Primary culture of gill epithelial cells from the sea bass Dicentrarchus labrax.

Abstract

We have developed the first explant technique that allows the in vitro study of gill physiology and biochemistry in marine species. Gill fragments were cultured at 17 degrees C, in atmospheric PCO2, with nutrient medium (Leibovitz L15), pH 7.8, supplemented with 10% fetal bovine serum and adjusted to the osmolarity of fish plasma (350 mOsm/liter). Coating plates with collagen, gelatin, or polylysin did not improve our results. Decrease in osmotic pressure, removal of bovine serum, or its replacement by fish serum inhibited growth from the explants. Approximately 50% of the explants produced cell growth, and after 4 days of culture a monolayer of contiguous cells was formed. This technique is rapid and does not require the use of enzymes. The cells appeared flat and thin with an epitheloid shape. They looked polygonal with a maximum length of 10 to 50 microm. Evidence that they are unique gill cells is the presence of polymorphic surface crenelations (microplicae), prominent Golgi apparatus, tight junctions and desmosomes. Comparison with in vivo tissue showed them to be epithelial cells having differentiated in a homogeneous population of respiratory-like (pavement) cells. They are polarized with their apical surface facing the culture medium. The development of this culture system represents a new tool for cellular approaches to determine precisely the functions and transport mechanism of gill cells.