Potassium secretion in rat distal colon during dietary potassium loading: role of pH regulated apical potassium channels

Abstract

Background—Chronic dietary K⁺ loading increases the abundance of large conductance (210 pS) apical K⁺ channels in surface cells of rat distal colon, resulting in enhanced K⁺ secretion in this epithelium. However, the factors involved in the regulation of these K⁺ channels are at present unclear. 
Aims—To evaluate the effect of dietary K⁺ loading on intracellular pH and its relation to large conductance apical K⁺ channel activity in surface cells of rat distal colon. 
Methods/Results—As assessed by fluorescent imaging, intracellular pH was higher in K⁺ loaded animals (7.48 (0.09)) than in controls (7.07 (0.04); p<0.01) when surface cells were bathed in NaCl solution, and a similar difference in intracellular pH was observed when cells were bathed in Na₂SO₄ solution (7.67 (0.09) and 6.92 (0.05) respectively; p<0.001). Ethylisopropylamiloride (EIPA; an inhibitor of Na⁺-H⁺ exchange; 1 µM) decreased intracellular pH when surface cells from K⁺ loaded animals were bathed in either solution, although the decrease was greater when the solution contained NaCl (ΔpH 0.50 (0.03)) rather than Na₂SO₄ (ΔpH 0.18 (0.02); p<0.05). In contrast, EIPA had no effect in cells from control animals. As assessed by patch clamp recording techniques, the activity of large conductance K⁺ channels in excised inside-out membrane patches from distal colonic surface cells of K⁺ loaded animals increased twofold when the bath pH was raised from 7.40 to 7.60. As assessed by cell attached patches in distal colonic surface cells from K⁺ loaded animals, the addition of 1 µM EIPA decreased K⁺ channel activity by 50%, consistent with reversal of Na⁺-H⁺ exchange mediated intracellular alkalinisation. 
Conclusion—Intracellular alkalinisation stimulates pH sensitive large conductance apical K⁺ channels in rat distal colonic surface cells as part of the K⁺ secretory response to chronic dietary K⁺ loading. Intracellular alkalinisation seems to reflect an increase in EIPA sensitive Na⁺-H⁺ exchange, which may be a manifestation of the secondary hyperaldosteronism associated with this model of colonic K⁺ adaptation. 



Keywords: colon; dietary potassium; pH; potassium channels

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Figure 1 .

Effect of pH on large conductance K⁺ channels in isolated surface cells from rat distal colon. Typical recordings from an inside-out patch of membrane from a distal colonic surface cell isolated from K⁺ loaded animals at different V_com values, referenced to the pipette interior (K₂SO₄ solution containing 32 nM Ca²⁺ in bath, K₂SO₄ solution containing 1.2 mM Ca²⁺ in the pipette). (A) and (B) Recordings at bath pH values of 7.40 and 7.60 respectively. The dashed lines indicate zero current levels, upward current deflections indicate K⁺ flow from the bath to the pipette, and downward current deflections indicate K⁺ flow from the pipette to the bath. 


Figure 2 .

Effect of pH on large conductance K⁺ channel activity in isolated surface cells from rat distal colon. P_o was determined over a range of V_com values using inside-out patches from distal colonic surface cells isolated from K⁺ loaded animals (K₂SO₄ solution containing 32 nM Ca²⁺ in bath, K₂SO₄ solution containing 1.2 mM Ca²⁺ in the pipette). Data at pH 7.40, 7.60, and 8.00 were obtained from five to eight patches. *p<0.05, **p<0.01, and ***p<0.001 compared with value at pH 7.40. NS, not significant compared with value at 7.40. 


Figure 3 .

Effect of 1 µM ethylisopropylamiloride (EIPA) on large conductance K⁺ channels in isolated surface cells from rat distal colon. Typical recording from a cell attached patch of membrane from a distal colonic surface cell isolated from K⁺ loaded animal (Na₂SO₄ solution in bath, K₂SO₄ solution in the pipette; V_com = 0 mV). (A) K⁺ channel activity on slow time base before and after the addition of 1 µM EIPA to the bath (denoted by arrow). Sections of (A) are presented on a faster time base to compare K⁺ channel activity before the addition of EIPA (B) and afterwards (C). (D) Restoration of channel activity after removal of EIPA. The dashed lines indicate zero current levels, and downward deflections indicate K⁺ flow from the pipette to the cell. 


Figure 4 .

Effect of 1 µM ethylisopropylamiloride (EIPA) on large conductance K⁺ channel activity in isolated surface cells from rat distal colon. (A) Summary of results from five cell attached patches of membrane from distal colonic surface cells isolated from K⁺ loaded animals (Na₂SO₄ solution in bath, K₂SO₄ solution in the pipette; V_com = 0 mV). *p<0.001 compared with the value before the addition of EIPA. (B) Uniform effect of EIPA on K⁺ channel activity at holding voltages between −60 and 40 mV (n = 3 patches). Error bars not shown were smaller than the symbols. Differences between the values before and after the addition of EIPA were highly significant (p<0.001) at each holding voltage.