title: Study on apoptotic effects of neurotoxin anatoxin-a on fish immune cells.
abstract: OBJECTIVE
  The aim of this study was to assess the possible in vitro apoptotic effects of anatoxin-a on the selected immune cells isolated from the blood of carp.
  
  
  METHODS
  In the experiments pure anatoxin-a was used at concentrations of 0.01, 0.05, 0.1 and 1 µg/ml RPMI-1640 medium. Apoptosis or necrosis of fish leukocytes (lymphocytes and phagocytes) induced by the toxin was determined by measurement of the activity of caspases-3/7 and the analysis of phosphatidylserine on the outer leaflet of apoptotic cell-membranes using Annexin-V-Fluorescein and Propidium Iodide. Moreover, fluorescent measurement of the release of lactate dehydrogenase from the cells with damaged membranes was done.
  
  
  RESULTS
  The viability of the lymphocytes exposed only to the highest concentration of anatoxin-a (1 µg/ml) was significantly decreased. The exposure to the toxin at higher concentrations (0.1 and 1 µg/ml) resulted in a significant increase of caspases 3/7 activity in phagocytes and lymphocytes. Moreover, fluorescent analysis with the use of annexin-V-fluorescein and propidium iodide staining showed more cells at the apoptotic stage than necrotic cells.
  
  
  CONCLUSIONS
  Our study showed that anatoxin-a is an inducer of apoptosis in fish immune cells.
authors.affiliation: Department of Physiology and Ecotoxicology, The John Paul II Catholic University of Lublin, Poland. anrym@kul.lublin.pl
authors.forename: Anna
authors.initials: A
authors.lastname: Rymuszka
authors.forename: Anna
authors.initials: A
authors.lastname: Sieroslawska
mesh_terms: D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D002347:Carps / Q000378:metabolism*; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D000087522:Cyanobacteria Toxins; D004305:Dose-Response Relationship, Drug; D007107:Immune System / Q000166:cytology*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008214:Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D023421:Models, Animal; D009498:Neurotoxins / Q000494:pharmacology*; D010586:Phagocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D014326:Tropanes / Q000494:pharmacology*
chemical_list: D017304:Annexin A5; D000087522:Cyanobacteria Toxins; D009498:Neurotoxins; D014326:Tropanes; C509783:anatoxin a; D007770:L-Lactate Dehydrogenase; D020169:Caspases