Identification of infant and adult swine susceptible to enterotoxigenic Escherichia coli by detection of receptors for F4(K88)ac fimbriae in brush borders or feces.

We attempted to determine F4(K88)-adhesive and non-adhesive phenotypes of infant (neonatal < 3 day old and weaned < 4 week old pigs) and adult (> 6 month old) swine by ELISA using immobilized F4(K88)ac fimbrial antigen or whole F4(K88) + E. coli cells (strains M1823 and 1476) and isolated small intestinal brush borders or easily-obtainable fecal samples from the same animals. Nineteen of 22 neonates (86%), 17 of 20 weaners (85%), and 26 of 39 adults (67%) were classified identically as F4(K88) receptor-positive or negative by the ELISA. The ELISA with feces from adult swine was found to be almost equally specific (87%) as that with feces from neonatal (90%) and weaned (91%) pigs. However, the sensitivity of the assay was low (38%), indicating that fecal samples from adults contained less receptor-material than necessary for comparable phenotyping. The receptor-positive brush borders from neonates and weaners reacted significantly better (P < 0.02, < 0.001 respectively) with purified F4(K88) antigen than did those from adults. There was good agreement between the average ELISA values for feces from infant and adult swine regardless the source of coating antigen applied. With this assay we can determine F4(K88) phenotypes of infant swine using easily-collected fecal samples rather than isolated brush borders. It was also concluded that tested feces is not an acceptable alternate source of the receptor-material to brush borders from F4(K88)-susceptible adult swine.


Occurrence of fimbriae and enterotoxins in Escherichia coli strains isolated from piglets in Poland.

1125 and 1146 E. coli strains isolated from suckling and weaned piglets with diarrhea, respectively, and 724 strains from healthy piglets were tested for the presence of fimbriae and production of enterotoxins. The fimbriae were determined by hemagglutination and slide agglutination tests, enterotoxins--by the use of ileal loop test in piglets (LT and STb enterotoxins) and suckling mouse assay (STa enterotoxin). It was found that 72.8 and 53.0% strains, isolated from diseased suckling and weaned piglets, respectively, possessed specific fimbrial hemagglutinins, in most cases with K88 antigen. Additionally, 987P fimbriae were detected in 14.0 and 0.7% strains isolated from piglets with diarrhea. Only 5 strains (0.7%) recovered from healthy piglets had specific fimbriae, usually with undetermined antigenic structure. F1 fimbriae (called common or unspecific) were found in strains isolated both from diseased (15.2 and 16.3% strains, respectively) and healthy piglets (27.1% strains). It was noted that the strains isolated from suckling and weaned piglets with diarrhea in most cases were enterotoxigenic (90.5 and 69.1% strains, respectively) and most frequently produced heat-labile toxin LT alone or with STb. 18.5% of enterotoxigenic strains isolated from healthy piglets produced STa toxin.


Characterization of chicken intestinal brush border membrane Na/H exchange.

1. Na/H exchange is the major pathway for Na uptake in brush border membrane vesicles from chicken small intestine. Hanes-Woolf analysis demonstrated that Na and H competed at the same extravesicular site. The KNa for Na+ at extravesicular pH 6.6 is 35 mM and at pH 7.4, 12 mM. 2. Similar to mammalian intestinal cells, the Na/H exchanger does not appear to have an internal proton modifier site. Varying intravesicular pH from 6.1 to 7.8 stimulates uptake, but a sigmoidal relationship is not observed. 3. The ability of several amiloride analogs to inhibit the exchanger was tested and the inhibitory profile was similar, but not identical to Na/H exchangers in mammalian tissues. The potency series (from most to least potent) is hexamethylamiloride approximately ethylisopropylamiloride > methylisobutylamiloride > dimethylamiloride >> amiloride.


Phosphatase activity in the hepatopancreas of Helix aspersa.

1. Phosphatase acid (PhA) activity in the digestive gland (hepatopancreas) of the common garden snail Helix aspersa has been investigated using cytochemical methods. 2. All the cells composing this gland show PhA activity, the distribution pattern differing according to the cell type. 3. The digestive cells show the most widely distributed reaction product (brush border, phagolysosomes, multivesicular bodies and autophagic vacuoles). 4. In the excretory cells this activity appears in large sacs, while in the calcium cells the reaction product is abundant in the calcium granules. 5. Cellular digestion processes performed by each of these cell types is discussed together with their role in the detoxification of heavy elements derived from the environment.


Anuran amphibia which are not acclimable to high salt, tolerate high plasma urea.

1. The capacity of five anuran Amphibians (Bufo viridis, B. regularis, Rana ridibunda, Hyla arborea and Pelobates syriacus) to acclimate to NaCl and urea solutions was investigated. 2. All species could be acclimated to relatively high concentrations of urea solutions, while only Bufo viridis and Hyla arborea could be acclimated to 500 mOsm/kg or higher NaCl solutions. 3. The plasma urea concentration in B. viridis and H. arborea was elevated to levels over 140 mmol/l. 4. The sum of plasma sodium and chloride concentrations did not increase over 400 mmol/l in any species. 5. Urine osmolality, which was normally low, increased, but never exceeded the plasma osmolality. 6. In the urea acclimation conditions, urine electrolytes diminished, similarly in all species in this study. 7. It is concluded that anuran Amphibians can tolerate high plasma urea concentrations, but only those species which can elevate it, either through retention or net synthesis, can be acclimated to high salt solutions.


Effects of a low-sodium diet on electrolyte transport in the proximal and distal colon of the guinea pig (Cavia porcellus).

1. Animals were randomly assigned to three different treatment groups. (I) Control group maintained on a standard guinea pig diet, (II) a group obtaining a low-Na diet (0.02% Na w/w) and (III) a group fed the low-Na diet and in addition being treated with K-canrenoate prior to the perfusion experiment. 2. Proximal and distal colonic segments of anesthetized guinea pigs were perfused simultaneously with Ringer's solution. 3. In the proximal colon of the control group Na and Cl were absorbed in equal quantity and K was secreted. 4. In the distal colon of the control group Na and K were absorbed and Cl was secreted. Na absorption was only approximately 1/10 of that in the proximal colon. 5. Feeding a low-Na diet for two weeks induced secondary hyperaldosteronism. Plasma aldosterone concentration increased 3.4-fold. Na and Cl absorption rose 34% and 27% above control in the proximal colon, respectively. In the distal colon Na absorption was elevated 3.7-fold above control. 6. Na absorption and transepithelial potential difference (PD) in the proximal colon were amiloride-insensitive in all groups. In the distal colon Na absorption was abolished by luminal amiloride and the transepithelial PD was reversed in polarity. 7. K-canrenoate treatment prevented the increase in Na transport in the distal colon.


Carbonic anhydrase activity in blood of early sheep fetuses.

1. The level of carbonic anhydrase activity in the red cells was measured in sheep fetuses at different times after conception: 39, 56, 77, 90 and 140 days, the last being close to full term. Measurements were also made on blood from four of the mothers. 2. There was a low level of the enzyme present in the 39 day fetuses (0.037 enzyme units (E.U.)/100 micrograms Hb) and its increase up to 90 days of gestation (0.19 E.U./100 micrograms Hb) had a form approximating exponential. 3. The earliest levels were only 11% of the full term levels and only 4% of the adult levels previously reported. 4. Even the earliest samples were of blood that was fetal rather than embryonic but these results are the earliest carbonic anhydrase activities reported in this mammal.


The transfer of thyroxine from the mother to the young of the marsupials, the bandicoot (Isoodon macrourus) and the possum (Trichosurus vulpecula).

1. The young of the bandicoot and possum are of similar weight at birth (200-250 mg) but by day 50 post-partum have weights of 100 and 20 g, respectively. The aim of this study was to determine whether the disparate growth rate was associated with a difference in the transfer of thyroxine from mother to young. 2. Radioactive thyroxine was injected intramuscularly into the lactating female and 24 hr later a blood sample was obtained from the mother and the pouch young removed. The amount of radioactive thyroxine remaining in the blood and within the young was determined. 3. The data obtained indicated that milk is a source of thyroid hormones in early pouch life. 4. The hypothesis that greater quantities of thyroxine are transferred in the milk of bandicoots than that of brushtail possums is not supported.


Alterations of cytoplasmic enzyme activities of liver in rats fed on diets containing orotic acid with dietary fibers.

1. Rats were fed with diets containing orotic acid with dietary fibers to induce alteration of lipid metabolism and cytoplasmic enzyme activities in the livers. Concentrations of blood glucose, serum insulin and free fatty acids (FFA) were measured. 2. In CP (cellulose + orotic acid) group compared to basal (cellulose - orotic acid) or CBH (corn bran hemicellulose + cellulose + orotic acid) groups, glucose-6-phosphate dehydrogenase, pyruvate kinase and ATP citrate lyase activities and FFA concentration were decreased according to lipid accumulation of livers. 3. In the CBH group, glycerokinase activity was higher than basal group.


Changes in body composition of germ-free and conventional chickens during starvation.

1. Changes in body composition during starvation were compared between germ-free (GF) and conventionalized (CVL) chicks in experiment 1. At 8 days of age, the GF birds were divided into two groups, i.e. GF and CVL groups. The CVL birds were inoculated with faeces from conventionally reared birds. Until 14 days of age, both birds were fed a diet ad lib. and thereafter starved for 6 days. 2. Nitrogen loss during starvation was significantly lower in CVL birds, though the reverse was true for water loss. Fasting heat production was comparable between two environments. 3. Influence of the gut microflora on body weight and nitrogen losses during starvation was investigated in birds prefed diets high or low in dietary protein in experiment 2. 4. No significant effect of the gut microflora was observed in body weight and nitrogen losses. Body weight was severely reduced in birds prefed the high protein diet and nitrogen loss was lower in birds prefed the low protein diet.


Gut microflora can modify fatty acid composition in liver and egg yolk lipids of laying Japanese quail (Coturnix coturnix japonica).

1. The influence of the gut microflora on lipid metabolism was investigated in germ-free (GF) and conventional (CV) laying Japanese quail. 2. Serum and egg yolk cholesterol concentrations showed comparable values in both GF and CV environments. 3. The fatty acid composition of liver lipids was modified by the presence of gut microflora. Notably, in the presence of the gut microflora, proportion of oleic acid was reduced and conversely, stearic and linoleic acids were enhanced. 4. In egg yolk lipids, the proportion of myristoleic and palmitoleic acids was significantly lowered and that of stearic acid was significantly enhanced by the presence of the gut microflora, though the difference was very small. 5. It was suggested that oleic acid could be easily either hydrogenated to stearic acid or desaturated to linoleic acid by the action of the gut microflora in Japanese quail.


Comparison of fibre digestion and digesta retention time between nutrias (Myocaster coypus) and guinea-pigs (Cavia porcellus).

1. Digestibilities of feed, and transit and retention time of fluid and particle digesta marker measured in nutrias (Myocaster coypus) and guinea-pigs (Cavia porcellus) fed on a diet containing 50% alfalfa. 2. The digestibility of fibre was higher in the nutria, along with the longer retention time of digesta. 3. The liquid and particle marker were similarly excreted, suggesting no separation mechanism in the gastrointestinal tract of both the animals. 4. The apparent digestibility of protein in the nutria was superior to the guinea-pig and other small hindgut fermenters, suggesting that the contribution of coprophagy on protein nutrition of nutrias is significant.


Comparative effects of the water accommodated fraction of three oils on mussels. 1. Survival, growth and gonad development.

1. Mussels, Mytilus galloprovincialis, were exposed to the water accommodated fraction (WAF) of two crude oils (types Ural and Maya) and of a commercial lubricant oil for 91 days. Synchronous spectrofluorometry showed that the aromatic hydrocarbon composition of the WAFs obtained from the three oils were significantly different (Maya-WAF heaviest, Ural-WAF lightest). 2. No significant mortality occurred in mussels exposed to the WAFs of the two crude oils, while all the animals exposed to lubricant-WAF died after 49 and 77 days in intermediate and high doses respectively. 3. Flesh and shell growth of mussels exposed to the WAFs was reduced significantly with respect to controls as demonstrated by several size, condition and allometric parameters. The response to petroleum hydrocarbon exposure is not dose-dependent, maximal responses being measured in low (Ural) and intermediate (Maya) doses. This seems to depend on a behavioural reaction (shell valve closure) against high doses of pollutants which may avoid the sublethal damage to some extent. 4. Shell abnormalities or changes in shell thickness were not detected. 5. The histological study of gonad tissue revealed that gamete release is retarded and hemocytic infiltration of mature gonad tissue is increased after exposure to low and intermediate doses of the WAFs. On the other hand, spawning is readily induced under exposure to high doses of the WAFs.


Comparative effects of the water accommodated fraction of three oils on mussels. 2. Quantitative alterations in the structure of the digestive tubules.

1. Mussels have been exposed to the water accommodated fraction (WAF) of Ural and Maya crude oils and of a lubricant oil for 91 days and the digestive gland structure has been studied (a) by planimetry, to calculate the mean epithelial thickness (MET), the mean diverticular radius (MDR) and the mean luminal radius (MLR) of the digestive tubules and (b) by subjective tubule grading, to determine the relative (%) occurrence of the different tubule types found in the digestive gland (holding, absorpting, disintegrating, reconstituting and necrotic tubules). 2. Exposure to the 3 types of WAF leads to lowered MET and MET/MDR values, and concomitant higher MLR/MET values. In the case of lubricant- and Ural-WAF exposures, MDR decreases significantly at increasing exposure-doses. 3. Exposure to oil WAFs leads to decreased percentages of holding and absorpting tubules. This decrease is concomitant with an increase of disintegrating tubules in lubricant- and Ural-WAF exposures, and with an increase of regenerating tubules in the Maya-WAF exposure. 4. The statistical significance of the differences found between control and exposed mussels is better at Day 21 than at any other further sampling day (35, 77 and 91). 5. Multiple correlation coefficients between all the percentages of tubule types and each planimetric parameter present an excellent statistical significance.


The involvement of collagenase in the necrosis induced by the bites of some spiders.

1. The midgut extracts of 13 Australian spider species produced cellular disruption in mouse skin in tissue culture conditions. 2. Microbial collagenase and the venoms of some of these species had similar effects. 3. Five venoms also caused severe dermonecrosis in living mice. 4. Pre-mixing the venoms with L-cysteine caused complete in vivo and partial in vitro inhibition of their effects. 5. It was concluded that collagenase is a major factor in the aetiology of necrotic arachnidism.


Mouse and rat strain variations in sensitivity to N-nitroso-diethylamine, hereditary transmission of the trait and the effect of 3-tert-butyl-4-hydroxyanisole on sensitivity.

1. Strain variations among male mice were studied in terms of the number of days of survival with chronic administration of N-nitroso-diethylamine (NDEA). Four inbred strains, two F1 progenies and one F2 progeny were tested. 2. BALB/c mice survived for the longest period, whereas C3H mice survived for the shortest time. Results of examinations of BALB/c-C3H-F1, -F2 and C57BL-CBA-F1 mice revealed that the hereditary trait could be adequately explained by postulating two loci of genes or gene clusters that regulate the sensitivity to NDEA. 3. Simultaneous chronic administration of 3-tert-butyl-4-hydroxyanisole (BHA) could prolong the survival period. 4. Preliminary histopathological examinations of the liver tissues revealed that the lesion at the time of death of the mice varied considerably depending on the strain and the length of survival. Evidence for hereditary transmission of the characteristics of histopathological changes, including development of liver hemangiosarcoma, is presented. 5. The strain variations among male and female rats were also studied in terms of the number of days of survival with chronic administration of NDEA. Five strains and one F1 progeny were tested. 6. From these and previous observations, the possible biochemical factors determining sensitivity to NDEA were discussed.


Cyclophosphamide and its metabolite acrolein. Some studies on their porphyrinogenic action in 17 day old chick embryo.

1. Some studies of cyclophosphamide (CP) and its metabolite acrolein in chick embryo liver were carried out in order to investigate the mechanism of the porphyrinogenic action of CP. 2. In vitro and in vivo studies revealed that CP induced but did not activate delta-aminolaevulinic acid (ALA) synthase. 3. Pretreatments with phenobarbital (PB) or SKF-525A did not modify ALA-synthase induction produced by CP. 4. Acrolein administration neither induced ALA-synthase activity nor increased cytochrome P-450 content or led to hepatic porphyrin accumulation. 5. Time course induction of cytochrome P-450 content after administration of CP or PB was similar. 6. The results obtained would indicate that CP is a strong inducer of both ALA-synthase activity and microsomal cytochrome P-450 content in liver of 17 day-old chick embryos and that its porphyrinogenic activity is not mediated by its metabolite acrolein.


The effects of acetylcholine, dopamine and noradrenaline on the visceral ganglion of Helix pomatia. I. Ongoing compound field potentials of low frequencies.

1. The question was addressed what effects do some neurotransmitters exert upon the ongoing compound activity of an organized invertebrate ganglion, such as the visceral ganglion of the gastropod mollusc, Helix, recorded with extracellular semimicroelectrodes in the neuropile, avoiding dominating single units. 2. Methods of analysis based on systems theory are used: the electrical activity of the snail brain is considered to be a summation of different measurable variables or subsystems (in this case, "frequency components"). Frequency components were analyzed by the Fourier transform and, for quantification of each component, root-mean-square (RMS)-voltage of digitally filtered signals. 3. Spontaneous discharge of the untreated ganglion shows RMS-voltage of less than 10 microV in the 1-50 Hz range, most frequently 3-7 microV. Energy extended far above 50 Hz, with less decline than in vertebrates, but was quantified only up to 80 Hz due to the wider fluctuations of RMS-voltage in the range greater than 50 Hz. 4. ACh (10(-5)-10(-3) M) induces strong, dose-related increase of activity with spontaneous frequency responses in a broad band between 2 and 20 Hz which accompany fluctuations of minor peakings frequently centering at 2-4, 4-8, 9-15 and 20 Hz. These frequency centers still remain but as an impression since a statistical study on the reality and position of the peaks has not yet been done. ACh-induced excitation seems to be rather transient: all frequency components except the 4-8 Hz component decrease within 4-6 min. 5. DA (10(-4)-10(-3) M) induces a dose-related increase in the low frequency range (1-15 Hz) which reaches a maximum within 4 min and remains relatively unchanged for as long as 10 min. 6. NA (10(-5)-10(-2) M) mainly depresses the ganglionic activity, though moderately, reducing all components (-25%). The action of NA on the Helix ganglion seems to incline more towards inhibition of firing than towards excitation, especially depressing the 15-48 Hz activity, whereas that on the mammalian brain is known to be enhancement of alpha (8-13 Hz)- and beta (15-30 Hz)-activities. 7. The activities observed in the present investigation at several lower frequency power peaks are also commonly found in the electrograms of vertebrates and the EEG of higher animals. The manner, in which ACh and DA influence these peaks in Helix pomatia, are very similar to what has been reported on the EEG of mammals.(ABSTRACT TRUNCATED AT 400 WORDS)


The effects of acetylcholine, dopamine and noradrenaline on the visceral ganglion of Helix pomatia. II. Stimulus evoked field potentials.

1. Neurophysiological effects of acetylcholine (ACh), dopamine (DA) and noradrenaline (NA) on stimulus evoked field potentials in the Helix pomatia visceral ganglion were investigated. Electrical stimuli were applied to nerve trunks. 2. For frequency analysis of evoked potentials (EPs) the Fourier transform was performed. For quantification of pre- as well as poststimulus activities, root-mean-square (RMS)-voltages of digitally filtered signals were used. 3. ACh at 10(-4)-10(-3) M selectively enhances the 4-15 Hz as well as the 30-70 Hz components and DA at 10(-5)-10(-3) M increases the 30-70 Hz activity. NA at 10(-5)-10(-4) M augments the 15-30 Hz response and only at 10(-5) M increases the 30-48 Hz response. 4. All transmitters at 10(-2) M show a strong tendency to suppression. 5. Prestimulus activities of ACh and DA are damped by 40-50% compared to the ongoing induced activities in the non-stimulus study (cf. Part I). These damped prestimulus activities may be considered as a late part of the prolonged evoked activity. 6. The 4-15, 15-30 and 30-70 Hz responses selectively increased by the transmitters may be compared to theta-alpha combined rhythm, beta-rhythm and 40 Hz rhythm in the mammalian EEG.


Dopamine inhibits transmission between the interneuron initiating pacemaker activity in a bursting neuron and the bursting neuron on the snail Helix pomatia.

1. The effect of external application of dopamine on connection between a peptidergic interneuron initiating bursting pacemaker activity in non-active burster RPa1 and the bursting neuron itself of the snail Helix pomatia has been investigated under clamp conditions. 2. External application of dopamine in a dose-dependent manner (Kd 15 microM) produces a reversible inhibition of slow inward current (SIC) in the RPa1 neuron evoked by stimulation of peptidergic interneuron. At the same time, an increase of duration and amplitude of interneuronal action potential was observed. 3. Stimulation of the anal nerve evokes in RPa1 neuron postsynaptic current consisting of at least three components: two fast ones with reversal potentials -50 and -70 mV and a long-lasting one with no reversal potential between -50 and -95 mV. 4. It is concluded that inhibition of SIC by dopamine is due to the processes occurring in the postsynaptic RPa1 neuron. 5. Based on the hypothesis that bursting activity of RPa1 neuron results from the activation of presynaptic unidentified peptidergic interneuron(s) with persistent activity and on data presented it is suggested that inhibition of bursting activity evoked by application of dopamine or anal nerve stimulation is a consequence of decreased efficiency of synaptic transmission between the interneuron initiating bursting activity and the burster.


In vivo neurotoxin administration alters immune responses in chickens (Gallus domesticus).

1. 6-Hydroxydopamine (6-OHDA) administered in ovo enhanced in primary immune response to sheep red blood cells (SRBC) in chicks 2. Splenic norepinephrine levels increased during the peak anti-SRBC response. 3. Cell-mediated immunity as measured by delayed-type hypersensitivity to phytohemagglutinin-P (PHA-P) was not affected by treatment with 6-OHDA.


Contractile responses to electrical field stimulation and ATP in guinea-pig urinary bladder.

1. The responses of guinea-pig urinary bladder smooth muscle to intramuscular nerve stimulation were investigated by isometric tension recording. 2. The nerve mediated contractions, evoked by field stimulation with a stimulus of 0.5 msec duration, a frequency of 15 Hz and a voltage of 50 V, were inhibited by atropine (10(-6) M) and alpha, beta-MeATP (10(-4) M) to about 80% and 20%, respectively, of control. 3. Simultaneous application of atropine (10(-6) M) and alpha, beta-MeATP (10(-4) M) abolished the contraction evoked by selective nerve stimulation. 4. Terodiline (10(-4) M), one of Ca2+ antagonists, inhibited the contraction evoked by both nerve selective stimulation and exogenously applied ATP. 5. Based on these results we suggested that in guinea-pig urinary bladder, ATP was a predominant excitatory transmitter and the nerve-mediated contraction and exogenous ATP were mainly dependent upon Ca2+ influx.


Effect of Galleria mellonella larvae preparation and honeybee products on cell cultures.

1. The action of the extract from larvae of the great wax moth Galleria mellonella L. used in Russian folk medicine was studied. 2. Two active components influencing the growth and morphological differentiation of cells in vitro were found. 3. The presence of these components in the extract was conditioned by consumption of honeybee products by Galleria mellonella larvae. 4. Cytotoxicity of honeybee products was studied.


Inhibition of the transepithelial potential difference and short circuit current in the isolated frog skin by alloxan.

1. Electrical parameters: short circuit current (SCC), transepithelial potential difference (PD) and electrical resistance (R) were measured in isolated frog skin (Rana pipiens) in the presence and in the absence of alloxan. 2. Alloxan decreased SCC and PD in a concentration-dependent pattern, while R remained unchanged. 3. The effect on SCC and PD was observed after 25 min of exposure to the drug. Maximal average effect was 20% in SCC and 17.5% in PD. 4. These results suggest that alloxan decreased epithelial sodium transport, through interference with the activity of the Na(+)-K(+)-ATPase.


Histamine activates whole cell K+ currents in swine carotid arterial smooth muscle cell.

1. The effects of histamine on whole cell K+ currents were evaluated in cultured smooth muscle cells that were derived from explants of swine carotid artery. These cells exhibited cytoplasmic protein and agonist-induced calcium responses typical of noncultured smooth muscle cells. 2. Upon obtaining whole cell configuration, depolarization from a holding potential of -80 mV elicited a slowly inactivating voltage-dependent outward current that was sensitive to tetraethylammonium and 4-aminopyridine. 3. These K+ currents were at least partially Ca2+ dependent since they were inhibited by extracellular bath application of verapamil or by substitution of external Ca2+ with Co2+. 4. The addition of extracellular histamine increased the outward current both in the presence and absence of extracellular Ca2+ and this response was blocked by (+/-)chlorpheniramine. 5. We conclude that swine carotid arterial smooth muscle cells exhibit voltage and calcium dependent potassium current and that these currents are modified by histamine stimulation.


Lead retards development of Drosophila melanogaster.

1. Lead (Pb) is a ubiquitous environmental toxicant which has been reported to have growth-retarding effects. That premise was examined in the current study of the effects of developmental exposure to Pb on the maturation of the fruitfly Drosophila melanogaster. 2. Flies were raised from egg to adulthood in media to which 0, 100, 250, or 500 ppm Pb acetate added. 3. There was a dose-dependent delay of maturation but no apparent effect on survival to adulthood. There were no significant differences in this effect between male and female flies. 4. Weights of fly offspring and their fecundity were not related to increasing exposure levels, suggesting that the delays were not due to gross nutritional deficits. 5. Analyses of Pb content indicated exposure-dependent body burdens of Pb in flies. Timed analyses of the Pb content of media itself indicated a heterogeneous distribution of Pb in the media, suggesting some precipitation of Pb at the highest exposure, occurring primarily during the first hour. 6. The mechanistic bases of the Pb-induced retardation of D. melanogaster development remain unknown, but it is concluded that because of the extensive body of knowledge on D. melanogaster genetics, molecular biology, and developmental biology, this procedure could serve as a model system for further study of the developmental consequences of exposure to Pb or other toxicants. 7. Environmental Pb exposure resulting in retarded development could have deleterious repercussions for insect populations exposed chronically to high levels of Pb.


Bioactive substances in nerve-cord extracts from an echiuroid, Urechis unicinctus--II. Peptidic substances.

1. Two peptides that showed an inhibitory effect on twitch contraction of the inner circular body-wall muscle of the echiuroid Urechis unicinctus were purified from extracts of the ventral nerve cords of the worms. 2. On the basis of the results of amino acid analysis, amino acid sequence analysis and FAB-MS measurement, the primary structures of the peptides were proposed to be H-Phe-Arg-Val-Phe-OH (FRVF) and H-Phe-Arg-Phe-OH (FRF). 3. FRVF and FRF were synthesized, and their behavior on HPLC and biological activities on the body-wall muscle were confirmed to be identical to those of the native peptides. 4. FRVF was also found to have a potentiating effect on spontaneous contractions of the intestine of the African clawed toad Xenopus laevis.


The comparison of the effects of heavy metal ions on the antioxidant enzyme activities in human and fish Dicentrarchus labrax erythrocytes.

1. The effect of the increasing concentrations of CuSO4 and HgCl2 (0.01-0.3 mmol/l) on erythrocyte haemolysis and the activities of peroxide metabolism enzymes: superoxide dismutase, catalase, peroxidase and glutathione peroxidase was investigated in human erythrocytes and the nucleated red blood cells of marine fish (Dicentrarchus labrax). 2. The results show that both heavy metal ions had only little effect on haemolysis and antioxidant enzyme activities in human erythrocytes; in contrast the effect of heavy metals on fish erythrocytes was statistically significant when compared to control values. 3. Copper was found to have more pronounced effect than mercury on the erythrocytes of Dicentrarchus labrax; otherwise there were no significant differences between the toxic effects of both ions on human erythrocytes. 4. We suggest that the mechanism of copper-induced haemolysis may be different from that of mercuric ion in the erythrocytes of Dicentrarchus labrax.


Effect of local hyperthermia and chemotherapy on MC29 virus-induced liver tumor transplanted into subcutis of chickens.

1. A transplantable liver tumor induced by MC29 virus, whose specific utilization of glutamine in growth of the tumor had been well-demonstrated, was implanted in the subcutis of day-old chickens, and after 7-8 days, when the tumor had reached a soybean size, local hyperthermia with microwave and/or chemotherapy with 6-diazo-5-oxo-1-norleucine (DON), an inhibitor of glutamine metabolism, were given once a day for 7 days successively. 2. At the 7th day of experiment, the tumor growth ratio, expressed by mean +/- standard deviation was, in the tumor control group, 14.3 +/- 1.81; in DON group, 6.93 +/- 0.95; in local hyperthermia group, 3.24 +/- 2.25, and in DON+hyperthermia group, 0.61 +/- 0.57. 3. The body weight-gain ratio in the local hyperthermia group was evidently higher than that in the tumor control group. DON suppressed body weight gain most remarkably. 4. In the present transplantable tumor system, microwave local hyperthermia was more effective in tumor suppression and less hazardous to the host than our former whole-body hyperthermia.


Separation of cytochrome P-450 containing vesicles from the midgut microsomal fraction of Manduca sexta.

1. Microsomes prepared from midgut tissue of Manduca sexta by differential centrifugation are a heterogeneous population of vesicles. 2. Upon centrifugation of microsomes in a sucrose gradient of 30%, 15% and 10%, specific activity of cytochrome P-450 catalyzed O-demethylation of p-nitroanisole (a marker enzyme for endoplasmic reticulum) was increased 2.5 to 3.5-fold in the 15% fraction. 3. Specific activities of alkaline phosphatase and leucine aminopeptidase (marker enzymes for brush border membranes) were increased 3.1 to 5.7-fold in the pellet. 4. Differential centrifugation is a useful step in the purification of cytochrome P-450 enzymes.


Amine oxidase enzymes of sheep blood vessels and blood plasma: a comparison of their properties.

1. A comparison between the biochemical properties of semicarbazide-sensitive amine oxidase (SSAO) activities has been made in sheep blood plasma and arterial wall. 2. The metabolism of benzylamine (BZ) by blood plasma was resolved into high affinity (Km 2.76 +/- 0.24 microM) and low affinity (Km 743 +/- 49 microM) activities. Spermidine metabolism was by a single component (Km 174 +/- 22 microM) and this amine reduced the metabolism of high concentrations of BZ. 3. A single component metabolised BZ in arterial homogenates (Km 11.3 +/- 1.3 microM) and which metabolised spermidine at a very slow rate. 4. Dopamine was deaminated by plasma SSAO and competed with both low and high concentrations of BZ. Dopamine also interfered with arterial metabolism of BZ. 5. These results suggest that there are two SSAO enzymes in sheep blood plasma, with one having similar properties to SSAO in the arterial wall.


The FMRFamide-related decapeptide of Mytilus contains a D-amino acid residue.

1. An FMRFamide-related decapeptide isolated from the anterior byssus retractor muscle (ABRM) of the bivalve mollusc, Mytilus edulis, was shown to have D-Leu as the second amino acid residue. 2. The excitatory effects of the peptide (Mytilus-FFRFamide) on the ABRM were not changed appreciably by substituting an L-Leu residue for the D-Leu residue.


Haematological effects of vanadium on living organisms.

Although vanadium has been of great interest for many researchers over a number of years, its biochemical and physiological role is not yet fully clear. There are many papers describing the haematological consequences of its excess in living organisms and most of their data are quoted in this mini-review. The authors of these papers used various laboratory animals, different vanadium compounds, frequently different routes of administration and duration of intoxication. Hence a checklist and comparison of the results are rather difficult. Vanadium reduces the deformability of erythrocytes, and such cells are rather frequently retained in the reticuloendothelial system of the spleen and eliminated faster from the blood stream (Kogawa et al., 1976). Vanadium produces peroxidative changes in the erythrocyte membrane, this leading to haemolysis. Therefore, the depressed erythrocyte count in animals intoxicated with vanadium may be the consequence of both the haemolytic action of vanadium and the shortened time of survival of erythrocytes. Changes of the haem precursor level in blood serum and urine observed in humans exposed occupationally to vanadium suggest an influence of this element on haem synthesis. This problem requires, however, further studies and observations. Changes occurring under the influence of vanadium on the leukocyte system of animals suggest the influence of this element on the resistance of the organism, but the mechanism of the action of vanadium still requires elucidation.(ABSTRACT TRUNCATED AT 250 WORDS)


Adrenergic mechanisms associated with the movement of platelets in iridophores from the freshwater goby, Odontobutis obscura.

1. Cultured iridophores from the freshwater goby, Odontobutis obscura, were used to investigate adrenergic mechanisms of movement of platelets in the iridophores. 2. Norepinephrine, which was assumed to be the transmitter of the iridophore nerves, induced dispersion of platelets within the cells. 3. The effect of norepinephrine was inhibited by an alpha-adrenergic antagonist, yohimbine, but not by a beta-adrenergic antagonist, propranolol. 4. Isoproterenol, a beta-adrenergic agonist, failed to bring about aggregation of platelets. 5. Forskolin, an activator of adenylate cyclase, was effective in inducing aggregation of platelets. 6. 8-Br-cAMP caused the aggregation of platelets and inhibited the norepinephrine-induced dispersion of platelets. 7. It appears that the adrenoceptors of the iridophores of this species are solely of the alpha type; they mediate the dispersion of platelets; and an increase in intracellular levels of cAMP induces the aggregation of platelets.


Isolation, primary structure and synthesis of neomyosuppressin, a myoinhibiting neuropeptide from the grey fleshfly, Neobellieria bullata.

1. An amidated decapeptide, showing strong inhibitory activity of spontaneous visceral muscle movement was isolated, from head extracts of 42 thousand fleshflies, Neobellieria bullata (Diptera, Sarcophagidae). 2. Amino acid sequencing and verification by peptide synthesis revealed the following primary structure: Thr-Asp-Val-Asp-His-Val-Phe-Leu-Arg-PheNH2. 3. The novel peptide was termed neomyosuppressin or Neb-MS. 4. During the process of consecutive high performance liquid chromatography (HPLC) purifications the biological activity of the samples was monitored using heterologous bioassay system. 5. The threshold level of synthetic Neb-MS was found to be 8.6 +/- 0.5 x 10(-11) M on the Leucophaea hindgut and 3.4 +/- 0.5 x 10(-10) M on the Locusta oviduct bioassay, respectively.


Changes in biochemical composition of gills, hepatopancreas and muscle of the red crayfish Procambarus clarkii (Girard) after sublethal exposure to mercury.

1. The changes in the biochemical composition of gills, hepatopancreas and muscle after exposure to 0.25 mg Hg/l were studied in Procambarus clarkii. 2. Sublethal exposure to mercury in P. clarkii resulted in significant decreases in protein concentration and caloric concentration in gills over the 96 hr period. Glycogen/lipid and glycogen/protein ratios increased after 48 and 96 hr of mercury exposure. 3. Lipid and caloric concentration in the hepatopancreas were significantly lower in 96 hr mercury exposed group. 4. Glycogen concentration in muscle was depleted as consequence of 96 hr mercury exposure.


Initial characterization of the organophosphate acid anhydrase activity of the chicken, Gallus domesticus.

1. Supernatant solutions from kidney and liver homogenates of the chicken, Gallus domesticus, were found to hydrolyze the organophosphate (OP) compound diisopropylfluorophosphate (DFP). The activity on DFP as substrate was heat-inactivated and was characterized for temperature and pH optima, enzyme kinetics, and requirements for manganous ion. 2. Gel column chromatography indicated that the DFPase in both tissues is in the range of 82,100 to 93,300 D. This activity is strongly inhibited by N,N'-diisopropylphosphorodia-midofluoridate (mipafox). 3. The chicken has organophosphate acid (OPA) anhydrase activity comparable to other eucaryotic sources in its ability to hydrolyze DFP. Although birds may not have paraoxonase activity comparable to mammalian species, they do not differ significantly in the ability to hydrolyze DFP and probably related compounds.


Inhibition of dipteran larval neuromuscular synaptic transmission by analogues of philanthotoxin-4.3.3: a structure-activity study.

1. The blocking action of delta-philanthotoxin (PhTX-4.3.3), a wasp toxin composed of a polyamine chain, an aromatic amino acid (tyrosyl) nucleus and an aliphatic (butanoic) acid was studied, together with 33 structural analogues, at the glutamatergic neuromuscular synapse in larvae of the house fly, Musca domestica. 2. Shortening of the polyamine or the aliphatic acid chain, or removing the aromatic nucleus caused a decrease in potency. 3. An increase in potency was obtained by elongating the molecule in any of several directions: elongation of the polyamine chain with amine containing groups, by iodination of the aromatic moiety and by aliphatic or aromatic elongation of the butanoic acid.


Some pharmacological properties of the oviduct muscularis of the stable fly Stomoxys calcitrans.

1. Spontaneous and rhythmic contractions were measured in 80% of the preparations of the stable fly oviduct which were separated from the central nervous system and other tissues. Measurements of the changes in the amplitude and frequency of contractions and changes in the baseline tonus were taken separately, even though they often occurred together during chemical treatments. 2. L-glutamate, at a concentration of 10(-4) to 10(-3) M, caused an increase in the frequency of contractions and in muscle tonus. Contraction frequency showed the highest percentage of change while amplitude and tonus had lower values. 3. The excitation threshold for proctolin on the oviduct was between 10(-13) and 10(-10) M. The initial responses to the peptide showed increases in the amplitude, frequency, and tonus of muscle contractions. However, tonic changes dominated the response profile when the concentration reached 10(-10) or 10(-9) M. In 12 experiments in which the oviduct was exposed to 10(-7) M leucomyosuppressin, no change in the amplitude, frequency, or tonus of muscle contractions was detected. 4. Octopamine caused an inhibition of spontaneous muscle contractions of the oviduct. Spontaneous contractions showed a graded drop in activity over the concentration range tested (10(-8)-10(-6) M). Also 57% of the preparations tested showed a substantial drop in baseline tonus after the addition of octopamine. 5. Stable fly oviducts exposed to extracts from the male reproductive tract showed a noticeable change in the amplitude frequency and tonus of muscle contraction at levels of 2 to 3 equivalents.


Effect of acrylamide monomer on hepatic CYP1A1 monooxygenase induction in rainbow trout.

1. Rainbow trout hepatic microsomes were pre-incubated in vitro with 1 pM, 1 nM, and 1 microM acrylamide for 20 or 30 min and ethoxyresorufin-O-deethylase (EROD) was measured. In vitro pre-incubation did not produce a significant decrease in EROD catalytic activity. 2. The effects of 50 ppm acrylamide monomer on hepatic CYP1A1 mRNA and CYP1A1 isozyme steady state levels in rainbow trout were determined after 6, 10, and 14 days of exposure. 3. Acrylamide monomer produced a 35% increase, 51% decrease and 140% increase in CYP1A1 mRNA at 6, 10 and 14 days, respectively, while at the same time CYP1A1 isozyme levels were decreased 12%, 67%, and 62% and EROD activity was decreased 33%, 90%, and 86%, respectively. 4. This indicates that acrylamide treatment may result in either a change in the translation of CYP1A1 mRNA or an isozyme selective inactivation of CYP1A1 resulting in loss of CYP1A1 apoprotein. 5. The effect of acrylamide treatment on hepatic CYP1A1 induction was determined using 10 or 14 day treatment with 50 ppm acrylamide monomer in a flowthrough exposure and induction with beta-naphthoflavone (beta-NF; 100 mg/kg i.p.) for 1 or 4 days. 6. Acrylamide and 1-day beta-NF had no effect on CYP1A1 mRNA levels when compared to 1-day beta-NF treatment alone, but acrylamide and 4-day beta-NF resulted in a 74% decrease in CYP1A1 mRNA compared to 4-day beta-NF treatment alone.(ABSTRACT TRUNCATED AT 250 WORDS)


Oxygen consumption and ammonia-N excretion of Penaeus chinensis juveniles exposed to ambient ammonia at different salinity levels.

1. Oxygen consumption rate (O2 mg/g/hr) and ammonia-N (un-ionized plus ionized ammonia as nitrogen) excretion rate (microgram/g/hr) of Penaeus chinensis juveniles (0.54 +/- 0.11 g) decreased with increase of salinity among 10, 20 and 30 ppt. 2. Oxygen consumption rate increased with increase of ambient ammonia-N at three salinity levels. 3. Ammonia-N excretion rate was highest (204 micrograms/g/hr) in P. chinensis exposed to 0.985 mg/l ammonia-N at 10 ppt, and lowest (-416 micrograms/g/hr) in that exposed to 9.897 mg/l ammonia-N at 30 ppt. 4. In the prawns exposed to 9.921 mg/l ammonia-N at 20 ppt, and to 5.087 and 9.897 mg/l ammonia-N at 30 ppt, ammonia-N excretion was heavily depressed and ammonia-N influx occurred (ammonia-N uptake was much higher than ammonia-N excretion).


A comparative study on the effects of diethyl ether and thiopentone sodium on plasma catecholamine levels in the rat.

1. The primary aim of this investigation was to ascertain the effects of thiopentone sodium and diethyl ether as anaesthetics on plasma catecholamine levels of Sprague-Dawley and Long-Evans rats. A secondary aim was to investigate the effect of norepinephrine on susceptibility to electrically induced arrhythmia in the Langendorff perfused isolated rat heart. 2. Thiopentone sodium anaesthesia resulted in significantly lower plasma norepinephrine and epinephrine levels when compared with diethyl ether anaesthesia. 3. Long-Evans rats exhibited significantly higher epinephrine levels regarding both anaesthetics in comparison with their Sprague-Dawley counterparts. 4. If plasma catecholamines are an indicator of the level of induced stress then diethyl ether anaesthesia resulted in more stress both of a psychological and physiological origin when compared with thiopentone sodium anaesthesia. 5. Long-Evans rats seem to be more prone to the physiologically-induced component of stress than Sprague-Dawley rats. 6. In isolated hearts, norepinephrine increased susceptibility (P less than 0.002) to electrically induced arrhythmia. This occurred at a concentration corresponding to measured upper levels of total catecholamines in diethyl ether anaesthetised animals.


The actions of phorbol esters upon isolated calcium currents of Helix aspersa neurones.

1. Voltage-clamp recordings have been made from identified neurones in the suboesophageal ganglia of Helix aspersa. 2. Calcium currents were isolated pharmacologically and studied under two-electrode voltage-clamp. 3. PdBu but not PMA caused a transient enhancement followed by an irreversible inhibition of the calcium current. 4. It is concluded that activation of protein kinase C within these neurones may show selectivity with respect to the exogenous activator used.


Cardiac activity of calcitonin gene-related peptide in amphibian species (Rana tigrina, Triturus sp).

1. Calcitonin gene-related peptide (CGRP) produced dose-related positive chronotropic and inotropic responses in isolated atria of a newt, Triturus sp. and a frog, Rana tigrina. It was apparent that CGRP was stimulatory on the heart of the amphibian species. 2. In the frog, these dose-related responses were attenuated in the presence of a CGRP antagonist (hCGRP 3 x 10(-8) M). This was indicative of the presence of cardiac CGRP receptors. 3. In the presence of beta-adrenergic (propranolol 10(-6) M) and calcium channel (verapamil 10(-8) M, but not nifedipine 10(-8) M) blocker, the basal AR in the frog were decreased. AR induced by CGRP were increased however. 4. The maximal AR and AT responses of the isolated atria to CGRP were not affected by the presence of propranolol. 5. In the presence of calcium channel blockers (verapamil and nifedipine), the maximal AT response was attenuated. These changes suggest the cardiac effects of CGRP are related to calcium mobilization.


Resumption of testicular activity in Gobius paganellus after administration of ethane 1,2-dimethane sulfonate (EDS).

1. The effect of a single injection of ethane-1,2-dimethane sulfonate (EDS) was studied in the teleost fish, Gobius paganellus in two different periods of the year. 2. During June EDS did not induce any change, while during December the drug was highly effective in promoting testicular activity. 3. Nucleus/cytoplasm ratio of interstitial cells strongly decreased concomitantly with the detection of high testicular androgen levels. 4. The germinal compartment was well developed showing the appearance of all spermatogenic stages and the cavity of lobular compartments filled of spermatozoa. 5. Our data are the first evidence of a stimulatory activity of EDS on testes of a vertebrate species.


Effects of copper, iron and zinc on oedema formation induced by phospholipase A2.

1. Intradermal or subplantar injection of soluble snake venom phospholipase A2 (PLA2) evoked a brisk inflammatory response, with cutaneous vascular permeability increase and paw oedema. 2. These inflammatory processes are mainly the result of arachidonic acid cascade activation and mast cell degranulation. 3. Copper, iron and zinc have an inhibitory effect on vascular permeability increase and paw oedema induced by PLA2. 4. Copper and iron could have not only a direct effect on PLA2 but on enzymes of arachidonic acid cascade. 5. However, zinc have a moderate antiinflammatory activity. This effect could be the result to inhibit PLA2 induced mast cell degranulation.


Gender influences the in vitro renin release responses to dopamine and adrenergic receptor blockade in the summer-active ground squirrel Spermophilus lateralis.

1. 10(-4) M Dopamine stimulated and 10(-7) M inhibited in vitro resting renin release (RR) only in the female animals. 2. Alpha- or beta-adrenergic blockade prevented the stimulation but not the inhibition of RR by dopamine in the female animals. 3. Membrane receptor mechanisms mediating these changes do not involve alterations in tissue cyclic AMP content. 4. The sensitivity of the juxtaglomerular cells to dopamine challenge is gender dependent during summer activity.


VX enhances neuronal excitability and alters membrane properties of Rana catesbeiana sympathetic ganglion neurons.

1. The effects of VX (10 microM) were examined on sympathetic ganglion neurons from the bullfrog using intracellular recording techniques. 2. VX significantly increased the amplitude of the residual EPSP from 4.8 +/- 0.86 mV (n = 4) to 13.7 +/- 1.23 mV (n = 4). 3. VX significantly decreased the membrane potential 5.2 +/- 0.75 mV (n = 6). The input resistance and the duration of the spike afterhyperpolarization (AHP) were also reduced 69.8% and 69.6% of control, respectively. 4. VX increased neuronal excitability greater than 200% (n = 5) of control. 5. The VX-induced neuronal excitability may result from a reduction in the duration of the AHP and contribute to the CNS toxicity.


Molecular genetic analysis in autosomal dominant keratoconus.

Members in three generations of a family whose propositus had keratoconus were examined by biomicroscopy, with a corneoscope and a computer-assisted videophoto-keratoscope. Keratoconus was detected in eight of 15 family members with vertical transmission consistent with autosomal dominant inheritance. Affected individuals displayed variable topographic features. Abortive "nipple-type" cones were identified in some individuals in successive generations using the computer-assisted videophotokeratoscope and more advanced nipple-type cones detected on biomicroscopy of other family members. We selected a COL6A1 cDNA (the gene encoding the alpha 1 chain of type VI collagen) as a "candidate gene" to determine cosegregation with the disease locus. Linkage analysis excluded a gene locus for keratoconus on the most telomeric region of chromosome 21 in this family.


Immunopathology and altered immunity in posterior uveitis in man: a review.

Posterior uveitis is thought to be a T-cell mediated disease since active foci of inflammation, identified in eyes enucleated for the complications of intraocular inflammation, are found to be predominantly composed of CD4+ T-cells. Few B-cells and little immunoglobulin are found suggesting that antibody and immune complex deposition do not play a major role in perpetuating the inflammatory process. As ocular biopsy is not a feasible method for monitoring disease activity and response to treatment, parameters of T-cell activation and retinal damage have been studied in the peripheral blood. These have included antibody and T-cell sensitisation to retinal S-antigen, serum soluble IL-2 receptors and IL-2 receptors on activated T-cells. none of these parameters, however, have been found to be useful in the monitoring of ocular disease activity alone or in the prediction of disease relapse.


Circulating ICAM-1 levels in serum of uveitis patients.

Intercellular Adhesion Molecule-1 (ICAM-1) is a cytokine-inducible adhesion molecule expressed on cells of multiple lineages at sites of inflammation. Recently a truncated form of ICAM-1 has been discovered to be circulating in serum. This study reports on circulating serum (cICAM-1) levels in 132 uveitis patients (HLA-B 27 pos. acute anterior uveitis (AAU); HLA-B27 neg. anterior uveitis (AU); intermediate uveitis (IU); heterochromic cyclitis Fuchs (HCF); sarcoidosis; Toxoplasmosis). Measurement of circulating ICAM-1 serum levels was performed using a monoclonal antibody based ELISA, with healthy blood donors serving as the control group. Applying multiple variance analysis and the Student Newmann-Keuls test we found a statistically significant elevation of serum cICAM-1 level in the HLA-B 27 neg. AU group (n:31), in the IU group (n:25) and in patients with sarcoidosis (n:18). Serum levels of HLA-B27 pos. AAU patients, patients with HCF and patients suffering from ocular toxoplasmosis did not differ significantly from levels of the control group.


Distribution of IL-2R and CD45Ro expression on CD4+ and CD8+ T-lymphocytes in the peripheral blood of patients with posterior uveitis.

Different lines of evidence support a major role for activated T-lymphocytes in the pathogenesis of posterior uveitis. The initial site of activation of these autoreactive T-cells, either locally in the eye or in the peripheral immune compartment, is still unknown. This study was undertaken to investigate whether with currently available techniques, it is possible to detect alterations in the levels and subsets of activated T-cells in the peripheral blood of patients with posterior uveitis. For this reason, 3-colour immunofluorescent staining was performed to assess the distribution of IL-2 receptors (IL-2R) and the CD45RO-antigen on CD4+ and CD8+ subsets of peripheral blood lymphocytes (PBLs) from patients with posterior uveitis (n = 29). Only the subgroup of patients with posterior uveitis as part of a systemic immune-mediated disease (sarcoidosis, Behçet's disease) (n = 9) showed a significant increase in IL-2R expression on peripheral blood lymphocytes (p less than 0.005) when compared to normals (n = 12). This increased expression was reflected much more significantly in the CD4+ (p less than 0.0005) rather than in the CD8+ subset (p less than 0.05) of lymphocytes. In contrast, no significant increase in CD45RO expression on either subset of T lymphocytes was found in any subgroup of posterior uveitis in comparison with normals.


Experimental autoimmune uveoretinitis: a model system for immunointervention: a review.

Experimental autoimmune uveoretinitis (EAU) is a useful model of human posterior uveitis and as such, permits the analysis of strategies for immuno-intervention. Modulation of the autoimmune response may be attempted at the stages of induction of EAU, during homing of autoreactive lymphocytes to the target organ, the retina, or during the effector stage of the disease. This paper presents a brief overview of current immuno-therapeutic modalities and assesses the usefulness for extrapolation to human disease.


Differential regulation of dopaminergic neurons in the mammalian brain: a brief review.

The mammalian brain contains anatomically distinct dopaminergic neuronal systems that subserve a variety of functions which include maintaining postural reflexes, modulating basic psychic processes, and controlling the secretion of hormones from the pituitary. In turn, these various dopaminergic neuronal systems are regulated by different mechanisms that are appropriate for the functions that they control. This is illustrated by comparing the responses to endocrinological and pharmacological manipulations of the nigrostriatal dopaminergic neuronal system that is involved with sensorimotor integration and the tuberoinfundibular dopaminergic neuronal system that tonically inhibits the release of prolactin.


Intrahypothalamic neurohormonal interactions in the control of growth hormone secretion.

The secretion of growth hormone (GH, somatotropin) is regulated by two neurohormones: one inhibitory, somatotropin release-inhibiting hormone (SRIH) or somatostatin, and one stimulatory, GH-releasing hormone (GHRH). There are several lines of evidence for reciprocal interactions between SRIH and GHRH neuronal networks. Anatomically, GHRH terminals contact SRIH-containing neurons in the periventricular nucleus and SRIH-containing fibres innervate GHRH-containing neurons in the arcuate nucleus. Physiologically, SRIH and GHRH are secreted into the portal blood in complementary oscillating patterns. Results from immunizations with anti-SRIH antisera suggest that endogenous SRIH blocks GHRH release from the median eminence. Intracerebroventricular injections of SRIH stimulate secretion of GHRH indirectly, probably via autoinhibition of SRIH neurons in the anterior periventricular region. High resolution autoradiography allowed us to visualize high affinity, specific [125]SRIH receptors on 30% of GHRH mRNA-containing cells in the ventrolateral portion of the arcuate nucleus. The functional importance of these receptors was demonstrated by blocking endogenous SRIH action with cysteamine, which resulted in an increase in GHRH mRNA levels and desaturation of SRIH receptors in the ventrolateral part of the arcuate nucleus. Alterations in GH production caused by hypophysectomy or acute and chronic GH hypersecretion have opposite effects on the synthesis of SRIH and GHRH, giving further evidence for reciprocal interactions between these two neurohormonal systems.


Erythromycin stimulates gallbladder emptying and motilin release by atropine-sensitive pathways.

The effect of administering different doses of erythromycin on gallbladder emptying and plasma concentrations of immunoreactive motilin was investigated in healthy volunteers. Erythromycin was infused for 30 min at four different doses: 20, 50, 100, and 1000 mg/hr. Gallbladder volume was determined by ultrasound scanning every 10 min for 60 min. All doses, except 20 mg/hr, provoked a significant reduction in gallbladder volume (P < 0.01). The gallbladder emptying peak occurred after 20 min infusion. It was approximately 40-45% of basal volume and 60-70% of the emptying observed after a standard meal. At 100 mg/hr, erythromycin caused a 2.5-fold increase in plasma motilin concentration, which reached a peak after 30 min infusion. Plasma motilin peaked following maximum gallbladder emptying in all subjects. To evaluate whether cholinergic pathways were implicated in the action of erythromycin, 100 mg/hr erythromycin was infused together with 6 micrograms/kg/hr atropine. Atropine inhibited both gallbladder emptying and motilin release (P < 0.001). Infusion of 1 microgram/kg/hr somatostatin had the same inhibitory effects (P < 0.001). Our results suggest that atropine acts by inhibiting an erythromycin-activated cholinergic neural mechanism. Somatostatin could exert its inhibitory effect by blocking the release of acetylcholine from neural terminations.


Controlled study of oxatomide vs disodium chromoglycate for treating adverse reactions to food.

A controlled, cross-over trial was carried out to compare the efficacy and safety of oxatomide vs disodium chromoglycate (DSCG) for treating food allergy and intolerance. Twenty patients (15F, 5M; mean age 36.6 years), with chronic urticaria (twelve patients) or eczema (eight patients) caused by food allergy (ten cases) or food intolerance (ten cases), were treated with oxatomide (60 mg/day in a single evening administration) and with DSCG (2000 mg/day) for six weeks. The two treatments were separated by a 3-week wash-out period. All the patients completed the trial. During the treatment, both drugs succeeded in controlling the symptoms. With oxatomide, the wheals totally disappeared from 75% of the patients (p = 0.00135), the eczematous lesions disappeared from 64% (p = 0.056), and the itching from 70% (p = 0.00012); the figures for DSCG were 33%, 50% (p = 0.038) and 50%, respectively. Both drugs were well tolerated and there was no need to discontinue the treatment of any of the patients.


Conjugation of dinitrofluorobenzene to plasma proteins in vivo in the rat.

The extent of protein dinitrophenylation was determined in plasma and other tissues of anesthetized rats after administration of the model immunogen [3H]dinitrofluorobenzene (DNFB) (25 mg/kg; 5-25 microCi). DNFB was given by the intravenous, intraportal, intramuscular, or oral route. Irreversible binding was determined radiometrically after exhaustive solvent extraction of plasma or organ proteins. The extent of binding was high in plasma after parenteral administration (approximately 1% dose/ml plasma), but less (approximately 0.1% dose/ml) if DNFB was given orally. Low levels of radioactivity were bound irreversibly in liver (0.01-0.13% dose/g) and kidney (0.03-0.10% dose/g) and only residual amounts in other organs. Western blotting was used to identify target proteins in plasma, liver, and kidney using a specific antidinitrophenyl antiserum. No dinitrophenylation could be detected in liver or kidney samples, but strong recognition of two protein bands was observed in plasma. Bands with the same apparent molecular masses (67 and 44 kDa) were seen when DNFB was incubated with rat plasma in vitro. Preliminary evidence for these proteins being albumin and alpha 1-acid glycoprotein, respectively, is presented. The latter may be important for interindividual variability in immune responsiveness, because it is an acute phase protein whose levels fluctuate widely during disease states.


Induction of CYP1A1 mRNA in rat epidermis and cultured human epidermal keratinocytes by benz(a)anthracene and beta-naphthoflavone.

Cytochrome P-4501A1 (CYP1A1) plays a major role in the bioactivation of procarcinogens in target tissues, including skin. However, the factors controlling CYP1A1 expression in mammalian skin are unknown. Utilizing the reverse transcriptase-linked polymerase chain reaction, we analyzed the effect of treatment with beta-naphthoflavone (beta-NF) and benz(a)anthracene on the expression of CYP1A1 mRNA in rat epidermis and normal human epidermal keratinocytes (NHEK). Inducer treatment of rats and NHEKs resulted in several-fold increases in aryl hydrocarbon hydroxylase activity. Following inducer treatment of adult or neonatal rats, increase in CYP1A1 gene message occurred as compared with that in controls. Higher basal level and inducibility were detected in mature rather than in 4-day-old rats. This induction occurred as early as 4 hr after beta-NF application. Exposure to beta-NF and benz(a)anthracene also resulted in substantial increases in gene message in NHEK. Northern blot analyses complemented the polymerase chain reaction data. These results indicate that CYP1A1 gene expression is increased in mammalian epidermis by inducers of epidermal aryl hydrocarbon hydroxylase activity.


Isolation, purification, and characterization of two new chemical decomposition products of methylazoxyprocarbazine.

We have previously reported that the antineoplastic agent, procarbazine, in aqueous solutions was chemically oxidized to its azoxy metabolites (methylazoxy and benzylazoxy). To determine if there was additional metabolism of the most active metabolite, methylazoxyprocarbazine, it was incubated in the presence and absence of CCRF-CEM human leukemia cells. Incubations were extracted, and potential metabolites were detected by HPLC with UV detection and by combined HPLC and thermospray mass spectrometric analysis. The major metabolite identified by HPLC with UV detection of the extracts was N-isopropyl-p-formylbenzamide; this was identified by comparison of its retention time with that of a synthesized standard. This identification was further corroborated by HPLC/thermospray mass spectrometry (LC/MS). Analysis of the extracts by LC/MS also showed the presence of a closely eluting peak that had a protonated molecular ion at m/z 207. This new metabolite was identified as N-isopropyl-(benzene-1,4-bis-carboxamide) by 1H NMR and gas chromatography/ion trap mass spectrometry. This metabolite is postulated to arise from breakage of the N-N bond in the hydrazine portion of the molecule. Reconstructed ion (m/z 236) current profiles from the analysis of the cell extracts indicated that there was only a trace amount of methylazoxyprocarbazine left after a 72-hr incubation. Interestingly, a peak with the same molecular weight as the parent compound (methylazoxyprocarbazine) was observed in the cellular incubations and also in extracts of control incubations in which methylazoxyprocarbazine was incubated in medium without cells. This unknown was silylated and identified as a hydroxyazo compound by an ion trap mass spectrometer operated under both single and multiple-stage mass analysis. Formation of this decomposition product appears to involve a novel intramolecular rearrangement of methylazoxyprocarbazine in solution. This pathway may be responsible for the formation of the ultimate cytotoxic species by chemical decomposition of procarbazine.


Effects of dose, strain, and dosing vehicle on methacrylonitrile disposition in rats and identification of a novel-exhaled metabolite.

Methacrylonitrile (MAN), an aliphatic nitrile used in the production of plastics and elastomers, is structurally related to the known animal carcinogen, acrylonitrile. Although MAN has potential to cause significant toxicity, minimal information is available on its toxicity or fate. Current studies were designed to investigate the biological fate of [2-14C]MAN in male F344 rats. Following gavage administration of 115, 11.5, or 1.15 mg MAN/kg in water, male F344 rats were placed in glass metabolism cages and urine, expired air, and feces were collected. Rats were sacrificed at various times, and the concentration of MAN-derived radioactivity in tissues was determined. MAN was rapidly absorbed from the gastrointestinal tract and distributed to all major tissues. After gavage administration of 1.15-115 mg/kg, [2-14C]MAN is primarily eliminated in the expired air. Sixty to 70% of the low and medium doses were exhaled as 14CO2 in 72 hr compared with 25% of the highest dose. Whereas 40% of the high dose was expired as organic volatiles in 72 hr, only 9-12% of the low and medium doses were exhaled as such. It is therefore apparent that saturation of MAN metabolism occurs at the high dose. HPLC analysis of expired organic volatiles from MAN-treated rats showed that it contained two components that were identified as unchanged MAN and acetone. The MAN:acetone ratio was directly proportional to dose and decreased as a function of time. Urinary excretion accounted for 20-30% of all MAN doses within 72 hr after dosing. Investigating the effect of dosing vehicle on MAN disposition in rats revealed that administration of 115 mg MAN/kg in oil resulted in the death of rats within 24 hr after treatment. Furthermore, monitoring the fate of MAN in these rats before death showed that a significantly higher percentage of the dose was eliminated in urine and expired air. Analysis of this expired air also revealed that significantly more acetone and less unchanged MAN were exhaled by these animals. It is apparent that administration of MAN to F344 rats in oil resulted in slower absorption, decreased elimination of unchanged MAN, and increased metabolism to acetone and/or decreased degradation of acetone to CO2. The combination of these effects of an oil vehicle may have contributed to the death of rats by MAN. Comparison of the metabolism and disposition of MAN in F344 and Sprague-Dawley rats showed minor differences between the two strains.(ABSTRACT TRUNCATED AT 400 WORDS)


Pharmacokinetics and renal handling of enprofylline in pregnant rats.

The pharmacokinetics and renal handling of enprofylline during pregnancy were investigated in Sprague-Dawley rats. Significant differences in the pharmacokinetic parameters of enprofylline were observed between nonpregnant rats and pregnant rats on the 20th day of gestation: volume of distribution was higher, and systemic clearance was lower in pregnant rats. Parameters obtained from rats at 7 days postpartum were the same as those obtained from nonpregnant rats. There were no significant differences in the fraction of urinary excretion of enprofylline between nonpregnant and pregnant rats. The protein binding of enprofylline in the plasma of pregnant rats was significantly lower than in nonpregnant rats, as a decrease in the albumin concentration consequentially reduced the binding capacity of enprofylline. The volume of distribution for unbound enprofylline in pregnant rats was not significantly different from nonpregnant rats, although a significant decrease was observed in pregnant rats in the systemic clearance for unbound enprofylline. In addition, the clearance ratio was lower in pregnant rats (2.8) when compared with nonpregnant rats (6.4). Pregnancy caused a decrease in the apparent maximum capacity of transport (Vmax) from 29.9 to 20.8 micrograms/min and in the Michaelis-Menten constant (KM) from 2.59 to 2.26 micrograms/ml, indicating that the tubular secretion ability of enprofylline becomes reduced during pregnancy. These results suggest that changes that occur in the plasma protein binding behavior and in renal handling as a result of pregnancy are primary factors influencing the disposition of enprofylline during pregnancy.


Species and tissue differences in the microsomal oxidation of 1,3-butadiene and the glutathione conjugation of butadiene monoxide in mice and rats. Possible role in 1,3-butadiene-induced toxicity.

Rat and mouse liver, lung, and kidney microsomes metabolized 1,3-butadiene to butadiene monoxide (BM), whereas microsomes from testis, one of the target organs of 1,3-butadiene toxicity in both species, were ineffective. 1,3-Butadiene metabolism was NADPH-dependent and inhibited by 1-benzylimidazole. With mouse microsomes, a 4-fold higher rate was measured with kidney compared with liver or lung, which exhibited similar rates. With rat microsomes, the rate obtained with liver was 2- and 6-fold higher than those of lung and kidney, respectively. Overall, oxidation rates by mouse tissues were higher than those of rat tissues. These results, along with the finding that BM was stable in the presence of rat plasma, provide evidence for the role of circulating metabolites in 1,3-butadiene-induced toxicity. Furthermore, crotonaldehyde, a known carcinogen, was detected with mouse tissues only. Thus, in addition to the greater ability of mouse tissues to produce BM, formation of crotonaldehye may contribute to the greater susceptibility of mice to 1,3-butadiene toxicity compared with rats. Nearly all rat liver glutathione S-transferase activity was localized to the cytosol (greater than 96%). BM glutathione conjugation rates with liver cytosol of both species were similar, whereas conjugation rates with mouse lung and kidney cytosol were 4- and 2-fold higher than those of rat lung and kidney, respectively. Thus, species differences in BM glutathione conjugation do not correlate with species susceptibility.(ABSTRACT TRUNCATED AT 250 WORDS)


Identification and comparison of the urinary metabolites of [1,2,3-13C3]acrylic acid and [1,2,3-13C3]propionic acid in the rat by homonuclear 13C nuclear magnetic resonance spectroscopy.

Acrylic acid (AA) and its esters are used extensively for the production of a variety of polymers. Despite their ubiquitous nature, little has been reported on the metabolism of the parent acid. The metabolites of AA may be volatile, unstable, polar, and thus difficult to isolate. Therefore, 13C NMR was used to help identify and compare directly the urinary metabolites of both 99% 13C-enriched AA and propionic acid (PA). Male Sprague-Dawley rats received [1,2,3-13C]AA (400 mg/kg in water p.o.) or an equimolar dose of [1,2,3-13C]propionate together with a radioactive tracer, [2,3-14C]AA, or [1-14C]propionate, respectively, and excreta were collected for 72 hr. For both acids, expiration of 14CO2 was the major route of elimination of radiolabel (approximately 80%). Approximately 6% of the dose was excreted in the urine. Urine was analyzed directly using proton-decoupled 13C and two-dimensional 13C homonuclear correlated NMR spectroscopy. The urine from AA-treated rats revealed major signals, the intensity of which was time-dependent, from at least five 13C-enriched metabolites of AA. Signals have been assigned to 3-hydroxypropionic acid, N-acetyl-S-(2-carboxyethyl)cysteine, and N-acetyl-S-(2-carboxyethyl)cysteine-S-oxide by comparison with spectra of authentic standards. No unchanged AA was detected. In contrast, the spectra of urine from a propionate-treated rat revealed only a few minor 13C-enriched signals that were assigned to methylmalonic acid. No unchanged PA was detected.(ABSTRACT TRUNCATED AT 250 WORDS)


Pharmacokinetics of intravenously injected [3H]tetrachloro(D,L-trans)1,2-diaminocyclohexane platinum (IV) (tetraplatin) in mice.

Platinum (Pt) complexes with diaminocyclohexane (DACH) carrier ligand are of interest because of their effectiveness against tumor cells resistant to cisplatin. We examined the pharmacokinetics and in vivo stability of Pt-N bond of tetraplatin using radiolabeled drug containing the 1,2-[4,5-3H]diaminocyclohexane ([3H]DACH) moiety and monitored the ratio 3H:Pt in biological samples. Following intravenous administration of [3H]tetraplatin (5 mg/kg and 163 microCi/kg) in mice, blood, urine, and tissue samples were collected and analyzed for both Pt level and radioactivity using flameless atomic absorption spectrophotometry and scintillation counting, respectively. In plasma, half-lives of 5 min (alpha-phase) and 1 hr (beta-phase) were generated for both "free" 3H and Pt. The 3H:Pt ratio of "free" levels did not change for at least 2 hr after intravenous administration. This indicated that the Pt-N bond was largely intact in the free drug fraction over this time period. In blood and plasma, however, there were decreases in "total" 3H:Pt ratios by 21 and 13% over 2 hr and 38 and 51% over 4 days, respectively, as a result of a greater rate of elimination of 3H than Pt. Urinary excretion in mice of 3H and Pt accounted for 55 and 43% of the administered dose, respectively, in 4 days, with 35 and 30% being excreted in the first 7 hr. The 3H:Pt ratio in urine increased 1.4-fold between 7 and 72 hr, and this was a further indication of dissociation of DACH ligand from Pt. Tissue distribution study revealed that the DACH moiety was also being removed more rapidly, and this was particularly evident in the liver and spleen than in other tissues.


Disposition of acrivastine in the male beagle dog.

Three male beagle dogs were given 10 mg/kg iv and oral doses of [14C]acrivastine, a novel nonsedating antihistaminic agent, in a nonrandomized crossover experiment. Urine and feces were collected for 72 hr after dosing. After iv dosing, a mean of 34% was recovered in the urine, and 63% was recovered in the feces. After po dosing, a mean of 29% of the radiocarbon was recovered in the urine, and 63% was recovered in the feces (dose adjusted for 14% lost in vomitus). Acrivastine and three major metabolites were detected in the excreta. The metabolites were identified as a side-chain-reduced analog of acrivastine (metabolite 3, 270C81), a gamma-aminobutyric acid analog of 270C81 (metabolite 2), and a benzoic acid analog of 270C81 (metabolite 1). After iv dosing, 34% of the dose was excreted as parent drug, 21% as metabolite 3, 15% as metabolite 2, and 6% as metabolite 1, while after po dosing, 35% of the dose was excreted as parent drug, 18% as metabolite 3, 11% as metabolite 2, and 7% as metabolite 1. Pharmacokinetic analysis of acrivastine plasma concentration-time curves after both routes of administration indicated a mean total body clearance of 17.3 ml/min/kg, a Vss of 0.93 liter/kg, a terminal half-life of 0.7 hr, and an oral bioavailability of 40%. The apparent plasma half-life of the metabolite, 270C81, was 1.5 hr. Analysis of AUC values indicated that greater amounts of 270C81 than acrivastine circulated in plasma after both iv and po dosing, and that first-pass metabolism of acrivastine to 270C81 occurred. The results indicated that acrivastine was extensively metabolized in the dog to 270C81 and suggested that 270C81 itself underwent further metabolism to metabolites 1 and 2.


Disposition of metabolically labeled recombinant soluble CD4 (sT4) in male Sprague-Dawley rats following intravenous and subcutaneous administration.

Soluble CD4 (sT4) has been metabolically labeled with [3H]leucine in Chinese hamster ovary cells and purified by S Sepharose chromatography. Over 250 microCi of high specific radioactivity [3H]sT4 (42 Ci/mmol) was prepared. The radiolabeled molecule was chemically and biologically representative of the unlabeled molecule and thus appropriate for in vivo metabolic investigations. To explore the biotransformation and disposition of a recombinant protein, this uniformly labeled [3H]sT4 was administered intravenously and subcutaneously to male Sprague-Dawley rats. Following a single dose of 0.3 mg/kg, blood samples were collected for 9 days and analyzed for total radioactivity, total plasma radioactivity, trichloroacetic acid-precipitable plasma radioactivity, sT4-related plasma radioactivity (by extraction with a Sepharose-bound polyclonal anti-sT4 antibody), and plasma sT4 concentration (by an N and C terminal-specific Leu3A/OKT4 ELISA). Excreta were analyzed for total radioactivity. The pharmacokinetic profiles of intact sT4 were as expected from the results of previous studies. sT4 was cleared rapidly from plasma with an elimination t1/2 of 7 min (intravenous), and low sT4 levels were observed following subcutaneous administration. Comparison of the kinetic profiles of total radiolabel, trichloroacetic acid-precipitable radiolabel, sT4-related radiolabel, and the isolation of plasma proteins containing tritium have led to the following conclusions. One of the major metabolic pathways for [3H]sT4 was the degradation of the polypeptide to its constituent amino acids, which were subsequently incorporated into endogenous proteins. Incorporation of tritium into blood cell proteins resulted in a prolonged radiolabel blood profile (t1/2 greater than 250 hr). Following subcutaneous administration, [3H] sT4 was significantly degraded before reaching the vascular circulation.


Bromo(monohydroxy)phenyl mercapturic acids. A new class of mercapturic acids from bromobenzene-treated rats.

Alkaline permethylation and GC/MS analysis of urinary mercapturic acids from rats given bromobenzene yielded several quinone-derived bromodimethoxythioanisole isomers as expected. Unexpectedly, seven bromomonomethoxythioanisole isomers were also observed, suggesting the presence of bromomonohydroxyphenyl mercapturic acids in the urine. Alkaline permethylation of synthetic 4- and 5-bromo-2-hydroxyphenyl mercapturic acid gave 4- and 5-bromo-2-methoxythioanisole, respectively, which were also observed after alkaline permethylation of urine from bromobenzene-treated rats, as was 2-bromo-4-methoxythioanisole. To explore the biosynthetic origin of the bromonohydroxyphenyl mercapturic acids, rats were separately dosed intraperitoneally with synthetic racemic 2-, 3-, or 4-bromophenyl mercapturic acid, or biosynthetic L-(-)-4-bromophenyl mercapturic acid, or a biosynthetic mixture of the 3,4- and 4,3-premercapturic acids from bromobenzene, and their urine (0-24 hr) analyzed by alkaline permethylation and GC/MS. The administered mercapturic acids and premercapturic acids were partly excreted unchanged (60-80% and 24%, respectively), but both gave rise to bromomonohydroxyphenyl mercapturic acids (0.1-5.2% of dose). Results indicated that the latter could be formed by 1) dehydrogenation of premercapturic acids and 2) hydroxylation of mercapturic acids (or their cysteine equivalents).


Exogenous glutathione decreases cellular cadmium uptake and toxicity.

The effect of intracellular glutathione (GSH) on cadmium metabolism and toxicity has been extensively investigated. However, little is known regarding the effect of extracellular GSH on cellular cadmium responses. Therefore, this study was conducted to investigate the effect of exogenously added GSH on cadmium toxicity in normal rat kidney fibroblasts (NRK-49F). Exponentially growing NRK-49F cells were arrested by serum deprivation and then stimulated with epidermal growth factor (EGF). CdCl2, at concentrations that range from 0.25 to 2 microM, was found to inhibit, in a dose-dependent fashion, the EGF-induced DNA synthesis (as judged by [3H]thymidine incorporation) in the cells. A long-term survival assay revealed that CdCl2 above 1 microM was toxic to the cells. Exogenous GSH had a dose-dependent antagonistic effect on cadmium inhibition of EGF-induced DNA synthesis, and 1 mM GSH was found to block completely cadmium inhibition of both EGF-induced DNA synthesis and cell survival. Exogenously added GSH did not increase intracellular GSH levels but decreased cadmium accumulation by the cells. This decrease was primarily caused by a reduced cadmium uptake. Further studies indicated that exogenous GSH would form a complex with cadmium outside of the cells preventing cellular cadmium uptake. This may explain the mechanism of action of the exogenous GSH in cytoprotection against cadmium. The results also suggested a practical potential for GSH as a cadmium chelator. If GSH were coadministered with a cadmium mobilizer and a gamma-glutamyl transpeptidase inhibitor, it could enhance cadmium excretion from the body.


Pharmacokinetics and pharmacodynamics of topotecan in patients with advanced cancer.

Topotecan, a semisynthetic water-soluble analog of camptothecin, is the first topoisomerase I-directed drug to enter clinical trial in the United States in over 20 yr. In this study, 30-min infusions of topotecan were administered daily for 5 days every 3 weeks at doses ranging from 0.5 to 2.5 mg/m2. Topotecan is reversibly hydrolyzed in a pH-dependent reaction in aqueous solutions to the ring-open hydroxy acid. The disposition of the closed ring lactone has been studied in 26 patients, and the disposition of both lactone and hydroxy acid has been studied in 12 patients. The clearance rate for topotecan lactone was 1220 ml/min/m2, with a range of 300-4760 ml/min/m2. The clearance rate for total topotecan (lactone and hydroxy acid) was 493 ml/min/m2, with a range of 163-815 ml/min/m2. A model for the disposition of lactone and hydroxy acid incorporating both reversible hydrolysis and elimination was developed. We have shown that topotecan is partially hydrolyzed prior to administration in parenteral solutions, and that clearance of the parent compound proceeds in vivo by conversion to hydroxy acid and elimination. Renal clearance accounted for 30 +/- 18% of drug elimination in patients. The relationship between topotecan dose and myelotoxicity is well fit by a sigmoidal Emax model, as is the relationship between total topotecan area under the concentration-time curve and myelotoxicity. The disposition of topotecan was also studied in mice. The clearance rate for total topotecan in mice was 330 ml/min/m2 after administration of topotecan lactone.(ABSTRACT TRUNCATED AT 250 WORDS)


Concomitant induction of microsomal epoxide hydrolase and UDP-glucuronosyltransferase activities by dipyridine compounds.

The coordinated response of the major rat hepatic phase II xenobiotic-metabolizing enzymes following 3-day exposure to diaryl compounds was investigated. Four diaryl compounds containing heterocyclic nitrogen atoms elevated microsomal epoxide hydrolase activity from 2- to 4-fold. Equivalent compounds lacking the heteroatom, when given in the same dosing regimen (75 mg/kg, ig, daily for 3 days), did not induce this or any other drug-metabolizing enzyme activity. Epoxide hydrolase activity closely paralleled UDP-glucuronosyltransferase activity toward three aglycones: 4-nitrophenol (r = 0.87), morphine (r = 0.84), and 1-naphthol (r = 0.78). There was less correlation (r = 0.60) between epoxide hydrolase activity and both UDP-glucuronosyltransferase activity toward testosterone and cytosolic glutathione S-transferase activity. There was no correlation between microsomal epoxide hydrolase activity and cytochrome P-450 or the monooxygenase reaction (4-nitrophenol hydroxylase) preferentially induced by pyridine-containing compounds. Induction of rat hepatic microsomal epoxide hydrolase activity by some pyridine-containing compounds appears coordinately regulated with glucuronidation rather than oxidation enzymes.


Comparative pharmacokinetics of cefoperazone and cephradine in untreated streptozotocin diabetic rats.

Experimental diabetes mellitus was induced in adult male rats by injecting streptozotocin (STZ; 60 mg/kg iv) for the purpose of surveying changes in the pharmacokinetics of biliary excretion after the intravenous administration of 40 mg/kg of cefoperazone (CPZ) or cephradine (CED). CPZ, CED, and other organic anions share affinity for the organic anion transport system in the bile canalicular membrane. The STZ treatment had a marked influence on the distribution and elimination of both cephalosporins. The blood levels of both cephalosporins at each time point after administration differed significantly between the STZ-treated and control rats. The values of mean residence time (MRT) of CPZ and CED were significantly decreased in the STZ-treated rats. Basal bile flow rates were increased after the administration of CPZ in the control and STZ-treated rats. Biliary clearance (CLbile) of CPZ was more than 60% of the CLtot, whereas CLbile of CED was less than 20% of CLtot in both groups of rats. The mean CLbile value of CPZ in the STZ-treated rats was 1.0 ml/min higher than that of the control rats, whereas the mean CLbile value of CED was almost the same as that of the control rats. The increased CLbile of CPZ suggested that diabetes alters the biliary excretion of CPZ. The changes in MRT of CPZ in the STZ-treated and control rats are mainly caused by an increase in the biliary excretory rate and renal clearance. The changes in MRT of CED in the STZ-treated and control rats are caused by a decrease in the apparent volume of distribution and increased renal clearance.


Enantioselectivity of microsomal and cytosolic esterases in rat intestinal mucosa.

Enantioselectivity of esterases, contained in 9,000g supernatant fraction (S9) prepared from homogenate of small intestinal mucosa of male Sprague-Dawley rats and the subsequent 105,000g supernatant (cytosol) and pellet (microsomes) prepared from S9, was studied using racemic oxazepam 3-acetate (rac-OXA) as the substrate. Esterases in S9 were enantioselective in hydrolyzing either S-OXA or R-OXA, depending on a particular subcellular preparation. Cytosolic and microsomal esterases had opposite enantioselectivity and selectively hydrolyzed S-OXA and R-OXA, respectively. Enantioselectivity of esterases solubilized from microsomes with Triton X-100 (0.1%, w/v) was identical to that of the membrane-bound microsomal esterases. Cytosolic esterases were more sensitive to temperature than either solubilized or membrane-bound microsomal esterases. In the presence of paraoxon (1 microM), the esterases selective toward R-OXA in both microsomes and S9 were completely inhibited, whereas the esterases selective toward S-OXA in cytosol were inhibited by approximately 6%. These results indicate that cytosolic and microsomal esterases in rat small intestinal mucosa are distinctly different enzymes, with opposite enantioselectivity in the hydrolysis of rac-OXA.


Pharmacokinetics in mice of the anti-retrovirus agent 9-(2-phosphonylmethoxyethyl)adenine.

The pharmacokinetics of 9-(2-phosphonylmethoxyethyl)adenine (PMEA), a potent inhibitor of retrovirus (i.e. human immunodeficiency virus) replication was determined in mice. Upon iv bolus administration of PMEA at 25, 100, or 500 mg/kg, PMEA was rapidly cleared from the plasma in a monoexponential and dose-independent manner (half-life, 7-12.5 min; distribution volume, 0.30-0.36 liter/kg; total body clearance, 1.21-2.41 liters/hr/kg). Irrespective of the initial PMEA dose, 67% of unchanged PMEA was recovered from the urine of mice within 24 hr after administration of PMEA. [3H]PMEA, administered as an iv bolus injection, mainly accumulated in the kidney, liver, and lungs. Significant amounts of monophosphorylated PMEA were detected in kidney and liver, but not other tissues, at 10, 30, and 60 min after iv administration of PMEA. Low but significant levels of PMEA were attained in the brain.


Benzodiazepine use and HIV risk-taking behaviour among injecting drug users.

This paper examines the prevalence of benzodiazepine use, and its relationship to other drug use and HIV risk-taking among a sample of 1245 injecting drug users (IDU). Approximately a third (36.6%) of the sample had used benzodiazepines during their last typical month of injecting. Benzodiazepine users had injected more frequently, injected more heroin and amphetamines, and had more poly-drug use than other IDU. They also had higher levels of HIV risk-taking, having shared injecting equipment more frequently and with more people. There were no differences between groups in number of sexual partners or condom use, although benzodiazepine users were more likely to have been paid for sex. The demographic and drug use variables indicate that benzodiazepine users are a more dysfunctional subgroup of IDU who require particular attention in HIV interventions.


Isolation and structural characterization by spectroscopic methods of two glucuronide metabolites of mexiletine after N-oxidation and deamination.

Urine samples from control and mexiletine-treated human subjects or rabbits (test group) were collected and passed through an ion exchange resin to isolate polar compounds. Methanolic eluates from control and test urines were analyzed by TLC. Exposure to p-dimethylaminocinnamaldehyde gave an additional intense pink band at Rt 0.40-0.45 in TLC analysis of test urine eluate when compared to control urine eluate. Non-exposed silica at this Rt was scraped and metabolites were extracted with methanol. Hydrolysis of this methanolic extract at 100 degrees C with hydrochloric acid released mexiletine. GC/MS and fast atom bombardment mass spectrometry analyses of nonhydrolyzed methanolic extracts evidenced the presence of two conjugated metabolites of mexiletine, namely, N-hydroxymexiletine glucuronide and mexiletine alcohol glucuronide. Synthetic compounds corresponding to these metabolites were obtained and spectra compared with those of isolated metabolites from urine. Definite structure assignment of N-hydroxymexiletine glucuronide was obtained from NMR spectrometry which confirmed the structure to be a hydoxylamine glucuronide (N-O-C link) and showed that the glycoside moiety was in the beta configuration. Thus, it is proposed that N-hydroxymexiletine glucuronide corresponds to mexiletine acid-labile conjugate and represents a major metabolic pathway in the disposition of mexiletine.


The pharmacokinetics of 1,2-diethyl-3-hydroxypyridin-4-one (CP94) in rats.

The pharmacokinetics of 1,2-diethyl-3-hydroxypyridin-4-one (CP94) and its 2-(1-hydroxyethyl) metabolite (metabolite A) were examined in male Wistar rats using a chronically cannulated conscious-rat model. Serial blood samples were assayed by a reversed phase HPLC method with UV detection. Following iv doses of 25, 50, and 100 mg/kg, the parent compound was eliminated from blood in a biexponential fashion with an average systemic clearance of 1.5 liters/hr/kg. The mean terminal elimination half-life was 2.02 hr and the mean volume of distribution at steady state was 2.69 liters/kg. The areas under the curve (AUCs) for the 25, 50, and 100 mg/kg iv doses were 15, 36, and 72 micrograms/ml/hr, respectively, suggesting that the disposition of CP94 in rats obeys linear kinetics. The oral bioavailability of CP94 (100 mg/kg) was about 53%. Peak blood concentration occurred at about 0.5 hr after oral administration. Following iv doses of CP94 at 25, 50, and 100 mg/kg, metabolite A peaked at about 0.75 hr.


[Type-2B multiple endocrine neoplasms with diffuse liver metastases as the cause of chronic diarrhea].

During examination before surgical correction of pes valgus a 20-year-old man reported having 3-5 pasty, foul smelling diarrhoeic motions per day for the past 3 years. He was noted to have rather thick lips and Marfan-like body build. Erythrocyte sedimentation rate was 18/34 mm, serum activity of GOT 22.5 U/l, GPT 35.7 U/l. Faecal weight was increased to 640 g/d, fat content to 12 g/d. Serum levels of the carcinoembryonic antigen (2494 ng/ml; normal: < 2.5) and of calcitonin (1,619,760 pg/ml; normal < 100) were elevated. Gastroscopy, partial coloscopy, colon-contrast imaging, ultrasonography and computed tomography of the neck and abdomen, as well as magnetic nuclear imaging of the neck were all normal. But laparoscopy revealed the liver to be infiltrated by small whitish nodules which immunohistologically proved to be metastases of a C-cell carcinoma. Total thyroidectomy was performed and the diagnosis of a C-cell carcinoma of the thyroid confirmed intra-operatively. After the operation the diarrhoea was stopped with codeine (9 mg/d). In case of tumour progression, therapy with octreotide, a somatostatin analogue, will be carried out. The concomitant occurrence of C-cell carcinoma, Marfan-like body build, thick lips and skeletal changes is typical of multiple endocrine neoplasia type 2B, which is caused by a chromosomal defect.


Metabolism of 7-(1,3-thiazolidin-2-ylmethyl)theophylline by rat liver microsomes. Evidence for a monooxygenase-dependent step in 1,3-thiazolidine ring cleavage.

Metabolic transformation of the mucoregulator and bronchodilator 7-(1,3-thiazolidin-2-ylmethyl)theophylline was studied in vitro with a rat liver microsomal preparation containing a NADPH-generating system. The only metabolite observed was 7-theophyllinacetaldehyde. In contrast to previous literature pointing out the chemical nature of 2-substituted thiazolidine ring cleavage, the formation of 7-theophyllinacetaldehyde was mediated by monooxygenase-dependent oxidation. Possibly an unstable sulfoxide was the first metabolic product, rapidly converted to 7-theophyllinacetaldehyde by hydrolysis. The sulfoxidation was apparently catalyzed mainly by flavin-containing monooxygenases, as selective thermal inactivation and methymazole significantly reduced the rate of formation of the metabolite. No N7-dealkylation pathway producing theophylline was detected, indicating a high regioselectivity in in vitro metabolism, due to the nucleophilicity of the sulfur atom.


Gonad formation and development requires the abd-A domain of the bithorax complex in Drosophila melanogaster.

The abdominal-A (abd-A) gene, a member of the bithorax complex, is required for the correct identity of parasegments (PS) 7 through 13. Mutations in iab-4, one of the cis-regulatory regions of abd-A, transform epidermal structures of PS 9 and also cause loss of gonads in adult flies. Here, we describe a developmental and molecular analysis of the role of iab-4 functions in gonadal development. In flies homozygous for a strong iab-4 allele, gonadogenesis is not initiated in the embryo because the mesodermal cells fail to encapsulate the pole cells. Flies homozygous for weaker iab-4 mutations sometimes form ovaries. The ovary-oviduct junctions are abnormal, however, and egg transfer from the ovary to the uterus is blocked in the adult. To localize the sites that require iab-4 function, we have analyzed animals chimeric for the mutant and wild-type cells. These chimeras were generated by three kinds of transplantation experiments: pole cells, embryonic somatic nuclei or larval ovaries. Our results suggest that iab-4 is required in the somatic cells of the gonadal primordia, but not the germ line. In addition, the formation of functional ovary-oviduct junctions and egg transfer also requires iab-4 functions in the somatic cells of the ovary and in at least one additional somatic tissue.