Metabolism of some possible ecdysone precursors in Calliphora stygia

A study has been made, using Calliphora stygia at the time of puparium formation, of the incorporation of a number of labelled sterols into β-ecdysone. [1-³H]-Cholesterol and [4-¹⁴C]-cholesterol are incorporated to a similar extent (0·01-0·02%). [1-³H]-7-Dehydrocholesterol is better incorporated (0·025%) than cholesterol while [1-³H]-cholesterol sulphate, (22R)-22-hydroxy-[22-³H]-cholesterol, and 25-hydroxy-[26-¹⁴C]-cholesterol are not incorporated to a significant extent.     

Increased biological potency of hexafluorinated analogs of 1,25-dihydroxyvitamin D3 on bovine parathyroid cells

1,25-dihydroxyvitamin D₃ (1,25-(OH)₂D₃) is known to be involved in regulating the proliferation of parathyroid cells and PTH synthesis through reactions involving its nuclear receptor. We evaluated the effects of 1,25-(OH)₂D₃ and its hexafluorinated analog, 26,26,26,27,27,27-hexafluoro-1,25-dihydroxyvitamin D₃ (26,27-F₆-1,25-(OH)₂D₃), on parathyroid cells. The 1,25-(OH)₂D₃ and 26,27-F₆-1,25-(OH)₂D₃ each inhibited [³H]thymidine incorporation and ornithine decarboxylase (ODC) activity, which is important in cell proliferation, in primary cultured bovine parathyroid cells. The inhibitory effect of 26,27-F₆-1,25-(OH)₂D₃ on PTH secretion from parathyroid cells was significantly more potent than that of 1,25-(OH)₂D ₃ between 10⁻¹¹ M and 10⁻⁸ M. Study of 26,27-F₆-1,25-(OH)₂D₃ metabolism in parathyroid cells in vitro elucidated its slower degradation than that of 1,25-(OH)₂D₃. After 48 h of incubation with [1β-³H]26,27-F₆-1,25-(OH)₂D₃, two HPLC peaks, one for [1β-³H]26,27-F₆-1,25-(OH)₂D₃, and a second larger peak for [1β-³H]26,27-F₆-1,23(S),25-(OH)₃D₃, were detected. No metabolites were detected after the same period of incubation with 1,25-(OH)₂[26,27-³H]D₃. We observed that 26,27-F₆-1,23(S),25-(OH)₃D₃ was as potent as 1,25-(OH)₂D₃ in inhibiting the proliferation of parathyroid cells.

Data suggest that the greater biological activity of 26,27-F₆-1,25-(OH)₂D₃ is explained by its slower metabolisms and by the retention of the biological potency of 26,27-F₆-1,25-(OH)₂D₃ even after 23(S)-hydroxylation.     

On the mechanism of biosynthesis of leukotrienes and related compounds

[10D-³H; 3-¹⁴C]- and [10L-³H; 3-¹⁴C]arachidonic acids were incubated with human polymorphonuclear leukocytes and with human platelets. Leukotriene B₄ and 5(S),12(S)-dihydroxy-6trans,8cis,10trans,14-cis-eicosatetraenoic acid (5,12-DHETE) were isolated and the ³H/¹⁴C ratios determined. It could be concluded that the 10D (pro-R)-hydrogen is eliminated in the conversion of 5(S)-hydroperoxy-6trans,8cis,11cis,14cis-eicosatetraenoic acid into leukotriene A₄ whereas in the conversion of arachidonic acid into 5,12-DHETE the 10L (pro-S)-hydrogen is lost. Incubation of the doubly labeled arachidonic acids with human platelets confirmed and extended previous data on the stereochemistry of the hydrogen removal from C-10 during the conversion into 12(S)-hydroperoxy-5cis,8cis,10trans,14cis-eicosatetraenoic acid, i.e., the 10L (pro-S)-hydrogen is eliminated and the 10D (pro-R)-hydrogen retained.     

Specificity of Muscarinic Acetylcholine Receptor Regulation by Receptor Activity

Abstract— Regulation of muscarinic acetylcholine receptor concentration by receptor activity in neuron-like NG108-15 hybrid cells is a highly specific process. Receptor levels, monitored by binding of [³H]quinuclidinyl benzilate ([³H]QNB), decreased 50-75% following 24-h incubation of cells with muscarinic agonists, but none of the following cellular processes was altered by this chronic receptor stimulation: (1) glycolytic energy metabolism, measured by [³H]deoxy- d -glucose ([³H]DG) uptake and retention; (2) rate of cell division; (3) transport, measured by [³H]valine and [³H]uridine uptake; (4) RNA biosynthesis, measured by [³H]uridine incorporation; (5) protein biosynthesis, measured by [³H]valine and [³⁵S]methionine incorporation into total protein and into protein fractions obtained by polyacrylamide gel electrophoresis. In contrast, chronic stimulation did cause a threefold decrease in the capacity of carbachol to stimulate phosphatidylinositol (PI) turnover, a receptor-mediated response. In addition to cholinomimetics, the neuroeffector adenosine (1 m m for 24 h) also caused a decrease in [³H]QNB binding levels, but chronic stimulation of α -adrenergic, opiate, prostaglandin E₁, and prostaglandin F_2α receptors found on NG108-15 cells caused no changes. The data indicate that loss of muscarinic receptors caused by receptor stimulation is not a consequence of fundamental changes evoked in overall cellular physiology but reflects a specific regulation of cholinoceptive cell responsiveness.     

Differentiation of [H]phencyclidine and ( + )-[H]SKF-10,047 binding sites in rat cerebral cortex

The potency of a series of opioid and non-opioid psychotomimetic drugs to inhibit the specific binding of [³H]PCP and ( + )-[³H]SKF-10,047 to rat cerebral cortical membranes was examined. ( + )-PCMP, the 3-methylpiperidino analog of PCP, was a potent inhibitor of the specific binding of both ligands. All of the other 12 compounds examined, however, displayed a 3-277-fold selectivity for either [³H]PCP or (+)-[³H]SKF-10,047 binding. These results suggest that although these opioid and non-opioid psychotomimetics bind to both sites, most have significantly different affinities. The binding sites for [³H]PCP appear to be distinct from the ‘sigma’ binding sites labeled with (+)-[³H]SKF-10,047.

SKF-10,047 Sigma receptor Phencyclidine Phencyclidine receptor Psychotomimetic activity     

Molecular structure of the carbon-bound phosphorus ylide complex trans-dichlorobis(benzoyltri-n-butylphosphorane)palladium(II)

The molecular structure of trans-[Pd(PhC(O)CHP(n-C₄H₉)₃)₂Cl₂] has been determined via a single crystal X-ray diffraction study: triclinic,P1,a = 8.876(2),b = 10.908(3),c = 11.938(4)Å, = 97.06(2)°, β = 102.79(2)°, γ = 100.51(2)°,V= 1092.1(5)ų,Z = 1 and R(F) = 4.61%. The phosphorus ylide molecules are bound to the palladium atom through their methine carbon atoms, the overall coordination geometry about the palladium being square planar. The protons in the ortho-positions of the two phenyl group are poised above and below the palladium atom, suggesting that the complex is a precursor of the ortho-metalated complex [Pd(μ-Cl)(C₆H₄C(O)CHP(n-C₄H₉)₃)]₂ synthesized earlier in our laboratory.     

Peroxidase-Mediated Oxidation, a Possible Pathway for Activation of the Fungal Nephrotoxin Orellanine and Related Compounds. ESR and Spin-Trapping Studies

Orellanine is the tetrahydroxylated and di-N-oxidized bipyridine toxin extracted from several Cortinarius mushrooms among them C. orellanus. The pathogenic mechanism involved in the C. orellanus-poisoning by orellanine leading to kidney impairment is not yet fully understood until now. Electron spin resonance (ESR) spectroscopy has been used to study the activation of orellanine by horseradish peroxidase/H₂O₂ system at physiological pH. Evidence for a one-electron oxidation of the toxin by this enzymatic system to an ortho-semiquinone radical intermediate is presented.

The orellanine ortho-semiquinone generated by the peroxidase/H₂O₂ system abstracts hydrogen from glutathione, generating the glutathionyl radical which is spin-trapped by 5,5'-dimethyl-1-pyrroline N-oxide (DMPO) and subsequently detected by ESR spectroscopy. Similarly, the ortho-semiquinone abstracts hydrogen from ascorbic acid to generate the ascorbyl radical which is detected by direct ESR. The peroxidatic oxidation of orellanine to semiquinone followed by its reduction by glutathione or ascorbic acid does not induce dioxygen uptake. The relationship between chemical structure and HRP oxidation of orellanine-related molecules, namely orelline and DHBPO₂ (the parent molecule lacking of hydroxyl groups in 3 and 3' position) has been investigated in absence or in presence of reducing agents. None of the orellanine-related compounds can be oxidized by the HRP/H₂O₂ system, showing that both catecholic moieties and aminoxide groups are necessary for observing the formation of the ortho-semiquinone form of orellanine. As shown for the (photo)chemical oxidation of orellanine, the mechanism of toxicity could be correlated with a depletion of glutathione and ascorbate levels which are implicated in the defence against oxidative damage.     

Prostanoid formation in primary astroglial cell cultures: Ca-dependency and stimulation by A 23187, melittin and phospholipases A2 and C

Prostaglandin (PG) and thromboxane B₂ (TXB₂) biosynthesis was studied in cultured astrocytes from neonatal rat brain hemispheres. After two weeks of cultivation, prostanoids were formed with the spectrum: PGD₂ > TXB₂ > PGF₂ > PGE₂, as measured by specific radioimmunoassays. Under basal conditions PGD₂ biosynthesis (9.55 ng/mg protein/15 min) was in the same order of magnitude as the sum of the other prostanoids. The formation of prostanoids was stimulated in a concentration dependent manner (up to 6–10 fold) by the calcium ionophore A 23187 (0.01–10 μM) as well as by melittin (0.01–5 μg/ml), phospholipase A₂ (10–40 U/ml) and phospholipase C (0.01–1 U/ml). Basal and evoked PG and TXB₂ biosynthesis depended on the availability of Ca²⁺, as demonstrated in Ca²⁺ free incubation medium containing Na₂EDTA (1 μM), or with verapamil (100 μM) and 3,4,5-trimethoxybenzoic acid-8-(diethylamino)-octylester-HCl (TMB-8, 1–100 μM). Indomethacin (10 μM), mepacrine (100 μM) and p-bromophenacylbromide (50 μ M) inhibited basal and evoked PG formation. Thin-layer chromatography (TLC) detection after incubation of the cells with [³H]arachidonic acid (1 μCi/ml, for 60 min) confirmed the results obtained by radioimmunoassay. Incubation of [³H]arachidonic acid labelled cells with inonophore or phospholipases, followed by lipid extraction and TLC, showed that A 23187 liberated [³H]arachidonic acid predominantly from phosphatidylethanolamine, whereas phospholipase A₂ and C reduced mainly the labelling of the phosphatidyl-inositol/-choline fraction. Potassium depolarization of the cells did not enhance prostanoid formation. Similarly, drugs with affinity to - or β-adrenoceptors, or to dopamine-, 5-hydroxytryptamine-, muscarine-, histamine-, glutamate-, aspartate-, GABA, adenosine- and opioid-receptors failed to stimulate prostanoid biosynthesis. Also compounds like angiotensin, bradykinin and thrombin were ineffective in this respect.

In conclusion, our results confirm that cultured astrocytes possess the complete pattern of enzymes necessary for prostanoid formation and hence might play a crucial role in brain prostanoid biosynthesis. Stimulation of prostanoid biosynthesis involves Ca²⁺-dependent activation of phospholipase A₂, cyclooxygenase reaction and further PG metabolism. However, the endogenous stimulus for enhanced prostanoid synthesis in the brain still has to be established.     

Temperature dependent binding of mebendazole to tubulin in benzimidazole-susceptible and -resistant strains of Trichostrongylus colubriformis and Caenorhabditis elegans.

The binding of [³H]mebendazole ([³H]MBZ) to tubulin in benzimidazole-susceptible (BZ-S) and benzimidazole-resistant (BZ-R) strains of Trichostrongylus colubriformis and Caenorhabditis elegans was examined in order to investigate the biochemical changes to tubulin that result in BZ resistance in parasitic and free-living nematodes. In both species the extent of [³H]MBZ binding to tubulin was significantly reduced in the BZ-R strain compared with the BZ-S strain. The decrease in [³H]MBZ binding in the BZ-R strain of each species was the result of a significant reduction in the amount of charcoal stable [³H]MBZ-tubulin complexes and was not related to a change in the association constant of the [³H]MBZ-tubulin interaction. [³H]MBZ binding to tubulin was temperature dependent, reaching maximum levels at 37°C in BZ-S T. colubriformis and 10°C in BZ-R T. colubriformis. Both the BZ-S and BZ-R strains of C. elegans displayed maximum [³H]MBZ binding at 4°C. Resistance ratios derived from the amount of [³H]MBZ binding in the BZ-S and BZ-R strains and in vitro development assays demonstrated that the temperature dependence and extent of drug binding was indicative of BZ resistance status and was species specific in the BZ-S isolates. These results indicate that biochemical differences exist in the binding of benzimidazole carbamates to tubulin in nematode species, and suggest that the susceptibility of the parasitic nematodes to the benzimidazole anthelmintics is the result of a unique high affinity and/or high capacity interaction ofbenzimidazole carbamates with tubulin.     

Characterization of [H]glutamate binding sites on frozen sections from rat adrenal

Some biochemical characteristics of [³H]glutamate (Glu) binding sites on frozen sections from the rat adrenal glands were studied. Adrenal frozen sections exhibited stereo-selective, saturable and temperature-dependent binding of [³H]Glu. An agonist for one of the subclasses of central Glu receptors, quisqualic acid (QA), elicited a significant inhibition of the binding, whereas neither N-methyl- -aspartic acid nor kainic acid, agonists for other subclasses of the receptors, had such a significant effect on the binding at the concentration range similar to QA. In vitro addition of sodium acetate (100 mM) resulted in a significant inhibition of [³H]Glu binding to frozen sections of the rat adrenal glands. It thus appears that there exist QA-sensitive binding sites of [³H]Glu in the rat adrenal glands which exhibit pharmacological characteristics distinctly different from those in the brain.     

人参连作根际土壤中酚酸物质对人参锈腐病菌的化感效应

采用高效液相色谱法对人参连作根际土壤中的酚酸物质进行了分离鉴定,检测发现人参根际土壤中含有没食子酸、水杨酸、3-苯基丙酸、苯甲酸和肉桂酸5种酚酸物质.采用外源添加法研究该5种酚酸物质对人参锈腐病菌的化感效应.结果表明: 5种酚酸对人参锈腐病菌的菌丝生长和孢子萌发都表现出高浓度抑制、低浓度促进的作用.没食子酸、水杨酸和苯甲酸在0.5 mmol·L^-1处理浓度下,3-苯基丙酸和肉桂酸在0.05 mmol·L^-1处理浓度下,均能够显著促进人参锈腐病菌菌丝生长和孢子萌发,并显著加重人参锈腐病病害严重度.     

Molecular pharmacology of the AMPA agonist, (S)-2-amino-3-(3-hydroxy-5-phenyl-4-isoxazolyl)propionic acid [(S)-APPA] and the AMPA antagonist, (R)-APPA

The heterocyclic analogue of (S)-glutamic acid, (S)-2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid [(S)-AMPA] is a potent and selective AMPA receptor agonist, whereas the enantiomeric compound, (R)-AMPA, is virtually inactive. We have previously characterized (RS)-2-amino-3-(3-hydroxy-5-phenyl-4-isoxazolyl)propionic acid [(RS)-APPA] as a partial AMPA receptor agonist showing about 60% of the efficacy of (RS)-AMPA. This partial agonism produced by (RS)-APPA is, however, only apparent, since resolution of (RS)-APPA has now been shown to provide the full AMPA receptor agonist, (S)-APPA, whereas (R)-APPA is a acid (non-NMDA) receptor antagonist showing preferential AMPA blocking effects. In agreement with classical theories for competitive interaction between agonists and antagonists, the efficacy of depolarizations produced by (S)-APPA in the rat cortical wedge preparation was shown to be progressively reduced with increasing molar ratios of (R)-APPA/(S)-APPA. These compounds and the competitive antagonists (RS)-2-amino-3-(3-carboxymethoxy-5-methyl-4-isoxazolyl)propionic acid [(RS)-AMOA], 6-cyano-7-nitroquinoxalin-2,3-dione (CNQX) and 6-nitro-7-sulfamoylbenzo(f)quinoxalin-2,3-dione (NBQX) were also tested in [³H]AMPA and [³H]CNQX binding systems, the latter ligand being used in the absence or presence of thiocyanate ions. On the basis of these studies it is suggested that (RS)-AMPA and the AMPA agonist (S)-APPA interact with a high-affinity receptor conformation, whereas the competitive antagonists (RS)-AMOA and (R)-APPA, derived from these agonists, preferentially bind to a low-affinity AMPA receptor conformation. The competitive antagonists, CNQX and NBQX which are structurally unrelated to (RS)-AMPA or (RS)-APPA, do not seem to discriminate between these two AMPA receptor conformations. The modified [³H]CNQX binding assay containing thiocyanate ions was shown to provide receptor affinity data for AMPA receptor agonists as well as antagonists, which correlate with the potencies of these compounds in the cortical wedge preparation. Using autoradiographic techniques, (S)- and (R)-APPA were shown to exhibit significantly different absolute potencies as inhibitors of [³H]AMPA binding in a number of regions of the rat brain.     

Synthesis of [2S-2-H]- and [2R-2-H]hexadecanoic acids

[2S-2-²H]- and [2R-2-²H]hexadecanoic acids were synthesized in overall yields of 59–67%. Methyl(2R)-2-hydroxyhexadecanoate, from the acid produced by Hansenula sydowiorum, was converted to the p-toluenesulphonate, reduced to trideutero alcohol with lithium aluminium deuteride and oxidized to [2S-2-²H]hexadecanoic acid. Methyl (2S)-2-chlorohexadecanoate, which was a by-product of tosylation and was also prepared by chlorinatioon of the hydroxy ester with thionyl chloride, on reduction and oxidation as before gave [2R-2-²H]-hexadecanoic acid. Intermediates were fully characterized, isotopic purity was 97% and optical purity was maintained throughout the syntheses. Attempts to reduce the tosyl or chloro groups, only, with sodium borodeuteride gave low yields probably due to preferential reduction of the ester group; 1,2-epoxyhexadecane was obtained from the tosylate and 2-chlorohexadecan-1-ol from the chloro ester.     

Aliphatic nitro-compounds in Astragalus canadensis

A reinvestigation of the alphatic nitro-compounds in Astragalus canadensis resulted in the identification of two new esters of glucose with 3-nitropropanoic acid and 5-oxotetrahydrofuran-3-acetic acid, together with six known conjugates of 3-nitropropanoic acid. ¹H and ¹³CNMR data are reported for the new compounds.     

Hinokiresinol is not a precursor of agatharesinol in the norlignan biosynthetic pathway in Japanese cedar

The biosynthetic relationship between the two norlignans agatharesinol and trans-hinokiresinol was investigated. Fresh sapwood sticks of Cryptomeria japonica were fed with stable isotope-labeled compounds, namely p-coumaryl alcohol-[9,9-²H], p-coumaryl alcohol-[9-¹⁸O] and trans-hinokiresinol-[1-²H], and then incubated under high-humidity for approximately 20 days, during which the two norlignans were produced simultaneously. While trans-hinokiresinol was strongly deuterium-labeled after feeding with p-coumaryl alcohol-[9,9-²H], agatharesinol was only lightly labeled after feeding with either p-coumaryl alcohol-[9,9-²H] or -[9-¹⁸O]. These results suggest that p-coumaryl alcohol, which is a precursor of hinokiresinol, is not involved in the biosynthesis of agatharesinol. Therefore, the norlignan carbon skeleton of agatharesinol must be framed from different types of phenylpropanoid monomers compared to those utilized by the trans-hinokiresinol pathway. The biosynthesis of these two norlignans seems to branch at an early stage, i.e., before the framing of the norlignan carbon skeleton. Furthermore, agatharesinol was not labeled with deuterium after feeding with ²H-labeled trans-hinokiresinol, which has the simplest norlignan structure. This result strongly supports the suggestion that the conversion of trans-hinokiresinol to agatharesinol is not part of the biosynthesis of norlignans and that early branching occurs instead.     

Lipoprotein-associated paf (LA-paf) was found in washed human platelets and monocyte/macrophage-like U937 cells

Beyond cholesterol, inflammatory ether phospholipids such as platelet-activating factor (paf) may play a role in atherogenesis. (1) We detected a paf-like compound (‘LA-paf’) associated with human serum lipoproteins, mainly in LDL but not with the lipoprotein-poor fraction. (2) LA-paf was also found in washed human platelets, from where it was partially released during platelet aggregation in response to paf (50 nM) or thrombin (1 U). In addition, resident monocyte/macrophage-like U937 cells carried huge amounts of LA-paf (41 ng per 10⁷ cells) and metabolized added [³H]paf to a labelled compound co-eluting with the retention time of LA-paf in standard HPLC. (3) Functionally, LA-paf had a comparable potency to synthetic paf, because LA-paf aggregated washed aspirin-treated platelets in a concentration-dependent manner. The specific paf receptor antagonist WEB2086 inhibited the platelet aggregation induced by three distinct LA-paf preparations as compared with synthetic paf with similar inhibitory concentrations (IC₅₀: 35.6 ± 12.8, 24.0 ± 4.0, 38.0 ± 15.8 nM for LA-paf, and 43.6 ± 6.5 nM for synthetic paf), indicating that LA-paf interacted with paf receptors. (4) However, LA-paf had a distinct retention time using high-pressure liquid chromatography (HPLC) as compared with synthetic paf. LA-paf eluted at 9–15 min and synthetic paf at 21–24 min. In addition, total and non-specific [³H]paf binding to intact washed human platelets was affected differently by the two unlabelled agonists: while LA-paf increased total and non-specific (but not specific) binding in a significant manner (P < 0.002 and P < 0.007) as LDL did (P < 0.006 and P < 0.03), synthetic paf decreased total binding (P < 0.03). Similarly, low-density lipoproteins (LDL) increased significantly the total [³H]paf binding. In contrast, paf did not affect specific [¹²⁵I]LDL binding to human fibroblasts. Our results show the presence of LA-paf in lipoproteins,     

Peripheral oxytocin treatment modulates central dopamine transmission in the mouse limbic structures

The effects of subcutaneous (s.c.) oxytocin treatment have been investigated on various parameters of dopaminergic neurotransmission in basal forebrain structures (nucleus olfactorius posterior + nucleus accumbens + septum) of the mouse. Acute oxytocin treatment failed to influence dopamine utilization in the basal forebrain. Following chronic injections of oxytocin (0.2 mg/kg) for 8 8 days, the neuropeptide decreased dopamine utilization. Neither in vivo nor in vitro oxytocin treatment was capable of influencing the in vitro uptake of [³H]dopamine in basal forebrain slices. The spontaneous release of [³H]dopamine (in the presence of 4.2 mM K⁺) from basal forebrain tissue slices was not affected by in vitro or acute or chronic in vivo oxytocin treatment. The stimulated release of [³H]dopamine (in the presence of 30 mM K⁺) was significantly inhibited by chronic in vivo oxytocin administration. Chronic oxytocin treatment decreased the B_max value of [³H]spiroperidol binding in the basal forebrain. The dissociation constant (K_d) of [³H]spiroperidol binding was not influenced by oxytocin. The data indicate that peripheral oxytocin treatment is capable of modifying dopaminergic neurotransmission in mouse basal forebrain regions.     

Synthesis, radiolabeling and receptor binding of [3H][(1S,2R)ACPC2]endomorphin-2

Previously, we have shown that substitution of Pro² for cis-2-aminocyclopentanecarboxylic acid, ACPC in endomorphin-2 results in an analogue with greatly augmented proteolytic stability, high μ-opioid receptor affinity and selectivity. We now report the synthesis and biochemical characterization of [³H][(1S,2R)ACPC²]endomorphin-2 with a specific activity of 1.41 TBq/mmol (38.17 Ci/mmol). Specific binding of [³H][(1S,2R)ACPC²]endomorphin-2 was saturable and of high affinity with an equilibrium dissociation constant, K_d = 1.80 ± 0.21 nM and receptor density, B_max = 345 ± 27 fmol × mg protein⁻¹ at 25 °C in rat brain membranes. Similar affinity values were obtained in kinetic and displacement assays. Both Na⁺ and Gpp(NH)p decreased the affinity proving the agonist character of the radioligand. [³H][(1S,2R)ACPC²]endomorphin-2 retained the μ-specificity of the parent peptide. The new radioligand will be a useful tool to map the topographical requirements of μ-opioid peptide binding due to its high affinity, selectivity and enzymatic stability.     

3-Mercaptopropionic acid administration increases the affinity of [H]Quinuclidinyl benzilate binding to membranes of the striatum and cerebellum

It is known that quinuclidinyl benzilate (QNB) binds specifically and with high affinity to the cholinergic muscarinic receptor and that behaves as a potent antagonist of this receptor.

We have analysed -[³H]QNB binding to rat CNS membranes after the administration of the convulsant 3-mercaptopropionic acid (MP) (150 mg·kg⁻¹, i.p.). The studies were done in rats killed at two stages: during and after seizures. No changes in [³H]QNB binding to hippocampus and cerebral cortex membranes were found. [³H]QNB binding increased about 40 and 80% in striatum and cerebellum membranes, respectively. The changes were observed both in seizure and postseizures states. The study was extended to the assay of [³H]QNB binding kinetic constants in the anatomical areas modified by the convulsant. The analysis of the saturation curves indicated an increase in the binding affinity but no change in the number of binding sites. Hill number values were near the unit suggesting a non-cooperative interaction between the ligand and the receptor, and the labelling of a homogeneous population of receptor sites.

The results suggest the participation of some cholinergic pathways in the development and maintenance of MP-induced seizures.