Snake Liver Research Articles

Hepatozoon cf. terzii (Sambon & Seligman, 1907) infection in the snake boa constrictor constrictor from north Brazil: transmission to the mosquito Culex quinquefasciatus and the lizard Tropidurus torquatus.

Specimens of Hepatozoon-infected Boa constrictor constrictor were obtained from localities in Pará State, north Brazil. Gametocytes in erythrocytes of the peripheral blood measured 10 x 2.5-16.2 x 3.7 microns. They were similar to those described as Haemogregarina terzii by Sambon & Seligmann (1907) in B. c. constrictor, in that they did not distort the infected erythrocyte, and their dimensions approximated those given by Carini (1947). Lungs and liver of infected snakes contained actively dividing meronts of a single type, and cysts containing two to six cystozoites were also present in the liver. Our initial feeding of Culex quinquefasciatus on infected snakes consistently resulted in a heavy death-rate of the engorged mosquitoes, with only a few surviving till the 9th day post feeding. These contained numerous oocysts which were undivided or in early stages of division. A fifth and final experiment, however, provided a few mosquitoes surviving up to 21 days post infection (dpi), and these contained fully sporulated oocysts measuring 190-200 microns in diameter and containing over 60 sporocysts of 19-30 microns in diameter. The number of sporozoites in each sporocyst was estimated as approximately 50. The nature of the parasite's sporogonic cycle in the mosquito thus justifies inclusion of this haemogregarine in the genus Hepatozoon. Two wild-caught specimens of the lizard Tropidurus torquatus were fed with mosquitoes containing fully developed oocysts (21 dpi). When sacrificed, three months later, large numbers of dizoic, tetrazoic and hexazoic cysts were demonstrated in their livers. Cystozoites released from these cysts were shown to possess a conspicuous refractile body.

IMP-GMP 5′-nucleotidase in reptiles: occurrence in a turtle, a tortoise and three species of snakes

IMP-hydrolyzing activity, which is reactive with goose anti-pig lung IMP-GMP 5′-nucleotidase (c-N-II: EC.3.1.3.5) serum, was detected in extracts from liver, heart, kidney, spleen, stomach, skeletal muscle and lung from several species of reptiles: The values found in liver (U/mg protein) of one animal were: 4.5 for an ammono-ureotelic turtle ( Trionyx sinensis japonicus); 3.7 for an ureo-uricotelic tortoise ( Testudo elegans); 13–23 for three species of uricotelic snakes: Elaphe quadrivirgata, Elaphe conspicillata and Elaphe climacophora. These findings suggest that in the liver of snakes, c-N-II may participate in the production of uric acid as an end product of amino acid metabolism.

Liver Histopathology of the Southern Watersnake, Nerodia fasciata fasciata, Following Chronic Exposure to Trace Element-Contaminated Prey from a Coal Ash Disposal Site

Previous studies have demonstrated the accumulation of arsenic, cadmium, selenium, strontium, and vanadium in livers of Southern Watersnakes fed fish from a coal-ash contaminated site. Our study is the first to investigate effects of trace element accumulation on cytology of snake liver. Snakes were born in the laboratory and raised for one or two years on diets consisting of varying proportions of contaminated fish. The majority (71%) of snakes fed contaminated prey did not exhibit any differences in liver histology when compared to control snakes fed an uncontaminated diet. In the remaining contaminant-exposed snakes, some aberrations were noted. The most prevalent pathology involved the proliferation of collagen fibers that resulted in narrowing or occlusion of sinusoids and increasing the mass of the intersinsuoidal parenychma. Fibrosis of the liver as a result of chronic injury has been reported previously in reptiles, but this is the first report that links such tissue damage to dietary contamination.

Structural flexibility of the small intestine and liver of garter snakes in response to feeding and fasting.

Garter snakes Thamnophis sirtalis parietalis feed frequently but also tolerate extended periods of fasting when food is unavailable. We studied the dynamics, reversibility and repeatability of size changes of the small intestine and liver using ultrasonography. We employed light and transmission electron microscopy and flow cytometry to study the tissue mechanism that drives this flexibility. We compared garter snakes that fed every other day, snakes that fed once a week and fasting snakes. In all feeding trials, the size of the small intestine and the liver increased rapidly after feeding. Constantly feeding snakes maintained an elevated level of organ size, while snakes that were fed only once a week showed a marked up- and downregulation of organ size. Histology revealed the mucosal epithelium to be a transitional epithelium that can change cell configuration considerably to accommodate organ size changes. Upregulation of small intestine and liver size was always associated with the incorporation of lipid droplets into enterocytes and hepatocytes. Cell proliferation was not involved in upregulation of organ size. In contrast, cell proliferation increased during downregulation of organ size, indicating that cells worn out during digestion were replaced. The dynamics of flexibility and the functional features of the tissue were the same as described for the Burmese python Python molurus bivittatus. We suggest that garter snakes employ the same energetically cheap mechanism of organ size regulation as pythons, which allows for rapid, repeated and reversible size changes with no cell proliferation involved. Comparative evidence suggests that the transitional mucosal epithelium is an ancestral character of snakes and that feeding ecology is not directly related to the cytological features of the mucosal epithelium.

Bumpus in the snake den: effects of sex, size, and body condition on mortality of red-sided garter snakes.

Huge breeding aggregations of red-sided garter snakes (Thamnophis sirtalis parietalis) at overwintering dens in Manitoba provide a unique opportunity to identify sources of mortality and to clarify factors that influence a snake's vulnerability to these factors. Comparisons of sexes, body sizes, and body condition of more than 1000 dead snakes versus live animals sampled at the same time reveal significant biases. Three primary sources of mortality were identified. Predation by crows, Corvus brachyrhynchos (590 snakes killed), was focussed mostly on small snakes of both sexes. Crows generally removed the snake's liver and left the carcass, but very small snakes were sometimes brought back to the nest. Suffocation beneath massive piles of other snakes within the den (301 dead animals) involved mostly small males and (to a lesser extent) large females; snakes in poor body condition were particularly vulnerable. Many emaciated snakes (n = 142, mostly females) also died without overt injuries, probably due to depleted energy reserves. These biases in vulnerability are readily interpretable from information on behavioral ecology of the snakes. For example, sex biases in mortality reflect differences in postemergence behavior and locomotor capacity, the greater attractiveness of larger females to males, and the high energy costs of reproduction for females.

Modulation of Snake Hepatic Cytochrome P450 by 3-Methylcholanthrene and Phenobarbital

Induction mode of the hepatic drug-metabolizing enzymes was studied in Corn snake ( Elaphe guttata emoryi). Treatment of snakes with 3-methylcholanthrene or phenobarbital produced no effects on liver weight and total content of cytochromes P450 and b 5. Treatment with 3-methylcholanthrene significantly induced the activities of arylhydrocarbon hydroxylase, 7-ethoxyresorufin O-deethylase and 7-pentoxyresorufin O-dealkylase, whereas those of ethoxycoumarin O-deethylase, benzphetamine N-demethylase, erythromycin N-demethylase and testosterone hydroxylases were not affected. 3-Methylcholanthrene-induced activities of 7-ethoxyresorufin O-deethylase and 7-pentoxyresorufin O-dealkylase were inhibited by 20 μM α-naphthoflavone by 98% and 73%, respectively. Phenobarbital-treatment caused a significant induction of the activities of erythromycin N-demethylase and testosterone 6 β-hydroxylase, but did not affect those of the other phase I enzymes and the other testosterone hydroxylases. The activities of UDP-glucuronyltransferase and glutathione S-transferase were not affected by either 3-methylcholanthrene or phenobarbital administration. Immunoblotting showed that 3-methylcholanthrene-treatment induced a protein band related to hamster CYP1A2, and decreased the intensity of the two bands detected with anti-rat CYP2B1. Phenobarbital-treatment did not affect the intensity of CYP2B-related proteins. The results suggest that snake liver has multiple forms of cytochrome P450, notably those inducible by 3-methylcholanthrene.

Liver microsomes from the yellow rat snake (Elaphe obsoleta) and American bullfrog (Rana catesbeiana) oxidize polyunsaturated fatty acids by NADPH-dependent hydroxylation and epoxidation.

Polyunsaturated fatty acids (PUFAs) can be oxygenated by mammalian hepatic P450s to a series of metabolites. The most prominent of these are formed by omega- and (omega-1)-hydroxylation, epoxidation of the double bonds or bisallylic hydroxylation. The object of the present investigation was to determine whether similar oxygenations are catalyzed by liver microsomes of the yellow rat snake (Elaphe obsoleta) and the American bullfrog (Rana catesbeiana). Liver microsomes were incubated with [1-14C]-labeled arachidonic (AA), eicosapentaenoic (EPA), and linoleic acids (LA) in the presence or absence of 1 mM NADPH, and the major metabolites were analyzed by reverse-phase and straight-phase high performance liquid chromatography and capillary gas chromatography-mass spectroscopy. No metabolites were produced in the absence of NADPH. Profiles of metabolites were different depending on the organism and the acclimation state. In all incubations, EPA was the most effective substrate utilized and LA the least effective. The major products from EPA were 19-HEPE, 13-HEPE, and 20-HEPE from cold-acclimated (5 degrees C), warm-acclimated (22 degrees C) frogs, and snakes (22 degrees C), respectively. In contrast, 20-HETE production from AA was greater than 19-HETE in all three. Cold-acclimated frog liver microsomes produced significantly more of all metabolites when compared with microsomes from warm-acclimated frogs. We conclude that amphibian and snake liver can catalyze epoxidation and hydroxylation of PUFAs and that products are species-specific and acclimation-state dependent.

Antioxidant defenses in the tolerance of freezing and anoxia by garter snakes.

The garter snake Thamnophis sirtalis parietalis can readily tolerate several hours of freezing or anoxia exposure. Both stresses halt oxygen availability to tissues and to endure these stresses snakes must cope with potential oxidative stress arising as a result of the ischemic/anoxic condition followed by reperfusion of aerated blood during recovery. To determine whether antioxidant defenses are important for freezing and anoxia survival, we monitored the activities of antioxidant enzymes and the levels of glutathione (GSH and GSSG) during freezing (5 h at -2.5 degrees C) and anoxia (10 h under N2 gas at 5 degrees C) exposures in three organs (muscle, liver, and lung) of snakes. Freezing resulted in a significant rise in the activity of muscle and lung catalase (by 183 and 63%) and in muscle glutathione peroxidase (52%). Anoxia enhanced muscle and liver superoxide dismutase activities (by 59 and 118%) and also caused a 57% increase in muscle GSH levels. The increase in muscle GSH concentration in anoxia (from 0.45 to 0.71 mM) could also stimulate muscle glutathione peroxidase activity in vivo by 1.5-fold because of its low affinity for GSH (Km = 11 mM). The ratio of GSSG/GSH was not affected by experimental state in any tissue, suggesting that oxidative stress did not occur during the freezing or anoxic exposure. Rather, H2O2- and O2(-)-detoxification systems may be activated in preparation for possible oxygen free radical overgeneration during thawing or reoxygenation. Antioxidant defenses appear to be part of the adaptive machinery for reptilian tolerance of freezing and anoxia.

Immunorecognition of estrogen receptors by monoclonal antibody H222 in reproductive tissues of the red-sided garter snake

A study was conducted to determine if a monoclonal antibody (MAB), H222, prepared against human breast cancer estrogen receptors (ER) would recognize ER in the oviduct and liver of garter snakes (Thamnophis sirtalis parietalis). Using sucrose gradient analysis of antibody-ER complexes bound to [3H]estradiol we have determined that the MAB H222 binds to an ER in the cytosolic and nuclear extracts of snake tissues. The snake ER is not bound by nonspecific MABs in the sucrose gradient analysis. Further, the snake ER does not bind to other steroids, including a synthetic progestin, R5020, or the androgens 5 alpha dihydrotestosterone and R1881. The quantity of ER in the snake oviduct (200-700 fmol/mg DNA) is within an order of magnitude of that found in the oviduct of the nonhuman primate. These results suggest that the MAB H222 and 17 beta-estradiol bind to an ER in the snake that shares common properties with mammalian ER.

Dolichols in the liver of some poikilothermic vertebrates and birds

1. 1. The liver tissue from nine species of amphibians, reptiles and birds were tested for dolichol concentration and isoprenologue distribution of dolichol family by high-performance liquid chromatography. In the case of chicken ( Gallus domesticus) and grass snake ( Natrix matrix L.) the liver dolichol content was determined in respect of sex. 2. 2. Great fluctuations in the total content of dolichols in the liver of reptiles could be observed–from 4.4 μg/g for grass snake liver (male) to 242.1/gmg/g for common wall lizard; a 50-fold increase. 3. 3. The average value of total dolichol content in the liver of adult birds tested ( x = 170.3 μg/g) was higher in comparison with that of adult individuals of amphibians and reptiles (with the exception of common wall lizard and grass snake (male), x = 61.4 μg/g, independently of variations in each group investigated. 4. 4. The dolichol amount in the liver of male animals tested was lower, compared to the respective female ones. In the case of grass snake, the liver dolichol concentration of female specimens was about five times higher in comparison with the male. 5. 5. The chain length of the different isoprene analogues of liver dolichol family varied more frequently between 16 and 22 isoprene units, but in some liver tissue dolichols, 15, 23 and 24 isoprene units could be detected. 6. 6. The main analogue component in the liver dolichols of most species of poikilothermic vertebrates and birds was dolichol-19.

Xanthine:NAD + oxidoreductase in the liver of the teleostean fish Cyprinus carpio

1. 1. The enzyme hydroxylating oxypurines in the liver of the teleostean fish Cyprinus carpio is found to be xanthine:NAD + oxidoreductase (EC 1.2.1.37) non-convertible to the oxidase form. 2. 2. The Michaelis constants for NAD + and xanthine are 24.7 and 4.8 μM respectively. At substrate concentrations lower than 10 μM the rates of hypoxanthine and xanthine utilization are similar whereas at higher concentrations xanthine is hydroxylated faster. 3. 3. The time course of hypoxanthine → xanthine → uric acid hydroxylation shows a strong dependence of the second hydroxylation rate on the initial concentration of hypoxanthine. 4. 4. The enzyme is slowly inhibited by NADH at high, non-physiological concentrations. 5. 5. The enzyme resembles in some respects xanthine oxidoreductase from rat liver, and in other respects the enzyme from snake liver.

Xanthine:NAD + oxidoreductase in the liver of grass snake Natrix natrix

1. 1. The enzyme hydroxylating oxypurines in the liver of grass snake ( Natrix natrix, Colubridae) was found to be a stable xanthine:NAD + oxidoreductase (EC 1.2.1.37). 2. 2. The Michaelis constants for NAD + and xanthine amounted to 14.4 and 12.3 μM, respectively. The enzyme affinity to hypoxanthine is lower than that to xanthine, but the former substrate is hydroxylated faster than the latter. 3. 3. The enzyme is only slowly and slightly (up to 22%) inhibited by NADH accumulating during xanthine hydroxylation. 4. 4. The above data and the time-course of hypoxanthine → xanthine → uric acid hydroxylation indicated that the kinetic properties of the snake liver enzyme provide in this uricotelic animal fast elimination of superfluous nitrogen derived from protein catabolism.

The effect of thyroid hormones on the oxygen consumption of the liver tissue in the squamate reptiles

1. 1. The effects of temperature and/or the thyroid hormone on the oxygen consumption of lizard ( Hemidactylus) and snake ( Ptyas) liver tissue were examined. 2. 2. Significant increases in the oxygen consumption rate were observed when these squamates were maintained at or above 30°C. 3. 3. At 30°C, experimentally induced hypothyroid condition (thyroidectomy) and hyperthyroid condition (thyroxine injection) resulted in an increase or decrease respectively in the oxygen consumption in the lizard. Such treatment however produced no apparent effect in the snake. 4. 4. The reason for this discrepancy between the 2 sub-orders of the squamate reptiles remains to be sought.