Nonselective cation transport in native esophageal epithelia

Abstract

Rabbit esophageal epithelia actively transport Na+ in a manner similar to that observed in classic electrically tight Na+-absorbing epithelia, such as frog skin. However, the nature of the apical entry step is poorly understood. To address this issue, we examined the electrophysiological and biochemical nature of this channel. Western blotting experiments with epithelial Na+ channel (ENaC) subunit-specific antibodies revealed the presence of all three ENaC subunits in both native and immortalized esophageal epithelial cells. The amino acid sequence of the rabbit α-ENaC cloned from native rabbit esophageal epithelia was not significantly different from that of other published α-ENaC homologs. To characterize the electrophysiological properties of this native apical channel, we utilized nystatin permeabilization to eliminate the electrical contribution of the basolateral membrane in isolated native epithelia mounted in Ussing-type chambers. We find that the previously described apical Na+ channel is nonselective for monovalent cations (Li+, Na+, and K+). Moreover, this channel was not blocked by millimolar concentrations of amiloride. These findings document the presence of a nonselective cation channel in a native Na+ transporting epithelia, a finding that hereto has been thought to be limited to artificial culture conditions. Moreover, our data are consistent with a potential role of ENaC subunits in the formation of a native nonselective cation channel.

The Pharmacological Profile of Brain Liver Intestine Na+ Channel: Inhibition by Diarylamidines and Activation by Fenamates

“…Those currents include a Na ϩ -selective channel from rabbit proximal tubules (Gögelein and Greger, 1986;Jacobsen et al, 1988) that is sensitive to Ca 2ϩ (Blokkebak-Poulsen et al, 1991) and a Na ϩ channel in LLC-PK1 cells (Raychowdhury et al, 2004), a cell line with proximal tubule characteristics derived from pig kidney. Furthermore, several studies describe an amilorideinsensitive and thus ENaC-independent pathway for Na ϩ entry in epithelial cells of various organs, such as lung (Shlyonsky et al, 2005), colon (Inagaki et al, 2004), and esophagus (Awayda et al, 2004). Because its affinity for amiloride is low, rBLINaC might be a candidate responsible for these currents.…”

Differentiation of Epithelial Na+ Channel Function

“…Sodium entry through amiloride-insensitive pathways has also been described in surface cells of rectal colon (35), esophageal epithelia (36), and keratinocytes (37). In patch clamp studies of rectal colon cells in situ, distinct populations of cells with either amiloride-sensitive or amiloride-insensitive sodium currents were described (35).…”

“…Results from the literature as well as from the present study indicate that amiloride-insensitive sodium absorption is the Na ϩ transport pathway found in cells that have not achieved complete maturation. The proteins involved in amiloride-insensitive Na ϩ transport include NSC channels, CNG channels, and possibly immature or "misassembled" ENaC (36). It was recently shown that immature ENaCs have very low activity at the cell membrane (39), whereas cells with amiloride-insensitive sodium currents express ␣, ␤, and ␥ ENaC subunits (36).…”

“…The proteins involved in amiloride-insensitive Na ϩ transport include NSC channels, CNG channels, and possibly immature or "misassembled" ENaC (36). It was recently shown that immature ENaCs have very low activity at the cell membrane (39), whereas cells with amiloride-insensitive sodium currents express ␣, ␤, and ␥ ENaC subunits (36). Whether ENaC subunits can misassemble or bind with other proteins to form conductive channels, has never been shown.…”

Effect of luminal acidity on the apical cation channel in rabbit esophageal epithelium

“…More recently, Awayda and colleagues (2, 3) used antibodies to specific EnaC subunits to show by Western blotting that the Na ϩ channel in native rabbit esophageal stratified squamous epithelium (ESSE) possessed the same three subunits and that the amino acid sequence for ␣-ENaC in ESSE was the same as that of the Na ϩ channel cloned from kidney. Despite structural similarities, the Na ϩ channel in ESSE, like that in sheep rumen, differs functionally from the classic Na ϩ channel in that it is neither Na ϩ selective nor amiloride sensitive (2,16). With respect to selectivity, the Na ϩ channel in ESSE was observed to support a similar short-circuit current (I sc ) when Na ϩ was substituted with either K ϩ or Li ϩ .…”