasma membranes of the rat uterus and duodenum. The bradykinin-degrading enzyme has been characterized as a kininase II with the aid of various inhibitors. The conformative and configurative prerequisites decisive for enzymatic degradation are others than those decisive for binding to the discriminator.--The changes in the activities of the membrane-bound adenylate and guanylate cyclases (produced by the bradykinin-discriminator complex) that take place at the rat duodenum and uterus in the presence of extracellular calcium ions contrast with each other: At the duodenum, the ratio between these two cyclic nucleotides is changed in favour of adenylate cyclase; and at the uterus, in favour of guanylate cyclase; Substances which increase or decrease the cAMP level may also potentiate or inhibit the relaxation of the duodenum. These bradykinin-induced changes in enzyme activity must be considered in connection with other effectors, e.g. prostaglandins and calcium ions.--The calcium-ion-dependence of the effect of bradykinin on the guinea-pig ileum and the rat uterus indicates the importance of these ions as additional second messengers. Bradykinin stimulates the influx of calcium ions into the ileum; it is ineffective if no extracellular calcium ions into the ileum; it is ineffective if no extracellular calcium ions are available. It seems that intracellular and membranal calcium is mobilized in the uterus, which is evidenced by results from experiments with EGTA on the isolated organ and by the release of calcium from plasma membranes after application of bradykinin. It is assumed that the observed changes in membrane functions are induced by the peptide-discriminator complex simultaneously and not in the form of a causal chain.

Effect of bradykinin on prostaglandin synthetase activity in microsomal fractions from rat duodenum and uterus.

The bioconversion of tritiated arachidonic acid by microsomal fractions from rat uterus and duodenum is described. In rat duodenum the formation of both prostaglandin E2 and F2alpha is enhanced by the peptide hormone bradykinin. In contrast, bradykinin inhibits the synthesis of PGE2 in rat uterus.

Effects of prostaglandins, calcium, and bradykinin on guanylate cyclase in different organs.

The effects of prostaglandins and bradykinin on the activity of guanylate cyclase in plasma membranes from rat uterus and duodenum have been found strongly dependent on calcium content.

Bradykinin action in the rat duodenum through the cyclic AMP system.

Activators of the adenylate cyclase or inhibitors of the cAMP-phosphodiesterase, respectively, potentiate the bradykinin-induced relaxation of the rat duodenum, whereas imidazole as a stimulator of the cAMP-phosphodiesterase reduces the relaxation. The experiments indicate a linkage between the adenylate cyclase system with the biological action of bradykinin on the rat duodenum. In contrast, no similar effect has been observed on the rat uterus.

[The effect of potentiating factors on the bradykinine effect in vitro]. / Der Einfluss potenzierender Faktoren (BPF) auf die Bradykininwirkung in vitro

Bradykinin potentiating factors from the venom of Bothrops jajaraca and Agkistrodon halys blomhoffii potentiate the action of bradykinin at several smooth muscles. This potentiation is specific for bradykinin and has to be distinguished from an unspecific potentiation. The potentiation induced by BPF is not due to an indirect cholinergic mechanism or to a kininase inhibition in vitro. The results suggest that there would be an allosteric transition of the bradykinin receptor.

[Calcium-dependence of bradykinin-induced contraction in the isolated longitudinal muscle of the guinea pig ileum]. / Befunde zur Kalziumabhängigkeit der Bradykininwirkung am isolierten Längsmuskel des M-erschweinchenileums

The calcium-dependence of the bradykinin-induced contraction was demonstrated on the isolated logitudinal smooth muscle of the guinea-pig ileum. Using the "La-method" for measuring 45Ca entry into smooth muscle cells we were able to show, that bradykinin significantly increases the intracellular Ca-concentration. This effect of bradykinin was dose-dependent.

[Stability of some bradykinin analogues against kininase II (author's transl)]. / Stabilität verschiedener Bradykininanaloga gegen Kininase II

Some analogues of bradykinin, especially with replacements by other amino acids of phenylalanine in position 8, have been investigated for enzymatic stability against kininase II from rat duodenum microsomes and rat uterus plasma membranes, respectively. As compared with bradykinin, two of the analogues, [8-erythro-beta-phenylserine]- and [8-erythro-alpha-Amino-beta-phenylbutyric acid]-Bradykinin were stable to enzymatic degradation. Therefore, the latter may be used for studies in hormone-receptor interaction.

[Studies on bradykinin-binding cell fractions. 1. Characterization of kininase activity of the microsomal and membrane fraction from the rat uterus]. / Untersuchungen an bradykininbindenden Zellfraktionen 1. Charakterisierung der Kininaseaktivität der Mikrosomen- und Membrankraktion aus dem Rattenuterus

In the membrane fraction of rat uterus, kinin-destroying activity was found and characterized as kininase II by means of several kininase inhibitors. It could be shown that sudies of bradykinin-receptor interactions requires the use of kininase inhibitors like phenanthroline or the use of a stable bradykinin analogue like [8-erythro-alpha-amino-beta-phenylbutyric acid]-bradykinin.

[Characterization of the effect of bradykinin on the smooth muscle with special reference to its latency]. / Charakterisierung der Bradykininwirkung am glatten Muskel unter besonderer Berücksichtigung der Latenzzeit

The biological activity of bradykinin on the isolated guinea-pig ileum is evident after a defined period of time - its latency. The latency of a bradykinin induced contraction is not caused by the strong protonization of the polypeptide. Analogues with the same or diminished basicity have a shorter latency than bradykinin. The latency is increased at high hydrogen concentration in comparison with the physiological pH-value. But this phenomenon is also observed at angiotensin, eledoisin, acetylcholine, histamine and barium chloride. The latency is dependent upon the temperature. Exogenous calcium ions are without demonstrable influence on the bradykinin induced latency. The membrane potential is not changed during the latency. The results are discussed in connection with the drug-receptor interactions.