On the natriuretic effect of verapamil: inhibition of ENaC and transepithelial sodium transport.

The natriuretic effect of Ca(2+) channel blockers has been attributed to hemodynamic changes and to poorly defined direct tubular effects. To test the possibility that verapamil may inhibit Na(+) reabsorption at the distal tubule, its effect on transepithelial Na(+) transport in aldosterone-stimulated A6 cells was determined. Cells were grown on permeable supports, and short-circuit current (I(sc)) measured in an Ussing chamber was used as a surrogate marker for transepithelial Na(+) transport. Application of 300 microM verapamil to the apical side inhibited I(sc) by 77% and was nearly as potent as 100 microM amiloride, which inhibited I(sc) by 87%. Verapamil-induced inhibition of I(sc) was accompanied by a significant increase in transepithelial resistance, suggesting blockade of an apical conductance. Its action on transepithelial Na(+) transport does not appear to occur through inhibition of L-type Ca(2+) channels, since I(sc) was unaffected by removal of extracellular Ca(2+). Verapamil also does not appear to inhibit I(sc) by modulating intracellular Ca(2+) stores, since it fails to inhibit transepithelial Na(+) transport when added to the basolateral side. The effect on Na(+) transport is specific for verapamil, since nifedipine, Ba(2+), 4-aminopyridine, and charybdotoxin do not significantly affect I(sc). A direct effect of verapamil on the epithelial Na(+) channel (ENaC) was tested using oocytes injected with the alpha-, beta-, and gamma-subunits. We conclude that verapamil inhibits transepithelial Na(+) transport in A6 cells by blocking ENaC and that the natriuresis observed with administration of verapamil may be due in part to its action on ENaC.

A mechanogated nonselective cation channel in proximal tubule that is ATP sensitive.

Ion channels that are gated in response to membrane deformation or "stretch" are empirically designated stretch-activated channels. Here we describe a stretch-activated nonselective cation channel in the basolateral membrane (BLM) of the proximal tubule (PT) that is nucleotide sensitive. Single channels were studied in cell-intact and cell-free patches from the BLM of PT cells that maintain their epithelial polarity. The limiting inward Cs+ conductance is ~28 pS, and channel activity persists after excision into a Ca2+- and ATP-free bath. The stretch-dose response is sigmoidal, with half-maximal activation of about -19 mmHg at -40 mV, and the channel is activated by depolarization. The inward conductance sequence is: NH ~ Cs+ ~ Rb+ > K+ ~ Na+ ~ Li+ > Ca2+ ~ Ba2+ > N-methyl-D-glucamine ~ tetraethylammonium. The venom of the common Chilean tarantula, Grammostola spatulata, completely blocks channel activity in cell-attached patches. Hypotonic swelling reversibly activates the channel. Intracellular ATP concentration ([ATP]i) reversibly blocks the channel (inhibitory constant approximately 0.48 mM), suggesting that channel function is coupled to the metabolic state of the cell. We conclude that this channel may function as a Ca2+ entry pathway and/or be involved in regulation of cell volume. We speculate this channel may be important when [ATP]i is depleted, as occurs during periods of increased transepithelial transport or with ischemic injury.

Regulation of the voltage-gated K+ channel KCNA10 by KCNA4B, a novel beta-subunit.

Voltage-gated K+ (Kv) channels are heteromultimeric complexes consisting of pore-forming alpha-subunits and accessory beta-subunits. Several beta-subunits have been identified and shown to interact with specific alpha-subunits to modify their levels of expression or some of their kinetic properties. The aim of the present study was to isolate accessory proteins for KCNA10, a novel Kv channel alpha-subunit functionally related to Kv and cyclic nucleotide-gated cation channels. Because one distinguishing feature of KCNA10 is a putative cyclic nucleotide-binding domain located at the COOH terminus, the entire COOH-terminal region was used to probe a human cardiac cDNA library using the yeast two-hybrid system. Interacting clones were then rescreened in a functional assay by coinjection with KCNA10 in Xenopus oocytes. One of these clones (KCNA4B), when injected alone in oocytes, produced no detectable current. However, when coinjected with KCNA10, it increased KCNA10 current expression by nearly threefold. In addition, the current became more sensitive to activation by cAMP. KCNA4B can be coimmunoprecipitated with the COOH terminus of KCNA10 and full-length KCNA10. It encodes a soluble protein (141 aa) with no amino acid homology to known beta-subunits but with limited structural similarity to the NAD(P)H-dependent oxidoreductase superfamily. KCNA4B is located on chromosome 13 and spans approximately16 kb, and its coding region is made up of five exons. In conclusion, KCNA4B represents the first member of a new class of accessory proteins that modify the properties of Kv channels.

Hyperkalemia: An adaptive response in chronic renal insufficiency.

BACKGROUND: Hyperkalemia is a common feature of chronic renal insufficiency, usually ascribed to impaired K+ homeostasis. However, various experimental observations suggest that the increase in extracellular [K+] actually functions in a homeostatic fashion, directly stimulating renal K+ excretion through an effect that is independent of, and additive to, aldosterone. METHODS: We have reviewed relevant studies in experimental animals and in human subjects that have examined the regulation of K+ excretion and its relation to plasma [K+]. RESULTS: Studies indicate that (1) extracellular [K+] in patients with renal insufficiency correlates directly with intracellular K+ content, and (2) hyperkalemia directly promotes K+ secretion in the principal cells of the collecting duct by increasing apical and basolateral membrane conductances. The effect of hyperkalemia differs from that of aldosterone in that K+ conductances are increased as the primary event. The changes in principal cell function and structure induced by hyperkalemia are indistinguishable from the effects seen in adaptation to a high K+ diet. CONCLUSIONS: We propose that hyperkalemia plays a pivotal role in K+ homeostasis in renal insufficiency by stimulating K+ excretion. In patients with chronic renal insufficiency, a new steady state develops in which extracellular [K+] rises to the level needed to stimulate K+ excretion so that it again matches intake. When this new steady state is achieved, plasma [K+] remains stable unless dietary intake increases, glomerular filtration rate falls, or drugs are given that disrupt the new balance.