Pulmonary clearance and inflammatory potency of intratracheally instilled or acutely inhaled nickel sulfate in rats

Rats were exposed to nickel sulfate (NiSO₄) either by intratracheal (IT) instillation or by acute aerosol inhalation, and pulmonary clearance of Ni and pulmonary inflammatory responses were studied. The half-time of Ni in the lung (initial lung burden = 50 μg Ni/rat) was about 32 h in both the IT instillation and inhalation groups. Ni retention in the lung tissue following IT instillation of NiSO₄ was saturable with reference to dose, suggesting that clearance rate of Ni from the rat lung depends on lung burden of Ni. Lung inflammatory responses were evaluated by biochemical, elemental and cytological indicators in bronchoalveolar lavage fluid (BALF) following IT instillation of NiSO₄. Activities of lactate dehydrogenase and β-glucuronidase, contents of lysozyme, protein, sulfur and calcium, and the number of polymorphonuclear leukocytes were increased with a peak at 2 – 3 days post-instillation, while BALF alkaline phosphatase (ALP) activity was significantly decreased after IT instillation of NiSO₄. Lung tissue ALP activity was also decreased by NiSO₄. Because Ni does not inhibit ALP directly, the decrease in ALP activity is probably due to functional changes of type II cells (a major source of BALF ALP). Thiobarbituric acid reacting substances in the lung tissue were not changed by NiSO₄, suggesting that lipid peroxidation plays a minimal, if any role, in the Ni-induced inflammation in the rat lung. Received: 1 December 1993/Accepted: 16 March 1994     

Inhalation toxicity of diborane in rats assessed by bronchoalveolar lavage examination

 Changes in bronchoalveolar lavage fluid (BALF) and blood were examined to assess the toxic effects of diborane (B₂H₆, CAS: 19287-45-7) on the lung. Male Wistar rats were exposed to diborane at 20 ppm (intended concentration) for 4 h (phase I study) to evaluate time-course changes up to 14 days, and at 10 or 1 ppm (intended concentrations) to assess the dose-effect relationship after 3 days (phase II study). BALF parameters [leukocyte counts, α₁-antitrypsin (α₁-AT), superoxide dismutase (SOD), total protein, phospholipids etc.] were examined and biochemical and histopathological studies were also carried out. In the phase I study, neutrophils (%) in BALF increased on the day of exposure and then decreased gradually for 3 days. Rapid and marked increases in α₁-AT and SOD activity in BALF were detected on the day of exposure, and phospholipids had sharply increased on day 3. After 14 days, these parameters in the exposed rats had returned to their background level and α₁-AT decreased significantly. In the phase II study, total protein, α₁-AT activity and phospholipids in BALF showed dose-dependent increases, and serum α₁-AT activity increased significantly. Alveolar capillary and alveolar cell damage were confirmed in rats exposed to 20 ppm, 10 ppm or 1 ppm diborane for 4 h by evaluating the parameters examined. The protection system appeared to start operating immediately after exposure, and the recovery mechanism seemed to start operating 1 day after exposure and cease by day 14. The no-observed-effect level could not be observed. Received: 14 February 1995 / Accepted: 30 May 1995     

Dopaminergic system activity and cellular defense mechanisms in the striatum and striatal synaptosomes of the rat subchronically exposed to manganese

In 6-month-old male Wistar rats, levels of dopamine (DA), dihydroxyphenylacetic acid (DOPAC), ascorbic acid (AA), dehydroascorbic acid (DHAA), uric acid and glutathione (GSH) were determined by HPLC in the striatum and striatal synaptosomes after subchronic oral exposure to MnCl₂ 50 – 100 – 150 mg/kg. Mn significantly decreased levels of DA and GSH and increased levels of DHAA and uric acid both in the striatum and synaptosomes. In synaptosomes, individual total Mn doses/rat were directly correlated with individual DOPAC/DA ratio values (r = +0.647), uric acid (r = +0.532) and DHAA levels (r = +0.889) and inversely correlated with DA (r = –0.757) and GSH levels (r = –0.608). In turn, GSH levels were inversely correlated with uric acid (r = –0.451) and DHAA levels (r = –0.460). In conclusion, the response of striatal cellular defense mechanisms (increase in AA oxidation, decrease in GSH levels) correlated well with changes in markers of dopaminergic system activity and increase in uric acid levels. The latter provides evidence of an Mn-induced oxidative stress mediated by xanthine oxidase. Received: 21 February 1994/Accepted: 18 April 1994     

Cardiotoxicity of adrenochrome in isolated rabbit hearts assessed by epicardial NADH fluorescence

Noradrenaline in a micromolar concentration has recently been shown to contribute to ischemic tissue injury by direct cardiotoxic effects independent of functional alterations. Oxygen free radicals, generated during the auto-oxidation of catecholamines, are important mediators of catecholamine cardiotoxicity. However, the role of the oxidative products (aminochromes) is still unclear. We examined the effects of adrenochrome on functional parameters and on regional myocardial ischemia (MI) in isolated electrically-driven rabbit hearts with depleted catecholamine stores (reserpine 7.0 mg/kg i.p. 16 – 24 h before preparation, Langendorff, constant pressure: 70 cm H₂O, Tyrode solution, Ca⁺⁺ 1.8 mmol/l, 37°C). Repetitive MI, separated by a reperfusion period of 50 min, was induced by coronary artery branch ligature, and MI was quantitated from epicardial NADH fluorescence photography. Adrenochrome-treatment (10^– 6 M or 10^– 4 M) was started after a reperfusion period of 20 min. The left ventricular pressure (LVP) was significantly enhanced by adrenochrome (p <0.05), but it fell thereafter to below its initial value in hearts treated with adrenochrome 10^– 4 M. The global coronary flow (CF) was not affected by adrenochrome 10^– 6 M (P >0.05), but it was significantly decreased by adrenochrome 10^– 4 M (P <0.05). The relative CF (= CF/LVP × heart-rate) was numerically decreased by adrenochrome 10^– 6 M (p >0.05) and more markedly by adrenochrome 10^– 4 M (p <0.05). Whereas epicardial NADH fluorescence was similar after repetitive coronary artery occlusions in controls and in hearts treated with adrenochrome 10^– 6 M (p >0.05), it was significantly enhanced by adrenochrome 10^– 4 M (p <0.05). In isolated rabbit hearts, adrenochrome possesses deleterious effects on MI only at a very high concentration but not in a micromolar concentration. Therefore, it seems that aminochromes may be less cardiotoxic than catecholamines. Received: 28 February 1994 / Accepted: 2 May 1994     

Biphasic effect of cadmium ions on the secretion of leukotriene B4 in rabbit alveolar macrophages

 One major role of alveolar macrophages is the production of eicosanoids, which modulate immune and inflammatory processes in the lung. In this study, the effects were investigated of cadmium ions on the secretion of leukotriene (LT)B₄ and prostaglandin (PG)E₂, predominant products of lipoxygenase and cyclooxygenase, respectively. Cd²⁺ had an inhibitory effect on the secretion of LTB₄ and PGE₂ in response to A23187 stimulation at concentrations >3× 10^-5 M. This effect can be explained by the inhibition of arachidonic acid (20 : 4) liberation from membrane phospholipids by Cd²⁺, because Cd²⁺ inhibits both [³H]arachidonic acid (20 : 4) liberation from [³H]20 : 4-prelabeled macrophages and the cytosolic phospholipase A₂ activity. At concentrations <3×10^-5 M, Cd²⁺ had no effect on PGE₂ secretion but showed an augmentation of LTB₄ secretion. In vitro study using macrophage lysate showed enhanced LTB₄ synthesis from arachidonic acid by Cd²⁺, which could be responsible for the augmentation of LTB₄ secretion in cells. These results indicate that Cd²⁺ may increase inflammation by increasing LTB₄ production in lung. Received: 13 February 1996/Accepted: 23 April 1996     

Biliary excretion of exogenous cadmium and manganese in Long-Evans Cinnamon (LEC) rats characterized by an inherently gross amount of copper-metallothionein in the liver

Long-Evans Cinnamon (LEC) rats are characterized by the sudden onset of hepatitis around 4 months after birth and the gross accumulation of hepatic copper (Cu) accompanied by metallothionein (MT). The biliary excretion of manganese (Mn) and cadmium (Cd) injected intravenously was studied in 3-month-old LEC rats without signs of hepatitis. Injected Mn was excreted into the bile in LEC and Fischer rats used for comparison. However, increased biliary excretion of Cd was found not in the LEC rat but in the Fischer rat. Excretion of horseradish peroxidase (HRP) injected along with the metal mixture was significantly lower in the LEC group than in the Fischer group. Our results suggest that Mn excretion is not related to the existence of a gross amount of Cu-MT. Reduced excretion of Cd may be partly due to binding to Cu-MT in the liver. Decreased excretion of HRP implies the existence of an inherent defect in the bile excretion route for endo- and exogenous substances. Received: 31 January 1994 / Accepted: 11 April 1994     

Toxicity of β-blockers in a rat whole embryo culture: concentration-response relationships and tissue concentrations

Beta-adrenoceptor blockers are widely used drugs for the treatment of cardiovascular diseases. Since β-blockers cross the placenta, it is essential to consider possible adverse effects on the embryo. Six β-adrenoceptor blockers were tested at various concentrations (10 – 5000 μM) in a rat whole embryo culture. Although inducing a very similar pattern of dysmorphogenetic effects (incomplete flexure, disturbed development of the neural tube, the head, the heart and the tail bud), the compounds exhibited a wide range of embryotoxic potency. Estimation of the EC₅₀ (median-concentration producing dysmorphogenesis in 50% of the embryos) for the six compounds revealed differences of more than two orders of magnitude: propranolol 25 μM, alprenolol 30 μM, metoprolol 100 μM, pindolol 150 μM, acebutolol 500 μM, atenolol 4000 μM. Measurements of the concentrations of the various drugs in the cultured embryos at corresponding EC₅₀ levels showed differing values: metoprolol 4.5 μM, propranolol 5.2 μM, alprenolol 8.4 μM, pindolol 9.0 μM, acebutolol 12.5 μM and atenolol 77.0 μM. With regard to the EC₅₀ and the degree of substance transfer to the embryo it can be stated that propranolol and metoprolol show a much higher intrinsic potency to interfere with normal in vitro embryonic development than, e.g. atenolol. Received: 1 September 1993 / Accepted: 16 February 1994     

Distribution and reactivity of inhaled 14C-labeled toluene diisocyanate (TDI) in rats

Inhalation exposure to toluene diisocyanate (TDI) can result in a variety of airway diseases. Concern has been expressed that a putative carcinogenic potential of TDI exists as a result of the formation of toluenediamine (TDA) by hydrolysis of the isocyanate in the body. Results from long-term bioassays (TDI inhalation versus gavage in rats and mice) are contradictory and discrepancies do exist concerning the interpretation of adverse effects. This study was performed to analyze the distribution and reactivity of radioactively-labeled TDI using vapor exposure in a rat model system. Rats were exposed to ¹⁴C-TDI vapors at concentrations ranging from 0.026 to 0.821 ppm for 4 h. All tissues examined showed detectable quantities of radioactivity, with the airways, gastrointestinal system and blood having the highest levels which increased with exposure concentration. The concentration of radioactivity in the bloodstream after exposure was linear with respect to dose. The majority (74 – 87%) of the label associated with the blood was recovered in the plasma, and of this, 97 – 100% of the ¹⁴C existed in the form of biomolecular conjugates. Analysis of stomach contents shows that the majority of the label is also associated with high (>10 kDa) molecular weight species. While a larger percentage (28%) of the label is found in the low molecular weight fraction relative to blood, this low molecular weight labeled material represents at least eight different components. Thus, over the vapor exposure concentrations and time tested, it appears that conjugation is the predominant reaction and that free TDA is not a primary in vivo reaction product under the conditions tested. Received: 7 December 1993/Accepted: 7 March 1994     

Prolactin lowering activity of the retinoid Ro 14-9706 affecting lactation and pup survival

The arotinoid Ro 14-9706, though devoid of any teratogenic potential, was found to reduce dose dependently the survival of pups when their mothers were treated with toxic doses during days 6 – 15 of gestation. The increased mortality was primarily seen during early lactation. When pups derived from treated mothers were nursed by control foster mothers unexposed to the drug, their survival was significantly improved indicating that the increased mortality was not solely due to fetal drug exposure. When pups derived from untreated mothers were fostered by dams that were exposed to the arotinoid during pregnancy, a significant pup mortality (p <0.01) was observed, suggesting that the nursing behaviour of lactating dams was seriously affected. This impairment could be linked to a prolactin-suppressive activity of the arotinoid during lactation which was also seen during pregnancy. Other pituitary hormones, however, were not affected by the compound. Although the drug induced pronounced structural alterations in mitochondria of adrenocortical cells, visualized by light microscopy as extended vacuolization in the zona fasciculata and reticularis, this pathological finding did not translate into functional impairment of steroidogenesis. Thus, the arotinoid Ro 14-9706 exhibits in rats a prolactin-suppressive activity which affects lactation and subsequently pup survival. This particular endocrinological interference is a new phenomenon and uncommon for retinoids. Received: 14 June 1993/Accepted: 11 January 1994     

Toxic effects and excretion in urine of 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX) in the rat after a single oral dose

Toxic effects and excretion in urine of 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), the potent mutagenic compound in chlorinated drinking water, was evaluated in male Wistar rats by the up-and-down method. MX was dosed by gavage in deionized water at doses between 200 mg/kg and 600 mg/kg, for one animal at a time, and effects were observed for 14 days. Urine was collected in metabolism cages up to 72 h after dosing for chemical analysis of MX in urine. The animals receiving 200 mg/kg did not display clear clinical signs but at higher doses the signs of ill effects included dyspnea, laborious, wheezing and gasping breathing, decreased spontaneous motor activity, ataxia, nostril discharges, catalepsia and cyanosis. In necropsy bronchi contained foamy liquid and the lungs appeared edematous and spongy. The stomach cavity was expanded due to accumulation of fluid and gas and the gastrointestinal tract from stomach to caecum was reddish. Microscopically, the main target organ of toxicity was the gastrointestinal tract (diffuse congestion and necrosis in the mucosa). Signs of toxicity were recorded also in lungs (slight edema) and kidneys (dilated tubules, thin tubular epithelium, brownish tubular and interstitial concretion). The LD₅₀ in 48 h was 230 mg/kg. Only 0.03 – 0.07% of the dose (200 mg/kg or 300 mg/kg) was excreted in urine as intact MX. The results indicate that at high doses MX has a strong local irritating effect in the gastrointestinal tract and it probably increases liquid permeability in lungs. MX may also cause tubular damage in kidneys. Data also indicate that after an oral dose only traces of MX are excreted in urine as intact compound, suggesting that MX is subjected to intense metabolism before excretion, even at lethal doses. Received: 14 December 1993/Accepted: 28 February 1994     

Effect of norathyriol,isolated from Tripterospermum lanceolatum,on A 23 187-induced pleurisy and analgesia in mice

A 23 187-induced pleurisy in the mouse was demonstrated in this study. The protein leakage, leukocyte accumulation, LTB₄ and PGE₂ production in the pleural cavity of mice were increased by A 23 187 in a dose-dependent manner. At 7.5 nmole A 23 187 intrapleural injection, the protein level peaked at 0.5–2 h, PMN leukocytes accumulation peaked at 3–4 h, and LTB₄ and PGE₂ production peaked at 0.5–1 h. In this in vivo model we investigated the anti-inflammatory effect of norathyriol, isolated from Tripterospermum lanceolatum. A 23 187-induced protein leakage was reduced by norathyriol (ID₅₀ was about 30.6 mg/kg i.p.), indomethacin and BW 755 C. A 23 187-induced PMN leukocytes accumulation was suppressed by norathyriol (ID₅₀ was about 16.8 mg/kg, i.p.) and BW 755 C, while enhanced by indomethacin. Like BW 755 C, norathyriol reduced both LTB₄ and PGE₂ production (ID₅₀ was about 18.6and 29.1 mg/kg i.p., respectively), while indomethacin reduced PGE₂ but not LTB₄ generation. We also demonstrated the analgesic effect of norathyriol on the acetic acid-induced writhing response. Acetic acid-induced writhing response was depressed by norathyriol (ID₅₀ was about 27.9 mg/kg i.p.), indomethacin and ibuprofen. These results suggest that norathyriol, like BW 755 C, might be a dual, yet weak, cyclooxygenase and lipoxygenase pathway blocker. The inhibitory effect of norathyriol on the A 23 187-induced pleurisy and acetic acid-induced writhing response in mice is proposed to be dependent on the reduction of eicosanoids mediators formation in the inflammatory site. Correspondence to: J.-P. Wang at the above address     

Genotoxic risk for humans due to work place exposure to ethylene oxide: remarkable individual differences in susceptibility

Single strand breaks of DNA of peripheral mononuclear blood cells from 97 male and female workers occupationally exposed to ethylene oxide were analysed by the alkaline elution method. These individuals were occupied with the sterilization of medical devices in hospitals and in commercial plants. Ethylene oxide in the air of the working areas was detected up to a maximal concentration of 16.5 mg/m³ calculated as 4-h time-weighted average (4h TWA). Mean value was 1.47±0.52 mg/m³ (1 mg/m³ = 0.55 ppm). Compared to the mean elution rate of the DNA from non-smoking workers exposed to air concentrations of ethylene oxide below the detection limit of 0.1 mg/m³ (4h TWA) the non-smokers working in rooms with a concentration of ethylene oxide between 0.5 mg/m³ and 2 mg/m³ showed a statistically significant (P <0.05) 119% higher mean elution rate and even for the non-smokers exposed to 0.1 – 0.5 mg/m³ of ethylene oxide a statistically significant (P <0.05) 53% higher mean elution rate was observed. For smokers a similar tendency was found but the increase in elution rates in response to the external exposure was smaller than in non-smokers and no statistical significance was obtained. According to their sensitivity to ethylene oxide the non-smoking workers could be classified into two subpopulations. In the majority of the non-smokers (67%) approximately 5-fold more DNA strand breaks were induced by ethylene oxide than in the other non-smokers. A lowest detectable effect level could only be specified for non-smokers. For the “higher sensitive” group the lowest detectable effect level in an examination of a single individual was calculated to be 0.6 mg/m³ ethylene oxide in the air (4h TWA). For the “lower sensitive” group a lowest detectable effect level was calculated to be 3.5 mg/m³. Received: 18 October 1993/Accepted: 16 February 1994     

Studies of the biochemical toxicology of uranyl nitrate in the rat

High resolution ¹H NMR spectroscopy of urine and plasma, conventional clinical chemical methods and histopathology have been applied to investigate the effects of uranyl nitrate (UN) on renal function and biochemistry in the Fischer 344 (F344) rat. Administration of UN (5 – 20 mg/kg) to male F344 rats resulted in a dose-related proximal nephropathy assessed conventionally by histopathology and urinary excretion of N-acetyl-β-d-glucosaminidase (NAG), and related to changes in the patterns of low MW metabolites observed in 400 MHz ¹H NMR spectra of urine. The changes in urinary metabolite profiles included elevations in glucose accompanied by minor elevations in certain amino acids (alanine, valine and glutamate). ¹H NMR urinalysis also revealed altered excretion of low MW metabolites which are not routinely measured, such as l-lactate, acetate, citrate, succinate and 2-oxoglutarate (2-OG). In addition, the striking appearance of high concentrations of 3-d-hydroxybutyrate (HB) in the urine was noted, in the absence of acetoacetate or acetone, and it is suggested that this may provide a new marker of proximal tubular damage for certain types of nephrotoxic mechanism. Broadening of the ¹H NMR signals of citrate following 10 mg/kg UN was shown to be due to a dynamic exchange process involving chelation with urinary Ca²⁺ and Mg²⁺ ions. Conventional biochemical analysis of plasma from UN-treated rats revealed dose-related increases in creatinine, urea and HB concentrations. ¹H NMR-detected evidence of raised alanine aminotransferase (ALT) levels in rats administered the highest dose of UN was indicated by the partial deuteration of alanine in lyophilised plasma reconstituted in ²H₂O. The degree of ¹H NMR-detected abnormalities agreed well with histopathological observations and conventional biochemical indices of nephrotoxicity and more fully characterised the renal changes produced by UN. The significance of HB-uria in UN-induced proximal nephropathy is discussed in relation to biochemical observations on other proximal nephrotoxins. Received 7 June 1993/Accepted 23 August 1993     

Comparative pharmacokinetics of dirithromycin and erythromycin in normal volunteers with special regard to accumulation in polymorphonuclear leukocytes and in saliva

Objective: In a randomized cross-over study, we assessed pharmacokinetics and intracellular concentrations in polymorphonuclear leukocytes (PMN) and saliva of erythromycin and erythromycylamine, the active metabolite of dirithromycin. Methods: Ten healthy volunteers received 1 g erythromycin b.i.d. or 500 mg dirithromycin qd for 5 days (wash out period, 35 days). Concentrations of erythromycin and erythromycylamine were measured in serum, urine, saliva, and granulocytes by bioassay and high-performance liquid chromatography (HPLC) on days 1, 3, and 5 of each study period, respectively. Results: While maximal serum concentrations (C_max) and the area under the data (AUD_tot) of erythromycin were significantly higher (C_max 1.44 mg · l⁻¹, AUD_tot 5.66 mg · h · l⁻¹) than those of erythromycylamine (C_max 0.29 mg · l⁻¹, AUD_tot 1.96 mg · h · l⁻¹), erythromycylamine had a significantly higher mean residence time (21 h) than erythromycin (5.5 h). Erythromycylamine accumulated significantly more in PMN than erythromycin;␣the accumulation factor of erythromycylamine was 100 with a maximal intracellular concentration of 13.4 mg · l⁻¹, whereas the maximal accumulation factor of erythromycin was 4 with a maximal intracellular concentration of 6.1 mg · l⁻¹. There were no significant differences in maximal saliva concentrations (erythromycin 0.35 mg · l⁻¹, erythromycylamine 0.31 mg · l⁻¹). Received: 16 September 1996 / Accepted in revised form: 12 February 1997     

Effect of bolesatine on phospholipid/calcium dependent protein kinase in vero cells and in rat thymus

 Bolesatine, a glycoprotein from Boletus satanas Lenz, has previously been shown to be mitogenic to rat and human lymphocytes at very low concentrations, whereas higher concentrations inhibit protein synthesis in vitro and in several in vivo systems. The mechanism whereby this mitogenic activity occurs was previously unknown. To elucidate this mechanism, the effects of bolesatine have been studied in a cell-free system, VERO cells in culture, and in rat thymus. Bolesatine was found to activate PKC, in vitro (cell free system), in VERO cells, and in vivo in rat thymus. In a cell-free system, bolesatine appears to be a direct effector of PKC. The activation is concentration dependent for 1 – 10 ng/ml. At the same time, VERO cells significantly proliferate when incubated with bolesatine (3, 5 and 10 ng/ml), since the DNA synthesis increases by 27, 48 and 59%, for respectively, 3, 5 and 10 ng/ml compared with control. Moreover, Bolesatine (5 and 10 ng/ml) induces InsP₃ release in a concentration-dependent manner (114 and 142%) as compared to control. In vivo, 24 h after oral administration of bolesatine to rats (20, 100 and 200 μg/kg), PKC activity is significantly increased in thymus. The most effective doses (100 and 200 μg/kg) give 590 – 620% increase in cytosolic PKC activity and 85 – 91% increase in total PKC activity as compared to control. This PKC activation by bolesatine in rat thymus is directly linked to the mitogenic activity observed in vivo. Bolesatine is thus capable of activating the PKC directly and/or indirectly (via InsP₃ release) during its mitogenic process. Received: 6 February 1995 / Accepted: 20 March 1995     

Effect of sodium pyridinethione on the uptake and distribution of nickel in rats,ferrets and guinea-pigs

Oral administration of sodium pyridinethione together with Ni²⁺ (using ⁶³Ni²⁺ as a tracer) to rats, ferrets and guinea-pigs produced highly increased tissue levels of the metal in several tissues in comparison with animals given the Ni²⁺ alone. Ni²⁺ forms a lipophilic complex with pyridinethione and it can be assumed that a facilitated passage of the Ni²⁺ across the cellular membranes of various tissues is important for the observed effects. Pigmented tissues (e. g. the eye melanin), the pancreatic islets, the nervous system and striated muscles showed high levels of Ni²⁺ in animals given sodium pyridinethione. However, in some instances marked species differences were observed. Thus, microautoradiography indicated an uptake of Ni²⁺ both in the β- and α-cells in the pancreatic islets in the rat, whereas in the guinea-pig only some cells (probably the α-cells) accumulated high levels of Ni²⁺. In the ferret sodium pyridinethione induced a high uptake of Ni²⁺ in the heart muscle, which was not seen in the other species. The Ni²⁺ is probably taken up in the various tissues complexed to pyridinethione. Within the tissues the complex may dissociate and the Ni²⁺ may bind to some endogeneous tissue components. The affinity of the Ni²⁺ for the endogeneous ligands in relation to the affinity for the pyridinethione may be of importance for the effects on the disposition of the Ni²⁺. The species variations may be related to differences in the structural conformations of the endogeneous Ni²⁺-binding ligands. Received: 25 October 1993/Accepted: 25 January 1994     

Influence of the route of administration and the chemical form (MnCl2, MnO2) on the absorption and cerebral distribution of manganese in rats

  The absorption and cerebral distribution of manganese (Mn) have been studied with respect to the route of administration and the chemical form of the Mn compound. Different groups of adult male rats received either MnCl₂ · 4H₂O or MnO₂ once a week for 4 weeks at a dose of 24.3 mg Mn/kg body wt. (b.w.) by oral gavage (g.) or 1.22 mg Mn/kg b.w. by intraperitoneal injection (i.p.) or intratracheal instillation (i.t.). Control rats were treated with 0.9% saline. Four days after the last administration the rats were killed and the concentration of Mn measured in blood, hepatic and cerebral tissues (cortex, cerebellum, and striatum). The liver Mn concentration was not affected by the treatments whatever the chemical form or the route of administration of the Mn compound. Administration of MnCl₂ by g., i.p., and i.t. routes produced equivalent steady-state blood Mn concentrations (about 1000 ng Mn/100 ml), representing increases of 68, 59, and 68% compared with controls, respectively. Mn concentrations were significantly increased in the cortex but to a lesser extent (g., 22%; i.p., 36%; i.t., 48%) and were higher in the cerebellum after i.p. and i.t. administrations than after oral gavage. Rats treated i.t. with MnCl₂ showed an elective increase of the striatal Mn concentration (205%). In contrast, MnO₂ given orally did not significantly increase blood and cerebral tissue Mn concentrations; the low bioavailability is most likely due to the lack of intestinal resorption. Administration of MnO₂ i.p. and i.t., however, led to significant increases of Mn concentrations in blood and cerebral tissues. These increments were not significantly different from those measured after MnCl₂ administration, except for striatal Mn after i.t. which was markedly less (48%) after MnO₂ administration. A comparison of the blood Mn kinetics immediately after g. and i.t. treatment with MnCl₂ or MnO₂ indicated that the higher elevation of blood Mn concentration ( > 2000 ng Mn/100 ml) after i.t. administration of MnCl₂ could account for the elective uptake of Mn in the striatum observed in repeated dosing experiments. It is concluded that the modulation of Mn distribution in brain regions according to the route of administration and the chemical form of the Mn compound may be explained on the basis of different blood Mn kinetics and regional anatomic specificities of the striatal region. Received: 2 May 1996 / Accepted: 21 August 1996     

Effect of Sodium Naproxen on Inflammatory Response Induced by Anterior Chamber Paracentesis in the Rabbit

This study evaluated the effect of sodium naproxen (a reversible competitive inhibitor of cyclo-oxygenase) and phenylephrine (a mydriatic α-adrenergic agent) eye drops in maintaining atropine mydriasis in the rabbit after paracentesis. Moreover, to assess the influence of these treatments on vascular and cellular inflammatory responses in the rabbit eye, several biochemical parameters were considered. Anterior chamber paracentesis significantly reduced atropine-induced mydriasis and a parallel elevation of proteins, polymorphonuclear leucocytes (PMNs), prostaglandin E₂ (PGE₂) and leukotriene B₄ (LTB₄) levels in the secondary aqueous humour (obtained 120 min later) was observed. A significant increase in PMNs in the aqueous humour and a parallel increase in myeloperoxidase activity, a measure of PMN infiltration, in the iris-ciliary body were detected. Atropine-induced mydriasis was maintained in rabbits treated with either sodium naproxen or phenylephrine eye drops. However, only in the former group were the inflammatory parameters significantly reduced, with the exception of aqueous LTB₄ levels. The inhibition of the protein influx in the aqueous humour and of the miosis produced by sodium naproxen can be related to the high drug levels in the aqueous humour that were effective in inhibiting the cyclo-oxygenase pathway of arachidonic acid metabolism, whereas the effects on PMN infiltration appear to be independent of significant release of the potent chemotactic agent LTB₄, synthesized via the 5-lipoxygenase pathway.     

Postnatal development and behaviour of Wistar rats after prenatal toluene exposure

Pregnant Wistar rats were treated with different concentrations of toluene by inhalation (300, 600, 1000 and 1200 ppm) from day 9 to day 21 of pregnancy for 6 h a day in a whole-body inhalation chamber (controls inhaled fresh air only). From day 22, rats were kept single-caged and were allowed to deliver. Besides a detailed evaluation of the physical development of the offspring we performed the following tests: forelimb-grasp reflex, righting reflex, cliff-drop aversion reflex, maintainance of balance on a rotating rod, measurement of locomotor activity and learning ability in a discrimination learning test. A toluene exposure of 1200 ppm resulted in a reduced body weight of rat dams and offspring and a higher mortality until weaning. The physical development (incisor eruption, eye opening and vaginal opening) was retarded in this group. There were no clear-cut and concentration-dependent differences in the development of reflexes, rota rod performance and locomotor activity between the offspring of animals exposed to toluene and the controls. Likewise, no effects were found on learning ability in the operant conditioning task. Compared to the controls there were no differences in mating, fertility and pregnancy indexes in the F₁-generation. The tests performed have provided no evidence that toluene exposures ≤ 1200 ppm induce adverse effects on the behaviour of rat offspring exposed during late embryonic and fetal development. Received: 17 July 1996 / Accepted: 20 September 1996     

Plasma membrane hyperpolarization by cyanide in chromaffin cells: role of potassium channels

Exposure of rat pheochromocytoma (PC12) cells to cyanide produces elevation of cytosolic calcium, impaired Na⁺-H⁺ exchange, membrane lipid peroxidation and release of neurotransmitters. Since these observations suggested cyanide alters plasma membrane function, the present study examined the effect of NaCN on the membrane potential of undifferentiated PC12 cells in suspension. In PC12 cells loaded with the voltage sensitive fluorescent dye, bis-oxonol, cyanide (2.5 – 10 mM) elicited an immediate (within seconds), concentration related decrease in fluorescence, indicating hyperpolarization of the plasma membrane. Increasing extracellular K⁺ concentration to 20 mM blocked the effect of cyanide (5 mM), suggesting cyanide increased K⁺ efflux. Pretreatment with quinine blocked the cyanide-induced hyperpolarization, whereas glyburide had little effect, showing the hyperpolarization produced by cyanide was due to activation of Ca²⁺ sensitive K⁺ channels. Removal of Ca²⁺ from the media did not influence cyanide-induced hyperpolarization. However, buffering intracellular Ca²⁺ by loading cells with the Ca²⁺ chelators, Quin II or BAPTA, abolished the cyanide effect, showing cytosolic Ca²⁺ is a key factor. These findings suggest that cyanide mobilizes Ca²⁺ from intracellular stores which leads to hyperpolarization via the activation of Ca²⁺ sensitive K⁺ channels. Received: 7 October 1993/Accepted: 11 January 1994