The effect of in vitro exposure to white spirit on [Ca2+i] in synaptosomes from rats exposed prenatally to white spirit.

Female rats were exposed to white spirit (400 and 800 ppm for 6 hr/day) at day 7-20 during pregnancy. Thirty-five days after birth all female offspring were sacrificed, the brains removed, and the synaptosomal fractions prepared for in vitro studies. The cytosolic calcium concentration was measured using the FURA-2 technique. The results show that cytosolic calcium was increased in synaptosomes from rats exposed to white spirit prenatally compared to synaptosomes from unexposed rats. When synaptosomes were exposed to white spirit in vitro, the cytosolic calcium concentration changes were identical in all groups of rats. The membrane leakage measured as FURA-2 leakage from the synaptosomes identical in all three groups of animals. The results suggest that prenatal exposure to white spirit induces long-lasting and possibly irreversible changes in calcium homeostasis in the rat nervous system.

The effect of ageing and in vitro exposure to xylene and KCl on [Ca2+]i in synaptosomes from rats exposed prenatally to xylene.

Female rats (Mol: WIST) were exposed prenatally to 500 p.p.m. of technical xylene on days 7-20. At the age of fourteen months the rats were sacrificed and the synaptosomal fraction prepared for in vitro studies. The cytosolic calcium concentration was measured using the FURA-2 technique. The cytosolic calcium was increased in synaptosomes from old rats compared to those from rats at the age of three months, but no effect of prenatal exposure was seen. When synaptosomes were incubated with xylene, potassium or both, the cytosolic calcium concentration was changed identically in all groups of rats. When synaptosomes were incubated simultaneously to xylene and potassium a dramatical leakage of FURA-2 was observed. The mechanisms behind the membrane leakage are discussed.

Late postnatal growth in the dentate area of the rat hippocampus compensates for volumetric changes caused by early postnatal toluene exposure.

The present report assesses the reversibility of previously described quantitative morphological alterations in the dentate area of the hippocampi of rats exposed to 500 ppm toluene for 12 h daily from postnatal day 1 (P1) to P28. Following the exposure, rats were allowed to survive until P120. The volumes of hippocampal components, as they appear in Timm-stained histological sections, were determined using the Cavalieri estimator. At P120, no differences were apparent in the volumes of the dentate components of control and experimental animals. The differences observed immediately after exposure were eliminated by significantly larger increases in the sizes of early affected components in experimental animals. Different periods of neurogenesis in rodents and primates are briefly discussed as possible confounders of rodent models of permanent toluene-induced morphological alterations in primates.

Effect of organic solvents on nervous cell membrane as measured by changes in the (Ca2+/Mg2+) ATPase activity and fluidity of synaptosomal membrane.

The effect of various solvents on the central nervous system was studied by using rat brain synaptosomal membranes as an in vitro model. The activity of (Ca2+/Mg2+) ATPase and the membrane fluidity was determined. The alteration of the ATPase activity depended on the physio-chemical characteristics of the solvent in question. Incubation with aliphatic alkanes caused a stimulation of the ATPase activity whereas mixed hydrocarbons as kerosene, white spirit and gasoline inhibited the enzyme. Incubation with chlorinated hydrocarbons caused a biphasic response dependent on the concentration. Oxygen-containing hydrocarbons exhibited various effects as found after incubation with hydrocarbons. The different effects of the solvents on the ATPase activity suggest that the lipophilicity of the solvents is one of more parameters affecting the membrane. Furthermore, the biphasic response following the incubation with chlorinated hydrocarbons indicates that more mechanisms are involved in the enzyme effect. The membrane fluidity is increased with higher concentrations of the solvents. From the results it is concluded that the ATPase activity depends not only on the membrane fluidity and volume, but also on the hydrophilic vicinity of the enzyme molecule.

Effects of simultaneous ethanol and toluene exposure on nerve cells measured by changes in synaptosomal calcium uptake and (Ca2+/Mg2+)-ATPase activity.

The effect of simultaneous exposure of rats to toluene and ethanol on synaptosomal calcium uptake and (Ca2+/Mg2+)-ATPase activity was studied. Rats were exposed to 500 p.p.m. toluene by inhalation for 12 hr a day during four weeks. During the exposure period, the rats had access to 5% sucrose solution containing 20% ethanol or to 5% sucrose solution alone. Rats drinking ethanol exhibited a smaller weight gain than rats drinking water alone. Furthermore, rats exposed simultaneously to toluene and ethanol had a higher ethanol intake than unexposed rats. The toluene exposure caused a higher synaptosomal calcium uptake in vitro. Ethanol intake did not change the synaptosomal calcium uptake in vitro. The synaptosomal calcium uptake in rats exposed to toluene and ethanol was nearly identical to that measured in control rats. In vivo exposure to toluene, or ethanol, or toluene/ethanol simultaneously did not affect the (Ca2+/Mg2+)-ATPase activity in vitro. Incubation with toluene in vitro decreased the (Ca2+/Mg2+)-ATPase activity in a concentration dependent manner. Ethanol had only a slight effect on the enzyme. Simultaneous incubation with toluene and ethanol showed an antagonistic effect of ethanol on the toluene inhibition of the ATPase activity.

The effects of alcohols in vitro on the nervous cell membrane measured by changes in the (Ca2+/Mg2+) ATPase activity and fluidity of the synaptosomal membrane.

The effect of various alkanols on the central nervous system was studied by using rat brain synaptosomal membranes as an in vitro model. The activity of (Ca2+/Mg2+)ATPase and the membrane fluidity were determined. The n-alkanols exhibited an increased molar inhibition of the ATPase activity with an increase in the carbon chain length up to 1-octanol. 1-octanol and 1-decanol caused a biphasic effect on the ATPase activity depending on the alkanol concentration, whereas 1-dodecanol caused a stimulation of the ATPase activity. All alkanols studied caused an increased fluidity of the membrane. Our results indicate that the effect of alkanols on the ATPase activity depends on changes in the border layer between the membrane and the surrounding medium and on a binding of the alkanols to the enzyme molecule. Furthermore, the two-way effect of 1-octanol and 1-decanol and the stimulatory effect of 1-dodecanol indicate that more mechanisms are involved. In addition, the results indicate that changes in the membrane fluidity do not seem to be a prerequisite of the ATPase inhibition.

The effect of low-level toluene exposure on the developing hippocampal region of the rat: histological evidence and volumetric findings.

With the intention of investigating possible morphological alterations effected by toluene in the developing CNS, rat pups were exposed to 100 ppm and 500 ppm of atmospheric toluene from postnatal day 1 until sacrifice at postnatal day 28, when the hippocampal region (area dentata, Ammon's horn, subiculum) was examined light microscopically and alterations in the volumes of the layers of the subdivisions were determined. The layers of Ammon's horn and the subiculum were not affected qualitatively or quantitatively by the 500 ppm exposure. Within the area dentata, the volume of the granule cell layer was 6% smaller in animals exposed to 100 ppm and 13% smaller in animals exposed to 500 ppm than they were in controls. The volumes of the hilus, which is a terminal field of granule cell axons, and the commissural-associational zone of the dentate molecular layer, which is the terminal field of the hilar projection to the granule cells, were smaller (12% and 19%) in animals exposed to 500 ppm than they were in controls. Argyrophilic cells were found in the granule cell layer of all animals exposed to 500 ppm. Pronounced granule cell degeneration was found in one animal exposed to 500 ppm. The granule cell layer of animals exposed to 100 ppm appeared qualitatively normal. The alterations reported here support the few earlier reports of morphological alterations in the CNS of adult laboratory animals. Effects of toluene similar to those described, that is alterations in specific neuron populations and their afferent and efferent terminal fields may complement changes in neurophysiology and behavior that have been observed in prenatally and perinatally exposed rodent pups. Causal relationships, however, remain to be elucidated.

The effect of toluene exposure for up to 18 months (78 weeks) on the (Ca2+/Mg2+)ATPase and fluidity of synaptosomal membranes isolated from rat brain.

The effect of toluene on the central nerve system was studied by using rat brain synaptosomal membranes as in vitro and in vivo models. The activity of Ca2+/Mg2+ ATPase and the membrane fluidity were determined. Short-term exposure in vivo to 500 p.p.m. of toluene had an inhibitory effect on the enzyme studied whereas long-term exposure to toluene caused an increased activity. Exposure to toluene had no effect at all on the membrane fluidity. The in vitro experiment showed an effect of toluene on both parameters. The alteration in the enzyme activity and membrane fluidity was parallel in the exposed animals as well as those of control. Our results show that long-term exposure to toluene affects nerve cell membranes by other mechanisms than those observed under in vitro conditions.

Calcium uptake in brain synaptosomes from rats exposed to daily toluene for up to 80 weeks.

Rats were exposed to toluene, 500 p.p.m., for 12 hrs/day for up to 80 weeks. The brains were removed and the synaptosomes were prepared. Potassium-stimulated and unstimulated synaptosomes were incubated with 45Ca2+ for 1/2, 2, 4, 8, and 16 min. Toluene exposure for 4 and 12 weeks caused a significant, approximately 20%, increase in 45Ca2+ uptake into unstimulated synaptosomes. The effect was of moderate quantity and transient, since it was not significant after 30 and 80 weeks of exposure. It seems doubtful whether the demonstrated change in calcium uptake should have any relevance in connection with the "organic solvent neurotoxicity syndrome".

Metabolism of 14C-codeine in the isolated, perfused rat liver.

The metabolism of 14C-codeine in the isolated rat liver was studied in single-pass and recirculation perfusion experiments. The perfusate, a semi-synthetic medium with bovine erythrocytes, was delivered at a constant rate (12 ml/min.) and contained codeine in the range of 3-57 nmol/ml. Samples of perfusate were collected and analyzed for codeine and its metabolites after extraction and thin-layer chromatographic separation. In single-pass perfusion, steady state was reached within 20 min. The codeine concentration in the effluent perfusate varied from 17 to 48% of that in the affluent corresponding to extraction ratios of 0.83 to 0.52. There was a significant negative correlation between codeine dose and extraction ratio (r = 0.86, P less than 0.05, N = 6). The steady state concentration of free and conjugated morphine made a total of 21 to 49% of the molar concentration of codeine at the inflow side. The recovery of radioactivity at the end of the perfusions was on an average 89%. In recirculation perfusion experiments the codeine extraction ratios varied from 0.65 to 0.35. The amount of free morphine in the reservoir increased to a maximum within 25-45 min. Our results suggest a relatively high hepatic first-pass metabolism of codeine in the rat which is apparently dose-dependent. The quantitatively most significant metabolites of codeine are morphine and conjugated morphine. The rate and extent of morphine formation is compatible with the hypothesis that metabolically produced morphine may be responsible for the analgesic effect of codeine.