Proceedings of the Physiological Society, 12‐13 June 1992, St Andrews Meeting: Poster Communications

Na-K-ATPase expression was investigated in the euryhaline European eel (Anguilla anguilla) during adaptation from freshwater (FW) to seawater (SW) environments. A partial Na-K-ATPase a-subunit cDNA was cloned using the polymerase chain reaction with oligonucleotide primers designed to take advantage of regions of homology present in the amino acid sequences of all known Na-K-ATPase a-subunit sequences. These primers contained neutral inosine nucleotide analogue bases at positions of uncertainty in the sequence. The nucleotide sequence of the partial eel cDNA corresponded to positions 509-707 of the rat Na-K-ATPase a-1 isoform amino acid sequence (Shull et al. 1986). The eel Na-K-ATPase a-partial cDNA demonstrated 87 and 78 %amino acid and nucleotide sequence homology respectively to the corresponding region of the rat a-1 isoform and also 82 and 74 % amino acid and nucleotide sequence homology respectively compared to that of the ray (Torpedo californica) Na-K-ATPase a-subunit sequence (Kawakami et al. 1985).

The partial eel Na-K-ATPase a--cDNA was used to study the effect of acclimation on the expression of Na-K-ATPase a-subunit mRNA in the branchial epithelium. During initial experiments, long term FW-adapted eels were transferred to SW for periods of up to a week, and at various time intervals gill tissues were either processed for total RNA extraction or were homogenized for the determination of maximal Na-K-ATPase activity. Total RNA from the gill tissue samples was electrophoresed, Northern blotted and hybridized to either the partial eel Na-K-ATPase a--cDNA probe, or a control 18 S ribosomal RNA (rRNA) antisense oligonucleotide probe. The results indicate that relative to the 18 S rRNA probe, the level of Na-K-ATPase a­-subunit mRNA was increased between 2 and 4 days following transfer to SW. This increase in the level of Na-K-ATPase a-mRNA expression was concur­rent with corresponding increases in branchial Na-K-ATPase activity during the period up to 4 days after SW transfer. This demonstrates that increases in Na-K-ATPase a-subunit mRNA may, at least in part, account for the increases in Na-K-ATPase activity found during the acclimation of eels to SW.