Distribution of Galanin mRNA Containing Cells and Galanin Receptor Binding Sites in Human and Rat Hypothalamus

The distribution of cells containing galanin mRNA and that of galanin receptor binding sites were investigated using in situ hybridization histochemistry and receptor autoradiography in male rat hypothalamus and in postmortem hypothalamic tissues from control human brains. Oligonucleotide probes labelled with 32P were used for hybridization experiments. The specificity of the hybridization signal was ascertained using several probes, competition assays and Northern blot analysis. High levels of hybridization were found in the paraventricular, supraoptic and arcuate nuclei of rat and human hypothalamus. Human intermediate nuclei and scattered cells of the posterior perifornical nucleus also contained galanin mRNA. Galanin mRNA was also found in the dorsomedial nucleus of the rat. The distribution of galanin receptor sites was investigated by receptor autoradiography using 125I-labelled porcine galanin. The specificity of the binding was assessed by competition with different neuropeptides. While galanin blocked the binding at nanomolar concentrations, the other neuropeptides examined were ineffective at 10-7 M concentrations. The highest densities of galanin binding sites were seen in the preoptic area, ventromedial and lateral nuclei, of rat and human hypothalamus. In contrast, very low densities of binding sites were observed in the paraventricular, supraoptic and arcuate nuclei. Our results show that the distribution of neurons expressing galanin is complementary to that of galanin receptors in the rat and human hypothalamus. This suggests that receptors for galanin are not located on the cell bodies of galaninergic neurons, but are probably presynaptic on or postsynaptic to the processes of these cells.     

The effect of flow and mass transport in thrombogenesis

The paper presents a mathematical analysis of the contributions of flow and mass transport to a single reactive event at a blood vessel wall. The intent is to prepare the ground for a comprehensive study of the intertwining of these contributions with the reaction network of the coagulation cascade. We show that in all vessels with local mural activity, or in “large” vessels (d>0.1 mm) with global reactivity, events at the tubular wall can be rigorously described by algebraic equations under steady conditions, or by ordinary differential forms (ODEs) during transient conditions. this opens up important ways for analyzing the combined roles of flow, transport, and coagulation reactions in thrombosis, a task hitherto considered to be completely intractable. We report extensively on the dependence of transport coefficient k_L and mural coagulant concentration C_w on flow, vessel geometry, and reaction kinetics. It is shown that for protein transport, k_L varies only weakly with shear rate in large vessels, and not at all in the smaller tubes (d<10⁻² mm). For a typical protein, k_L∼10⁻³ cm s⁻¹ within a factor of 3 in most geometries, irrespective of the mural reaction kinetics. Significant reductions in k_L (1/10–1/1,000) leading to high-coagulant accumulation are seen mainly in stagnant zones vicinal to abrupt expansions and in small elliptical tubules. This is in accord with known physical observations. More unexpected are the dramatic increases in accumulation which can come about through the intervention of an autocatalytic reaction step, with C_w rising sharply toward infinity as the ratio of reaction to transport coefficient approaches unity. Such self-catalyzed reactions have the ability to act as powerful amplifiers of an otherwise modest influence of flow and transport on coagulant concentration. The paper considers as well the effect on mass transport of transient conditions occasioned by coagulation initiation or pulsatile flow. During initiation, instantaneous flux varies with diffusivity and bulk concentration, favouring the early adsorption/consumption of proteins with the highest abundance and mobility. This is akin to the ‘Vroman effect’ seen in narrow, stagnant spaces. The effect of flow pulsatility on k_L has the potential, after prolonged cycling, of bringing about segregation or accumulation of proteins, with consequences for the coagulation process.     

Häufigkeit pathologischer Veränderungen des oberen Gastrointestinaltraktes bei Patienten vor Herzoperationen

Resümee Schwerwiegende gastroduodenale Erkrankungen sind bei fast der Hälfte aller Patienten, die sich einer Operation am offenen Herzen unterziehen müssen, auch bei Fehlen von Symptomen nachweisbar. Das erhebliche Überwiegen von Magenläsionen spricht dafür, daß die arteriosklerotisch bedingte Perfusionsminderung der Schleimhaut die entscheidende Ursache hierfür ist. Routinemäßige präoperative Ösophago-Gastro-Duodenoskopien können die nicht unerhebliche durch gastrointestinale Komplikationen bedingte postoperative Mortalität senken helfen.

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Pathological changes in the upper gastrointestinal tract in patients awaiting open heart surgery

Summary While waiting for open heart surgery, in 153 patients (104 male, 49 female, 22–76 years of age) without gastrointestinal symptoms and/or history esophago-gastro-duodenoscopy was performed. 124 patients suffered from coronary heart disease, 29 from valvular defect, aneurysm of the sinus of Valsalva or tumor of the heart.In 47.1% endoscopy revealed serious abnormal findings: in 16.3% gastric ulcer, in 20.9% erosive gastritis, duodenal ulcer and erosive duodenitis in 5.2%, respectively, 1 case of gastric carcinoma, 2 of large polyps and 3 of reflux esophagitis of higher degree (totally 3.9%).In patients with coronary artery disease, the relation of erosive and ulcerous gastric lesions as compared with those of duodenal origin was 41, in patients with other cardiac diseases it was 21, respectively (p<0,001).Compared with a normal population, the incidence of pathological gastric findings was 54-fold higher in our patients, and 1.7-fold concerning duodenal lesions, respectively (p<0.001).51 patients on acethylsalicylic acid (160 mg/ die) showed pathologic findings in 41.2%, and 96 patients without ulcer-inducing therapy in 51%. Thus, low-dose Aspirin does not seem to have serious gastric side effects.The results of the study stress the necessity of routinely performed endoscopy of the upper gastrointestinal tract in patients awaiting open heart surgery. This will lead to a lower incidence of serious gastrointestinal complications postoperatively, which are known to have a high mortality.

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Abkürzungen ASS Azetylsalizylsäure - J. Jahre - GI-Trakt Gastrointestinaltrakt     

Hydrolysis of Carbonates,Thiocarbonates, Carbamates,and Carboxylic Esters of α-Naphthol, β-Naphthol,and p-Nitrophenol by Human,Rat, and Mouse Liver Carboxylesterases

Thirty carbonates, thiocarbonates, carbamates, and carboxylic esters of -naphthol, -naphthol, and p-nitrophenol were synthesized and tested as substrates for liver carboxylesterases from the crude microsomal fractions of human and mouse, and purified isozymes, hydrolases A and B, from rat liver microsomes. The carbonates, thiocarbonates, and carboxylic esters of -naphthol were cleaved more rapidly than the corresponding -naphthol isomers by the mammalian liver esterases. -Naphthyl esters of acetic, propionic, and butyric acids were among the best substrates tested for these enzymes. The majority of the substrates was consistently hydrolyzed at higher rates by hydrolase B compared with hydrolase A, although the Michaelis–Menten constant (K _m) values of selected substrates differed widely with these two isozymes. Malathion was a 15-fold better substrate for hydrolase B than for hydrolase A. Compared with the corresponding carboxylates, the carbonate moiety of - and -naphthol and p-nitrophenol lowered the specific activities of the enzymes by about fivefold but improved stability under basic conditions. The optimum pH of mouse liver esterase with the acetate, methylcarbonate, and ethylthiocarbonate of -naphthol was between pH 7.0 and pH 7.6. Human and mouse liver microsomal esterase activities were about five orders of magnitude lower than the esterase activities of purified rat liver hydrolase B. A relationship between the catalytic activity of the enzymes and the lipophilicity of the naphthyl substrates indicated that (i) in the - and -naphthyl carbonate series, an inverse relationship between enzyme activity and lipophilicity of the substrates was observed, whereas (ii) in the -naphthyl carboxylate series, an increase in enzyme activity with increasing lipophilicity of the substrates up to a log P value of about 4.0 was observed, after which the enzyme activity decreased.     

Serotonergic inhibition of the mossy fibre—granule cell glutamate transmission in rat cerebellar slices

Summary The glutamatergic mossy fibre granule cell pathway has been investigated in rat cerebellar slices. Exposure to 35 mM KCI, a concentration of K⁺ known to elicit Ca²⁺-dependent releases of excitatory amino acids from cerebellar slices, raised cGMP levels. The cGMP response was decreased in a concentration-dependent manner by D-(–)-2-amino-5-phosphonopentanoic acid (D-AP5) and by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) indicating the involvement of ionotropic glutamate receptors of both the N-methyl-D-aspartate (NMDA) and the non-NMDA type. The K⁺-evoked production of cGMP was potently inhibited (EC₅₀ = 1.21 nM) by 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), a selective 5-HT₂ receptor agonist. The effect of DOI (0.01 M) was antagonized by 0.03 M of the 5-HT₂ receptor antagonists ketanserin and methiothepin. At concentrations higher than 0.1 M, both antagonists increased on their own the cGMP response elicited by high-K⁺. This effect was insensitive to tetrodotoxin.It had been previously shown that rat mossy fibre endings release glutamate upon depolarization and that such release can be inhibited by activation of 5-HT₂ receptors sited on the mossy fibre endings. Altogether the available data suggest the following conclusions: (a) the glutamate/aspartate endogenously released in cerebellar slices during K⁺ depolarization increase cGMP synthesis through the activation of both NMDA and non-NMDA receptors; (b) a portion of the cGMP response can be prevented by 5-HT₂ receptor activation and may reflect the activity of the mossy fibre-granule cell pathway. Thus serotonin is likely to exert a potent inhibitory control of the excitatory mossy fibre input to the cerebellum by acting at receptors of the 5-HT₂ type. Correspondence to M. Raiteri at the above address     

Effects of 6-hydroxydopamine on pre- and post-junctional 5-HT1-like receptor-mediated responses in dog saphenous vein

Summary To investigate whether 5-HT₁-like receptor-mediated inhibition of adenosine 3 : 5-cyclic monophosphate (cyclic AMP) accumulation occurs in nerves or smooth muscle of saphenous vein, infusions of 6-hydroxydopamine (6-OHDA) were administered to dogs with the aim of inducing sympathetic nerve damage. The effects of 6-OHDA on other 5-HT₁-like receptor-mediated responses at the pre- and post-junctional level were investigated for comparison by studying 5-hydroxytryptamine (5-HT)-induced inhibition of ³H-noradrenaline release and contraction of smooth muscle respectively.Disruption of nerve function by 6-OHDA was revealed by the lack of catecholaminergic fluorescence and neurogenic contractile responses in saphenous veins from dogs treated with 6-OHDA. In addition, severe impairment of neuronal uptake mechanisms were apparent since basal efflux of ³H-noradrenaline, electrically-evoked release of ³H-noradrenaline and remaining ³H-noradrenaline content were considerably reduced. Some ³H-noradrenaline was taken up and released in 6-OHDA treated tissues which is consistent with the existence of nerve varicosities resistant to the present dosing regime of 6-OHDA, an observation substantiated by electron microscopy studies showing inconsistent lesions of nerve terminals.6-OHDA pre-treatment potentiated the smooth muscle contractile responses mediated by 5-HT₁-like receptors as well as potentiating 5-HT-evoked inhibition of prostaglandin E₂-stimulated cyclic AMP accumulation. It did not, however, affect 5-HT-induced inhibition of ³H-noradrenaline release. The present results suggest that inhibition of cyclic AMP accumulation by 5-HT occurs predominantly in smooth muscle. Correspondence to A. J. Kaumann at the above address     

Modulation of N-methyl-d-aspartate (NMDA)-stimulated noradrenaline release in rat brain cortex by presynaptic α2-adrenoceptors

Summary Rat brain cortex slices and synaptosomes preincubated with [³H]noradrenaline were used to investigate whether the NMDA-evoked noradrenaline release is modulated by agonists or antagonists at presynaptic ₂-adrenoceptors.In experiments on slices, noradrenaline and the preferential ₂-adrenoceptor agonists talipexole (former B-HT 920) and clonidine inhibited the NMDA evoked tritium overflow whereas the selective ₁-adrenoceptor agonists cirazoline and methoxamine were ineffective. The ₂-adrenoceptor antagonists rauwolscine and idazoxan facilitated the NMDA-evoked tritium overflow whereas the preferential ₁-adrenoceptor antagonist prazosin was ineffective. The concentration-response curve of talipexole for its inhibitory effect on NMDA-evoked overflow was shifted to the right by idazoxan (apparent pA₂ = 7.5). The EC₅₀ of NMDA (97 mol/l) for its stimulating effect on tritium overflow was not substantially changed by blockade of ₂-autoreceptors with 1 mol/l rauwolscine (EC₅₀ of NMDA in the presence of the ₂-adrenoceptor antagonist, 155 mol/l), but the maximal overflow of tritium was increased 2.5 fold by this rauwolscine concentration. In experiments on synaptosomes, talipexole and noradrenaline inhibited the NMDA-evoked tritium overflow. The inhibitory effect of talipexole was abolished by idazoxan which, given alone, was ineffective, as was prazosin. Talipexole did also not produce an inhibition when tritium overflow was evoked by NMDA in the presence of -conotoxin GVIA 0.1 mol/l; the latter, by itself, decreased the response to NMDA by about 55%. It is concluded that the NMDA-evoked noradrenaline release in the cerebral cortex is modulated via presynaptic ₂-adrenoceptors on the noradrenergic neurones. Stimulation of these autoreceptors in slices by endogenous noradrenaline does not result in a decreased potency of NMDA, but in a decreased maximum effect, in stimulating noradrenaline release. The inhibitory effect of ₂-adrenoceptor agonists on the NMDA-evoked release is at least partially due to a functional interaction between the NMDA receptors and ₂-autoreceptors at the level of the same varicosities. The results obtained with -conotoxin GVIA suggest that Ca²⁺ influx via the N-type voltage-sensitive calcium channel (VSCC) occurs in response to NMDA receptor stimulation and contributes substantially to the induction of NMDA-evoked noradrenaline release. The inhibitory effect of ₂-adrenoceptor stimulation on this release appears to be ultimately due to an inhibition of the influx of Ca²⁺ via the N-type VSCC. Correspondence to: M. Göthert at the above address     

Autoradiographic characterisation and localisation of 5-HT1D compared to 5-HT1B binding sites in rat brain

Summary The regional distribution and the pharmacology of the binding sites labelled with the novel 5-hydroxytryptamine (serotonin) 5-HT_1B/1D selective radioligand serotonin-O-carboxy-methyl-glycyl-[¹²⁵I]tyrosinamide (abbreviated [¹²⁵I]GTI for the sake of simplicity) was determined using quantitative autoradiography in rat brain. The distribution of [¹²⁵I]GTI binding sites was largely comparable to that of [¹²⁵I] iodocyanopindolol ([¹²⁵I] ICYP) which labels 5-HT_1B binding sites (in the presence of 8-OH-DPAT (8-hydroxy-[2N-dipropylamino]tetralin) and isoprenaline, to prevent binding to 5-HT_1A and -adrenoceptor binding sites), although a detailed analysis revealed differences.The pharmacology of the [¹²⁵I]GTI binding sites was analysed using compounds known to display high affinity for and/or distinguish between 5-HT_1B and 5-HT_1D sites: 5-carboxamidotryptamine (5-CT), sumatriptan, CP 93129 (5-hydroxy-3(4-1,2,5,6-tetrahydropyridyl)-4-azaindole), (–)pindolol, PAPP (4[2-[4-[3-(trifluoromethyl)phenyl]-1-piperazinyl]ethyl] benzeneamine), rauwolscine, and 8-OH-DPAT. The displacement of [¹²⁵I]GTI by 5-CT was monophasic. By contrast, the selective 5-HT_1B compound CP 93129 and (–)pindolol produced biphasic curves showing a majority of high affinity sites in the globus pallidus and the substantia nigra, whereas PAPP and sumatriptan (which are somewhat 5-HT_1D selective) produced biphasic curves indicating a minority of high affinity sites in these areas. In addition, by blocking the 5-HT_1B sites with 100 nM CP 93129, the remaining population of [¹²⁵I]GTI binding sites could be studied and was found to have high affinity for PAPP, rauwolscine and 8-OH-DPAT. The pharmacological profile of the major binding component was typical of the 5-HT_1B type: 5-CT > CP 93129 (–)pindolol > sumatriptan >/ PAPP > rauwolscine. The profile of the minor component of [¹²⁵I] GTI binding is best characterised as that of a 5-HTID site: 5-CT > PAPP sumatriptan > rauwolscine > (–)pindolol CP 93129.The localisation of the non 5-HT_1B [¹²⁵I]GTI binding sites was characterised by blocking the 5-HT_1B receptors with 100 nM CP 93129. Low densities of the 5-HT_1D recognition sites were found to be present in globus pallidus, ventral pallidum, caudate-putamen, subthalamic nucleus, entopeduncular nucleus, substantia nigra (reticular part), nuclei of the (normal and accessory) optic tract, different nuclei of the geniculate body and frontoparietal cortex, although higher densities of 5-HT_1B sites were always observed in the same structures. Thus, in agreement with the recent cloning of a rat 5-HT_1D receptor cDNA, the presence and the distribution of 5-HT_1D sites could be documented in rat brain. However, when compared to 5-HT_1B sites, 5-HT_1D sites represent only a minor component of the [¹²⁵I]GTI binding in the rat brain structures studied.Correspondence to: D. Hoyer at the above address     

Effects of blood pressure changes on the catecholamine release in the locus coeruleus of cats anaesthetized with pentobarbital or chloralose

Summary Effects of carotid occlusion and drugs applied intravenously on the release of endogenous catecholamines in the locus coeruleus of cats anaesthetized with pentobarbital or chloralose were investigated. The locus coeruleus was superfused bilaterally with artificial cerebrospinal fluid through push-pull cannulae inserted stereotaxically. Dopamine, noradrenaline and in some experiments also adrenaline were determined radioenzymatically in the superfusate.Under pentobarbital anaesthesia, a bilateral carotid occlusion increased the release rate of noradrenaline in the locus coeruleus, while the release of dopamine was decreased. These changes were due to the fall of blood pressure in the carotid sinus caused by the occlusion. Loading of baroreceptors by elevating blood pressure with phenylephrine (10 g·kg^–1·min^–1, i.v. infusion) was accompanied by a decreased release of noradrenaline in the locus coeruleus. This decrease in noradrenaline release was not detected in the caudal aspect of the locus coeruleus. Under chloralose anaesthesia, phenylephrine diminished the release rate of noradrenaline to about the same extent as under pentobarbital anaesthesia. The release rate of adrenaline was also decreased. A prolonged infusion of phenylephrine led to a prolonged pressor response associated with a sustained decrease in the noradrenaline release rate. Intravenous injection of chlorisondamine (3 mg·kg^–1) did not change the release of noradrenaline, while dopamine release was reduced.It is concluded that the release of catecholamines in the locus coeruleus is influenced by signals originating from peripheral baroreceptors. The influences are similar under pentobarbital and chloralose anaesthesia. Noradrenergic neurons responding to haemodynamic signals are not uniformly distributed within the locus coeruleus. It is suggested that noradrenergic and possibly dopaminergic and adrenergic neurons of the locus coeruleus are involved in the baroreceptor reflex, thus contributing to central homeostasis of blood pressure.     

Antihypertensive effect of once daily sustained release isradipine: a placebo controlled cross-over study

Summary To evaluate the magnitude and duration of the antihypertensive effect of sustained release (SRO) isradipine, 37 uncomplicated essential hypertensive patients (diastolic blood pressure 100–115 mm Hg after a one month run-in on placebo) were randomised to receive, according to a double-blind cross-over design, isradipine SRO 5 mg once daily and the corresponding placebo for 1 month. At the end of each treatment period, sitting blood pressure and heart rate were measured immediately before and every hour for 6 h after the last dose. Thirty-four patients [16 m, age 54 (7) y] completed the study.As compared to randomised placebo, isradipine SRO significantly reduced the systolic (SBP) and diastolic (DBP) blood pressure. Absolute DBP decrements versus placebo peaked 6 h after dosing (-8.8 mm Hg) and were not significantly lower (-8.2 mm Hg) at the end of the dose interval. At the same times, the absolute decrements in SBP were -9.8 mm Hg and -9.7 mm Hg, respectively.DBP was normalised in 19 patients (56%) at peak and in 17 (50%) at trough time. The trough to peak efficacy ratio in patients with peak DBP 90 mm Hg was 70%. Heart rate was slightly increased by isradipine SRO. Adverse effects monitored with a check-list occurred in 8/36 patients (22%) on isradipine SRO and in 4/35 (11%) on randomized placebo.The data suggest that isradipine SRO is an effective antihypertensive drug, with a trough to peak efficacy ratio supporting once daily administration in most mild to moderate essential hypertensives.