Ethanol-induced conditioned place preference, but not aversion, is blocked by treatment with d-penicillamine, an inactivation agent for acetaldehyde

Rationale There is evidence to suggest that acetaldehyde is involved in the control of ethanol-seeking behavior and reward. d-penicillamine, a thiol amino acid, is a highly selective agent for the inactivation of acetaldehyde. Previous studies from our laboratory have demonstrated that d-penicillamine prevents both behavioral stimulation induced by ethanol and acetaldehyde-produced locomotor depression in mice. Objectives The contribution of ethanol-derived acetaldehyde to the affective effects of ethanol (preference and aversion) was assessed using an unbiased place conditioning design. Methods Male mice received four pairings of a distinctive floor stimulus (CS+: GRID+ or HOLE+) with injections of saline and ethanol (2 g/kg) given before (preference) or after (aversion) the 5-min exposure to the place conditioning apparatus. A different floor stimulus (CS−: GRID− or HOLE−), associated with saline-saline injections on alternate days, was presented. For a different group of animals, the pairings with the CS+ were associated with saline and ethanol injections, but on alternate days, they received d-penicillamine (50 or 75 mg/kg) and ethanol injections paired with the CS−floor stimulus. A 60-min preference test was carried out 24 h after the last conditioning trial. A similar procedure was followed to test the effect of d-penicillamine on morphine (16 mg/kg) and cocaine-induced (20 mg/kg) conditioned place preference (CPP). Results CPP and conditioned place aversion (CPA) were observed for ethanol, but d-penicillamine only blocked CPP. d-penicillamine, by itself, did not produce either rewarding or aversive effects. CPP observed for morphine and cocaine was unaffected by d-penicillamine pretreatment. Conclusions The results of the present study suggest that the selective inactivation of acetaldehyde blocked the rewarding, but not aversive, effects of ethanol and support the role of this ethanol metabolite in the affective properties of ethanol.     

Biliverdin reductase: Characterization in the rat kidney and the inhibition of activity by mercuric chloride

The effects of metal ions on the activities of biliverdin reductase in the rat kidney and liver were examined; the pH optimum and the cofactor requirement for the enzyme activity in the kidney were also studied. The reduction of biliverdin IXα by biliverdin reductase in the rat kidney cytosol fraction could be supported by NADPH and NADH. The activity was optimal around pH 8.7 when NADPH was the cofactor. The activity with NADH was undetectable at this pH. NADH-dependent biliverdin reductase was optimal at pH 7.0, where the NADPH-dependent activity was negligible. Biliverdin reductase activity was not inducible in the kidney or liver in response to treatment of rats with metal ions—Co^2?, Ni^2?, Pb²⁺, Sn^2?, Zn^2?, Cd²⁺, and Cu^2- or sodium selenite. Rather, both NADPH- and NADH-dependent activities in the kidney were decreased markedly in a time- and dose-related manner following the adminstration of HgCl₂ (10–30 μmoles/kg, 24 hr). The pretreatment of rats (30 min) with sodium selenite (5 μmoles/kg) effectively blocked the Hg²⁺ (20 μmoles/kg. 24 hr) inhibition of the kidney cytosol biliverdin reductase activity. Similarly, in vitro Hg²⁺ was an effective inhibitor of the kidney biliverdin reductase. In addition, highly purified biliverdin reductase also was extremely sensitive to Hg²⁺ and the thiol reagent, 5, 5'-dithiobis-(2-nitrobenzoic acid). The inhibition of purified reductase by 5,5'-dithiobis-(2-nitrobenzoic acid), but not by Hg^2-, could be reversed by dithiothreitol.     

Heavy metal modulation of lymphocyte activities. 1. In vitro effects of heavy metals on primary humoral immune responses

The ability of heavy metals to modulate in vitro primary humoral immune responses has been investigated. The relative immunosuppresive activities of the heavy metals tested were Hg²⁺ > Cu²⁺ > Mn²⁺ > Co²⁺; Cd²⁺ > Cr³⁺; Sn²⁺; Zn²⁺. Fe²⁺ had no significant effect on the development of SRBC-specific plaque-forming cells (PFC), and Pb²⁺ and Ni²⁺ enhanced the PFC response. The immunosuppressive activities of the heavy metals usually correlated with their toxicity and their inhibition of lymphocyte proliferation. The immunopotentiating metals, Pb²⁺ and Ni²⁺, alone induced lymphocyte proliferation, and they increased the proliferative response induced by LPS and 2-ME but not Con A and PHA. Although Pb²⁺ and Ni²⁺ were the only two heavy metals tested which enhanced lymphocyte reactivity, they did not appear to function via similar mechanisms. Pb²⁺ appeared to enhance the development of PFC by directly interacting with lymphocytes and altering their activity since lymphocyte preincubation with Pb²⁺ was sufficient for enhancement; whereas, lymphocyte preincubations with Ni²⁺ did not induce enhancement. Pb²⁺ did not appear to alter the immunogenicity of the antigens employed; however, pretreatment of SRBC with Ni²⁺ eliminated their ability to stimulate a SRBC-specific response in vitro. Inhibition of the PFC response by the chelator EGTA could be reversed by Ca²⁺ or Ni²⁺, but not by Cu²⁺, Hg²⁺, Pb²⁺, or Zn²⁺. The stimulatory activity of Pb²⁺ and Ni²⁺ could not be accounted for by significant alteration of the antigenicity of the lymphocytes, because heavy metal preincubated syngeneic cells were not stimulatory for untreated syngeneic cells. On the other hand, Pb²⁺ and Ni²⁺ did slightly enhance the mixed lymphocyte culture (MLC) response to allogeneic cells; Hg²⁺ inhibited the MLC response.     

Studies with tryptophan metabolites in vitro. effect of zinc,manganese, copper and cobalt ions on kynurenine hydrolase and kynurenine aminotransferase in normal mouse liver

The possible mechanisms by which increasing concentrations of Zn²⁺, Mn²⁺, Cu²⁺, or Co²⁺ may affect the vitamin B₆-dependent enzymes kynurenine hydrolase and kynurenine aminotransferase. were studied in normal mouse liver homogenates. It was found that Zn²⁺ inhibited kynurenine hydrolase. whereas Mn²⁺ activated this enzyme, but both Zn²⁺ and Mn²⁺ activated kynurenine aminotransferase. Co²⁺ and Cu²⁺ inhibited both enzymes. This inhibition is attributed to the blocking and inactivation of the -SH groups of these enzymes and may be due to the adequate sequence of the -SH groups in both enzymes for Cu²⁺ or Co²⁺ action. This is in contrast to the inadequate sequence of these groups in kynurenine aminotransferase for Zn²⁺ action. The decreasing order by which these metal ions inhibit (a) kynurenine hydrolase is Cu²⁺ > Co²⁺ > Zn²⁺, and (b) kynurenine aminotransferase is Cu²⁺ > Co²⁺. The decreasing order of the per cent activation of the aminotransferase enzyme is Mn²⁺ > Zn²⁺. These decreasing orders fall into a more reasonable order approximating the order of complex stability of these metal ions.     

Proline Prodrug of Melphalan Targeted to Prolidase,a Prodrug Activating Enzyme Overexpressed in Melanoma

Purpose To determine the bioactivation and uptake of prolidase-targeted proline prodrugs of melphalan in six cancer cell lines with variable prolidase expression and to evaluate prolidase-dependence of prodrug cytotoxicity in the cell lines compared to that of the parent drug, melphalan. Materials and Methods Hydrolysis, cell uptake, and cell proliferation studies of melphalan and the L- and D-proline prodrugs of melphalan, prophalan-L and prophalan-D, respectively, were conducted in the cancer cell lines using established procedures. Results The bioactivation of prophalan-L in the cancer cell lines exhibited high correlation with their prolidase expression levels (r ² = 0.86). There were no significant differences in uptake of melphalan and its prodrugs. The cytotoxicity of prophalan-L (GI₅₀) in cancer cells also showed high correlation with prolidase expression (r ² = 0.88), while prophalan-D was ineffective at comparable concentrations. A prolidase targeting index (ratio of melphalan to prophalan-L cytotoxicity normalized to their uptake) was computed and showed high correlation with prolidase expression (r ² = 0.82). Conclusions The data corroborates the specificity of prophalan-L activation by prolidase as well as prolidase-targeted cytotoxicity of prophalan-L in cancer cell lines. Hence, prophalan-L, a stable prodrug of melphalan, exhibits potential for efficiently targeting melanoma with reduced systemic toxicity.     

Studies on the reaction catalyzed by transport (Na, K) adenosine triphosphatase. I. Effects of divalent metals

The effects of divalent metals (Cu²⁺, Zn²⁺, Fe²⁺ and Pb²⁺) on a microsomal preparation of NaK-ATPase (ATP phosphohydrolase, EC 3.6.1.3) from beef cerebral cortex were studied. These metals are all potent inhibitors of the enzyme with I₅₀ values of 1 μM for Cu²⁺ and Zn²⁺, 3 μM for Fe²⁺ and 20 μM for Pb²⁺. Kinetic studies examining the effect of low concentrations of divalent metals on K_m and V for MgATP are reported. The results indicate that Fe²⁺ and Pb²⁺ are competitive inhibitors of NaK-ATPase with K_i values of 1.60 μM and 1.90 μM respectively. Cu²⁺ and Zn²⁺ are noncompetitive inhibitors of NaK-ATPase with K_i values of 1.18 μM and 3.48 μM respectively.     

Effect of Heavy Metals on Dopamine,Noradrenaline and Serotonin Uptake and Release in Rat Brain Synaptosomes

Abstract: The effects of some heavy metals on the initial high affinity uptake and spontaneous release of tritiated dopamine (³H-DA), noradrenaline (³H-NA) and 5-hydroxytryptamine (³H-5-HT) were studied in vitro in rat striatal, cortical and hypothalamic synaptosomes, respectively. As uptake inhibitors, metals were quite inactive in these conditions. At 10 μM Cu²⁺ was most potent, inhibiting ³H-DA and ³H-5-HT uptake nearly completely while inhibition of ³H-NA uptake varied. ³H-DA uptake was in addition inhibited slightly by Zn²⁺, sometimes by Sn²⁺ but never by Co²⁺, Hg²⁺ or Mn²⁺. Unexpectedly Pb²⁺ and Cd²⁺ tended to increase the synaptosomal ³H-DA uptake. ³H-5-HT uptake was affected least while that of ³H-NA showed some diversity. Zn²⁺, Pb²⁺ and Sn²⁺ induced inhibition of ³H-NA uptake possibly by direct interference with ³H-NA. As to the spontaneous release of tritiated amines during short incubation from preloaded synaptosomes, Cd²⁺ decreased that of ³H-DA at high concentrations but Hg²⁺, Pb²⁺, Sn²⁺ and Zn²⁺ were ineffective. The results suggest that in vitro the uptake and the release of ³H-DA are more affected than those of other amines. The inhibitory mechanisms of monoamine uptake may include both direct effects on synaptosomes and indirect ones by interference with the amines themselves.     

2,9-Dimethyl-1,10-phenanthroline (neocuproine): a potent,copper-dependent cytotoxin with anti-tumor activity

2,9-Dimethyl-1,10-phenanthroline (2,9-DMP), a copper-specific chelator, was a potent cytotoxin against L1210 cells in vitro; its activity was dependent upon available Cu²⁺ in the medium. Other divalent ions, Fe²⁺ and Zn²⁺, were ineffective as promotors of growth inhibition. As the copper chelate. a 4μM solution produced a 4 log kill after a 1-hr incubation. This was in marked contrast to 1.10-phenanthroline, whose inhibition of cell growth was overcome by added Cu²⁺, Fe^2- and Zn^2-. Cellular uptake of labeled 2,9-dimethyl-1,10-phenanthroline also required added Cu²⁺ in the medium. This transport was energy dependent, and the drug was concentrated over 200-fold by the cells. In preliminary evaluations, copper-2,9-DMP showed significant chemotherapeutic activity against the P388 murine lymphoma in vivo.     

Morphine,d-Pen2,d-Pen5 enkephalin and U50,488H differentially affect the locomotor activity and behaviours induced by quinpirole in guinea-pigs

The effects of morphined-Pen²,d-Pen⁵ enkephalin (DPDPE) and U50,488H on the behavioural syndrome elicited by the dopamine (DA) D-2 agonist quinpirole, were investigated. Morphine (1, 5 and 15 mg/kg SC) and morphine administered intracerebroventricularly (ICV) (2×5 µl, 10^–3 M; total dose = 10 nmol) produced piloerection and sedation. DPDPE-ICV (2×5 µl and 2×10 µl, 10–³ M; total doses = 10 and 20 nmol) produced piloerection and sedation similar to morphine. U50,488H (1 mg/kg SC) induced locomotor activity and some stereotyped behaviour, whereas U50,488H (5 and 10 mg/kg SC) induced muscle rigidity and dystonic-like movements. The locomotor and behavioural response elicited by quinpirole (3 mg/kg IP) was attenuated in guinea-pigs pretreated with morphine (1, 5 and 15 mg/kg SC), morphine-ICV (2 × 5 µl, 10–³ M), and DPDPE-ICV (2 × 5 µl and 2 × 10 µl, 10^–3 M). These effects were reversed by naloxone (15 mg/kg SC). U50,488H (1 mg/kg SC) increased the quinpirole-induced locomotor activity, whereas U50,488H (5 and 10 mg/kg SC) decreased the locomotor activity and stereotyped behaviours produced by quinpirole. These results indicate that the gross behavioural effects of mu, delta and kappa opioids differ in guinea-pigs compared to other rodent species, and suggest differential involvement of these opioid receptor subtypes with DA D-2 receptor-mediated activity.     

Comparison of some biochemical effects of teratogenic doses of mercuric mercury and cadmium in the pregnant rat

Mercuric mercury (Hg²⁺), like cadmium (Cd²⁺), interferes with the transport of certain essential metals to the conceptus in the pregnant Wistar rat and, at 48 h after the IV injection of a teratogenic dose (0.79 mg Hg²⁺/kg body weight) on day 12 of gestation, the foetal concentrations of Zn²⁺, Cu²⁺ and Fe³⁺, but not of Mg²⁺, are reduced significantly. Both Hg²⁺ and Cd²⁺, at teratogenic dose levels, inhibit the placental and foetal uptake of ⁶⁵Zn²⁺ and ⁶⁷Cu²⁺, but possibly by different mechanisms. In addition, the effects of Hg²⁺, at different times after dosing, on the uptake of these labelled tracers and of ⁵⁹Fe³⁺, administered as 15-min pulses, do not parallel the changes in the placental and foetal concentrations and contents of the endogenous, stable metallic ions. The teratogenic dose of Hg²⁺ inhibits the placental and foetal uptake of L-[4,5-³H]-leucine, but not the incorporation of the labelled amino acid into foetal protein. In contrast, the corresponding dose of Cd²⁺ inhibits both leucine uptake and protein synthesis in the placenta and foetus. Similarly, Cd²⁺ inhibits the uptake of [2-¹⁴C]-thymidine and its incorporation into foetal DNA, whereas Hg²⁺ reduces the placental and foetal uptake, but has little or no effect on the utilization of the nucleoside. Since both Cd²⁺ and Hg²⁺ reduce the foetal uptake of ⁶⁵Zn and the foetal concentration of Zn, but only Cd²⁺ interferes with DNA synthesis, it is unlikely that the inhibition of the metabolism of thymidine can be attributed to reduction in thymidine kinase activity in consequence of foetal Zn deficiency. It is concluded that the small amount of Cd²⁺ that is taken up by the foetus has a direct effect on the synthesis of DNA and protein, whereas Hg²⁺ primarily affects placental transport processes.     

Enhancement of blood-brain barrier permeability to sodium fluorescein by stimulation of µ opioid receptors in mice

Summary The effects of opioids on the permeability of the blood-brain barrier (BBB) were examined in mice with sodium fluorescein as an indicator of the permeability. The brain was perfused with saline 30 min after injection of sodium fluorescein (40 mg/kg, i. v.) and examined by fluorometry. Morphine hydrochloride (0.3–10 mg/kg, s. c.) markedly increased the brain level of sodium fluorescein dose-dependently without influencing the plasma level, when administered 20 min before sodium fluorescein injection. Intracerebroventricularly (i. c. v.) injected morphine hydrochloride (0.5 and 1.0 Erg) increased the brain sodium fluorescein level. Buprenorphine (0.1 and 0.5 mg/kg, s. c.) was also effective. However, pentazocine, ethylketazocine, U-50488H and SKF-10047 had no significant influence. The i.c.v. administration of [D-Ala², McPhe⁴, Gly(ol)⁵]enkephalin (0.1 g) and [D-Ala², D-Leu⁵]enkephalin (0.5 g) but not of [D-Thr², Leu⁵]enkephalin-Thr increased the brain level of sodium fluorescein significantly. A small dose of naloxone (i. p.) significantly inhibited the effects of morphine, buprenorphine, [D-Ala², McPhe⁴, Gly(ol)⁵]enkephalin and [D-Ala³, D-Leu⁵]enkephalin. ICI-174864 co-administered i. c. v. with [D-Ala², D-Leu⁵]enkephalin was ineffective in antagonizing the effect of the latter. These findings suggest that the stimulation of µ opioid receptors results in an increase in BBB permeability to sodium fluorescein. Send offprint requests to K. Saeki     

Correlation of the physicochemical properties of metal ions with their activation and inhibition of human erythrocytic δ-aminolevulinic acid dehydratase (ALAD) in vitro

Using normal adult human whole blood hemolysates this study determined, in a dose-response fashion, the in vitro effects of Na⁺, Mg²⁺, Al³⁺, Mn²⁺, Cu²⁺, Ag⁺, Zn²⁺, Cd²⁺, Hg²⁺, Ga³⁺, In³⁺, Sn²⁺, Sn⁴⁺, and Pb²⁺ on normal erythrocytic ALAD. The effects of these 14 metal ions on erythrocytic ALAD 50% inhibited by Pb²⁺ were also determined as was the ability of a maximum stimulatory concentration of Zn²⁺ to prevent or reverse the effects of these metal ions on erythrocytic ALAD. The effects of these metal ions were then classified in terms of their oxidation state, characteristic coordination number, coordination geometry, and hard and soft Lewis acid characteristics in order to determine the physical and chemical properties associated with the ability of a metal ion to activate or inhibit erythrocytic ALAD and whether these properties are unique to a single metal ion. Preincubation studies established Zn²⁺ to be the only metal ion to both activate erythrocytic ALAD and to prevent or reverse the Pb²⁺-induced inhibition of erythrocytic ALAD in vitro even after prolonged contact with the enzyme. Therefore future investigations of the use of nontoxic salts of Zn²⁺ as a prophylactic agent or therapeutic adjunct in the prevention or treatment of lead poisoning with its possibly toxic accumulation of ALA are proposed.     

Biochemical and pharmacological properties of a peripherally acting catechol-O-methyltransferase inhibitor entacapone

Summary Entacapone, OR-611, was found to be a potent peripherally acting inhibitor of catechol-O-methyl-transferase (COMT). IC₅₀ values of 10 nmol/1 and 160 nmol/1 were obtained for rat duodenum and liver-soluble COMT, respectively. There were no effects on other catecholamine metabolizing enzymes. Entacapone showed reversible, tight-binding type of inhibition of soluble rat liver COMT with a K; value of 14 nmol/1 and it also caused 50% inhibition of rat duodenal, erythrocyte, liver and striatal COMT activity 1 h after oral dosing with 1.1, 5.4, 6.7 and 24.2 mg/kg, respectively. However, penetration of entacapone into the brain was poor, since the formation of homovanillic acid (HVA), the O-methyl metabolite of dopamine in the striatum, was not reduced, even after the highest dose of 30 mg/kg.In rat blood serum, the concentration of 3-0-methyldopa (3OMD), the O-methylated product of l-dopa, was reduced in a dose-dependent manner, and the concentration of l-dopa was increased after the administration of entacapone (3 - 30 mg/kg p. o.) together with l-dopa + carbidopa. These changes were reflected, in the striatum, by a significant rise in the dopamine concentration and a reduction in the 30MD concentration.Consequently, when entacapone was added to the treatment with l-dopa + carbidopa, the dose of l-dopa could be lowered from 50 mg/kg to 15 mg/kg in order to produce the same striatal dopamine concentrations as with 50 + 50 mg/kg of l-dopa + carbidopa alone.Correspondence to E. Nissinen at the above address     

EFFECTS OF ENDURANCE TRAINING ON SUPEROXIDE DISMUTASE ACTIVITY,CONTENT AND mRNA EXPRESSION IN RAT MUSCLE#

1. The purpose of the present study was to investigate the changes in superoxide dismutase (SOD) isoenzyme (Mn²⁺-SOD and Cu²⁺, Zn²⁺-SOD) activities, contents and mRNA expressions in rat skeletal muscle during endurance training and a single bout of exercise. 2. Thirty-eight male Wistar rats were divided into untrained (U) and trained (T) groups. The T group rats were treadmill-trained for 9 weeks. The activity, content and mRNA expression of Mn²⁺-SOD and Cu²⁺, Zn²⁺-SOD were determined in the soleus muscle of each rat. 3. Mn²⁺-SOD activity and content in the T group were significantly higher than in the U group, both at rest (22 and 21%, respectively) and after exercise (24 and 46%, respectively), while a single bout of exercise affected neither the activity nor content of Mn²⁺-SOD in either group. 4. The content of Cu²⁺, Zn²⁺-SOD in both groups was not different at rest and after exercise, although its activity at rest was significantly higher in the T group than in the U group (by 29%). 5. After exercise, the expression of Mn²⁺-SOD mRNA was markedly attenuated only in the U group (49%); the expression of Cu²⁺, Zn²⁺-SOD mRNA was not influenced by exercise. 6. Our results suggest that adequate endurance training increases both the activity and content of Mn²⁺-SOD and that untrained rats are rather susceptible to oxidative stress during physical exercise. It thus appears that Mn²⁺-SOD provides a reliable index of physical training. 7. The results obtained in the present study also suggest that muscle has the capacity of responding to training in such a manner as to reduce the potential harm arising from the accumulation of oxygen free radicals resulting from enhanced metabolic activity.     

The effect of l-dopa on young patients with simple schizophrenia,treated with neuroleptic drugs

Thirteen out of 18 young out-patients with simple schizophrenia under neuroleptic treatment completed a double-blind cross-over trial with Madopar® [l-Dopa + benserazid (a peripheral decarboxylase inhibitor)] and placebo. Nine patients were given 900 mg l-Dopa + 225 mg benserazid daily, 1 patient received 600 mg l-Dopa + 150 mg benserazid, and 3 patients, 300 mg l-Dopa + 75 mg benserazid. In these doses, l-Dopa was effective against emotional withdrawal, blunted affect, tendency to isolation and apathy, without inducing or aggravating productive, accessory symptoms. The activity score, according to the specific activity-withdrawal scale, was significantly increased (P<0.05), whereas the total BPRS score (Brief Psychiatric Rating Scale) was slightly, but significantly reduced (P<0.05). In cases where l-Dopa had to be limited to 600 and 300 mg daily, a tendency to anxiety, distortion of thinking, and a sense of unreality were observed, depending on the dose of l-Dopa. In no case were gastrointestinal, cardiovascular or neurological sideeffects observed.     

Inhibition of N-methyl-d-aspartate (NMDA) and l-glutamate-induced noradrenaline and acetylcholine release in the rat brain by ethanol

Summary The influence of ethanol on stimulation-evoked ³H-transmitter release was examined in slices of the rat brain cortex and corpus striatum preincubated with ³H-noradrenaline and ³H-choline, respectively. ³H-Transmitter release was stimulated by NMDA, l-glutamate, electrical impulses, reintroduction of Ca²⁺ ions (Ca²⁺-evoked release; after superfusion with Ca²⁺-free, K⁺-rich solution) or veratridine. In cortical slices preincubated with ³H-noradrenaline and superfused with Mg²⁺-free, otherwise physiologically composed salt solution, ethanol inhibited the NMDA- or l-glutamate-induced tritium overflow (IC₅₀ 45 and 37 mmol/l, respectively). In contrast, the tritium overflow in response to electrical stimulation, reintroduction of Ca²⁺ ions or veratridine was not affected by ethanol at concentrations up to 320 mmol/l; these experiments were carried out in cortical slices superfused with solution containing a physiological Mg²⁺ concentration. Ethanol also failed to inhibit Ca²⁺-evoked release in the absence of Mg2⁺ ions. In the presence of 1 mol/l veratridine, but not in its absence, NMDA induced tritium overflow even when cortical slices were superfused with salt solution containing a physiological Mg²⁺ concentration; again, ethanol inhibited this NMDA-evoked tritium overflow (IC₅₀ 73 mmol/l). In striatal slices preincubated with ³H-choline and superfused with Mg²⁺-free physiological salt solution, the NMDA-evoked tritium overflow was also, although at lower potency, inhibited by ethanol (IC₅₀ 192 mmol/l).In spite of the differences between the IC₅₀ values of ethanol determined for the inhibition of cortical noradrenaline and striatal acetylcholine release, it may be concluded that the NMDA receptor-ion channel complex is one of the sites of action underlying the ethanol-induced inhibition of neurotransmitter release. Since in the brain cortex the NMDA-induced ³H-noradrenaline release appears to be mediated by an excitatory interneurone activated by NMDA, this neuronal system may be involved in the cortical actions of ethanol.     

Effects of L-ascorbic acid supplementation on dieldrin toxicity in rats

Chronic dieldrin administration to rats (5 mg/kg/day) produced pathological changes in liver and kidney tissues. Dieldrin treated rats showed high levels of liver ascorbic acid and increased activities of inorganic pyrophosphatase in brain and glucose-6-phosphatase in liver. The activities of Mg²⁺-ATPase in liver and acetylcholinesterase in brain were decreased under toxic doses of dieldrin. L-Ascorbic acid supplements in treated animals could partially prevent the pathological alterations, as observed histologically in liver and kidney tissues. Administration of this vitamin could also prevent alterations in some enzyme activities produced by toxic dieldrin doses.     

Characterization of nicotinic acetylcholine receptors on cultured bovine adrenal chromaffin cells using modified l-[3H]nicotine binding assay

Summary To characterize the properties of nicotinic acetylcholine receptors (nAChRs) in autonomic ganglia, we examined l-[³H]nicotine binding to membrane fraction prepared from cultured bovine adrenal chromaffin cells, using a modified filtration method. Binding of l-[³H]nicotine to non-treated glass fiber filters interfered with the detection of specific binding to the membrane fraction. Presoaking glass fiber filters in 3% or higher concentrations of polyethyleneimine (PEI) solution (sixty times higher than earlier used concentration) for at least 5 h could reduce the binding of l-[³H]nicotine to the filters to the background level. Specific l-[³H]nicotine binding to the membrane fraction was detected only when the membrane fraction was prepared in Ca²⁺- and Mg²⁺ (EDTA, EGTA and protease inhibitors were added)-free buffer. Specific binding of l-[³H]nicotine was saturable and reversible. Both computer program and Scatchard analysis revealed a single class of high affinity binding sites with an average K_d of 8.9 nM and a B_max of 42.5 fmol/mg protein. The Hill coefficient was 0.98. In inhibition studies, both cholinergic agonists (carbachol and l-nicotine) and ganglionic agonists (lobeline and 1,1-dimethyl-4-phenylpiperazinium iodide) were much effective in inhibiting l-[³H]nicotine binding, whereas both neuromuscular blocking (-bungarotoxin and d-tubocurarine) and ganglionic blocking agents were less effective. These results suggest that high affinity nicotinic binding sites on adrenal chromaffin cells are nAChRs of the ganglion-type, which have properties different from nAChRs on the neuromuscular junction but similar to nAChRs in the brain. Send offprint requests to K. Lee at his present address     

Effects of methysergide,bromocriptine and naloxone on prolactin,growth hormone and TSH release induced by d-Met2,Pro5-enkephalinamide in man

d-Met²,Pro⁵-enkephalinamide (EA) 10 mg, given SC, induced a dramatic rise in serum prolactin (PRL) and growth hormone (GH) levels in healthy male volunteers. The TSH content was also moderately elevated. Naloxone 0.8 mg administered IV abolished these effects. Bromocriptine 2.5 mg given per os also antagonized EA-induced PRL and TSH release but potentiated the GH surge. Methysergide 2.0 mg administered orally partially reversed EA-elicited PRL release, further augmented GH liberation and did not modify TSH output. The data indicate that inhibition of the dopaminergic tone and/or activation of certain serotonergic mechanisms play an important role in the EA-induced release of PRL and TSH. However, primarily other neurotransmitters might mediate the GH liberation elicited by this opioid peptide. Offprint requests to: J. Földes     

Induction of the Conversion of Xanthine Dehydrogenase to Oxidase in Rabbit Liver by Cu2+, Zn2+ and Selenium Ions

Abstract— Effects of Cu²⁺, Zn²⁺, Fe²⁺ and selenium ions on the conversion of xanthine dehydrogenase to oxidase in rabbit liver were examined. Under basal conditions, xanthine oxidase activity represented only 16% of the total xanthine oxidase plus dehydrogenase activity. Cu²⁺ (2–10 μm ), Zn²⁺ (5–30 μm ) and selenium ions (5–100 μm ) brought about the conversion of xanthine dehydrogenase to oxidase in a dose-dependent manner. The concentrations of Cu²⁺, Zn²⁺ and selenium ions required for increasing xanthine oxidase activity by 50% was approximately 4, 10 and 20 μm , respectively. On the other hand, Fe²⁺ had no effect on the conversion of the enzyme up to 100 μm . These results suggest that Cu²⁺, Zn²⁺ and selenium ions have the potential to modulate the conversion of xanthine dehydrogenase to oxidase in rabbit liver.