Claudins in a primary cultured puffer fish (Tetraodon nigroviridis) gill epithelium model alter in response to acute seawater exposure

Abstract

Gill epithelium permeability and qualitative/quantitative aspects of gill claudin (cldn) tight junction (TJ) protein transcriptomics were examined with a primary cultured model gill epithelium developed using euryhaline puffer fish (Tetraodon nigroviridis) gills. The model was prepared using seawater-acclimated fish gills and was cultured on permeable cell culture filter supports. The model is composed of 1–2 confluent layers of gill pavement cells (PVCs), with the outer layer exhibiting prominent apical surface microridges and TJs between adjacent cells. During development of electrophysiological characteristics, the model exhibits a sigmoidal increase in transpithelial resistance (TER) and plateaus around 30kΩcm2. At this point paracellular movement of [3H]polyethylene glycol (PEG) 4000 was low at ~1.75cms−1×10−7. When exposed to apical seawater (SW) epithelia exhibit a marked decrease in TER while PEG flux remained unchanged for at least 6h. In association with this, transcript encoding cldn TJ proteins cldn3c, -23b, -27a, -27c, -32a and -33b increased during the first 6h while cldn11a decreased. This suggests that these proteins are involved in maintaining barrier properties between gill PVCs of SW fishes. Gill cldn mRNA abundance also altered 6 and 12h following abrupt SW exposure of puffer fish, but in a manner that differed qualitatively and quantitatively from the cultured model. This most likely reflects the cellular heterogeneity of whole tissue and/or the contribution of the endocrine system in intact fish. The current study provides insight into the physiological and transcriptomic response of euryhaline fish gill cells to a hyperosmotic environment.