Characterization of ion transport in the isolated epipodite of the lobsterHomarus americanus

Abstract

SUMMARYUnfolded epipodite isolated from American lobsters (Homarus americanus) acclimated to dilute seawater was mounted in an Ussing-type chamber for ion transport studies. The split epipodite is an electrically polarized, one-cell-layer epithelium supported with cuticle. Under open-circuit conditions, the transepithelial potential was –4.2±1.0 mV (N=38). In the short-circuited epithelium, the current averaged over all of the preparations was –185.4±20.2 A cm–2 (N=38) with a high conductance of 55.2±11.4 mS cm–2 (N=38), typical for a leaky epithelium. The Na:Cl absorptive flux ratio was 1:1.6; ion substitution experiments indicated that the transport of Na+ and Cl– is coupled. Basolateral application of the Cl– channel blockers 5-nitro-2-(3-phenylpropylamino) benzoate (NPPB) and niflumic acid (NFA) dose-dependently inhibited short-circuit current (ISC). Secretory K+ (Rb+) fluxes exceeded influxes and were inhibited by the Na+/K+-ATPase inhibitor ouabain and the K+ channel blocker cesium. Western blot analysis showed that Na+/K+-ATPase α-subunit protein was more highly expressed in the epipodite of lobsters acclimated to 20 p.p.t. compared with animals acclimated to seawater (34 p.p.t.). 3-Isobutyl-1-methyl-xanthine (IBMX) stimulated a negative ISC and enhanced apical secretory K+ flux. Basolateral application of NPPB inhibited JRbB→A fluxes, suggesting the interaction of K+ channels with NPPB-sensitive Cl– channels. The results are summarized in a transport model, suggesting apical Na+/K+/2Cl– co-transport, a dominant apical K+-secreting channel and basolaterally located Cl– and K+ channels. This study represents the first comprehensive characterization of ion transport processes across the lobster epipodite epithelium and indeed in any tissue within the branchial cavity of the American lobster.