Dietary omega-3 polyunsaturated fatty acids induce plasminogen activator activity and DNA damage in rabbit spermatozoa.

The aim of this study was to determine the effect(s) of dietary omega-3 polyunsaturated fatty acids (ω-3 PUFA) on rabbit semen. Adult rabbit bucks were assigned to two groups that were given two diets, a standard diet (control) and a diet supplemented with ω-3 PUFA. Sperm samples were collected from all bucks with the use of an artificial vagina in 20-day intervals, for a total period of 120 days. The enrichment of membranes in ω-3 PUFA was manifested by the elevation of the 22:5 ω-3 (docosapentaenoic acid [DPA]) levels within 40 days. This increase in DPA content did not affect semen characteristics (i.e., concentration, motility and viability). However, it was associated with the induction of lipid peroxidation in spermatozoa, as determined on the basis of the malondialdehyde content. Lipid peroxidation was associated with DNA fragmentation in ω-3 PUFA-enriched spermatozoa and a concomitant increase in plasminogen activator (PA) activity. The effects of ω-3 PUFA on sperm cells were evident within 40 days of ω-3 PUFA dietary intake and exhibited peack values on day 120. Our findings suggest that an ω-3 PUFA-rich diet may not affect semen characteristics; however, it may have a negative impact on the oxidative status and DNA integrity of the spermatozoa, which was associated with an induction of PAs activity.

Effects of sub-lethal neurite outgrowth inhibitory concentrations of chlorpyrifos oxon on cytoskeletal proteins and acetylcholinesterase in differentiating N2a cells.

Previous work in our laboratory has shown that sub-lethal concentrations (1-10 µM) of chlorpyrifos (CPF), diazinon (DZ) and diazinon oxon (DZO) inhibit the outgrowth of axon-like neurites in differentiating mouse N2a neuroblastoma cells concomitant with altered levels and/or phosphorylation state of axonal cytoskeleton and growth-associated proteins. The aim of the present work was to determine whether chlorpyrifos oxon (CPO) was capable of inhibiting N2a cell differentiation in a similar manner. Using experimental conditions similar to our previous work, sub-lethal concentrations (1-10 µM) of CPO were found to inhibit N2a cell differentiation. However, unlike previous studies with DZ and DZO, there was a high level of sustained inhibition of acetylcholinesterase (AChE) in CPO treated cells. Impairment of neurite outgrowth was also associated with reduced levels of growth associated protein-43 and neurofilament heavy chain (NFH), and the distribution of NFH in cells stained by indirect immunofluorescence was disrupted. However, in contrast to previous findings for DZO, the absolute level of phosphorylated NFH was unaffected by CPO exposure. Taken together, the findings suggest that sub-lethal concentrations of CPO inhibit axon outgrowth in differentiating N2a cells and that this effect involves reduced levels of two proteins that play key roles in axon outgrowth and maintenance. Although the inhibition of neurite outgrowth is unlikely to involve AChE inhibition directly, further work will help to determine whether the persistent inhibition of AChE by CPO can account for the different effects induced by CPO and DZO on the levels of total and phosphorylated NFH.

Cisapride reverses the anticholinergic effect of disopyramide on the isolated guinea-pig urinary bladder.

The present investigation aims to examine whether the prokinetic agent cisapride is able to reverse disopyramide's anticholinergic effect on the isolated guinea-pig urinary bladder. Acetylcholine, at concentrations ranging from 10(-7) to 10(-3) M, produced a stimulatory effect on the urinary bladder (pEC(50) value=5.1). Disopyramide competitively antagonized the contractile effect of acetylcholine with an ID(50)=4.4 x 10(-6) M. Although cisapride by itself had either no intrinsic contractile action or a modest effect on the urinary bladder, at concentrations ranging from 3 x 10(-7) to 10(-6) M, it significantly reversed the above inhibitory effect of disopyramide, and produced a parallel leftward shift of the concentration-response curve for acetylcholine in the presence of disopyramide. The pEC(50) values for acetylcholine in the presence of 3 x 10(-6) M and 10(-5) M disopyramide were 4.7 and 4.2, respectively, while in the presence of 10(-5) M disopyramide, after pretreatment with 5 x 10(-7) M cisapride, the pEC(50) value for acetylcholine was 4.6. It is concluded that cisapride is effective in reversing the anticholinergic activity of disopyramide on the isolated guinea-pig urinary bladder, probably by facilitating cholinergic neurotransmission.

Biphenylacetic acid enhances the antagonistic action of fluoroquinolones on the GABA(A)-mediated responses of the isolated guinea-pig ileum.

This paper examines the effect of biphenylacetic acid on the antagonistic action of norfloxacin and enoxacin on the GABA(A)-mediated responses of the isolated guinea-pig ileum. GABA produced transient contractions followed by relaxation. The contractile effect of exogenously applied GABA was concentration-dependent with EC(50)= 9.8 x 10(-6) M. This contractile effect was not significantly modified by biphenylacetic acid, and the EC(50) value for GABA in the presence of 10(-5) M biphenylacetic acid was 1.15 x 10(-5) M. The GABA contractile effect was inhibited, dose-dependently, by either norfloxacin or enoxacin, but only at concentrations higher than 10(-5) M. The response of the ileum to GABA (at EC(50)) was reduced to 35 and 36% by pretreatment with 10(-5) M norfloxacin or enoxacin, respectively. However, in the presence of 10(-5) M biphenylacetic acid, the response of the ileum to GABA was reduced to 2.2% by pretreatment with 10(-5) M enoxacin, while it was completely abolished by pretreatment with 10(-5) M norfloxacin and the IC(50) values were 5.5 x 10(-7) and 1.5 x 10(-6) M for norfloxacin and enoxacin, respectively. These data show that biphenylacetic acid whilst having no effect at the GABA(A)-mediated contractile response of the guinea-pig ileum, enhances the antagonistic effect of both enoxacin and norfloxacin. This suggests that combined administration of fluoroquinolones and biphenylacetic acid synergistically inhibits GABA(A)-receptors at the intestinal level.

Synergistic and antagonistic pharmacodynamic interaction between ranitidine and cisapride: a study on the isolated rabbit intestine.

The present study examines the pharmacodynamic interaction between the H(2)-receptor antagonist ranitidine and the prokinetic agent cisapride on the isolated rabbit intestine. Ranitidine produced a concentration-dependent contractile effect on the duodenal, ileal and ascending colon preparations, with EC(50)values of 1.35 x 10(-4)M for the duodenum, 1.2 x 10(-4)M for the ileum and 1.15 x 10(-4)M for the ascending colon. The effect of cisapride on the ranitidine contractile effect was dependent on the cisapride concentration used. Thus, cisapride, at concentrations from 10(-10)up to 5 x 10(-7)for the duodenum and the ascending colon and up to 10(-6)M for the ileum, potentiated the contractile responses of the preparations to ranitidine. However, at higher concentrations cisapride produced a non-competitive inhibition of the intestinal contractile responses to ranitidine with IC(50)values of 4.2 x 10(-5)M for the duodenum, 1.65 x 10(-5)M for the ileum and 3.2 x 10(-6)M for the ascending colon. These data show that cisapride may modify the contractile responses of the isolated rabbit intestine to ranitidine, having a potentiating effect up to a certain concentration and an antagonistic one at higher concentrations. In conclusion, co-administration of the above drugs may lead to enhanced or reduced intestinal motility.

Inhibition of rat platelet 5-hydroxytryptamine uptake by chlorpyrifos and carbaryl.

The organophosphate insecticide chlorpyrifos and the carbamate insecticide carbaryl were investigated in adult male rats in terms of their effects on the activity of brain monoamine oxidase-A (MAO-A) activity and on the platelet uptake of 5-hydroxytryptamine (5-HT). The activities of brain acetylcholinesterase (AChE) and plasma butyrylcholinesterase (BuChE) were also determined. For each compound two different dosage regimens were employed. In the acute study, chlorpyrifos or carbaryl was administered at a single intraperitoneal dose of 100 mg/kg or 50 mg/kg, respectively In the subacute study, chlorpyrifos was injected at a daily dose of 20 mg/kg for 7 days, while carbaryl was given at a daily dose of 10 mg/kg for 14 days. Acute chlorpyrifos administration produced a 85.01% inhibition of AChE and a 43.4% inhibition of BuChE but had no effect on MAO-A activity and 5-HT uptake. In contrast, subacute chlorpyrifos exposure caused a 94.96% inhibition of AChE and a 85.8% inhibition of BuChE and, also, elicited a significant (35.02%) reduction in the platelet uptake of 5-HT. MAO-A activity was not affected. Acute carbaryl administration produced a 56.38% AChE inhibition and a 55.95% BuChE inhibition and also caused a significant (26.36%) decrease in 5-HT uptake but no change in MAO-A. Subacute carbaryl exposure failed to affect significantly any of the biochemical parameters determined. Interference with the 5-HT system by chlorpyrifos and carbaryl could contribute to the toxicity of these pesticides.

Propofol's biphasic effect on GABA(A)-receptor-mediated response of the isolated guinea pig ileum.

This study examines the effect of the anaesthetic agent propofol on the guinea pig ileum and whether this effect derives from an interaction of the drug with GABA receptors. Propofol produced a biphasic effect consisting of a dose-dependent contractile effect (EC(50)=2.2x10(-5)m) followed by an 'after relaxation'. This propofol effect was similar to the one produced by GABA and was bicuculline-sensitive (ID(50)=3.2x10(-7)m). Propofol, at concentrations of 10(-7)and 10(-6)m, potentiated the ileum contractile responses to GABA, but only at the lower dose range of applied GABA, while at a concentration of 10(-5)m, it inhibited the contractile effect over the entire dose range of applied GABA. In addition, while the contractile response of the ileum to exogenously applied acetylcholine was not influenced by propofol at concentrations of up to 7x10(-6)m, it was antagonised by higher concentrations of propofol. In conclusion, the above data permit us to suggest that propofol's contractile effect on the guinea pig ileum is mediated by an interaction of the drug with GABA(A)-receptors.

The influence of Datura ferox alkaloids on egg-laying hens.

Seeds of the weed Datura ferox are frequent contaminants of raw materials used for animal feed. In this study a mixture of scopolamine and hyoscyamine (98:2), the 2 main alkaloids of Datura ferox seeds, was incorporated at 4 total alkaloid levels (1.5, 15, 75 or 150 mg/kg feed) into a control diet fed to 100 egg-laying hens for 3 mo. Alkaloid doses of 150 mg/kg feed reduced egg production for the first 5-6 w of feeding, whereas lower doses had no effect. Egg weight, eggshell thickness and body weight of hens were unaffected at all doses. Doses of 150 mg/kg feed produced significant increases in the cardiac rate of hens after 5 w. Breathing frequency at all doses was unaffected. Determination of plasma aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and leucine aminopeptidase activities, as well as autopsy and histological examinations, revealed no obvious alkaloid-related toxic effects. It was concluded that a total alkaloid dose as high as 75 mg/kg feed can be safely administered to egg-laying hens.

Toxicity study of the main alkaloids of Datura ferox in broilers.

Seeds of the weed Datura ferox are frequent contaminants of raw materials used for animal feed. These seeds produce various toxic effects and contain mainly the alkaloids scopolamine and hyoscyamine. In this 3-month toxicity study, a mixture of scopolamine and hyoscyamine (98:2) was incorporated at four total alkaloid levels (1.5, 15, 75 or 150 mg/kg feed) into a control diet fed to 100 broilers. Alkaloid feeding caused significant reductions in the body weight gain of birds, especially of those fed a dose of 150 mg alkaloid/kg feed. Growth-retarding effects, however, were transient, as no changes in body weight gain were noted after 52 days of alkaloid feeding. Alkaloid-treated broilers showed no significant differences from controls with respect to the cardiac rate and breathing frequency nor in relation to plasma aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase activities. In contrast, plasma leucine aminopeptidase activity was significantly reduced after 3 months in all alkaloid-fed birds. Autopsy and histological examination of tissues by light and electron microscopy revealed no pathological changes associated with alkaloid feeding. Broilers appeared generally healthy and behaved normally throughout. These data should be considered in the formulation of new, improved regulations defining the maximum allowable alkaloid content of D. ferox seeds contaminating raw materials destined for use as broiler feed.

Effects of polychlorinated biphenyls with Ah receptor affinity on lymphoid development in the thymus and the bursa of Fabricius of chick embryos in ovo and in mouse thymus anlagen in vitro.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and its congeners bind to the Ah receptor and are known to cause thymic atrophy in most experimental animal species and also to inhibit lymphoid development in the embryonic thymus (T-cells) and in the bursa of Fabricius of chick embryos (B-cells). The coplanar polychlorinated biphenyls (PCBs) 3,3',4,4'-tetrachlorobiphenyl (TCB), 3,3',4,4',5-pentachlorobiphenyl (PeCB), and 3,3',4,4',5,5'-hexachlorobiphenyl (HCB) (relatively strong Ah receptor ligands) and the mono-ortho-chlorinated analogues of TCB and PeCB (relatively weak Ah receptor ligands) were administered to chick embryos by air chamber injection on Day 13 of incubation. The numbers of lymphoid cells (on Day 19) in the thymus and the bursa of Fabricius were lower, in a dose-dependent manner, in embryos treated with the coplanar PCBs compared with controls. Approximate ED50 values for inhibition of bursal cell development were 4 micrograms for PeCB, 50 micrograms for TCB, and 300 micrograms/kg egg for HCB. The most immunotoxic of the mono-ortho-chlorinated analogues of TCB and PeCB were about 1000 times less potent than PeCB. The in vitro effects of the PCBs were studied in organ cultures of thymi from 15-day-old mouse fetuses. The three coplanar chlorobiphenyls inhibited lymphoid development in this culture system in a dose-dependent manner. PeCB was only about 10 times less potent (EC50 approximately 2 x 10(-9) M) than than TCDD (EC50 approximately 2 x 10(-10) M), whereas HCB and TCB were about 100 times less toxic than PeCB. No inhibition of lymphoid development by the mono-ortho-chlorinated PCBs was observed using concentrations as high as 10(-6) M.

TCDD inhibits the support of B-cell development by the bursa of Fabricius.

The toxic effects produced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and its congeners include inhibition of lymphoid development. We have previously found an inhibition of B-cell development in the bursa of Fabricius of chick embryos treated with TCDD congeners in ovo. In the present study, the bursae of ten-day-old chick embryos were removed and cultured on filter paper for 24 hr in media with or without TCDD or 3,3',4,4'-tetrachlorazoxybenzene (TCAOB). Following culture, the bursae were transplanted onto the chorioallantoic membrane (CAM) of ten-day-old eggs of the same strain or of a strain expressing a different B-cell surface alloantigen. After 5 days on the CAM the number of B-cells was determined or the grafts were sectioned for subsequent immunohistochemistry. Results were as follows: 1) A lower number of lymphoid cells (dose dependent) was observed in the TCDD-treated transplants amounting to 40-50% of that in the controls at 10(-9) M TCDD. Higher concentrations of TCDD compromised survival of the grafts. A single concentration of TCAOB (10(-8) M) was tested, resulting in a lymphoid cell number of 60% of that of the controls. 2) The bursal epithelium showed relatively normal development even in cases where B-cell development was affected. 3) Lymphoid cells in the grafted bursae originated from the embryo of the host egg. These findings suggest that the TCDD congeners had a direct effect on the bursa of Fabricius, leading to an inhibition of lymphoid development. It is likely that the microenvironment is affected by these compounds, thus resulting in a decrease in the attraction of stem cells and/or in the capacity to induce proliferation of the colonizing cells.

Effects of TCDD and its congeners 3,3',4,4'-tetrachloroazoxybenzene and 3,3',4,4'-tetrachlorobiphenyl on lymphoid development in the thymus of avian embryos.

Thymus anlagen from 11-day-old chick and 14-day-old turkey and duck embryos were cultured in media containing 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) for 5 days. The maximal TCDD-induced decrease in lymphoid cell number of chick embryo thymus (to about 60% of the control number) occurred at concentrations of 10(-10) M and above. To produce the same effect on lymphoid cell number in the cultures of thymus anlagen from turkey and duck embryos, about a 100-fold higher concentration of TCDD was needed. The toxicity of the TCDD congeners 3,3'4,4'-tetrachloroazoxybenzene (TCAOB) and 3,3',4,4'-tetrachlorobiphenyl (TCB) to embryonic chicken thymus was tested in vitro and in ovo. In chick embryo thymus cultures, TCAOB and TCB were about two orders of magnitude less toxic than TCDD. Injection of TCAOB and TCB into chicken eggs preincubated for 11 days resulted in a dose-dependent decrease in thymic lymphoid cell number 5 days later, declining to about 14% of the controls at 10 micrograms TCAOB/kg egg. The ED50 value was estimated to be 3.6 and 60 micrograms/kg egg for TCAOB and TCB, respectively.

Effects of the TCDD congeners 3,3',4,4'-tetrachlorobiphenyl and 3,3',4,4'-tetrachloroazoxybenzene on lymphoid development in the bursa of Fabricius of the chick embryo.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and its congeners, such as 3,3',4,4'-tetrachlorobiphenyl (TCB) and 3,3',4,4'-tetrachloroazoxybenzene (TCAOB), act on targets in the immune system, probably by interacting with the Ah-receptor, causing a characteristic pattern of effects typified by inhibition of lymphoid development in the thymus. There are, however, also reports of effects on B cells. Birds have a well-defined site of B-lymphocyte development, the bursa of Fabricius, analogous to the thymus for T cells. Thus, we wanted to determine whether this organ is suitable for studying effects on B-cell development. TCB and TCAOB were administered by injection into the air sacs of White Leghorn eggs on Day 13 of incubation. Effects observed on Day 19 were as follows: (1) There was a reduction in bursal dry weight but not in body weight in the treated groups. (2) The number of lymphoid cells in the bursae decreased in a dose-dependent manner, and bursae from high-dose groups (300 and 30 micrograms/kg egg of TCB and TCAOB, respectively) were almost completely devoid of lymphoid cells. For the reduction of lymphoid cell number, the ED50 for TCB and TCAOB was approximately 45 and 1.4 micrograms/kg egg, respectively. (3) Histological sections showed that embryos from treated egges contained fewer bursal follicles and that follicles contained fewer lymphoid cells compared with controls. (4) Aryl hydrocarbon hydroxylase activity was 30 and 50 times that of the control at high doses of TCB and TCAOB (estimated ED50: 200-300 and 4 micrograms/kg egg, respectively). These findings suggest that lymphoid development in the bursa of Fabricius of the chick embryo is inhibited by TCDD congeners.

The relationship of histologic and clinical factors in laryngeal papillomatosis.

Laryngeal papillomatosis is a disease that can lead to many surgical procedures, especially in children, and is potentially lethal. In this study, we reviewed 83 cases that yielded 902 separate laryngoscopy specimens to determine if any clinical or histologic findings were prognostic. "Juvenile" (n = 73) and "adult" (n = 10) cases were classified according to the number of separate lesions and the number of recurrences, but not necessarily according to age. Four prognostic indicators were constructed for the juvenile group and were analyzed by linear regression. Three new microscopic classifications were used: papillary, acanthomatous, and angiokeratotic. In juvenile cases, the last two categories appeared to be somewhat prognostic, along with several other factors. Some histopathologic findings contradicted classic descriptions. Differences in clinical manifestation and similarities in histopathology may suggest differential responses to the same causal agent.