The dopamine D2 agonists RU 24213 and RU 24926 are also kappa-opioid receptor antagonists.

The di-(phenethyl)-amine derivatives, RU 24213 and RU 24926 are widely used as selective dopamine D2-receptor agonists. Binding studies now show that they also have affinity for the kappa-opioid receptor. Their affinity is not greatly reduced in the presence of NaCl/GTP, suggesting an antagonist action. This is confirmed for RU 24926, the more active of the two, using the field-stimulated rabbit vas-deferens. With respect to the mu-receptor, RU 24926 shows low binding affinity and also an antagonist effect. These results demonstrate kappa- as well as mu-antagonist activity in a novel chemical series. Moreover, since the kappa-receptor may regulate dopamine release, the results of experiments using these compounds as dopamine D2 agonists should be interpreted with caution.

GABA release from mouse striatal neurons in primary culture as a test for the functional activity of N-methyl-D-aspartate (NMDA) antagonists.

N-Methyl-D-aspartate (NMDA)-induced release of [3H]GABA from mouse striatal neurons in primary culture has been evaluated as a screening method for demonstrating the functional activity of potential NMDA antagonists with respect to a cellular response. Antagonists were chosen for their specificity towards each of the three principal binding sites which have been characterised on the NMDA-receptor complex: the glutamate site, the ion-channel and in particular the glycine regulatory site where several novel halogenated derivatives of kynurenic acid have been tested. All the compounds were effective in blocking [3H]GABA release and their activity was related to their potency in displacing the binding of specific ligands for each of the three sites in rat cortex membrane preparations. This was confirmed by a correlation curve for the series of kynurenate derivatives (correlation coefficient r = 0.96). The specificity of these latter compounds for the glycine site was demonstrated by the addition of excess glycine which totally reversed their inhibition but not that of antagonists acting at the glutamate or ion-channel sites. Within the kynurenate series the 5,7-dichloro derivative was shown to be more active than the 7-chloro derivative, the most active glycine antagonist previously described. These results show that this is a simple and reliable system for demonstrating a functional effect of NMDA antagonists.

Lesion of noradrenergic neurones with DSP4 does not modify benzodiazepine receptor binding in cortex and hippocampus of rat.

The effect of lesioning noradrenergic pathways on the benzodiazepine receptor has been studied using a novel neurotoxic agent, N(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4) which has better selectivity than the classical 6-hydroxydopamine (6-OHDA) towards noradrenergic neurones, and which has the added advantage of being injected systemically rather than intracerebrally. Three different radioligands were used: an agonist, [3H]flunitrazepam, an antagonist, [3H]Ro 15-1788, and a partial agonist [3H]RU 43028. Binding was measured using membrane homogenates from the cortex and hippocampus of the rat, two regions of the brain which receive an extensive noradrenergic innervation. In contrast to previous reports of a decrease in the binding of [3H]flunitrazepam following lesioning with 6-OHDA, no significant change was observed in either the affinity (KD) or the number of sites (Bmax) of any one of these ligands after lesioning with DSP4. While the reasons for this discrepancy are not clear, these results do not confirm that destruction of noradrenergic afferents to the cortex or hippocampus leads to any modification of the benzodiazepine receptor as demonstrated by ligand binding.

Carcinogenic potential of hydrotreated petroleum aromatic extracts.

Five experimental petroleum extracts were produced from luboil distillates derived from Middle East paraffinic crude by solvent extraction and severe hydrotreatment. The polycyclic aromatic content (PCA) of the extracts was determined by dimethyl sulphoxide extraction and ranged from 3.7-9.2% w/w. The five extracts were evaluated for their potential to induce cutaneous and systemic neoplasia in female mice derived from Carworth Farm No 1 strain (CF1). The test substances were applied undiluted (0.2 ml per application) to the shorn dorsal skin twice weekly for up to 78 weeks, with 48 mice in each treatment group and 96 in the untreated control group; two further groups, each of 48 mice, were similarly treated either with a non-hydrotreated commercial aromatic extract (PCA content, 19.7% w/v) or with a low dose of benzo(a)pyrene (12.5 micrograms/ml acetone). The mice were housed individually in polypropylene cages in specified pathogen free conditions. The incidence of cutaneous and systemic tumours was determined from histological analysis of haematoxylin and eosin stained tissue sections. The results were correlated with the PCA content of the extracts and compared with those from female mice exposed to a non-hydrotreated commercial aromatic extract. Four of the hydrotreated extracts were carcinogenic for murine skin; the two products with the lower PCA contents were less carcinogenic than the products with the higher PCA contents and all were less carcinogenic than the commercial extract. One extract with the lowest PCA content was non-carcinogenic. Thus refining by severe hydrotreatment was an effective method of reducing the carcinogenic potential of petroleum aromatic extracts. Although other physicochemical properties may influence the biological activity of oil products, the PCA content determined by dimethyl sulphoxide extraction may be a useful indicator of the potential of oil products to induce cutaneous tumours in experimental animals. There was no evidence that the commercial or hydrotreated extracts increased the incidence of systemic neoplasms when applied twice weekly to the dorsal skin.

The carcinogenic potential of twelve refined mineral oils following long-term topical application.

Twelve mineral oils, originating from naphthenic and paraffinic stocks and variously refined, were evaluated for their potential to induce cutaneous neoplasia in female CF1 mice. The oils were applied to the shorn dorsal skin for up to 78 weeks, using several different treatment regimes. The sole acid/earth refined naphthenic spindle oil was a moderately potent cutaneous carcinogen. By comparison, the 11 oils, processed by other refining routes, were less carcinogenic or non-carcinogenic to murine skin. Two of the 11 oils were weak cutaneous carcinogens viz, a naphthenic spindle oil refined only by mild hydrotreatment and a paraffinic spindle oil refined by mild solvent extraction and 'Ferrofining'. All 9 remaining oils had been solvent-extracted as part of the secondary refining process; none induced malignant tumours, although solitary benign tumours of the treated site were recorded after exposure to 3 oils. The cutaneous carcinogenic potential of the test oils did not correlate well with their potential to induce epidermal hyperplasia at the treated site. Consequently, hyperplasia caused after short term exposure is of little value for distinguishing between carcinogenic and non-carcinogenic oils.

The carcinogenic response in mice to the topical application of propane sultone to the skin.

The carcinogenic effects of limited and repeated skin applications of propane sultone were investigated in three strains of mice, CF1, C3H and CBah (a hairless strain). Propane sultone was shown to be carcinogenic when given as a single application of a 25% w/v solution in toluene and also following twice weekly application of a 2.5% w/v solution for up to 58 weeks. More limited exposure to 2.5% w/v solutions of propane sultone resulted in a few skin tumours, although the incidences were not statistically significant. Most neoplasms were papillomas or carcinomas, although a small number of mesenchymal tumours of dermal origin also developed. No skin neoplasms were found in any control mice. The skin application of propane sultone was associated with a statistically significant increase in the incidence of systematic neoplasia in CFl and C3H mice. The exposed CFl mice had a higher incidence of neoplasms of lymphoreticular and lung origin, while female C3H mice showed a higher incidence of mammary gland and uterine tumours. In mice exposed to beta-propiolactone as a positive control, neoplasms developed at the site of application but, there was no evidence of increased systemic neoplasis in contrast to the findings with ptopane sultone.

Dichlorvos -- a 2-year inhalation carcinogenesis study in rats.

To determine the effects of dichlorvos vapour on the tumour incidence in rats, 5 week old Carworth Farm E strain rats weighing between 94 and 150 g were exposed to 0, 0.05, 0.5 and 5.0 mg/m3 in a 2-year inhalation study. The growth rate of all treated rats was depressed, particularly in the males. There was increased survival of the rats exposed to 5 mg/m3. There were no consistent differences in food intakes, organ weights, haematological or blood chemistry estimations, except in cholinesterase activites, amongst the various groups of rats. No compound-related differences were seen in acetylcholine and choline estimations carried out on a small number of female rats' brain tissues after two years' exposure. There were no gross or microscopical compound-related changes in the rats' tissues. Ultrastructural examination of the respiratory tissues of the rats from the control and 5 mg/m3 group showed no changes attributable to dichlorvos. The results of a relative risk analysis of the tumour data showed that no dose-related increase in tumour risk was established for rats of either sex. These data confirm the results of earlier st.udies supporting the safety of insecticidal uses of dichlorvos.

Comparative action of fenfluramine on the uptake and release of serotonin and dopamine.

The anorectic agent, fenfluramine, proves to be a good inhibitor of serotonin uptake in vitro, in synaptosomes from rat whole brain (IC50 = 8.5 +/- 0.6 X 10(-7) M). After administration in vivo, its inhibitory activity in vitro equals that of chlorimipramine and in contrast to the latter, its effect is of long duration. Fenfluramine is also effective in promoting the release of serotonin from pre-loaded synaptosomes. In comparison, the structurally related compound, amphetamine, has little activity with respect to these serotonin mechanisms. It is, however, active both in inhibiting the uptake of dopamine and in promoting its release, whereas fenfluramine is inactive. The implication of these mechanisms in the serotonin-depleting capacity as well as in the anorectic activity of fenfluramine is discussed.

Studies on the biosynthesis of the ergosterol side chain.

1. A convenient synthesis of 24-methylene[23,25-(3)H(3)]dihydrolanosterol is described. 2. A general anaerobic-aerobic method for the incorporation of sterols into whole yeast cells is also described and illustrated by experiments with (3)H-labelled lanosterol. 3. The method was used to convert labelled 24-methylene-dihydrolanosterol into ergosterol, in good yield, by Saccharomyces cerevisiae. 4. Degradation of the biosynthetic ergosterol provided confirmation of the conversion, which supports the proposed mechanism for the biosynthesis of the ergosterol side chain. 5. Mechanisms for the further conversion of the 24-methylene side chain into the ergosterol side chain are discussed and it was shown that a compound, [3alpha-(3)H(1)]-ergost-7-en-3beta-ol, with a fully saturated side chain, can also be efficiently incorporated into ergosterol. 6. This result was confirmed by a procedure involving formation of the 5,8-epidioxide and subsequently the 5,8-epidioxy-22,23-epoxide of the biosynthetic ergosterol.

The introduction of the C-22-C-23 ethylenic linkage in ergosterol biosynthesis.

Methods for the chemical synthesis of [23-(3)H(2)]lanosterol, [23,25-(3)H(3)]24-methyldihydrolanosterol and [24,28-(3)H(2)]24-methyldihydrolanosterol are described. It is shown that, in the biosynthesis of ergosterol from [26,27-(14)C(2),23-(3)H(2)]lanosterol by the whole cells of Saccharomyces cerevisiae, one of the original C-23 hydrogen atoms is lost and the other is retained at C-23 of ergosterol. It is also shown that 24-methyldihydrolanosterol is converted into ergosterol in good yield and without prior conversion into a 24-methylene derivative. On the basis of these results possible pathways for the formation of the ergosterol side chain from a 24-methylene side chain are discussed.

The transfer of hydrogen from C-24 to C-25 in ergosterol biosynthesis.

1. A convenient synthesis of 3-hydroxytrisnorlanost-8-en-24-al and its conversion into [24-(3)H]lanosterol and [26,27-(14)C(2)]lanosterol is described. 2. A method for the efficient incorporation of lanosterol into ergosterol by the whole cells of Saccharomyces cerevisiae is also described. 3. It is shown that in the biosynthesis of ergosterol from doubly labelled lanosterol the C-24 hydrogen atom of lanosterol is retained in ergosterol. 4. On the basis of unambiguous degradations it is shown that the C-alkylation step in ergosterol biosynthesis is accompanied by the migration of a hydrogen atom from C-24 to C-25. 5. The mechanism for the biosynthesis of the ergosterol side chain is presented. 6. Mechanisms of other C-alkylation reactions are also discussed.