A short-term in vitro gill culture system to study the effects of toxic (copper) and non-toxic (cortisol) stressors on the rainbow trout, Oncorhynchus mykiss (Walbaum).
A short-term (24 h) method of gill filament culture system was developed to predict the effects of environmental contamination and stress in fish. Gill culture system containing two or three rainbow trout gill filaments in sterile glutamine supplemented Leibovitz 15 (L-15) media was submitted for 24 h to six different treatments: (i) CONT (control, medium only); (ii) CORT (cortisol, 0.28 microM cortisol); (iii) BLOCK (glucocorticoid receptor blocker, 14 microM RU 486); (iv) CORT+BLOCK (cortisol and blocker, 0.28 microM cortisol+14 microM RU 486); (v) CORT+CU (cortisol and copper, 100 microM CuSO4+0.28 microM cortisol); (vi) CU (copper, 100 microM CuSO4). After 24 h, the overall gill structure and cellular components resembled those of salmonids in vivo. Lactate dehydrogenase (LDH) activity in the culture media increased in the CORT+CU and CU groups but was significantly lower in the CORT+CU compared to CU group. Apoptotic cells increased in the CORT and CORT+BLOCK. The numbers of glucocorticoid (GR) receptor-positive cells were lower in the CU group. This short-term culture system seems to be suitable for studying the effects of both external and internal stress effectors (toxicants and hormones respectively), as it contains all cell types found in the gills and the cells give similar biological response as in vivo.
Mazon
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Department of Physiological Sciences, Federal University of São Carlos, Rod. Washington Luiz, km 235, P.O. Box 676, 13565-905 São Carlos, SP, Brazil.
Nolan
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DT
Lock
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Fernandes
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MN
Wendelaar Bonga
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D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003300:Copper / Q000633:toxicity*; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D004338:Drug Combinations; D005880:Gills / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D006854:Hydrocortisone / Q000494:pharmacology*; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D015735:Mifepristone / Q000494:pharmacology; D017686:Oncorhynchus mykiss; D009924:Organ Culture Techniques; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000378:metabolism
D017077:Culture Media, Conditioned; D004338:Drug Combinations; D018809:Proliferating Cell Nuclear Antigen; D011965:Receptors, Glucocorticoid; D015735:Mifepristone; D003300:Copper; D007770:L-Lactate Dehydrogenase; D006854:Hydrocortisone