PERSPECTIVE
Basolateral Potassium Channels and Epithelial Ion Transport

Calvin U. Cotton

Departments of Pediatrics, Physiology, and Biophysics, Case Western Reserve University, Cleveland, Ohio

The two-membrane hypothesis, proposed by Koefoed-Johnsen and Ussing (1) in 1958, formalized the concepts that the inner and outer membranes of the frog-skin epithelium are functionally different and that both membranes participate in active sodium absorption. The model described the frog skin as a homogenous intracellular compartment bounded by an apical and basolateral membrane with different ion transport properties. The apical membrane is sodium conductive (sodium channels), and the basolateral membrane contains Na⁺,K⁺-adenosine triphosphatase (ATPase) pumps and is potassium conductive (potassium channels) (Figure 1). Approximately 20 years later (2), a similar model was developed to explain Cl secretion by the dogfish rectal gland (Figure 2). Important modifications included (1) addition of a basolateral membrane Cl entry step, and (2) replacement of the apical membrane Na⁺ channel by a cyclic adenosine monophosphate (cAMP)-activated Cl channel. Again, the apical and basolateral cell membranes are functionally different, and both membranes participate in active Cl secretion. Several years earlier, Olver and colleagues (3) showed that canine airway epithelium absorbed Na⁺ and secreted Cl. A model that incorporates each of the transport elements found in the frog skin and shark rectal gland can explain both sodium absorption and chloride secretion by the airway epithelium (4).

View larger version (14K): [in this window] [in a new window]   Figure 1.   Schematic representation of the two-membrane hypothesis (1) for transepithelial sodium absorption by frog skin. The apical membrane contains sodium channels, and the basolateral membrane contains Na+,K+-ATPase pumps and potassium channels.

View larger version (17K): [in this window] [in a new window]   Figure 2.   Transport model for epithelial chloride secretion. The basic model proposed by Silva and colleagues (2) is modified to include a basolateral Na+,K+,2Cl cotransporter, rather than a NaCl cotransporter, as originally described.

A major impetus for the study of ion transport in nonrenal tissues is the desire to understand and treat secretory diarrheal diseases and cystic fibrosis (CF), disorders that result from abnormal salt and fluid secretion (5, 6). The discovery that CF is caused by the loss of an apical membrane, cAMP-activated Cl conductance (7, 8), coupled with the knowledge that cholera toxin activates adenylate cyclase and stimulates intestinal secretion, focused attention on apical membrane Cl channels in gastrointestinal and airway epithelia. The first epithelial ion channel to be identified and cloned was the CF transmembrane conductance regulator (CFTR) (9). Accordingly, we now know a great deal about the expression, regulation, and function of apical CFTR Cl channels and their role in Cl secretion (10). Several other Cl channels have been cloned during the past decade, but the importance of the so-called ClC protein family members in epithelial Cl secretion is uncertain (11). There are, however, other non-CFTR apical membrane Cl channels that do participate in epithelial Cl secretion, but the molecular identity and many of the details of regulation of these transport proteins are yet to be established (12, 13, 14). The sodium channel that is present in the apical membrane and that mediates amiloride-sensitive sodium absorption by frog skin, colon, airway, and numerous other epithelia was recently cloned and is known as ENaC (15). The molecular identity of the basolateral isoform of the Na⁺,K⁺,2Cl cotransporter, responsible for accumulation of intracellular Cl for secretion, is also known (16). The Na⁺,K⁺-ATPase, located at the basolateral membrane of nearly all epithelial cells, is crucial for generating and maintaining appropriate intracellular activities of sodium and potassium. The molecular identity of Na⁺,K⁺-ATPase, as well as detailed knowledge of the regulation and mechanism of transport by the pump, are well established (17). The transport models that describe sodium absorption and Cl secretion (Figures 1 and 2, respectively) both include basolateral potassium channels; however, the molecular identity of the proteins (channels) responsible for basolateral potassium conductance in epithelial cells remains largely unknown.

The basolateral K⁺ conductance pathway is critical in both secretory and absorptive epithelia, and inhibition of basolateral K⁺ conductance with nonselective K⁺ channel blockers (e.g., barium) will reduce transepithelial transport. Basolateral K⁺ channels recycle the potassium that enters the cell in exchange for sodium via the Na⁺,K⁺-ATPase (Figures 1 and 2) and also the potassium that enters the cell across the basolateral membrane coupled to Na⁺ and Cl (Figure 2) in Cl-secreting epithelia. Not only do K⁺ channels recycle potassium, but they contribute to the membrane potential of the cell, an important determinant of the electrochemical driving force for passive ion movement across the plasma membrane. In the two transport models depicted above, Na⁺ influx and Cl efflux are both driven in part by the electrical potential difference that exists across the apical membrane. Although it is not necessarily obvious, the apical and basolateral membranes in epithelia are electrically coupled by an intraepithelial current loop that arises due to the existence of a finite ionic conductance across the paracellular pathway (18). In other words, depolarization or hyperpolarization of the basolateral cell membrane will influence the opposite membrane and vice versa. It has long been argued that sustained Cl secretion requires activation of basolateral K⁺ conductance to facilitate K⁺ recycling and to hyperpolarize the apical cell membrane away from the chloride equilibrium potential (E^Cl). Intracellular microelectrode measurements of canine and human airway epithelia demonstrated that the initial depolarization of the apical membrane potential upon stimulation with beta agonists was followed by partial repolarization (8, 19, 20). The accompanying changes in membrane resistance and apical/basolateral resistance ratio suggested that the secondary repolarization resulted from activation of basolateral K⁺ conductance. The properties of basolateral K⁺ channels expressed in airway epithelial cells have received little attention, and the genes that encode these proteins are not known.

Since the first K⁺ channel was cloned a little more than a decade ago from Drosophila (21), molecular techniques have provided us with a plethora of potassium ion channel genes (22). Genomic sequence data will undoubtedly reveal additional K⁺ channel genes. Four broad classes of mammalian K⁺ channels have been identified: (1) voltage-gated K⁺ channels, (2) calcium-activated K⁺ channels, (3) "leak" K⁺ channels, and (4) inward rectifier K⁺ channels (22). Several of the cloned potassium channels from these families have properties similar to native epithelial K⁺ channels. For example, the Kir1.1 (ROMK1) inward rectifying K⁺ channel shares many of the features of the native small conductance secretory K⁺ channel, located in the apical membrane of cortical collecting-duct principal cells and the thick ascending limb of Henle cells (23). An ATP-sensitive K⁺ channel found in the basolateral membrane of mammalian proximal tubule cells is thought to respond to changes in intracellular ATP and thereby coordinate apical solute entry, Na⁺ pump activity, and basolateral K⁺ conductance (24, 25). The molecular identity of this channel is not yet established; however, several members of the Kir family of channels acquire ATP sensitivity when coexpressed with the sulfonylurea receptor (22). A calcium-activated K⁺ channel found on the basolateral membrane of the T84 colonic cell line is activated by 1-ethyl-2-benzimidazolinone (1-EIBO) and inhibited by clotrimazole (26). A channel with identical properties is encoded by the SK4 (also known as hIK1) gene, and mRNA for SK4 is expressed in T84 cells (27, 28). Thus, it appears that SK4 potassium channels comprise at least a portion of the basolateral K⁺ conductance in colonic epithelial cells. Greger and coworkers described a K⁺ channel in the basolateral membrane of colonic crypts that resembles the SK4 gene product (29). In addition, they found a cAMP-regulated small-conductance K⁺ channel that was too small to be resolved by single channel recording (probably < 3 pS) (30). The conductance was inhibited by chromanol 293B and is likely due to voltage-gated, K_vLQT1 channels (22).

In this issue, Mall and associates (31) show that cAMP-dependent stimulation of human airway epithelial Cl secretion causes parallel activation of basolateral K⁺ conductance. The secretory response, as well as the conductance increase, are blocked by chromanol 293B, a compound known to inhibit K_vLQT1 K⁺ channels (32). Furthermore, they demonstrate by RT-PCR that mRNA for K_vLQT1 is expressed in airway epithelium. Although K_vLQT1 mRNA was detected in CF nasal polyp cells, chromanol 293B did not inhibit transport in the CF epithelium. These results suggest that the depolarization of the apical and basolateral cell membranes upon activation of CFTR is necessary to stimulate basolateral, chromanol 293B-sensitive K⁺ channels. The authors conclude that K_vLQT1 potassium channels are important for maintaining cAMP-dependent secretion in the human airway and that pharmacologic activators of these channels may be useful for the treatment of CF patients. However, activation of basolateral potassium conductance in native CF airway epithelium may be counterproductive. Since CF airway epithelia exhibit excessive sodium absorption, activation of basolateral K⁺ channels would tend to increase sodium absorption and may further deplete airway-surface liquid volume (33, 34). Therefore, it may be necessary to fully inhibit sodium absorption prior to activation of basolateral K⁺ conductance in the CF airway. In contrast, CF epithelia, in which loss of cAMP-stimulated anion secretion is the dominant phenotype (e.g., pancreas, biliary epithelium, intestine), may prove to be better targets for activation of basolateral K⁺ conductance as a mechanism promoting anion secretion. Residual activity of mutant CFTR and/or stimulation of Cl secretion via non-CFTR apical membrane Cl channels might be enhanced in this way.

It has recently been demonstrated that isoflavonoids and flavonoids activate normal and mutant CFTR, and a phase I clinical trial of genistein is currently underway (35, 36). One potential problem associated with the use of isoflavonoids or flavonoids to stimulate CFTR-dependent anion secretion is that genistein inhibits basolateral K⁺ channels in a colonic epithelial cell line, HT29 (37). We have recently found that genistein inhibits cAMP- and calcium-stimulated anion secretion in immortalized, murine, pancreatic-duct epithelial cells (38). The inhibition of anion secretion appears to result from an effect of genistein on several different potassium channels present in the basolateral membrane of these cells. These observations, along with the findings of Mall and colleagues (31), highlight the necessity of considering ion transport pathways in the context of a polarized epithelial cell rather than in isolation.

	Footnotes

Address correspondence to: Calvin U. Cotton, Ph.D., Case Western University, Dept. of Pediatrics, 825 Biomedical Research Bldg., 2109 Adelbert Rd., Cleveland, OH 44106. E-mail: cuc{at}po.cwru.edu

(Received in original form July 10, 2000).

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