Expression and water calcium dependence of calcium transporter isoforms in zebrafish gill mitochondrion-rich cells

Abstract

Background: Freshwater fish absorb Ca 2+ predominantly from ambient water, and more than 97% of Ca 2+ uptake is achieved by active transport through gill mitochondrion-rich (MR) cells. In the current model for Ca 2+ uptake in gill MR cells, Ca 2+ passively enters the cytosol via the epithelium Ca 2+ channel (ECaC), and then is extruded into the plasma through the basolateral Na + / Ca 2+ exchanger (NCX) and plasma membrane Ca 2+ -ATPase (PMCA). However, no convincing molecular or cellular evidence has been available to support the role of specific PMCA and/or NCX isoforms in this model. Zebrafish (Danio rerio) is a good model for analyzing isoforms of a gene because of the plentiful genomic databases and expression sequence tag (EST) data.

Involvement of calcitonin and its receptor in the control of calcium-regulating genes and calcium homeostasis in zebrafish (Danio rerio)

“…Recently in zebrafish ECaC has been cloned and characterized to be responsible for Ca 2þ uptake in the gill MR cells. (6) Our results lead us to conclude that there is a hypocalcemic role for CT, probably through the inhibition of ECaC synthesis. Furthermore, this study suggests that this hypocalcemic role of CT could be conserved throughout zebrafish development and still be present in adults.…”

“…In zebrafish, epithelial Ca 2þ channels (ECaC), NCX1b, and PMCA2 have been shown to be specifically expressed in a group of ionocytes (mitochondria-rich [MR] cells) and be involved in transepithelial Ca 2þ uptake. (3)(4)(5)(6)(7) Different hormones participate in the control and regulation of calcium homeostasis. In mammals, the three main hormones involved in the calcium regulation are parathyroid hormone (PTH), vitamin D, and calcitonin (CT).…”

Ion uptake and acid secretion in zebrafish (Danio rerio)

“…All these suggest that ECaC is the rate-limiting step and the gatekeeper channel for active Ca 2+ transport in fish as in mammals (Vennekens et al, 2000). Acclimation to low-Ca 2+ FW caused upregulation of zECaC but not zPMCA2 or zNCX1b, implying that the steady-state expression of zPMCA2 and zNCX1b may fulfill the requirement for the transepithelial transport machinery under all situations (Liao et al, 2007). This further supports a previous enzyme kinetic study, in which the basolateral Ca 2+ extrusion mechanisms by PMCA and NCX were found to be far below their maximum capacity in rainbow trout (Oncorhynchus mykiss) gills (Flik et al, 1997).…”

“…Taken together, it may be that only PNA + cells express ECaC and basolateral transporters (PMCA and NCX) and thus can achieve the entire transepithelial Ca + pathway, although no data for the existence PMCA and NCX in trout gill cells are available so far. Indeed, it was also found that not all ECaC-expressing cells coexpressed both zPMCA2 and zNCX1b in zebrafish gill NaR cell populations (Liao et al, 2007), which may be analogous to trout PNA + cells (see below). Fish gill cells that express only one or two (never all) of the three major Ca 2+ transporters may be in the process of terminal differentiation as Hsiao and coworkers (Hsiao et al, 2007) reported in zebrafish embryonic skin ionocytes.…”

“…Using similar approaches to those used in the studies on zebrafish NHE and CA (Yan et al, 2007;Lin et al, 2008), one isoform of ECaC, six isoforms of PMCAs (zPMCA1a, zPMCA1b, zPMCA2, zPMCA3a, zPMCA3b, and zPMCA4) and seven isoforms of NCX (zNCX1a, zNCX1b, zNCX2a, zNCX2b, zNCX3, zNCX4a and zNCX4b) were identified in zebrafish, and triple fluorescence ISH and ICC were conducted to demonstrate the colocalization of zECaC, zPMCA2 and zNCX1b mRNAs in a portion of skin/gill NaR cells (Pan et al, 2005;Liao et al, 2007). Acclimation to a low-Ca 2+ environment caused stimulation of zebrafish Ca 2+ -uptake function and a concomitant upregulation in skin/gill zECaC expression, but no effect on the expression of either zPMCAs or zNCXs (Pan et al, 2005;Liao et al, 2007), supporting the current Ca 2+ -uptake model, in which ECaC may act as a major regulatory target for this mechanism during environmental challenge. In another study by Craig and colleagues (Craig et al, 2007), both ECaC mRNA and ECaC protein levels in gills were higher in zebrafish acclimated to soft FW (0.051 mmol l -1 Ca 2+ ) than those in control FW (0.946 mmol l -1 Ca 2+ ).…”