Integrated responses of Na+/HCO3− cotransporters and V-type H+-ATPases in the fish gill and kidney during respiratory acidosis

Abstract

Using degenerate primers, followed by 3' and 5' RACE and "long" PCR, a continuous 4050-bp cDNA was obtained and sequenced from rainbow trout (Oncorhynchus mykiss) gill. The cDNA included an open reading frame encoding a deduced protein of 1088 amino acids. A BLAST search of the GenBank protein database demonstrated that the trout gene shared high sequence similarity with several vertebrate Na(+)/HCO(3)(-) cotransporters (NBCs) and in particular, NBC1. Protein alignment revealed that the trout NBC is >80% identical to vertebrate NBC1s and phylogenetic analysis provided additional evidence that the trout NBC is indeed a homolog of NBC1. Using the same degenerate primers, a partial cDNA (404 bp) for NBC was obtained from eel (Anguilla rostrata) kidney. Analysis of the tissue distribution of trout NBC, as determined by Northern blot analysis and real-time PCR, indicated high transcript levels in several absorptive/secretory epithelia including gill, kidney and intestine and significant levels in liver. NBC mRNA was undetectable in eel gill by real-time PCR. In trout, the levels of gill NBC1 mRNA were increased markedly during respiratory acidosis induced by exposure to hypercarbia; this response was accompanied by a transient increase in branchial V-type H(+)-ATPase mRNA levels. Assuming that the branchial NBC1 is localised to basolateral membranes of gill cells and operates in the influx mode (HCO(3)(-) and Na(+) entry into the cell), it would appear that in trout, the expression of branchial NBC1 is transcriptionally regulated to match the requirements of gill pHi regulation rather than to match trans-epithelial HCO(3)(-) efflux requirements for systemic acid-base balance. By analogy with mammalian systems, NBC1 in the kidney probably plays a role in the tubular reabsorption of both Na(+) and HCO(3)(-). During periods of respiratory acidosis, levels of renal NBC1 mRNA increased (after a transient reduction) in both trout and eel, presumably to increase HCO(3)(-) reabsorption. This strategy, when coupled with increased urinary acidification associated with increased vacuolar H(+)-ATPase activity, ensures that HCO(3)(-) levels accumulate in the body fluids to restore pH.

A nose-to-nose comparison of the physiological and molecular responses of rainbow trout to high environmental ammonia in seawater versus freshwater

“…These include the higher expression of two basolateral Na + transporters, the 'freshwater' isoform NKA-1a of Na Fig.9F) (Richards et al, 2003;Shrimpton et al, 2005;Bystriansky et al, 2006) as well as NBC1 in FW (Fig.9C). The latter appears to play a major role in Na + uptake in FW fish (Perry et al, 2003;Parks et al, 2007). Surprisingly, baseline expression of the 'seawater' isoform NKA-1b of Na + ,K + -ATPase (Richards et al, 2003;Bystriansky et al, 2006) did not differ between the two salinities ( Fig.9G), but there are two other reports where this was also the case (Shrimpton et al, 2005;Kiilerich et al, 2007).…”

Interaction of osmoregulatory and acid–base compensation in white sturgeon (Acipenser transmontanus) during exposure to aquatic hypercarbia and elevated salinity

“…the first bout of pH e recovery) can be attributed to the HCSW sturgeon having accumulated relatively more Na + than Cl − during this initial salt-loading period of the SW acclimation. The apparent association between pH e recovery and a relative increase of Na + accumulation during this time may be the result of inward Na + transport being tied to outward H + transport via an apical NHE or an apical H + -ATPase with associated Na + channel (Claiborne and Heisler, 1984;Iwama and Heisler, 1991;Dymowska et al, 2014), or the co-transport of Na + and HCO 3 − into the extracellular compartment via a basolateral Na + /HCO 3 − co-transporter (NBC1) (Hirata et al, 2003;Perry et al, 2003).…”

A new paradigm for ammonia excretion in aquatic animals: role of Rhesus(Rh) glycoproteins

“…This may be sufficient because approximately 10 times more CO 2 is excreted across the gills compared with ammonia . There is now an emerging role for the Na + /HCO 3 -co-transporter, NBC-1, as a key contributor to basolateral Na + transport, in addition to the traditional role of Na + /K + -ATPase in this regard (Hirata et al, 2003;Perry et al, 2003;Scott et al, 2005;Parks et al, 2007). NBC-1 would probably act in the electrogenic '3 HCO 3 -+1 Na + ' efflux mode in this circumstance, moving Na + and HCO 3 -from the gill cells to the blood plasma, and must also be fuelled by cytoplasmic CA-2.…”