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Traditional herbal medicine in the Zagori region of Greece was studied by obtaining information from users and collecting herbs. In addition, traditional remedies found in old manuscripts and books were compared with those currently used in modern literature. The 36 medicinal plants used today are listed, including their common and scientific names, indications (some new) and recipes for preparations. J Ethnopharmacol

Increasing body of evidence indicates that early life stressful events may induce permanent alterations in neurodevelopment, which in turn, could lead to the development of psychopathologies in adulthood. In particular, maternal deprivation (MD) for 24h in rats has been associated with several abnormalities in brain and behaviour during adulthood, relevant to the neurobiological substrate of anxiety disorders. The aim of the present study was to clarify the long-term effects of MD, on hypothalamo-pituitary-adrenal (HPA) axis activity and serotonergic (5-HT) function, in adulthood, subjects that have not been yet thoroughly investigated. For this purpose, Wistar rat pups were deprived from their mothers for a 24-h single period at postnatal day 9 (pnd 9) and were examined when aged 69-90 days. Plasma corticosterone and ACTH levels along with the animal's behaviour in an open field were used as indices of stress. Moreover, serotonergic activity was estimated in hypothalamus and hippocampus, key structures in the coordination of neuroendocrine and behavioural responses to stress. Interestingly, in adulthood, MD rats compared to controls, displayed decreased body weight, increased serotonergic activity and "anxiety" related behaviour, as well as elevated plasma corticosterone and ACTH levels. The findings of this study showed that MD results in long-term modifications in HPA axis and serotonergic activity indicating a clear relationship between early life stressful events and the development of anxiety-like disorders later in adulthood. Neurosci Lett

Aims This study investigated the role of the cannabinoid CB1 receptor agonist, WIN 55,212-2, on motor activity. Subsequently, the effects of a low, stimulatory dose of WIN 55,212-2 and cocaine, as a positive control, were evaluated using a conditioned place preference (CPP) procedure. Upon completion of CPP, in rats that had been treated with WIN 55,212-2, dopaminergic status and spontaneous and d -amphetamine-induced motor activity were assessed. Main methods Sprague–Dawley rats were evaluated for habituated motor activity following WIN 55,212-2 (0, 0.1, 0.3, 1 mg/kg, i.p.) administration. A stimulatory dose of WIN 55,212-2 (0.1 mg/kg, i.p.) and cocaine (20 mg/kg, i.p.) was selected to assess CPP behaviour. Upon completion of CPP, in one group, tissue levels of dopamine and its metabolites were measured in distinct brain regions (dorsal striatum, nucleus accumbens, prefrontal cortex, amygdala, hippocampus) using High Performance Liquid Chromatography with electrochemical detection. In another group, spontaneous and d -amphetamine-induced motor activity was evaluated in an open-field apparatus. Key findings The lowest dose of WIN 55,212-2 increased motor activity but did not produce CPP. As expected, cocaine induced clear CPP. Dopaminergic status was increased in a region-specific way and motor activity was enhanced following a challenge of d -amphetamine in rats that had been administered with WIN 55,212-2 during conditioning. Significance A stimulatory effect of WIN 55,212-2 on motor activity was not accompanied by place preference. Upon completion of the CPP procedure, this dose was found to induce region-specific hyperdopaminergia along with a greater sensitivity to a subsequent challenge dose of d -amphetamine.

The cytosolic activity of aldehyde dehydrogenase (ALDH) was studied in the rat liver, after acute administration of various carcinogenic and chemically related compounds. Male Wistar rats were treated with 27 different chemicals, including polycyclic aromatic hydrocarbons, aromatic amines, nitrosamines, azo dyes, as well as with some known direct-acting carcinogens. The cytosolic ALDH activity of the liver was determined either with propionaldehyde and NAD (P/NAD), or with benzaldehyde and NADP (B/NADP). The activity of ALDH remained unaffected after treatment with 1-naphthylamine, nitrosamines and also with the direct-acting chemical carcinogens tested. On the contrary, polycyclic aromatic hydrocarbons, polychlorinated biphenyls (Arochlor 1254) and 2-naphthylamine produced a remarkable increase of ALDH. In general, the response to the effectors was disproportionate between the two types of enzyme activity, being much in favour for the B/NADP activity. This fact resulted to an inversion of the ratio B/NADP vs. P/NAD, which under constitutive conditions is lower than 1. In this respect, the most potent compounds were found to be polychlorinated biphenyls, 3-methylcholanthrene, benzo(a)pyrene and 1,2,5,6-dibenzoanthracene. Our results suggest that the B/NADP activity of the soluble ALDH is greatly induced after treatment with compounds possessing aromatic ring(s) in their molecule. It is not known, if this response of the hepatocytes is related with the process of chemical carcinogenesis. Chem Biol Interact

Almost 200 pharmaceutical chemicals and groups of drugs have been evaluated for their carcinogenic properties by working groups convened by the International Agency for the Research on Cancer. Therapeutic agents are exceptional environmental carcinogens in that humans are exposed to relatively pure substances at well-defined dosages. Of those evaluated, 20 are conclusively carcinogenic to humans and 52 are probably or possibly carcinogenic. The human tissues most often affected are bone marrow, skin, urinary bladder, liver, lymphatic tissue and endometrium. In cases in which there is sufficient evidence for carcinogenicity from both epidemiological and experimental studies, the similarity between humans and animals with regard to the target organs involved is close (85%). Since data on carcinogenicity exist for several groups of pharmaceuticals, risk versus benefit evaluations should be made carefully in relation to possible clinical applications. Carcinogenesis

BACKGROUND: : Lipoplatin is a new liposomal cisplatin that already has been tested in solid tumors, with encouraging results. The purpose of the current study was to determine the maximum tolerated dose (MTD) and the dose-limiting toxicity (DLT) of a 21-day regimen of lipoplatin plus a fixed dose of gemcitabine in patients with refractory or resistant nonsmall cell lung carcinoma (NSCLC) with an Eastern Cooperative Oncology Group (ECOG) performance status of

Increased activity of CYP2E1 has been associated with increased risk of chemically-mediated cancers, through enhanced activation of a variety of procarcinogens. In this context, inhibition of CYP2E1 is potentially of significance in xenobiotic toxicity. The aim of the present study was to test the hypothesis that quinacrine inhibits hepatic CYP2E1. For this purpose, disulfiram (75 mg/kg i.p) as an inhibitor and isoniazid (100 mg/kg i.p) as an inducer of CYP2E1, as well as quinacrine (50 mg/kg i.p) were administered to Wistar rats and the hepatic activity of CYP2E1 was measured. The expression of CYP2E1 was further assessed by Western blot analysis. As expected, disulfiram inhibited, while isoniazid induced the activity and expression of the enzyme. Interestingly, treatment with quinacrine resulted in a significant decrease of CYP2E1 activity and expression. To investigate any similarities in the inhibition of CYP2E1 by quinacrine and disulfiram, molecular modeling techniques were adopted and revealed that quinacrine molecule anchors inside the same binding pocket of the protein where disulfiram is also attached. Finally, as assessed by the sister chromatid exchanges (SCE) assay, quinacrine was demonstrated to reduce the mutagenic effects of the tobacco-specific N-nitrosamine 4-(methyl nitrosamino)-1-(3-pyridyl)-1-butanone (NNK), which is known to be converted to active mutagen in the liver principally through CYP2E1. We suggest that these antimutagenic effects of quinacrine could be possibly attributed, at least in part, to its ability to block the bioactivation of NNK, mainly by the inhibition of CYP2E1. Our results, even preliminary, indicate that quinacrine as an inhibitor of CYP2E1 might be protective against chemically-induced toxicities such as NNK-induced mutagenicity.

Dopamine (DA) replacement therapy with L-3,4-dihydroxyphenylalanine (L-DOPA) remains the most effective medication for the symptomatic relief of Parkinson’s disease (PD). However, chronic L-DOPA treatment is often complicated by a variety of involuntary movements, termed L-DOPA-induced dyskinesia (LID), which represents one of the major limitations in the treatment for PD. Depending on clinical presentation and chronology after a dose of L-DOPA, LID can be classified in peak-dose, diphasic, and off period dyskinesias. Peak-dose dyskinesia occurs at the time of the highest brain concentration of LDOPA and represents the most common type of LID. The current pharmacological treatment of this form of dyskinesia involves, as a first measure, the reduction of individual L-DOPA doses, which indeed can lead to improvement of dyskinesia, exacerbating however the symptoms of the disease. Slow-release L-DOPA preparations or spreading of the daily dose of L-DOPA into smaller but more frequent doses has been also used; however, only limited improvements have been reported so far. On the other hand, the only currently available drug with an evidence-based recommendation for dyskinesia is the glutamate antagonist amantadine, even though no large scale study of this drug has been conducted. Moreover, only a number of patients can tolerate effective doses of amantadine, and its effects decline over time. Finally, dyskinesias that become disabling must often be managed surgically, despite the high cost and risk for serious adverse events of such intervention. Thus, it is a common belief that the development of new safe and effective therapies that treat dyskinesias without aggravating parkinsonism are urgently needed. A rapid and excessive increase in extracellular DA, after L-DOPA administration, is considered one of the major causes for peak-dose LID. This is due to the fact that oral formulations of L-DOPA have short plasma half-lives, so each dose causes a rapid increase followed by a rapid decrease in striatal dopamine levels, leading to intermittent stimulation of striatal dopamine receptors. Interestingly, in experiments with rats, administration of 8-hydroxy-2-(d-npropylamino) tetralin (8-OH-DPAT), a selective 5-HT1A receptor agonist, attenuated the increase in L-DOPA-induced extracellular DA [1] while alleviated L-DOPA induced motor complications, including LID [2]. In support of a role in receptor specificity, it was shown that these effects of 8-OH-DPAT were mediated mainly via stimulation of 5-HT1A receptors, since co-administration of a 5HT1A receptor antagonist (WAY100635) abolished the above mentioned effects of 8-OH-DPAT [1]. Moreover, in compliance with these animal studies, the use of the clinically available 5-HT1A agonist sarizotan, in the context of a multicenter trial, was proved to be effective in the treatment of LID in PD patients [3]. Taken together, all these data suggest that pharmaceutical agents capable of stimulating 5-HT1A receptors could avert the induction of L-DOPA induced motor complications in patients with PD. P. N. Karamanakos P. Pappas M. Marselos Department of Pharmacology, Medical School, University of Ioannina, Ioannina 451 10, Greece

This study investigated the role of catecholamine-related signaling pathways in the regulation of hepatic cytochrome P450 (CYP2E1). Central and peripheral catecholamine depletion with reserpine down-regulated CYP2E1. On the other hand, selective peripheral catecholamine depletion with guanethidine increased CYP2E1 apoprotein levels. Enrichment of peripheral catecholamines with adrenaline suppressed p-nitrophenol hydroxylase activity (PNP). PNP activity was also markedly suppressed by l-DOPA. Stimulation of D(2)-receptors with bromocriptine up-regulated CYP2E1, as assessed by enzyme activity and protein levels, whereas blockade of D(2)-dopaminergic receptors with sulpiride down-regulated this isozyme. These findings indicate that central and peripheral catecholamines have different effects on CYP2E1. Central catecholamines appear related to the up-regulation, whereas the role of peripheral catecholamines is clearly related to the type and location of adrenoceptors involved. D(2)-receptor-linked signaling pathways have an up-regulating effect on CYP2E1, while D(1)-receptor pathways may down-regulate this isozyme. It is worth noting that the widespread environmental pollutant benzo(alpha)pyrene (B(alpha)P) altered the modulating effect of catecholaminergic systems on CYP2E1 regulation. In particular, whereas stimulation or blockade of adrenoceptors had no effect on constitutive PNP activity, exposure to B(alpha)P modified the impact of central and peripheral catecholamines and alpha(2)-adrenoceptors on CYP2E1 expression. It appears that under the influence of B(alpha)P, alpha(2)-adrenergic receptor-linked signaling pathways increased CYP2E1 apoprotein levels. Given that a wide range of xenobiotics and clinically used drugs are activated by CYP2E1 to toxic metabolites, including the production of reactive oxygen species (ROS), it is possible that therapies challenging dopaminergic receptor- and/or alpha(2)-adrenoceptor-linked signaling pathways may alter the expression of CYP2E1, thus affecting the progress and development of several pathologies. Life Sci

Several pharmaceutical agents produce ethanol intolerance, which is often depicted as disulfiram-like reaction. As in the case with disulfiram, the underlying mechanism is believed to be the accumulation of acetaldehyde in the blood, due to inhibition of the hepatic aldehyde dehydrogenases. In the present study, chloramphenicol, furazolidone, metronidazole, and quinacrine, which are reported to produce a disulfiram-like reaction, as well as disulfiram, were administered to Wistar rats and the hepatic activities of alcohol and aldehyde dehydrogenases (1A1 and 2) were determined. The expression of aldehyde dehydrogenase 2 was further assessed by Western blot analysis, while the levels of brain monoamines were also analyzed. Finally, blood acetaldehyde was evaluated after ethanol administration in rats pretreated with disulfiram, chloramphenicol, or quinacrine. The activity of aldehyde dehydrogenase 2 was inhibited by disulfiram, chloramphenicol, and furazolidone, but not by metronidazole or quinacrine. In addition, although well known for metronidazole, quinacrine also did not increase blood acetaldehyde after ethanol administration. The protein expression of aldehyde dehydrogenase 2 was not affected at all. Interestingly, all substances used, except disulfiram, increased the levels of brain serotonin. According to our findings, metronidazole and quinacrine do not produce a typical disulfiram-like reaction, because they do not inhibit hepatic aldehyde dehydrogenase nor increase blood acetaldehyde. Moreover, all tested agents share the common property to enhance brain serotonin, whereas a respective effect of ethanol is well established. Therefore, the ethanol intolerance produced by these agents, either aldehyde dehydrogenase is inhibited or not, could be the result of a “toxic serotonin syndrome,” as in the case of the concomitant use of serotonin-active medications.

Background Endogenous and exogenous compounds as well as carcinogens are metabolized and detoxified by phase I and II enzymes, the activity of which could be crucial to the inactivation and hence susceptibility to carcinogenic factors. The expression of these enzymes in human brain tumor tissue has not been investigated sufficiently. We studied the association between tumor pathology and the expression profile of seven phase I and II drug metabolizing genes (CYP1A1, CYP1B1, ALDH3A1, AOX1, GSTP1, GSTT1 and GSTM3) and some of their proteins. Methods Using qRT-PCR and western blotting analysis the gene and protein expression in a cohort of 77 tumors were investigated. The major tumor subtypes were meningioma, astrocytoma and brain metastases, -the later all adenocarcinomas from a lung primary. Results Meningeal tumors showed higher expression levels for AOX1, CYP1B1, GSTM3 and GSTP1. For AOX1, GSTM and GSTP1 this could be verified on a protein level as well. A negative correlation between the WHO degree of malignancy and the strength of expression was identified on both transcriptional and translational level for AOX1, GSTM3 and GSTP1, although the results could have been biased by the prevalence of meningiomas and glioblastomas in the inevitably bipolar distribution of the WHO grades. A correlation between the gene expression and the protein product was observed for AOX1, GSTP1 and GSTM3 in astrocytomas. Conclusions The various CNS tumors show different patterns of drug metabolizing gene expression. Our results suggest that the most important factor governing the expression of these enzymes is the histological subtype and to a far lesser extent the degree of malignancy itself.

Background: Intrapericardial drug delivery is a promising new technique, but the pharmacologic properties of various agents delivered via this route are not known. Furthermore, the long-term safety of intrapericardial catheters has not been previously examined. Methods: Using a pericardial access device, a catheter connected to a drug-delivery system was implanted in five pigs. Plasma levels and electrocardiographic measurements were obtained after intravenous and intrapericardial administration of digoxin and procainamide. Histological examination was performed after the device had been implanted for a total of 6 months. Results: The QTc interval did not change significantly after digoxin or procainamide intravenous administration. QTc decreased by 47±23 ms ( p =0.046) 8 h after digoxin intrapericardial administration and increased by 128±60 ms ( p =0.002) 1 h after procainamide intrapericardial administration. The QRS duration did not change significantly after intravenous administration of either agent, but it increased by 17±9 ms ( p =0.004) 1 h and by 15±4 ms ( p =0.01) 8 h after procainamide intrapericardial administration. After intravenous procainamide the RR interval decreased, but it did not change significantly after intrapericardial administration of either agent. Histology showed moderate inflammatory infiltration and fibrosis adjacent to the catheter. Conclusions: Intrapericardial delivery of digitalis and procainamide produces unique electrophysiological properties. In contrast to satisfactory success of the implantation technique, long-term dwell of the catheter in the pericardium induces moderate, albeit probably clinically significant, fibrosis.

This study was undertaken to investigate the use of the in vitro test WST-1, an assay of cell proliferation and viability, for a preliminary safety evaluation of topical ophthalmic preparations. The cytotoxicity of two surfactants, benzalkonium chloride (BAC) and polyoxyethylene-20-stearyl ether (Brij78, PSE) was independently investigated in four laboratories in the EU by using an immortalized human corneal epithelial (HCE) cell line. The HCE cells were exposed to BAC and PSE for 5 min, 15 min, and 1 hour, and the results of the HCE-WST-1 tests were collected and compared. After one-hour exposure, the EC(50) values in BAC-treated cells in the presence of serum ranged between 0.0650 +/- 0.0284 (mean +/- SD) mM, and those in the absence of serum 0.0296 +/- 0.0081 mM. The corresponding values for PSE were 0.0581 +/-.0300 mM and 0.0228 +/-.0063 mM. There were variations in the results between different laboratories, with coefficients of variation ranging from 31 to 121%, mean 58%. The use of one-hour exposure time is to be preferred, and the elimination of serum in the culture medium is recommended to avoid both underestimation of toxic effects and variability of the test results. J Ocul Pharmacol Ther

BACKGROUND: Gemcitabine and oxaliplatin have broad antineoplastic activity and favorable toxicity. We conducted a phase I study to determine the maximum tolerated doses (MTDs) and dose-limiting toxicities (DLTs) of the combination in patients with advanced solid tumors. PATIENTS AND METHODS: Sixty-eight patients with advanced stage solid tumors were enrolled. Treatment was first-line for 35% of patients, second-line for 27%, and third-line for 38%. Gemcitabine was administered at escalating doses of 1000-2000 mg/m(2) as a 30-min intravenous (i.v.) infusion on days 1 and 8 and oxaliplatin at 60-130 mg/m(2) as a 4-h i.v. infusion on day 8 every 21 days without growth factor support. RESULTS: The MTD was defined at gemcitabine 1800 mg/m(2) on days 1 and 8 and oxaliplatin 130 mg/m(2) on day 8. Twelve dose levels were evaluated and DLTs occurring during the first cycle consisted of grade 4 neutropenia, grade 3 asthenia or mucositis and grade 1-3 neutropenia or thrombocytopenia resulting in treatment delays. A total of 266 cycles were administered with only one episode of febrile neutropenia and no toxic deaths. Seven (3%) and 26 (10%) cycles were complicated by grade 4 and 3 neutropenia, respectively, three (1%) and 13 (5%) by grade 4 and 3 thrombocytopenia, and eight (3%) by grade 3 anemia. The most common non-hematological toxicity was grade 2/3 asthenia observed in 23% of cycles. Responses were observed in patients with a variety of epithelial neoplasms. The pharmacokinetic study revealed no significant interaction between the two drugs. CONCLUSIONS: The combination of gemcitabine and oxaliplatin has excellent tolerability and promising activity in patients with advanced solid tumors. As the MTD exceeds the recommended single-agent dose for gemcitabine, and a dose-response effect has not been established, we recommend using both drugs at full doses, e.g. gemcitabine 1200-1400 mg/m(2) on days 1 and 8 and oxaliplatin 130 mg/m(2) on day 8 for further phase II studies. Ann Oncol

The aldehyde dehydrogenase-3A1 (ALDH3A1) enzyme, encoded by a member of the [Ah]-gene family, is dramatically increased (more than 100-fold) by benzo[a]pyrene (BaP) and other polycyclic hydrocarbons. Although much is known regarding the mechanism for the drug-metabolizing enzymes up-regulated by the Ah receptor, the physiological role of that tremendously increased ALDH3A1 enzyme activity is not yet fully clarified. The aim of this study was to identify a possible acute-phase response to different classes of xenobiotics affecting the metabolic capacity of the hepatocyte, by studying possible changes of serum acute-phase proteins (APPs) of hepatic origin, before and after BaP administration. Male Wistar rats were used in different series of experiments. The effects of BaP were estimated in terms of dose-response and time-response, with regard to the serum level of several APPs such as alpha-1-acid-glycoprotein (AAG), alpha-1-antitrypsin (AAT), C-reactive protein (CRP), and haptoglobin (HPT). In parallel experiments, levels of the same proteins have been determined after a time-dependent treatment with lipopolysaccharide (LPS). The changes in serum proteins were compared with the results of BaP or LPS administration on both hepatic ALDH3A1 and total ALDH enzyme activities. The results showed that BaP induced CRP and HPT in a time-dependent way, proportional to that caused by LPS. Additionally, ALDH3A1, CRP, and HPT were induced by BaP subacute treatment, whereas another type of ALDH inducer, phenobarbital, did not affect the levels of APPs or ALDH3A1, but did increase ALDH1A3 activity. Former studies of our group have shown that the inhibitory effects of different non-steroidal anti-inflammatory drugs (NSAIDs) on the ALDH3A1 induction were most possibly due to a decreased formation of arachidonic products like prostaglandins. Considering the changes of APPs caused by BaP, this study further supports the suggestion that the induction of ALDH3A1 is related to an atypical hepatocyte inflammation produced by xenobiotics. Chem Biol Interact

The cytotoxicity of the selected systemic and intravitreally dosed drugs tamoxifen, toremifene, chloroquine, 5-fluorouracil, gentamicin and ganciclovir was studied in retinal pigment epithelium (RPE) in vitro. The cytotoxicity was assayed in the human RPE cell line D407 and the pig RPE cell culture using the WST-1 test, which is an assay of cell proliferation and viability, The effects of experimental conditions on the WST-1 test (cell density. serum content in the culture medium, the exposure time) were evaluated. The EC50 Values in tamoxifen-treated D407 cells ranged between 6.7 and 8.9 mumol/l, and in pig RPE cells between 10.1 and 12.2 mumol/l, depending on the cell density used. The corresponding values for toremifene were 7.4 to 11.1 mumol/l in D407 cells and 10.0 to 11.6 mumol/l in pig RPE cells. In chloroquine-treated cells, the EC50 values were 110.0 mumol/l for D407 cells and 58.4 mumol/l for pig RPE cells. Gentamicin and ganciclovir did not shock any toxicity in micromolar concentrations. The exposure time was a significant factor, especially when the drug did not induce cell death, but was antiproliferative (5-fluorouracil). Serum protected the cells from the toxic effects or the drugs, Both cell cultures were most sensitive to tamoxifen and toremifene, and next to chloroquine. The drug toxicities obtained in the present study were quite similar in both cell tyupes. that is, the pig RPE cells and the human D 407 cell line, despite the differences in, for example, the growth rate and melanin contents Of the cell types. Owing to the homeostatic functions important for the whole neuroretina, RPE is an interesting in vitro model for the evaluation of retinal toxicity, but, in addition to the WST-1 test, more specific tests and markers based on the homeostatic functions of the RPE are needed. (C) 2002 Published by Elsevier Science Ltd. Toxicology in Vitro

Aldehyde dehydrogenases (ALDHs) are a group of enzymes which catalyze the conversion of aldehydes to the corresponding carboxylic acids in a NAD(P)(+)-dependent reaction. In mammals, different ALDHs are constitutively expressed in liver, stomach, eye and skin. In addition, inducible ALDH-isoenzymes are detectable in many tissues; apart from other physico- and immuno-chemical differences, two cytosolic ALDHs (ALDH1A3 and ALDH3A1) are known to be activated in rat liver, by different types of inducers of drug metabolism. Phenobarbital-type inducers increase the ALDH1A3, while polycyclic hydrocarbons (such as BaP and TCDD) increase the expression of the two members of ALDH3A subfamily (3A1 and 3A2). In this study, we used two Wistar rat substrains which have been well-characterized for different inducibility of ALDH1A3 enzyme activity after treatment with phenobarbital. Animals that respond (RR) or do not respond (rr) to treatment have been inbred for almost 25 years, offering a useful experimental model. Apart from the level of ALDH1A3 induced enzyme expression after phenobarbital treatment, no other differences between the two substrains have been noticed, as far as drug metabolizing enzyme activities (like the pentoxy- and ethoxy-O-dealkylation rate) are concerned. According to the present results, the ALDH1A3 expression is still the only difference between the two substrains. Immunoblotting experiments with polyclonal antibodies raised against CYP2B1 or/and CYP1A1/1A2 showed no differences between the two substrains. Additionally, data concerning time- and dose-response induction of ALDH1A3 after phenobarbital and griseofulvin treatment are presented. It is concluded that these two Wistar rat substrains represent a unique animal model for studying what seems to be the only difference between these substrains - the genetic basis of the phenobarbital induction. Chem Biol Interact

The effect of restraint stress on central neurotransmission was evaluated in mice and rats. Noradrenaline (NA), dopamine (DA) and serotonin (5-HT) levels and their primary metabolites were measured in discrete brain regions following exposure to stress. Mice and rats demonstrated a similar response to stress in some brain regions. Both species responded to stress with lower NA and 5-HT in the locus coeruleus compared to non-stressed controls. Dopaminergic activity, assessed by DA turnover, was elevated in the hypothalamus. While DA turnover was suppressed in the amygdala, 5-HT turnover was similarly elevated in both species. In most cases, however, there were differences in biogenic neurotransmission between mice and rats in response to stress. In particular, NA levels were suppressed by stress in the dorsal cortex of mice, but in the rats NA levels were decreased in the hypothalamus. While stress produced lower DA levels in the hypothalamus, DA levels demonstrated a marked increase in the amygdala of mice. Stress was also associated with a decrease in DA levels in the rat striatum and with an increase of DA turnover in the locus coeruleus of mice. On the other hand, 5-HT was suppressed in the mouse striatum and in the rat hypothalamus and amygdala, while 5-HT turnover was markedly decreased in the hippocampus and dorsal cortex of rats alone. In conclusion, the changes in the central neurotransmission which are evoked by stress appear to be species-specific in most cases, a fact which may trigger discrete alterations in homeostatic mechanisms. Pharmacol Res

The peracetylated derivative of kaempferol-3-O-beta-D-(6''-E-p-coumaroyl) glycopyranoside (tiliroside) (1a) was tested for its cytotoxic and cytostatic activity against several human leukemic cell lines. The significant cytotoxic activity of this derivative, prompted to an additional examination on some of the cell lines used. The effect on the uptake of [3H]thymidine as a marker of DNA synthesis and on the cell proliferation, was investigated as well as the morphology of the cells and the kind of death induced, using the Wright-Giemsa dye and horizontal agarose-gel electrophoresis. Flow cytometric experiments of 1a on some leukemic cell lines was also performed. Compound 1a showed a significant antiproliferative effect as soon as 1 h of continuous incubation at all cell lines tested. Cells were killed, through the process of apoptosis and the appearance of the apoptotic signs was time and dose-dependent, while from the flow cytometric experiments, a synchronisation (through a delay probably in the G(0/1) phase) of the cells seems to take place. Leuk Res

Gonadotrophin surge attenuating factor (GnSAF) is a new non-steroidal ovarian substance, different from inhibin, which attenuates the pre-ovulatory luteinizing hormone (LH) surge in superovulated women. Human follicular fluid (FF) was used as a source for the isolation of GnSAF, the activity of which was monitored in an in-vitro pituitary bioassay. Primary rat pituitary cells were incubated with test substances for 48 h and subsequently washed and incubated with 0.1 micromol/l gonadotrophin releasing hormone (GnRH) plus test substances for 4 h. GnSAF activity was expressed as the reduction of GnRH-induced LH secretion in the 4 h incubation. GnSAF was purified from 250 ml of FF which was heat-treated at 80 degrees C for 5 min. Heparin-sepharose chromatography, Con-A sepharose chromatography, reversed-phase high-performance liquid chromatography (HPLC) and preparative native gel electrophoresis were used for GnSAF fractionation. Using these purification steps, we have obtained an apparently homogeneous preparation that stains as a single band on sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis. GnSAF has an apparent molecular weight of 12.5 kDa and was identified by amino acid sequence (mass spectrometry) to be the C-terminal fragment of human serum albumin. Hum Reprod

Benzo[alpha]pyrene (B[a]P) is a product derived from incomplete combustion of organic material and is considered responsible for chemically-induced cancer in humans. In the present study, the levels of noradrenaline (NA), dopamine (DA), dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytryptamine (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) were measured in the brains of female Wistar rats 6, 12, 24 and 96 h after a single dose of B[alpha]P (50 mg kg(-1) b.w., i.p.), and also after repeated administration of B[alpha]P (50 mg kg(-1) b.w., i.p., 2 x wk, 1 mo). The brain regions studied were the striatum, hypothalamus, midbrain and cortex. Catecholamines were measured using high performance liquid chromatography (HPLC) and electrochemical detection. Significant changes were observed in the striatum where NA, DA, DOPAC were decreased after 24 h and HVA was decreased after 6 h. In contrast, no major alterations occurred in 5-HT and 5- HIAA. In the hypothalamus, a significant decrease in NA was observed after 96 h. In the midbrain, the most important change observed was the decrease in NA after 24 h. A trend toward an increase in 5-HIAA was observed in the cortex after 6 h. The results demonstrate that B[alpha]P induces alterations in the dopaminergic and serotoninergic systems throughout the brain. These alterations may lead to behavioural and hormonal disturbances. Eur J Drug Metab Pharmacokinet

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants formed primarily from the incomplete combustion of organic material. They are present in tobacco smoke, exhaust fumes and urban air. PAHs are also found in water and soil as a result the rain and gravitational influence providing other possible routes for human exposure (Wills and Weisburger, 1986). Benzo[a]pyrene (BP) is a potent inducer of drug- metabolizing enzymes. BP up-regulates the expression of [Ah] battery genes, by binding to the aryl-hydrocarbon receptor (AHR) (Nebert et al., 1993). The [Ah] battery consists of at least six genes: two Phase I (CYP1 Al and CYP1A2) and four Phase II genes (ALDH3, NQO-1,T1*A6 and GSTA-1). Induction of CYP1A1 and CYP1A2 enzymes provokes increased metabolic activation of PAHs to mutagenic derivatives that may activate pro- or inactivate tumor or inactivate tumor supressor genes (Nebert, 1989; Hankinson, 1995). On the other d, induction of the Phase II enzymes of the [Ah] gene battery may be beneficial, since se enzymes are involved in detoxification pathways.

Aldehydehydrogenase is a NAD(P)-dependent enzyme with wide distribution virtually in all animal tissues (Vasiliou and Marselos, 1989; Lindahl, 1992). Different consti- tutiveexpressed ALDHs are found in liver, stomach, brain, kidney, skin and eye (Vasil and Marselos, 1989; Pappas et al., 1997). In general, ALDHs are located especially in org with a high content of epithelial cells. An increased interest has been shown r the last years for aldehyde dehydrogenase-3 (ALDH3) activity in the cornea where h higher constitutive specific activity is detected compared to the liver cells (Boesct al., 1996). High levels of ALDH3 activity occurs in the cornea from baboon, cow, hn, opossum, pig and sheep (King and Holmes, 1997). The same study reports also tpresence of ALDH1 as the 1-2% of human lens soluble protein. Furthermore, retina and real pigment epithelial (RPE) cells have been shown to play an important protective role fthe photosensitive cells of the eye against photo- and chemical toxicity.

Clin Toxicol (Phila)

Although open-field behaviour has been considered a valid and reliable index of locomotor activity in rodents, the simple measures traditionally recorded in this test do not readily allow for differentiation between compounds of the same general class, e.g. psychostimulants. The present methodology was developed to facilitate detailed and continuous observations on the behaviour of drug-treated rats. In addition to an automated (photocell) measure of general locomotor activity, ethological techniques were used to record the frequency and duration of standing, moving, sniffing, rearing, grooming, scratching, sniffing air, freezing, head-swinging and licking. A series of factor analyses was also performed in order to further characterize treatment-induced changes in the structure of behaviour. Compounds studied were d-amphetamine (0.5, 1.5, 3, 6 mg/kg), cocaine (5, 10, 20, 50 mg/kg) and caffeine (5, 10, 20, 40 mg/kg). Although all three psychostimulants increased the automated measure of general locomotor activity, cocaine (which produced the largest effects) monotonically increased general activity over the dose range tested, whereas the stimulant effects of the other two compounds were either reduced (d-amphetamine) or eliminated (caffeine) at higher doses. More detailed observations provided confirmation of the differences in effect produced by these compounds. For example, the frequency and duration of 'moving' dose-dependently increased after cocaine, while d-amphetamine and caffeine again produced bell-shaped dose-response curves. However, whereas low-intermediate doses of d-amphetamine reduced the mean duration of moving, sniffing and rearing, no such effect was observed at the highest dose tested. This finding, together with the appearance of licking in the behavioural repertoire, suggests a stereotyped character to responses seen at high doses of this compound, though neither cocaine nor caffeine induced stereotypy. As factor analyses also revealed quite different behavioural structures associated with these three drugs, present findings demonstrate that detailed observation of behaviour represents a useful approach to research on the behavioural pharmacology of psychostimulants. Neurosci Biobehav Rev

Aldehyde dehydrogenase (ALDH) is involved in the metabolism of endogenous and exogenous aldehydes originating from biogenic amines, lipids, food and drugs. Rat liver contains at least two cytosolic ALDHs that can be stimulated by inducers of drug metabolism. Phenobarbital- type inducers increase ALDH1 activity while polycyclic aromatic hydrocarbons (such as benzo[alpha]pyrene) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) increase ALDH3c isoenzyme activity. Two rat substrains were isolated according to a different induction of hepatic ALDH after treatment with phenobarbital (PB). Animals that responded to treatment (RR) and those that did not respond (rr) were inbred and divided into two homogenous groups. These animals constituted an ideal experimental model due to their common origin. Apart from the dramatic induction of cytosolic ALDH1 and ALDH3c, the effects of PB on pentoxy-, ethoxy- and methoxy-resorufin-O-dealkylase (P-, E-, and MROD) between the two substrains were also studied. 3-Methylcholanthrene (3MC) greatly increased ALDH3c levels in both substrains, although it was slightly more pronounced in the rr rats, in which it was assessed either as ALDH3c or as total cytosolic ALDH. A similar trend was also noted in EROD, PROD and MROD activities. Dealkylation of the methoxy group was found to be statistically different between the two substrains (rr > RR). The relevance of the biochemical findings with the in vivo hepatic capacity for drug metabolism was investigated by measuring the duration of zoxazolamine paralysis. Both animal substrains were tested with zoxazolamine either without pretreatment or after administration of PB or 3MC: the paralysis produced by zoxazolamine lasted for a longer period in rr than in RR rats. After pretreatment with PB, the duration of paralysis was greatly reduced, but the differences between the two substrains remained. Pretreatment with various doses of 3MC produced differences in the duration of paralysis in RR and rr rats, although the time period was much shorter than that observed in control animals. Eur J Drug Metab Pharmacokinet

Repeated exposure of adult female Wistar rats to 3-methylcholanthrene (MC) (25 mg kg(-1) b.w., i.p., 2xwk, 1 mo) was associated with a significant increase in estrus cycle length. In addition, an increased frequency of females with constant diestrus and abnormal cycles was observed. Young females which had been exposed to MC prepubertally or whose parents had been treated with MC before and during mating also demonstrated cycle prolongation and an increased incidence of constant diestrus and abnormal cycles. These changes in female reproductive function were not associated with measurable changes in plasma sex hormone levels. In contrast, MC exposure in adult males was associated with significant reductions in circulating plasma testosterone levels. The present data also suggest that the offspring of parents who had been exposed repeatedly to MC before and during mating are also affected. Although the central nervous system in offspring of MC-treated parents appeared to be intact, their oral body temperature was significantly lower. Pharmacol Res

As the body’s first line of defense against external insult, both the eye and the skin are exposed routinely to chemical agents, serving as a portal of entry for topical contactants. In addition, global atmospheric changes like ozone depletion in the stratosphere contribute to enhanced chronic exposure of the skin and the eye of human tissue to UV light (Hendee, 1989). The hazardous effects of UV light on the eye were already recognized in 1920 by van der Hoeve. By the 1920s, basal cell carcinomas of the skin were also noted.

A Class 3 aldehyde dehydrogenase (ALDH3c; EC 1.2.1.3), present in rat liver cytosol is highly inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or polycyclic hydrocarbons such as 3-methylcholanthrene (3MC) and aromatic amines (Torronen et al., 1981; Vasiliou et al, 1988; Marselos and Vasiliou, 1991). Induction of ALDH3c has also been observed in a number of extrahepatic rat tissues including lungs, spleen, urinary bladder, heart, and brain (Dunn et al., 1988; Vasiliou and Marselos, 1989). Induction of ALDH3c by either TCDD or 3MC has also been found in cultures of human hepatoma cell lines (HepG2), normal human hepatocytes, mouse and rat hepatoma cells and (Marselos et al., 1987; Vasiliou et al., 1992, 1993a). This induction process requires a functional aromatic hydrocarbon (Ah) receptor present in the cytosol (Vasiliou et al., 1992; 1993a; Nebert et al., 1993). After binding, the inducer-receptor complex translocates into the nucleus, forms a heterodimer with the Ah receptor nuclear translocator (ARNT), and binds to one or more aromatic hydrocarbon-responsive elements (AhREs) identified upstream of murine and rat ALDH3c genes (Asman et al., 1993; Vasiliou et al., 1994). In this chapter, we address questions regarding possible parameters regulating the induction of rat liver ALDH3c by 3MC., such as (i) glutathione depletion, (ii) sex hormones, (iii) non steroid anti-inflammatory drugs (NSAIDs) and (iv) the overall drug metabolizing capacity.

One of the rat liver "Class 3" cytosolic aldehyde dehydrogenases (EC 1.2.1.3), ALDH3c, is known to be markedly induced by polycyclic aromatic hydrocarbons and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin). In the present study we examined whether hepatic ALDH3c induction is a general response to toxicity. Treatment of Wistar rats for 4 days with known toxic doses of hepatotoxic agents--carbon tetrachloride, dimethylnitrosamine, diethylnitrosamine, aflatoxin B1, and D-ethionine--did not induce ALDH3c enzyme activity. Whereas dimethylaminoazobenzene at 100 mg/kg/day for 4 days did not increase ALDH3c, a 10-fold lower dose of dimethylaminoazobenzene for 4 days produced a 20-fold increase in ALDH3c activity. Treatment with phorone, diethylmaleate or L-buthionine-S,R-sulfoximine--which deplete reduced glutathione (GSH) by different mechanisms--did not affect ALDH3c activity. One dose of benzo[a]pyrene for 24 hr increased ALDH3c activity by 25-fold. Treatment with both the GSH-depleting chemicals and benzo[a]pyrene inhibited ALDH3c induction by 45% to 75%, suggesting a role for GSH during ALDH3c induction. After ALDH3c activity had already been induced by benzo[a]pyrene, however, the GSH-depleting chemicals did not affect ALDH3c activity. No changes in ALDH3c activity were seen 24 or 48 hr after partial hepatectomy, on the fifth day following surgical cholestasis, or after guanethidine-induced sympathectomy. These data indicate that hepatic ALDH3c inducibility in the rat is not a general or direct response to chemical toxicity, or to conditions of GSH depletion or other forms of stress. Biochem Pharmacol

Various xenobiotics, also known for their ability to induce a number of different cytochrome P-450 isozymes, can increase the activity of aldehyde dehydrogenase (ALDH, EC 1.2.1.3) in the soluble fraction of rat liver (Deitrich, 1971; Marselos and Haninnen, 1974). It is well known that this induction involves at least two different isozymes which, depending on the type of the inducer, selectively react (Deitrich et al., 1977; Marselos et al., 1979).

Diethylstilboestrol (DES) exerts several toxic effects in experimental animals, by mechanisms which are still unclear. The genotoxicity of the drug has been attributed to a quinone metabolite and is mainly clastogenic, including sister chromatid exchange, unscheduled DNA synthesis, chromosomal aberrations, disruption of mitotic spindle and aneuploidy. There is evidence that genotoxic effects may occur also transplacentally. Intrauterine and early postnatal exposure to DES can cause a variety of dysplasias. In the offspring of female mice exposed to DES during pregnancy, histological changes are observed in the vaginal and cervical epithelium, the endometrium, the ovary, the testis and the epididymis. Prenatal exposure of rats to DES led to decreased litter size and to urethrovaginal cloaca, penile and testicular hypoplasia, and cryptorchidism. Vaginal ridging, vaginal adenosis, testicular hypoplasia and cryptorchidism have been observed in rhesus monkeys following prenatal exposure. There is sufficient evidence that diethylstilboestrol is carcinogenic in experimental animals, after either prenatal or postnatal exposure. Mice show a similar type of carcinogenicity to that observed in humans, target organs being vagina, cervix, uterus, ovary, mammary gland and testis. In rats, prenatal exposure to DES produces mostly mammary and pituitary tumours, but also some tumours of the vagina. Hamsters develop tumours of vagina, cervix, endometrium, epididymis, testis, liver and kidney. DES induces ovarian papillary carcinomas in dogs, and malignant uterine mesotheliomas in squirrel monkeys. Some experimental evidence points to the possibility of a transgenerational carcinogenic effect, since prenatal treatment of mice with DES is followed by an increased incidence of uterine and ovarian carcinomas in the second-generation descendants. Experimental results could have been used to predict the adverse effects of DES observed in humans in the early 1970s: DES had been reported to be carcinogenic in mice in the 1930s, while experiments in the 1960s had provided evidence that exposure during pregnancy could result in an increased cancer risk in the progeny. Eur J Cancer

Diethylstilboestrol is still used as an adjunct palliative treatment in certain patients with breast and prostate cancer. Its pharmacological, toxicological and carcinogenic properties are reviewed. In addition to the usual untoward effects following subacute or chronic administration of oestrogens, treatment with diethylstilboestrol has been associated with serious cardiovascular sequelae. Most characteristic are, however, the carcinogenic properties of this drug. Many epidemiological data provide evidence that prenatal exposure to diethylstilboestrol is causally associated with vaginal and cervical clear-cell adenocarcinomas, a very rare type of cancer in the unexposed female population. The intrauterine exposure of males leads to an increased risk of testicular cancer, although the data are less conclusive in this respect. There is some evidence that administration of diethylstilboestrol in large doses to adult women during pregnancy increases the risk of subsequent breast cancer and it probably increases the incidence of endometrial carcinoma, as has been shown with other similar oestrogens given chronically for menopausal symptoms. Eur J Cancer

Rat liver contains at least two cytosolic aldehyde dehydrogenases (ALDH 1.2.1.3) which are inducible by inducers of drug metabolism (Deitrich, 1971; Marselos and Hanninen, 1974; Deitrich et al., 1977).Phenobarbital (PB) type inducers induce the φ-isozyme in certain rat strains with a genetic predisposition (RR) (Deitrich, 1971; Deitrich et al., 1972; Marselos, 1976; Nakanishi et al., 1978). Polycyclic aromatic hydrocarbons or other type II inducers of drug metabolism can induce another isozyme (τ-ALDH), an isozyme which has been detected in all rat strains tested to date (Deitrich et al., 1978; Marselos et al., 1979; Vasiliou et al., 1988). An increase in ALDH activity was also found in primary cultures of human or rat hepatocytes, after in vitro exposure to PB or methylcholanthrene (Marselos and Michalopoulos, 1986; Marselos et al., 1987).

A survey was conducted in a population of about 8000 Greek high school students (grades 9 and 12) in order to investigate factors possibly contributing to alcohol consumption. A self-rating multiple choice questionnaire was used. The present report focuses on students with alcoholic parents. Parameters studied include students' alcohol and drug use, school performance, family relationships, socioeconomic factors and parental control. In 3.3% of the sample parental alcoholism was reported. The results indicated that parental alcoholism is a strong predictor variable for the student's alcohol and drug use. Furthermore, predictability of the student's alcohol and drug use is increased when relatives' and friends' alcoholism are taken into account. Thus, family and/or peer actual alcohol use patterns appear to be a strong influence on adolescent alcohol and drug use. School performance is worse in children of alcoholics, and family situations (eg. divorces) as well as family dynamics (relationships with the family) are disturbed, in comparison with families of students with nonalcoholic parents. These results are discussed in terms of 'modelling' for drinking and drug behaviour and the socialisation process within the family. Int J Soc Psychiatry

The potential involvement of catecholamines and in particular of alpha(2)-adrenoceptor-related signalling pathways, in the regulation of drug-metabolizing enzymes by stress was investigated in Wistar rats after exposure to the environmental pollutant benzo(alpha)pyrene. For this purpose, total cytochrome P450 content, the CYP1A2 mRNA levels, 7-methoxyresorufin-O-dealkylase (MROD), 7-pentoxyresorufin-O-dealkylase (PROD) and p-nitrophenol hydroxylase activity levels were determined in the livers of rats exposed to repeated restraint stress after treatment with benzo(alpha)pyrene coupled with pharmacological manipulations of peripheral and/or central catecholamines and alpha(2)-adrenoceptors. The data show that stress is a significant factor in the regulation of CYP1A2 induction and that catecholamines play a central role in the stress-mediated modulation of hepatic CYP1A2 inducibility by benzo(alpha)pyrene. The up-regulating effect of stress on benzo(alpha)pyrene-induced CYP1A2 gene expression was eliminated after a generalized catecholamine depletion with reserpine. Similarly, in a state where only peripheral catecholamines were depleted and central catecholamines remained intact after guanethidine administration, the up-regulating effect of stress was eliminated. It is apparent that stress up-regulates the induction of CYP1A2 by benzo(alpha)pyrene mainly via peripheral catecholamines, while central catecholamines hold a minor role in the regulation. Pharmacological manipulations of alpha(2)-adrenoceptors appear to interfere with the effect of stress on the regulation of CYP1A2 inducibility. Either blockade or stimulation of alpha(2)-adrenoceptors with atipamezole and dexmedetomidine respectively, eliminated the up-regulating effect of stress on CYP1A2 benzo(alpha)pyrene-induced expression, while it enhanced MROD activity. In contrast, stress and pharmacological manipulations of catecholamines and alpha(2)-adrenoceptors did not affect total P450 content, the CYP2B1/2-dependent PROD and the CYP2E1-dependent p-nitrophenol hydroxylase activities. In conclusion, stress is a significant factor in the regulation of the CYP1A2 inducibility by benzo(alpha)pyrene, which in turn is involved in the metabolism of a large spectrum of toxicants, drugs and carcinogenic agents. Although the mechanism underlying the stress effect on CYP1A2 induction has not been clearly elucidated, it appears that peripheral catecholamines hold a predominant role, while central catecholamines and in particular, central noradrenergic pathways hold a minor role.

Drug hydroxylation and glucuronidation enzyme levels were measured in the liver and small intestinal mucosa of male rats after starvation for 3 days and after starvation combined with phenobarbital treatment (80 mg/kg, 3 days). After simple starvation liver microsomal cytochrome P-450 content and NADPH cytochrome c reductase activity were unaffected, while p -nitroanisole demethylase activity was increased. Specific activities of the UDPglucose dehydrogenase, total β-glucuronidase and 3-hydroxy-acid dehydrogenase were increased, UDPglucuronosyltransferase was unaffected and glucuronolactone dehydrogenase was decreased. When activities were calculated per whole liver, all enzymes tested were decreased during starvation due to the reduction of the liver weight. In the small intestinal mucosa specific enzyme activities were lower in the starved animals, with the exception of UDPglucuronosyl-transferase which was not changed. The excretion into the urine of d -glucaric and l -ascorbic acid, two final products of the glucuronic acid pathway in the rat, was decreased by fasting. Phenobarbital treatment proved more effective in inducing several enzymes in the starved animals than in those fed ad lib . This compensated for the reduction of total activities due to the loss of liver weight. Despite fasting, the excretion into the urine of d -glucaric and l -ascorbic acid was enhanced after treatment with phenobarbital, and d -glucaric acid reached the levels found in normally fed rats. These findings suggest that starvation impairs drug-metabolism in the rat liver and small intestinal mucosa. Inducibility of the drug-metabolizing enzymes, however, is not depressed by this condition, but on the contrary it is markedly enhanced.

Three different fractions of d -glucuronolactone dehydrogenase were isolated from rat liver supernatant by means of CM and DEAE Sephadex ion exchange chromatography. These fractions also had acetaldehyde dehydrogenase activity. Intraperitoneal administration of phenobarbital, p , p ′-DDT and 3-methylcholanthrene increased the enzyme activity in two of the fractions, while the third remained unaffected by these drugs. Enhanced activity was obtained with both substrates, but it varied in degree within the group of animals. The rats could be divided into poor, medium and high response groups. A change in the ratio of the d -glucuronolactone and acetaldehyde dehydrogenase activities was also found after the drug treatment. Spironolactone, which is also an inducer of drug metabolism, did not alter the activity of any of the enzyme fractions, at the dose used.

A hepatic soluble aldehyde dehydrogenase (ALDH), inducible by polycyclic aromatic hydrocarbons, was studied in Wistar rats in connection with substances known to affect drug metabolism or aldehyde dehydrogenase activity, such as phenobarbital (PB), disulfiram (DS), β-diethylaminoethyl diphenylpropylacetate (SKF 525A) and calcium cyanamide (CC). 3-Methylcholanthrene (MC) was given as a model inducer of ALDH (100 mg/kg, i.p., as a single dose) and the animals were killed after 3 days. Pretreatment with PB (1 g/l drinking water, for 2 weeks) enhanced the inducing effect of MC. On the contrary, pretreatment with DS (100 mg/kg, i.p., daily x4) reduced by 70% the expected increase in ALDH activity. Neither SKF 525A (25 mg/kg, i.p., daily x4), nor CC (5 mg/kg, i.p., daily x4) could affect the action of the inducer. At the above doses, basal ALDH activity was inhibited by DS (30%) and CC (70%), but was not affected at all by PB or SKF 525A. The results were somewhat different when the various effectors tested were administered to animals already treated with MC (20 mg/kg, i.p., daily x6). In this case, DS did not affect the already induced ALDH activity. On the contrary, CC was still an effective inhibitor. Unexpectedly, post-treatment with SKF 525A further enhanced the initial induction brought about by MC. Our findings show that substances affecting microsomal drug metabolism can interfere with the process of ALDH induction by MC. The additive result of PB pretreatment is probably due to the enhanced accumulation of an active metabolite of MC. The opposite effect of DS on drug metabolism could explain the decreased ability of MC to induce ALDH activity. The MC-inducible ALDH isozyme can be effectively inhibited with CC, but not with DS.

Aldehyde dehydrogenase (ALDH) activity was measured in primary cultures of normal human hepatocytes and of the human hepatoma cell line HepG2 after application of phenobarbital (PB) or 3-methylcholanthrene (MC) for 5 days. Treatment with PB alone resulted in a significant increase in both protein and DNA content at concentrations of 2 and 3 mM. Treatment with MC at a concentration as low as 5 microM led to a significant loss of cells when it lasted more than 5 days. Concentrations of 3-5 mM of PB in the media of HepG2 cell cultures caused a 2-fold enhancement of the activity of ALDH, as measured with NAD and propionaldehyde (P/NAD) or benzaldehyde (B/NAD). On the other hand, MC-treated cultures (5 microM) showed a 20-fold increase in enzyme activity measured with NADP and benzaldehyde (B/NADP), and a 2-fold increase in B/NAD activity. Combined treatment with both PB and MC led to an effect of dynamic synergism as far as B/NAD and B/NADP activities are concerned, suggesting a metabolite of MC as the mediator for the increase of ALDH activity. Normal human hepatocytes in primary cultures responded to PB (3 mM) in a similar way as HepG2 cells as far as DNA and protein content and ALDH activity are concerned. It is concluded, that HepG2 hepatoma cells behave similar to the normal hepatocytes in terms of ALDH regulation and can be used for studies on the activity of ALDH as modified by added xenobiotics.

1. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) was administered to rats to study its effects on the enzyme activities of the D-glucuronic acid pathway in the liver, small intestine and kidney. 2. The UDP-glucuronosyl transferase activity of male albino rats given TCDD (80 mug/kg, one dose, i.p.) 6 days before killing was significantly increased in all tissues examined, and UDP-glucuronic acid pyrophosphatase activity was markedly decreased in the liver. D-Glucuronolactone and L-gulonate dehydrogenase activities in the liver and small intestine were slightly decreased after TCDD treatment. 3. The activities of UDP-glucose dehydrogenase and beta-glucuronidase were unchanged. 4. The 24 h urinary excretion of L-ascorbic acid was enhanced 8-fold, although no difference was detected in the excretion of D-glucaric acid between the control and experimental animals. 5. These results suggest an increased capacity for glucuronide conjugation after treatment with TCDD. 6. The lack of increase in the urinary excretion of D-glucaric acid further challenges its use as a reliable indicator of enhanced drug metabolism. Xenobiotica