Intra- and Interlaboratory Reliability of a Cryopreserved Trout Hepatocyte Assay for the Prediction of Chemical Bioaccumulation Potential

2014 ◽  
Vol 48 (14) ◽  
pp. 8170-8178 ◽  
Author(s):  
Kellie A. Fay ◽  
Robert T. Mingoia ◽  
Ina Goeritz ◽  
Diane L. Nabb ◽  
Alex D. Hoffman ◽  
...  
Keyword(s):  
Trout Hepatocyte

Evidence for the role of a Na(+)/HCO(3)(−) cotransporter in trout hepatocyte pHi regulation

2000 ◽  
Vol 203 (14) ◽  
pp. 2201-2208 ◽  
Author(s):  
M. Furimsky ◽  
T.W. Moon ◽  
S.F. Perry
Keyword(s):  
Membrane Potential ◽  
Anion Transport ◽  
Fluorescent Dye ◽  
Proton Extrusion ◽  
Pharmacological Agents ◽  
Rainbow Trout Oncorhynchus Mykiss ◽  
Trout Hepatocyte ◽  
Acid Loading ◽  
Na Uptake ◽  
Phi Regulation

The mechanisms of intracellular pH (pHi) regulation were examined in hepatocytes of the rainbow trout Oncorhynchus mykiss. pHi was monitored using the pH-sensitive fluorescent dye BCECF, and the effects of various media and pharmacological agents were examined for their influence on baseline pHi and recovery rates from acid and base loading. Rates of Na(+) uptake were measured using (22)Na, and changes in membrane potential were examined using the potentiometric fluorescent dye Oxonol VI. The rate of proton extrusion following acid loading was diminished by the blockade of either Na(+)/H(+) exchange (using amiloride) or anion transport (using DIDS). The removal of external HCO(3)(−) and the abolition of outward K(+) diffusion by the channel blocker Ba(2+) also decreased the rate of proton extrusion following acid load. Depolarization of the cell membrane with 50 mmol l(−)(1) K(+), however, did not affect pHi. The rate of recovery from base loading was significantly diminished by the blockade of anion transport, removal of external HCO(3)(−) and, to a lesser extent, by blocking Na(+)/H(+) exchange. The blockade of K(+) conductance had no effect. The decrease in Na(+) uptake rate observed in the presence of the anion transport blocker DIDS and the DIDS-sensitive hyperpolarization of membrane potential during recovery from acid loading suggest that a Na(+)-dependent electrogenic transport system is involved in the restoration of pHi after intracellular acidification. The effects on baseline pHi indicate that the different membrane exchangers are tonically active in the maintenance of steady-state pHi. This study confirms the roles of a Na(+)/H(+) exchanger and a Cl(−)/HCO(3)(−) exchanger in the regulation of trout hepatocyte pHi and provides new evidence that a Na(+)/HCO(3)(−) cotransporter contributes to pHi regulation.

Rainbow trout hepatocyte beta-adrenoceptors, catecholamine responsiveness, and effects of cortisol

1992 ◽  
Vol 262 (5) ◽  
pp. R794-R799 ◽  
Author(s):  
S. D. Reid ◽  
T. W. Moon ◽  
S. F. Perry
Keyword(s):  
Plasma Cortisol ◽  
Acute Stress ◽  
Glucose Production ◽  
Beta Adrenoceptors ◽  
Surface Receptors ◽  
Trout Hepatocyte ◽  
Rainbow Trout Hepatocyte ◽  
Beta 2 ◽  
Total Glucose

Hepatocyte beta-adrenoceptors were characterized in trout with chronically raised plasma cortisol levels (132.1 +/- 14.8 ng/ml, n = 8, 11-12 days) and compared with shams (1.8 +/- 0.9 ng/ml, n = 8) using radioreceptor assay techniques. Hepatocytes prepared from sham trout possessed 1,696 +/- 179 beta-adrenoceptors/cell. These receptors were characterized as beta 2-adrenoceptors based on the potency order of specific inhibition of CGP binding. The number of putative surface beta 2-adrenoceptors significantly increased to 6,005 +/- 1,165 receptors/cell, in hepatocytes from trout exposed to the elevated plasma cortisol concentration. The physiological significance of the increase in hepatocyte surface receptors was assessed by the in vitro responsiveness of hepatocytes to a range of epinephrine concentrations (10-1,000 nmol/l). Both total glucose production and intracellular adenosine 3',5'-cyclic monophosphate content, indicators of epinephrine responsiveness, were enhanced as a result of chronically raised levels of plasma cortisol. We suggest that these cortisol-mediated alterations in the adrenergic responsiveness of trout hepatocytes may ultimately enhance the ability of the liver to supply glucose to the fish during acute stress after extended periods of chronic stress.

Screening for oestrogenic activity of plant and food extracts usingin vitro trout hepatocyte cultures

2004 ◽  
Vol 15 (1) ◽  
pp. 40-45 ◽  
Author(s):  
C. Bennetau-Pelissero ◽  
K. Gontier Latonnelle ◽  
V. Lamothe ◽  
S. Shinkaruk-Poix ◽  
S. J. Kaushik
Keyword(s):  
Hepatocyte Cultures ◽  
Trout Hepatocyte

Comparison of trout hepatocyte culture on different substrates

1986 ◽  
Vol 22 (6) ◽  
pp. 360-362 ◽  
Author(s):  
Michael M. Lipsky ◽  
Talia R. Sheridan ◽  
Richard O. Bennett ◽  
Eric B. May
Keyword(s):  
Hepatocyte Culture ◽  
Trout Hepatocyte

Influence of xenobiotics on rainbow trout liver estrogen receptor and vitellogenin gene expression

1995 ◽  
Vol 15 (2) ◽  
pp. 143-151 ◽  
Author(s):  
G Flouriot ◽  
F Pakdel ◽  
B Ducouret ◽  
Y Valotaire
Keyword(s):  
Gene Expression ◽  
Estrogen Receptor ◽  
Rainbow Trout ◽  
Expression Vector ◽  
Mrna Accumulation ◽  
Trout Hepatocyte ◽  
Rainbow Trout Hepatocyte ◽  
Vitellogenin Gene ◽  
Rainbow Trout Liver ◽  
Hepatocyte Primary Culture

ABSTRACT Rainbow trout hepatocyte primary culture was used to test the influence of some xenobiotics on the expression of two genes implicated in reproduction, those for the estrogen receptor (ER) and vitellogenin (Vg). We showed that chlordecone, nonylphenol, a polychlorobiphenol (PCB) mixture (Aroclor 1245) and lindane were able to induce ER and Vg mRNA accumulation. Antiestrogens, 4-hydroxytamoxifen and ICI 164,384, prevented the effects of the xenobiotics, indicating that the induction of gene expression is mediated by the ER. Among these four xenobiotics, only chlordecone and nonylphenol were able to displace the binding of [3H]estradiol to ER-enriched COS-1 extracts, and to activate an estrogen-dependent reporter gene (ERE-TK-CAT) cotransfected with an expression vector containing ER cDNA. The results suggest that chlordecone and nonylphenol are direct inducers of rainbow trout ER and Vg gene expression, whereas PCBs and lindane act through their hepatic metabolites. Moreover, pentachlorophenol acts as an antagonist of the induction by estradiol of rainbow trout ER and Vg gene expression.

Trout hepatocyte culture: Isolation and primary culture

In Vitro ◽  
1985 ◽  
Vol 21 (4) ◽  
pp. 221-228 ◽  
Author(s):  
James E. Klaunig ◽  
Randall J. Ruch ◽  
Peter J. Goldblatt
Keyword(s):  
Primary Culture ◽  
Hepatocyte Culture ◽  
Trout Hepatocyte

Two complementary bioassays for screening the estrogenic potency of xenobiotics: recombinant yeast for trout estrogen receptor and trout hepatocyte cultures

1997 ◽  
Vol 19 (3) ◽  
pp. 321-335 ◽  
Author(s):  
F Petit ◽  
P Le Goff ◽  
JP Cravedi ◽  
Y Valotaire ◽  
F Pakdel
Keyword(s):  
Estrogen Receptor ◽  
Polychlorinated Biphenyl ◽  
Estrogenic Activity ◽  
Wide Spectrum ◽  
Direct Interaction ◽  
Recombinant Yeast ◽  
Estrogenic Compounds ◽  
Hepatocyte Cultures ◽  
Trout Hepatocyte

A relation between the chemical structure of a xenobiotic and its steroidal action has not yet been clearly established. Thus, it is not possible to define the estrogenic potency of different xenobiotics. An assessment may be accomplished by the use of different bioassays. We have previously developed a yeast system highly and stably expressing rainbow trout estrogen receptor (rtER) in order to analyze the biological activity of the receptor. The recombinant yeast system appears to be a reliable, rapid and sensitive bioassay for the screening and determination of the direct interaction between ER and estrogenic compounds. This system was used in parallel with a more elaborate biological system, trout hepatocyte aggregate cultures, to examine the estrogenic potency of a wide spectrum of chemicals commonly found in the environment. In hepatocyte cultures, the vitellogenin gene whose expression is principally dependent upon estradiol was used as a biomarker. Moreover, competitive binding assays were performed to determine direct interaction between rtER and xenobiotics. In our study, 50% of the 49 chemical compounds tested exhibited estrogenic activity in the two bioassays: the herbicide diclofop-methyl; the fungicides biphenyl, dodemorph, and triadimefon; the insecticides lindane, methyl parathion, chlordecone, dieldrin, and endosulfan; polychlorinated biphenyl mixtures; the plasticizers or detergents alkylphenols and phthalates; and phytoestrogens. To investigate further biphenyl estrogenic activity, its principal metabolites were also tested in both bioassays. Among these estrogenic compounds, 70% were able to activate rtER in yeast and hepatocytes with variable induction levels according to the system. Nevertheless, 30% of these estrogenic compounds exhibited estrogenic activity in only one of the bioassays, suggesting the implication of metabolites or different pathways in the activation of gene transcription. This paper shows that it is important to combine in vivo bioassays with in vitro approaches to elucidate the mechanism of xenoestrogen actions.

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