Two distinct adrenoceptor-biophases in the vasculature: One for α-and the other for β-agonists

Summary Strips of two canine vessels with different patterns of sympathetic innervation were used: the mesenteric artery which has an adventitio-medial plexus and the saphenous vein in which nerve terminals are distributed throughout the media.The pD₂ of (–)-adrenaline for -and -adrenoceptors was determined for both vessels (in the presence of 0.5 M propranolol and 7 M phentolamine, respectively; in the latter case the strips were contracted by 0.28 M prostaglandin F2) and found to be very similar: 6.96 and 7.01 in the saphenous vein and 6.77 and 6.91 in the mesenteric artery, respectively. The similarity of pD₂ for adrenaline acting on -and -adrenoceptors in both preparations allowed us to compare the effect of inhibition of neuronal uptake by 12 M cocaine with that of inhibition of COMT by 50 M dihydroxy-2-methyl propiophenone (U-0521) on: a) the potency of adrenaline (noradrenaline was used in the saphenous vein only) acting on -and -adrenoceptors and b) the time required by the strips to recover 50% in oil (t ₅₀) after contractions or relaxations caused by 0.23 M adrenaline. In the saphenous vein cocaine increased the potency of both adrenaline and noradrenaline for -effects more than for -effects (2.8 times vs. no increase for adrenaline; 7.1 vs. 1.9 times for noradrenaline) and U-0521 increased the potency of both adrenaline and noradrenaline for -effects more than for -effects (4.1 vs. 2.6 times for adrenaline; 1.8 times vs. no increase for noradrenaline); regarding the termination of action of adrenaline, cocaine prolonged the t ₅₀ 1.6 times after contraction and did not change it after relaxation, whereas U-0521 prolonged the t ₅₀ 6.9 times after relaxation and only 1.8 times after contraction.In the mesenteric artery only sensitivity experiments were done. Cocaine increased the potency of adrenaline by a factor of 2.1 for the -effects while there was no influence on its potency with regard to its -effects, and U-0521 increased the potency of adrenaline for the -effects more than for -effects (3.9 vs. 1.8 times, respectively).These results show that there are two different biophases for sympathomimetic agonists in the vasculature: one for -adrenoceptors which is more under the influence of the neuronal uptake and one for -adrenoceptors which is more under the influence of COMT activity. We conclude that -adrenoceptors are situated close to the nerve endings and -adrenoceptors close to COMT sites. Since these results do not differ qualitatively in two vessels with different patterns of innervation, we conclude that this asymmetry in the distribution of -and -adrenoceptors may be due to either an uneven distribution of cells with only one type of receptors each or due to an uneven distribution of receptors on the same cell.     

The effect of phenytoin and metamphetamine on spinal motor activity

Summary The investigation was initiated by the idea that reserpine might exert its action on spinal motor activity through facilitatory impulses in descending pathways to the spinal motoneurones. Therefore, the effect of conditioning by repetitive stimulation of descending and segmental pathways on and reflex discharges was studied in the rat. Conditioning by stimulation of the dorsolateral funiculus increased a and decreased reflex discharge. Conditioning by stimulation of the dorsal funiculus increased reflex discharge without changing reflex activity. In partially or totally spinalized preparations, conditioning by repetitive stimulation of the same dorsal root to which the test stimulus was applied, produced an increase in a reflex activity which was associated with a decrease in motor activity. In the preparations with an intact neuraxis, repetitive stimulation of the dorsal root facilitating reflex discharge did not change motor activity. motor activity also remained unchanged when reflex discharge was inhibited by repetitive stimulation of the dorsal root in the intact or totally spinalized preparation.Phenytoin 50 mg/kg increased the unconditioned response in all types of preparations, apart from that in which the dorsal funiculus was isolated. Facilitation of reflex discharge elicited by repetitive stimulation of segmental and descending pathways was depressed by phenytoin, whereas inhibition remained unchanged.Metamphetamine 2 mg/kg increased the unconditioned reflex discharge in all types of preparations, and decreased the response in the preparations with an intact neuraxis and also in those in which the dorsal funiculus was isolated. Metamphetamine did not depress the facilitatory or inhibitory effects produced by repetitive stimulation of descending or segmental pathways.From the results obtained it is concluded that reserpine rigidity in the rat may be mediated by an increased facilitatory impulse input reaching the a motoneurones by way of descending pathways in the dorsolateral column of the spinal cord, and that phenytoin may antagonize, at least in part, the effect of reserpine by a depressant action on facilitatory processes at the spinal level, whereas abolition of reserpine rigidity produced by metamphetamine must be due to a supraspinal site of action of this drug.Supported by a grant of the Deutsche Forschungsgemeinschaft.     

α-Adrenoceptor activity of arylalkylimidazoles is improved byα-methylation and impaired byα-hydroxylation

Summary To investigate the effects of hydroxyl and methyl substitution of the alkyl bridge bond on the-adrenoceptor activity of arylalkylimidazole derivatives, the cardiovascular effects of the molecules were studied in anaesthetized and pithed rats. The compounds studied were 4(5)-substituted imidazole derivatives with a methano, ethano or etheno bridge between the imidazole and the 2-, 2,3- or 2,6-methyl substituted phenyl rings. The hypotensive and bradycardic activities of the molecules in the anaesthetized rat were always reduced by-hydroxylation and usually augmented by-methylation of the bridge between the imidazole and phenyl rings. Hydroxylation was associated with a consistent, marked decrease in vasopressor and sympatho-inhibitory activity in the cardiovascular system of the pithed rat, but a methyl moiety as a bulky substituent in the-position of the alkyl bridge did not decrease but even caused an increase in-adrenoceptor activity in this test system. The detrimental effect of-hydroxylation of the compounds at ₁- and ₂-adrenoceptors supports the notion that the interaction of the imidazoles at-adrenoceptor is different from that of the classical, noradrenaline-like phenethylamines. The results also suggest that the alkyl bridge between the phenyl and imidazole rings of the imidazoles may contribute directly to the binding process.     

Different effects of vinblastine on the polymerization of isotypically purified tubulins from bovine brain

Vinblastine, a highly successful antitumor drug, targets the tubulin molecule. Tubulin, the subunit protein of microtubules, consists of an - and a -subunit, both of which consist of isotypes encoded by different genes. We have purified three isotypes of bovine brain tubulin, namely, _II, _III and _IV. Microtubule associated protein-2 (MAP2) and Tau-induced assembly of these isotypes were compared in the presence and absence of vinblastine. MAP2-induced assembly of unfractionated tubulin and all the isotypes except _II tubulin was resistant to 1M vinblastine. Vinblastine at low concentrations (<10M) progressively inhibited the assembly of all of the isotypes but the vinblastine concentration required for inhibition of MAP2-induced microtubule assembly was minimal for _II. The tau-induced assembly of unfractionated tubulin and _III were equally sensitive to 1M vinblastine whereas _II and _IV were much more sensitive to vinblastine. The microtubules obtained in the presence of tau from unfractionated tubulin, _II and _IV could be easily aggregated by 20M vinblastine whereas such as aggregation of microtubules obtained from _III and tau required approximatedly 40M vinblastine. Our results suggest that among the tubulin isotypes, _II is the most sensitive to vinblastine in the presence of MAPs while _III is the most resistant and this intrinsic resistance of _III dimers persists in the polymeric form of _III tubulin as well. These results may be relevant to the therapeutic and toxic actions of vinblastine.     

Isoprenaline-induced increase in mRNA levels of inhibitory G-protein α-subunits in rat heart

Summary Long-term -adrenergic stimulation has been shown to desensitize the -adrenoceptor/adenylyl cyclase signalling pathway at both the receptor and the G-protein level. To further elucidate the cellular mechanism of G-protein regulation we investigated the influence of prolonged infusion of isoprenaline (2.4 mg/kg·d) on myocardial mRNA levels of different G-protein -subunits in rats. For comparison rats were treated with triiodothyronine (T₃; 0.5 mg/kg·d) which induces cardiac hypertrophy like isoprenaline but has different effects on the adenylyl cyclase system. Isoprenaline- and T₃-treated animals developed an increase in heart/body weight ratio of 41±3% and 27±4%, respectively (P<0.05). Isoprenaline increased myocardial total RNA concentration by 39±6% (P<0.05). Hybridization with ³²P-labeled rat cDNAs demonstrated an expression rank order of G_s-mRNA>G_i-2-mRNA>G_i–3-mRNA and no detectable expression of G_i–1-mRNA in rat myocardium. mRNA levels of G_s G_i–2 and G_i–3 were 36.9±1.28, 10.7±1.07 and 3.7±0.19 pg/g total RNA, respectively. Isoprenaline increased G_i–2 ^– and G_i–3-mRNA concentrations per g total RNA by 49±18% and 27±710, respectively (P<0.05). This effect was abolished by simultaneously administered propranolol (9.9 mg/kg·d), indicating a,-adrenoceptor-mediated mechanism. In contrast, T₃-induced cardiac hypertrophy was not accompanied by changes in G_i-mRNA expression. G_sa-mRNA levels were unaffected by either treatment.In conclusion, long-term stimulation with isoprenaline in vivo induces a -adrenoceptor-mediated increase in myocardial G_i–2 ^– and G_i–3-mRNA without affecting G_s-mRNA. These results suggest that similar increases in myocardial G_i–2-mRNA in end-stage human heart failure may be at least partly explained by increased -adrenergic stimulation due to increased sympathetic activity.Parts of this work were presented at the wintermeeting of the Deutsche Gesellschaft fur Pharmakologie und Toxikologie in Hannover, 1990 (Eschenhagen et al.), Naunyn-Schmiedebergs Arch Pharmacol 342 (Suppl):R8. The work was supported by the Deutsche Forschungsgemcinschaft Send offprint requests to: T. Eschenhagen at the above address     

Effect of ethanol on the metabolism of lithocholic acid in rat liver

Summary Lithocholic acid 24-C¹⁴ is converted by 20000x g-supernatant of rat liver homogenate into several products of higher polarity. 3-6-dihydroxy-5-cholanic acid is the main metabolite, whereas 7-hydroxylation occurs only to a small extent. Besides of the hydroxylations conjugation with taurine and the formation of a 3-sulfate ester of lithocholic acid can be demonstrated. Addition of ethanol to the enzymatic system results in an inhibition of the formation of 3,6-dihydroxy-5-cholanic acid, whereas no effect can be observed with respect to the formation of the other products. This inhibition is present also in 20000x g-supernatant obtained from rats pretreated with ethanol 100 min before being preparation from ethanol-pretreated rats amounts to 68% of the controls. The chrome P-450 mediated oxidative mechanism, which has been shown to be involved in microsomal 6-hydroxylation of bile acids.This work was supported by the Deutsche Forschungsgemeinschaft.     

Stimulation of DNA repair synthesis of rat thymocytes by novobiocin and nalidixic acid in vitro without detectable DNA damage

Scheduled (SDS) and unscheduled (UDS) DNA synthesis as well as nucleoid sedimentation was investigated in vitro under the influence of novobiocin (NB) and nalidixic acid (NA) using intact thymic (T-cells) and splenic (S-cells) rat cells and cells which were exposed to X-rays, UV irradiation, methyl methanesulfonate (MMS), and DNA polymerase inhibitors. At concentrations of 56.25 (S-cells) and 225 g/ml (T-cells), respectively, NB inhibited SDS in a dose-dependent manner. Within a concentration range of 225–900 g NB/ml, UDS of S-cells decreased to values far below the tracer ([³H-methyl]-thymidine) incorporation of control cells, whereas UDS of T-cells increased by at least 200%. Within a concentration range of 450–1800 g/ml, NA enhanced SDS and UDS by about 30% in S-cells and by 100% in T-cells. The stimulating activity of NB and/or NA could be eliminated specifically by the DNA polymerase inhibitor 2',3'-dideoxythymidine. Enhanced nucleoid sedimentation was observed at NB concentrations 750 g/ml; S-cells revealed a higher sedimentation rate than T-cells. It is suggested that NB (and NA) influence DNA topology in a rather cell specific manner, stimulating UDS of T-cells by a DNA polymerase — dependent repair-like mechanism.     

Über den nachweis und die bedeutung von α-methylnoradrenalin im harn von hypertonikern bei verabreichung von α-methyldopa

Zusammenfassung An 13 Hypertonikern wurde die Harnausscheidung von -Methyldopa, -Methyldopamin und Katecholammen während einer Dauerbehandlung oder nach einer Einzeldosis von -Methyldopa untersucht. Die im Harn enthaltenen Substanzen werden durch Ionenaustausch- und Papierchromatographie voneinander getrennt und fluorometrisch oder biologisch bestimmt. — Von alien Patienten wurde außer freiem und konjugiertem -Methyldopa und -Methyldopamin auch -Methylnoradrenalin ausgeschieden. Gleichzeitig nahm die Ausscheidung von Noradrenalin gegenüber der Vor-und Nachperiode bei fast allen Patienten ab. Es kann ausgeschlossen werden, daß die bei der Behandlung durch die Nieren ausgeschiedenen Mengen von -Methyldopa, -Methyldopamin oder -Methylnoradrenalin die renale Ausscheidung von Noradrenalin herabsetzten. Die während der Behandlung ausgeschiedene Menge von Noradrenalin, Adrenalin und -Methylnoradrenalin zusammengenommen unterschied sich nicht von der Katecholamin-Ausscheidung vor und nach der Behandlung. — Das biosynthetisch vom Menschen gebildete -Methylnoradrenalin verhielt sich nach dem Ergebnis chemischer und pharmakologischer Tests wie Corbadrin, welches die Erythro-Konfiguration aufweist; -Methylnoradrenalin ließ sich eindeutig von dem Diastereomeren von Corbadrin differenzieren, welches die Threo-Konfiguration besitzt. Im Harn der Patienten konnte -Methyladrenalin nicht nachgewiesen werden. Gegen eine Beteiligung von -Methyladrenalin, welches nach Tierversuchen durch Erregung von adrenergen -Receptoren blutdrucksenkend wirkt, an der hypotensiven Wirkung von -Methyldopa spricht ferner, daß die Blutdruckabnahme nach -Methyldopa durch Propranolol nicht aufgehoben, sondern eher sogar verstärkt wurde. — Die systolische Blutdrucksenkung war sowohl der Abnahme der renalen Ausscheidung von Noradrenalin als auch dem Prozentsatz von -Methylnoradrenalin an der gesamten Katecholamin-Ausscheidung significant korreliert; auch im Zeitverlauf ergab sich bei drei Patienten nach einer Einzeldosis von -Methyldopa eine Korrelation zwischen hypotensivem Effekt einerseits und Abnahme der Noradrenalin- bzw. Zunahme der -Methylnoradrenalin-Ausscheidung andererseits. — Die Ergebnisse machen es wahrscheinlich, daß der früher in Tierversuchen nach Verabreichung von -Methyldopa nachgewiesene teilweise Ersatz des sympathischen Überträgerstoffs Noradrenalin durch -Methylnoradrenalin auch unter therapeutischer Dosierung am Menschen stattfindet. Die Korrelationen von hypotensiven Effekten und Veränderungen im Aminstoffwechsel lassen den SchluB zu, daß -Methyldopa den Blutdruek von Hypertonikern durch Freisetzung von -Methylnoradrenalin als falscher Überträgersubstanz und entsprechende Verminderung der Noradrenalin-Freisetzung aus sympathischen Nervenfasern herabsetzt.

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Detection and significance of -Methylnoradrenaline in the urine of hypertensive patients after administration of -Methyldopa

Summary In 13 hypertensive patients the urinary excretion of -methyldopa, -methyldopamine und catechol amines was analyzed either during continuous administration or after a single dose of -methyldopa. The compounds contained in the urine samples were separated by ion exchange and paper chromatography and determined fluorimetrically and biologically. — All patients excreted both free and conjugated -methyldopa, -methyldopamine, and small amounts of -methylnoradrenaline. Compared with the pre- and postdrug period, in nearly all patients the noradrenaline excretion was decreased while -methyldopa was administered. The experimental data obtained exclude the possibility that the noradrenaline excretion was diminished by the simultaneous excretion of -methylnoradrenaline or of excessive amounts of -methyldopa and -methyldopamine. The combined quantities of noradrenaline, adrenaline and -methylnoradrenaline excreted daily during administration of -methyldopa did not differ significantly from the amount of noradrenaline and adrenaline excreted daily before drug treatment was started and after it was discontinued. —Evidence was obtained that -methylnoradrenaline isolated from urine of patients given -methyldopa behaved chemically and biologically in a way similar to (–)-corbadrine (erythro-configuration) but differed markedly from the diastereomer of corbadrine (threo-configuration). Hence, biosynthesis of -methylnoradrenaline leads to the levorotatory erythro-isomer. — In animal experiments carried out previously -methyladrenaline was found as another metabolite of -methyldopa which decreased blood pressure by activation of adrenergic -receptors. The patients given -methyldopa did not excrete -methyladrenaline. The method employed was sensitive enough to detect amounts of -methyladrenaline less than 3 per cent of the -methylnoradrenaline present. Furthermore, involvement of a -adrenergic component in the response of the blood pressure to -methyldopa in hypertensive patients was made unlikely by the observation that propranolol did not antagonize but rather enhanced the hypotensive effect of -methyldopa. — There was a significant correlation between the fall of systolic blood pressure and both the decrease in urinary excretion of noradrenaline and the increase in the percentage of -methylnoradrenaline of the total catecholamine output. Likewise, the time course of the depressor response to a single dose of -methyldopa closely corresponded to the decrease in noradrenaline and the increase in -methylnoradrenaline excretion. Conversely, the return of the blood pressure to the initial level was reflected by an increase in noradrenaline and a decrease in -methylnoradrenaline excretion. — In animal experiments it was previously found that administration of -methyldopa caused a partial displacement of noradrenaline by -methylnoradrenaline which subsequently was released by sympathetic nerve stimulation. The present findings demonstrate that -methylnoradrenaline is formed in man as well. It is concluded that -methyldopa lowers the blood pressure of hypertensive patients by the release of -methylnoradrenaline as a false neurotransmitter and the concomitant decrease in noradrenaline liberation from sympathetic nerve fibres.

     

Protein Aggregates Seem to Play a Key Role Among the Parameters Influencing the Antigenicity of Interferon Alpha (IFN-α) in Normal and Transgenic Mice

Purpose. During long-term treatment of various malignant or viral diseases with IFN- up to 20% of patients develop anti-IFN- antibodies for as yet unknown reasons. Methods. To address this issue, a mouse model using Balb/C mice was established and the relevance of several potentially anti-IFN- antibodies inducing factors was studied. Results. The model revealed that both a higher frequency of injections and a higher dosage of IFN- were more immunogenic and that the route of administration affected the antibody response to IFN-. The intrinsic immunostimulatory activity of IFN- itself also enhanced the immune response. IFN- protein aggregates (IFN--IFN- and human serum albumin (HSA)-IFN- aggregates), which were recently identified in all marketed IFN- products, were significantly more immunogenic than IFN- monomers. These aggregates broke the tolerance against human IFN- monomers in human IFN- transgenic mice. Conclusions. Based on these animal studies it is proposed that the immune response to IFN- in humans is most probably elicited by a combination of several factors among which IFN- protein aggregates seem to play a key role.     

Binding of aprindine and moxaprindine to human serum, α1-acid glycoprotein and serum of healthy and diseased humans

Summary A comparison was made between the binding of the anti-arrhythmic agents aprindine and moxaprindine to human serum, to human serum albumin (HSA), to ₁-acid glycoprotein (₁-AGP) and to a mixture of HSA and ₁-AGP. In serum from healthy volunteers (n=4) the binding of aprindine-HCl 5 µg/ml (13.8 µM) was 93.8% (SD±1.0), and that of moxaprindine-HCl 5 µg/ml (12.8 µM) was 94.1% (SD±1.1). Their binding to the mixture of ₁-AGP and albumin approximated their binding to serum. For ₁-AGP, the binding was similar for both compounds, whereas for HSA the binding of aprindine was more pronounced than that of moxaprindine: for both products the affinity coefficient for binding to ₁-AGP was about 100 times greater than that for binding to albumin. In serum from rheumatoid patients and from patients with renal failure a small but significant increase in binding of aprindine and moxaprindine was observed, approximately 1%. Increased and decreased binding was seen in serum from cirrhotic patients; for example, for aprindine the range in cirrhosis was 96.7%–79.8%, and the range in controls was 95.0%–92.4%. Free drug fraction and ₁-AGP concentration were inversely correlated. The results show that ₁-AGP plays an important role in the binding of aprindine and moxaprindine, and that alteration in the binding of the two compounds in disease states to a large extent can be explained by changes in serum ₁-AGP concentration.     

Studies on the chemical constituents from the roots of <Emphasis Type="Italic">Platycodon grandiflorum</Emphasis>

Three known monodesmosidic saponins: 3-O--d-glucopyranosyl-2,3,16,23,24-pentahydroxyolean-12-ene-28-oic acid, 3-O--d-glucopyranosyl polygalacic acid, and 3-O--d-glucopyranosyl-(13)--d-glucopyranosyl polygalacic acid; and two known nonsaponin compounds: a mixed compound of n-tetracosanoic acid (lignoceric acid), n-hexacosanoic acid (cerotic acid), and n-octacosanoic acid, and -monopalmitin; were isolated for the first time from the root of Platycodon grandiflorum A. DC. together with another seven known compounds: platycoside G₁ (3-O--d-glucopyranosyl-(16)--d-glucopyranosyl-(16)--d-glucopyranosyl-2,3,16,23,24-pentahydroxyolean-12-ene-28-oic acid 28-O--d-xylopyranosyl-(14)--l-rhamnopyranosyl-(12)--l-arabinopyranoside), deapio-platycodin D, Polygalacin D, deapio-platycodin D₃, platycoside A, -spinasterol, and -spinasteryl-3-O--d-glucopyranoside. Alkaline hydrolysis of platycoside G₁ afforded a new monodesmosidic prosaponin: 3-O--d-glucopyranosyl-(16)--d-glucopyranosyl-(16)--d-glucopyranosyl-2,3,16,23,24-pentahydroxyolean-12-ene-28-oic acid. Their chemical structures were elucidated on the basis of their spectral data and chemical evidence.     

Possible subdivision of postsynaptic α-adrenoceptors mediating pressor responses in the pithed rat

Summary Additional evidence has been obtained indicating a possible subclassification of postsynaptic -adrenoceptors into ₁ and ₂-subtypes. The pressor responses to the -adrenoceptor agonists L-phenylephrine and guanfacine were quantified after i.v. administration to pithed rats. The -sympatholytic drug yohimbine (1 mg/kg) displaced both dose-response curves to the right, but the effect was greatest for guanfacine. After prazosin (0.1 mg/kg) a 53-fold shift to the right was noticed for the dose-response characteristic of L-phenylephrine. Prazosin antagonized the effect of only the higher doses of guanfacine. The findings indicate that L-phenylephrine and prazosin preferentially interact with ₁-adrenoceptors as agonist and antagonist, respectively. Yohimbine proved less selective than prazosin, but preferentially blocks postjunctional ₂-adrenoceptors in the vascular wall. The results obtained with guanfacine may be interpreted to indicate that this drug acts on ₂-adrenoceptors at lower doses and additionally stimulates ₁-adrenoceptors at higher ones. Preliminary findings with corynanthine and rauwolscine support this interpretation.     

Frequency- and train length-dependent variation in the roles of postjunctional α1- and α2-adrenoceptors for the field stimulation-induced neurogenic contraction of rat tail artery

Summary The present paper examines the roles of postjunctional ₁- and ₂-adrenoceptors for the noradrenaline (NA)-induced neurogenic contractile response to field stimulation mainly with 1–100 pulses at 2 or 20 Hz, in the tail artery of adult normotensive rats. Pharmacological tools were employed to isolate and characterize the ₁- and ₂-adrenoceptor-mediated components of this response. The degree to which the drugs influenced NA release or reuptake was assessed by their effects on the electrochemically determined, stimulation-induced rise in the NA concentration at the innervated outer surface of the media. This response was unaffected by ,-methylene ATP (10 M) or suramin (500 M), added to desensitize or block P₂-purinoceptors, respectively prazosin (0.1 M) or SK&amp;F 104078 (6-chloro-9-[(3-methyl-2-butenyl)oxyl]-3-methyl-1H-2, 3, 4, 5-tetrohydro-3-benzazepine, 0.1 M), used to block postjunctional ₁- and ₂-adrenoceptors respectively, nifedipine (10 M), blocker of Ca²⁺ influx through L-type channels, and ryanodine (10 M), which blocks mobilization of Ca²⁺ from intracellular stores; it was moderately enhanced by yohimbine (0.1 M), blocker of pre- and postjunctional ₂-adrenoceptors, and strongly enhanced by cocaine (3 M) or desipramine (1 M), blockers of NA reuptake. Judging from their inhibitory effects on the contractile responses to the ₁- and ₂-adrenoceptor agonists, phenylephrine andxylazine, prazosin (0.1 M)and SK & F 104078 (0.1 M) could be used to selectively block ₁- and ₂-adrenoceptors respectively, while yohimbine (0.1 M) was less selective, strongly depressing ₂- and slightly depressing ₁-adrenoceptor-mediated responses. The ₁-adrenoceptor-mediated component of the contractile response to short trains at 20 Hz was fast in onset, brief in duration and abolished by ryanodine; that mediated by ₂-adrenoceptors was more delayed, prolonged and insensitive to ryanodine. Both components were dose-dependently depressed by nifedipine (0.1–10 M). The small contractile responses to single pulses, or up to 50 pulses at 2 Hz, or short train (< 4 pulses) at 20 Hz, were more markedly depressed by 0.1 M yohimbine or SK & F 104078 than by 0.1 M prazosin and, hence, mediated mainly by ₂-adrenoceptors. The reverse was true of the much larger response to longer trains at 20 Hz, which thus probably was mediated mainly by ₁-adrenoceptors. Cocaine or desipramine, as well as ,-methylene ATP or suramin, amplified both components of the NA induced contractile response especially that mediated via a₁-adrenoceptors and caused by single pulses or short trains.The main conclusions are (i) that the small NA-induced contractile responses of this artery to single pulses, or pulses at low frequency, or in short trains at high frequency, are mediated mainly via ₂-, and the larger responses to longer trains at high frequency increasingly via ₁-adrenoceptors, (ii) that the ₁- and ₂-adrenoceptor-mediated components interact cooperatively, probably at least in part by utilizing two different pathways to increase the intracellular Ca²⁺, (iii) that neuronal reuptake of NA strongly restricts both components of the NA-induced contraction, especially the ₁-adrenoceptor-mediated response to single pulses or short trains, and (iv) that both components of the NA-induced contraction, especially that mediated by ₁-adrenoceptors, may be depressed by ATP released by field stimulation and acting via P_2x-purinoceptors on smooth muscle. Based on these results a novel working hypothesis is proposed, in which it is assumed that the geometry of NA-mediated neuromuscular transmission in this vessel varies with the frequency and number of impulses in a stimulus train.Correspondence to J.-X. Bao at the above address     

Inhibitory influences of the adrenal steroid, 3α, 5α-tetrahydroxycorticosterone on aggression and defeat-induced analgesia in mice

The effects of intraperitoneal administrations of the deoxycorticosterone metabolite, 5-pregnane-, 21 diol-20-one (3, 5-tetrahydrodeoxycorticosterone; -THDOC) on the responses to aggression and defeat-induced analgesia were examined in subordinate intruder male mice in resident-intruder pairings. -THDOC reduced in a dose-dependent mannter (1–20 mg/kg) the number of bites and time to obtain defeat in subordinate mice during the agonistic encounters, as well as attenuating defeat-induced analgesia. These inhibitory effects of -THDOC were separate from its sedative actions at 20–30 mg/kg. In addition, the stereo-isomer, 3-pregnane-3, 21 diol-20-one (20 mg/kg) had no significant effects on the agonistic encounters and defeat, indicating that the inhibitory effects of -THDOC on agonistic interactions are stereospecific. Pretreatment with the benzodiazepine antagonist Ro 15-1788 (5 and 10 mg/kg) attenuated the inhibitory effects of -THDOC on defeat-induced analgesia. Ro 15-1788 (5, 10 mg/kg) by itself, however, had minimal effects on these agonistic interactions and subsequent defeat-induced analgesia. These results indicate that the naturally occurring steroid, -THDOC, has significant effects on responses to aggression and defeat-induced analagesia.     

Über die Wirkung von α-Methyl-Dopa auf den Brenzcatechinamingehalt von Meerschweinchenorganen

Zusammenfassung 1. Versuche über die Wirkung von -Methyl-Dopa auf den Brenzcatechinamingehalt von Meerschweinchenorganen haben ergeben, daß Noradrenalin im Herzen, sowie Dopamin und Noradrenalin im Gehirn durch äquimolare Mengen von -Methyl-Noradrenalin bzw. -Methyl-Dopamin ersetzt werden.2. Die Adrenalinverarmung der Nebennieren nach Vorbehandlung mit -Methyl-Dopa wird demgegenüber nur zu einem geringen Teil durch die methylierten Brenzcatechinamine -Methyl-Noradrenalin und -Methyl-Dopamin ausgeglichen. Ein weiteres Brenzcatechinamin konnte nicht nachgewiesen werden, so daß -Methyl-Adrenalin wahrscheinlich nicht gebildet wird.3. Im Meerschweinchenherzen gespeichertes -Methyl-Noradrenalin wird durch Reserpin in vivo schlechter freigesetzt als Noradrenalin.4. Das in isolierten Herzgranula (Sediment 100 000·g) gespeicherte -Methyl-Noradrenalin wird durch Segontin bei der Inkubation in vitro ebenfall schlechter freigesetzt als Noradrenalin aus normalen Herzgranula.5. Der Mechanismus der -Methyl-Dopa-Wirkung wird diskutiert.6. Es wird eine fluorimetrische Methode zur Bestimmung von -Methyl-Noradrenalin angegeben.

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Summary 1. The catecholamine content of guinea pig organs after administration of -methyldopa has been determined. The results show that the loss of noradrenaline from heart and that of noradrenaline and dopamine from brain is compensated by a stoichiometric uptake of -methylnoradrenaline and -methyldopamine respectively.2. On the contrary the loss of adrenaline from the suprarenal medulla induced by pretreatment with -methyldopa is not completely restored by the -methylated amines, -methyldopamine and -methylnoradrenaline. -methyladrenaline could not be detected even by paperchromatography.3. The release of the stored -methylnoradrenaline from guinea pig heart by reserpine is very small in comparison with that of noradrenaline.4. Furthermore only small amounts of the stored -methyl-noradrenaline are released from isolated granules of the guinea pig heart after incubation in vitro with segontin. Whereas the same amount of segontin depleted completely the noradrenaline content of the granules isolated from normal hearts.5. The mechanism of action of -methyldopa is discussed.6. A method for the fluorimetric determination of -methylnoradrenaline is described. It is at first separated from -methyldopamine, dopamine and histamine by column chromatography, condensed with o-phthaldialdehyd and the obtained fluorescense is measured by using an Aminco-Bowman spectrophotofluorimeter.

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Mit 6 Textabbildungen

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Herrn Dr. H. Blaschko zum 65. Geburtstag gewidmet.

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Über einen Teil der Ergebnisse wurde auf der Frühjahrstagung der Deutschen Pharmakologischen Ges. in Mainz, April 1964, berichtet. Naunyn-Schmiedebergs Arch. exp. Path. Pharmak. 247, 297 (1964).

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Ausgeführt mit Unterstützung der Deutschen Forschungsgemeinschaft.     

Complex information processing by the transmembrane signaling system involving G proteins

Much of the information cells receive is transduced by a membranous signaling system that uses heterotrimeric guanine nucleotide binding proteins (G proteins) to functionally couple cell surface receptors to a variety of effectors. During recent years it has been shown that receptors, G protein , and subunits as well as effectors involved in this signaling system exhibit a remarkable structural diversity and that the interactions of these components display a bewildering complexity.Even though many questions remain to be answered, it is becoming obvious that G proteins form the basis of a complex membranous signaling network which allows the cell to coordinate and to process incoming signals already on the level of the plasma membrane.Abbreviations G protein regulatory heterotrimeric guanine nucleotide-binding protein - IP₃ inositol trisphosphate - PLC phospholipase C - G_x and _x nomenclature to describe a particular G protein and its subunit - PACAP pituitary adenylyl cyclase-activating polypeptide - TSH thyroidea stimulating hormone - fMLP formyl-methionyl-leucylphenylalanine - LTB₄ leukotriene B₄ - Sst somatostatin - _{2 2} adrenergic - _{1 1} adrenergic - PGE₂ prostaglandin E₂ - A₂ adenosine A₂ - PKC protein kinase C - [Ca⁺]_i cytosolic Ca²⁺ concentration - AC adenylyl cyclase - CaM calmodulin Correspondence to: G. Schultz at the above address     

Bildung und Speicherung von α-Methylnoradrenalin

Zusammenfassung An Meerschweinchen wurde die Wirkung von Reserpin auf die Bildung von -Methylnoradrenalin aus -Methyldopa untersucht. Im Herzen und Vas deferens verursacht Reserpin eine starke Verminderung der -Methylnoradrenalin-Bildung, im Gehirn hat es nur einen geringen Einfluß.Bei der Perfusion isolierter Meerschweinchenherzen mit Dopamin bzw. -Methyldopamin wird wesentlich mehr -Methyldopamin in das Herz aufgenommen als Dopamin. Reserpin-Vorbehandlung vermag weder die Dopamin- noch die -Methyldopamin-Aufnahme zu hemmen; es vermindert jedoch die Bildung von Noradrenalin bzw. -Methylnoradrenalin aus ihren Vorstufen, wobei die Bildung von Noradrenalin am stärksten reduziert wird.Bei der Inkubation isolierter Meerschweinchenherz-Granula mit -Methyldopamin oder -Methylnoradrenalin vermindert Reserpin die Aufnahme der beiden Amine in die Granula.Nach Perfusion von Kaninchenherzen mit Dopamin bzw. -Methyldopamin wurde die subcelluläre Verteilung dieser beiden Amine untersucht. Während -Methyldopamin hauptsächlich partikulär gebunden vorliegt, befindet sich Dopamin vorwiegend im Cytoplasma.

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Summary Experiments have been carried out in order to study the influence of reserpine pretreatment on the storage of -methylnoradrenaline and its synthesis from -methyldopa (3×400 mg/kg).Reserpine diminishes in heart and vas deferens of guinea-pigs the synthesis of -methylnoradrenaline from -methyldopa while its effect in brain is not significant.During infusion of isolated guinea-pig hearts with -methyldopamine or dopamine (0.6 moles/min, 30 min) much more -methyldopamine is taken up into the hearts than dopamine. Pretreatment with reserpine (4×25 g/kg) has no effect on the uptake of these amines; it causes, however, a decrease of synthesis of noradrenaline and -methylnoradrenaline from their precursors. The inhibitory effect on the synthesis of noradrenaline is much more pronounced than that on the synthesis of -methylnoradrenaline.In incubation experiments with storage granules isolated from guinea-pig hearts the uptake of -methyldopamine and -methylnoradrenaline (0.25 moles/ml) is significantly reduced by reserpine (40 g/ml).After infusion of rabbit hearts with -methyldopamine or dopamine (1.5 moles/min, 60 min) the subcellular distribution of these amines was studied. The main part of -methyldopamine is located in the particulate fractions whereas dopamine is predominantly found in the cytoplasma.

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Herrn Prof. Dr. Peter Holtz zum 65. Geburtstag am 6.2.1967 gewidmet.

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Ausgeführt mit der Unterstützung der Deutschen Forschungsgemeinschaft.     

Synthesis of aldosterone antagonists

Conclusions Two antagonists of aldosterone were synthesized based on ethynyl androstenediol: the -lactone of 3-(3-keto-17-hydroxy-4-androsten-17-yl)-propionic acid (SC-5233) and its 7-thioacetate (aldactone or SC-9420).DeceasedTranslated from Khimiko-Farmatsevticheskii Zhurnal, No. 1, pp. 10–13, January, 1969.     

Motor disturbance induced by tremorine and oxotremorine

Summary The action of tremorine and oxotremorine on spinal motor activity was studied in the rat. Both tremorogenio agents increased a reflex activity and spontaneous activity. The increase in spontaneous activity consisted of rhythmic bursts of discharges. The increase in a reflex activity was accompanied by rigidity, which manifested itself by the appearance of tonic muscle activity.Tremorine and oxotremorine-induced tremor was depressed by the antitremor agents metixene and Kr 339, the antiparkinson drugs atropine and biperiden, and the adrenergic receptor blocking agents propranolol and pronethalol. The adrenergic. receptor blocking agents azapetine, dihydroergotamine, haloperidol, phenoxybenzamine and phentolamine failed to inhibit tremor activity. Chlorpromazine, however, as well as procaine, verapamil and DOPA, diminished the intensity of tremor activity without blocking the generation of tremor bursts.Drugs which depressed tremor activity also antagonized the effect of oxotremorine on and motor activity, whereas the drugs, which only diminished tremor intensity, depressed the increased reflex discharge without reducing spontaneous activity. Rigidity disappeared when reflex discharge was normalized.It is concluded that experimental parkinsonlike rigidity may be interpreted in terms of a disturbed balance between monoaminergic and cholinergic mechanisms in the brain.The authors are grateful to KnolI-Ludwigshafen for the support of the investigation.     

Neonatal exposure to zearalenone causes persistent anovulatory estrus in the rat

The effect of neonatal administration of zearalenone on the female reproductive system was studied in the rat. A single subcutaneous injection of 1.0 mg zearalenone to 3- or 5-day-old rats caused persistent vaginal estrus in adulthood. Ovaries in these animals contained many large follicles but no newly formed corpora lutea. The same effects were observed in rats which had received 100 g estradiol-17 in the neonatal period. Most rats which had received 100 g zearalenone or 10 g estradiol-17 showed regular 4-day estrous cycles and had newly formed corpora lutea in their ovaries. These results demonstrate that neonatal exposure to zearalenone produces persistent anovulatory estrus in the rat, the potency being about one tenth that of estradiol-17.