Henning Anhut

Radioimmunological determination of thromboxane release in cardiac anaphylaxis.

A sensitive and specific radioimmunoassay for thromboxane B2 were detected in perfusates of anaphylactic guinea pig hearts. Indomethacin decreased thromboxane B2 levels in the perfusates to below the detection limit of the radioimmunoassay and concomitantly delayed the onset of coronary vasoconstriction after antigenic challenge.

Regional distribution of arachidonic acid metabolites in rat brain following convulsive stimuli

Seizures were induced in female Wistar rats by electroconvulsive shock (ECS) or administration of pentetrazole (PTZ). Brain content of various prostanoids measured by radioimmunoassay showed time-dependent changes after the induction of convulsions; highest levels were found for PGD2 followed by PGF2 alpha, PGE2, TXB2 and 6-keto-PGF1 alpha. Analysis of the various arachidonic acid metabolites in seven parts of the rat brain dissected according to the method of Glowinski and Iversen revealed the largest increases in hippocampus and cerebral cortex and smaller ones also in hypothalamus and corpus striatum both after ECS and PTZ. The ratios of the different cyclo-oxygenase products remained virtually the same in whole brain as well as in those regions where the formation of prostaglandins was markedly elevated. 15-keto-13,14-dihydro-PGF2 alpha also increased simultaneously in parallel to its parent compound, PGF2 alpha and was detected in significant amounts only in hippocampus and cerebral cortex. However, concentrations of 15-keto-13,14-dihydro-PGF2 alpha in these brain regions as well as in whole brain represented only 3-10% of the amounts found for PGF2 alpha. Thus, the metabolizing enzymes 15-hydroxy-PG-dehydrogenase and delta 13-PG-reductase seem to be of minor importance for the inactivation of prostanoids in brain tissue.

Pharmacological modification of thromboxane and prostaglandin release in cardiac anaphylaxis.

Isolated perfused sensitized guinea pig hearts release relatively large amounts of radioimmunologically measurable thromboxane B2 (TXB2) as well as smaller amounts of prostaglandin (PGs) after antigenic challenge. Using thin layer chromatography the major PG released was shown to cochromatograph with PGD2, while smaller amounts of immunoreactive PGF2alpha were found. The TX-synthetase inhibitor imidazole (100 microgram/ml) significantly decreased TXB2 release and simultaneously increased PG release during cardiac anaphylaxis. On the other hand, the beta-sympathomimetic drug isoproterenol decreased both TXB2 and PG release from the anaphylactic hearts. While isoproterenol significantly diminished anaphylactic coronary flow reduction, imidazole was without effect in this respect. PGD2 (0.5 microgram/min and 5.0 microgram/min) infused intraaortally into non-sensitized guinea pig hearts reduced coronary flow dose-dependently. These results are compatible with the view that release of TX and PGs might contribute to coronary flow reduction in cardiac anaphylaxis.

Release of 15-keto-13, 14-dihydro-thromboxane B2 and prostaglandin D2 during anaphylaxis as measured by radioimmunoassay

Summary1.A specific radioimmunoassy for 15-keto-13, 14-dihydro-thromboxane B2 was developed employing a polyvalent 125I-labelled tracer.2.Using the radioimmunoassay large amounts of immunoreactive 15-keto-13, 14-dihydro-thromboxane B2 were found in perfusates of anaphylactic guinea pig lungs, but not in perfusates of anaphylactic guinea pig hearts. On the other hand, both lungs and hearts released immunoreactive thromboxane B2 after challenge, lungs being, however, much more active.3.Using a radioimmunoassay for prostaglandin D2 variable amounts of this compound were detected in perfusates of anaphylactic guinea pig lungs.4.While the cyclo-oxygenase inhibitor indomethacin (1 μg/ml) abolished release of thromboxane B2, 15-keto-13, 14-dihydro-thromboxane B2 and prostaglandins from anaphylactic guinea pig lungs, the thromboxane synthetase inhibitor imidazole (100 μg/ml) was without significant effect.5.The results indicate that in anaphylactic guinea pig lungs, contrary to anaphylactic guinea pig hearts, a major fraction of the total thromboxane released into the perfusates is in the form of the biologically inactive 15-keto-13, 14-dihydro-thromboxane B2. On the other hand, prostaglandin D2, which can be formed enzymatically or non-enzymatically from the prostaglandin endoperoxide PGH2, might contribute to vascular and bronchial smooth muscle contraction observed in anaphylaxis.

Cholecystokinin-Like Immunoreactivity of Rat Medial Basal Hypothalamus:Investigations on a Possible Hypophysiotropic Function

Studies on the cholecystokinin-like immunoreactivity (CCK-IR), of the medial basal hypothalamus (MBH) were performed in rats. In immunohistochemical studies, a dense CCK-IR-positive staining was found in the external layer of the median eminence adjacent to portal capillaries. Either high potassium (56 mM) or veratridine (1-50 micrograms/ml) stimulated the release of CCK-IR from MBH incubated in vitro. This increase in CCK-IR release depended on the presence of calcium. In vivo, the content of CCK-IR in the MBH was reduced after bilateral adrenalectomy and this effect of adrenalectomy was reversed by dexamethasone treatment. The content of CCK-IR in the neurointermediate lobe of the pituitary gland was not changed under these conditions. These results raise the possibility that after activation of the hypothalamo-pituitary-adrenal axis cholecystokinin may be secreted from nerve terminals in the external layer of the median eminence into the hypophyseal portal blood.

Radioimmunological determination of prostaglandin D2 synthesis in human thrombocytes

A specific radioimmunoassay for prostaglandin D2 was developed. Using the radioimmunoassay, prostaglandin D2 synthesis by human thrombocytes was measured. While the cyclooxygenase inhibitor indomethacin inhibits formation of prostaglandin D2, increased formation of prostaglandin D2 was observed in the presence of the thromboxane synthetase inhibitor imidazole.

Vasopressin and β-endorphin release after osmotic and non-osmotic stimuli: effect of naloxone and dexamethasone

The purpose of this study was to determine whether or not vasopressin release in response to various stimuli in the conscious rat is controlled by endogenous opioid peptides, in particular beta-endorphin. Naloxone (1 mg.kg-1 i.m.) promoted vasopressin release in response to both an angiotensin II infusion (500 ng . kg-1 . min-1) or an isosmolar, nonhypotensive hypovolaemia achieved by polyethylene glycol injection (PEG, 20% solution i.p.); however, naloxone was without effect when vasopressin release was induced by hypertonic saline injection (2.5% solution i.p.) or a severe fall in arterial blood pressure following trimethidinium (10 mg . kg-1 i.m.) induced ganglionic blockade. Vasopressin release was accompanied by an increase in plasma beta-endorphin-like immunoreactivity (beta-EI) following an angiotensin II infusion of PEG administration, but not after hypertonic saline or trimethidinium injection. Dexamethasone pretreatment (0.5 mg . kg-1 twice i.p.) prevented the increase in plasma beta-EI following an angiotensin II infusion or PEG administration. The simultaneous angiotensin II- or PEG-induced increase in vasopressin release was unaffected or potentiated, respectively, by the glucocorticoid. In contrast, vasopressin release in response to hypertonic saline or trimethidinium injection was significantly inhibited by dexamethasone. We conclude that an inhibitory control by endogenous opiates is involved in some, but not all of the different pathways leading to vasopressin release. The results obtained do not prove but can be reconciled with the proposal that hypophyseal beta-endorphin is the compound responsible.

Vasopressin stimulates release of ?-lipotropin and ?-endorphin in conscious rats as measured by radioimmunoassay of unextracted plasma

SummaryUsing a newly developed radioimmunoassay to determine the β-endorphin-like immunoreactivity (β-EI) in unextracted plasma, the effect of vasopressin injections on plasma β-EI was investigated in conscious rats. Arginine vasopressin caused a dose-dependent increase of plasma β-EI from 34.5±7.8 fmol ml−1 (n=6) in vehicle-treated animals to 205.0±36.1 fmol ml−1 (n=7) after injection of the highest vasopressin dose employed (486 ng/100 g b.w.). In view of the appreciable cross-reactivity of β-lipotropin (β-LPH) in the radioimmunoassay used, plasma was extracted and subjected to gel chromatography on a Sephadex G-50 column. On average, about 70% of the β-EI co-eluted with human β-LPH and about 30% with human β-endorphin in plasma extracts obtained from both control and vasopressin-treated rats. No peripheral conversion of human β-LPH occurred under the experimental conditions, since after i.v. bolus injection of human β-LPH 97% of the β-EI comigrated with human β-LPH during gel filtration. A similar blood pressure increase to that induced by the vasopressin injections, when elicited by noradrenaline or angiotensin II i.v., was not followed by an elevation of plasma β-EI.These data indicate that vasopressin stimulates β-lipotropin and β-endorphin release into the systemic circulation in vivo.

β-endorphin release by angiotensin II: studies on the mechanism of action

Blood-borne angiotensin II induces release of beta-endorphin-like immunoreactivity (beta-EI) from rat anterior pituitary gland. To study the mechanism of action we investigated in rats the effect of transection of subfornical organ efferent projections on angiotensin-induced beta-EI release in vivo and also the direct action of angiotensin II on beta-EI release from isolated adenohypophyses in vitro. (i) No effect of transection of subfornical organ efferents on the increase in plasma beta-EI following intravenous infusions of angiotensin II was found. (ii) When anterior pituitary quarters were continuously superfused in vitro, angiotensin II (1-10 nM) caused release of beta-EI into the superfusion medium in a dose-dependent manner. The stimulatory effect of angiotensin II (3 nM) was blocked by the receptor antagonist saralasin (300 nM). We conclude that beta-endorphin release by blood-borne angiotensin II, in contrast to other central effects of angiotensin, is not mediated by the subfornical organ; instead a direct action of angiotensin II on the adenohypophysis could be a mechanism of action responsible.

Formation and elimination of prostacyclin metabolites in the cat in vivo as determined by radioimmunoassay of unextracted plasma.

Using newly develop radioimmunoassay for 6-keto-prostaglandin F1 alpha and 6,15-diketo-13,14-dihydro-prostaglandin F1 alpha, the plasma concentrations of these two prostacyclin derivatives were measured in anaesthetized cats. After the administration of angiotensin II, which releases prostacyclin into the circulation, concentrations of both derivatives rose simultaneously, the major immunoreactivity being 6,15-diketo-13,14-dihydro-prostaglandin F1 alpha. Angiotensin II-induced prostacyclin release was not caused by vasoconstriction alone, since comparable vasopressor responses to noradrenaline and vasopressin were not accompanied by increases in prostacyclin plasma levels. Injection of exogenous prostacyclin resulted in a shortlasting peak of 6-keto-prostaglandin F 1 alpha, which rapidly declined (t 1/2: 1.29-1.52 min). 6,15-diketo-13,14-dihydro-prostaglandin F1 alpha appeared with an t 1/2 of 0.48-1.38 min and was eliminated with a t 1/2 of 8.0-9.0 min. Due to its longer half-life in the circulation 6,15-diketo-13,14-dihydro-prostaglandin F 1 alpha again was the predominant derivative after 3 min. These data suggest that in vivo prostacyclin is mainly inactivated by the 15-hydroxy-PG-dehydrogenase-, delta 13-reductase-pathway, rather than by hydrolysis. Therefore, 6,15-diketo-13,14-dihydro-prostaglandin F1 alpha seems to be a better indicator of prostacyclin plasma levels than 6-keto-prostaglandin F1 alpha, although under certain conditions the additional determinations of this product of hydrolysis can be valuable.