Electrophysiological studies of forskolin-induced changes in ion transport in the human colon carcinoma cell line HT-29 cl.19A: Lack of evidence for a cAMP-activated basolateral K+ conductance
Journal of Membrane Biology , Volume 122 - Issue 3 p. 239- 250
Forskolin (i.e, cAMP)-modulation of ion transport pathways in filter-grown monolayers of the Cl--secreting subclone (19A) of the human colon carcinoma cell line HT29 was studied by combined Ussing chamber and microimpalement experiments. Changes in electrophysiological parameters provoked by serosal addition of 10-5m forskolin included: (i) a sustained increase in the transepithelial potential difference (3.9±0.4 mV). (ii) a transient decrease in transepithelial resistance with 26±3 Ω · cm2 from a mean value of 138±13 Ω · cm2 before forskolin addition, (iii) a depolarization of the cell membrane potential by 24±1 mV from a resting value of -50±1 mV and (iv) a decrease in the fractional resistance of the apical membrane from 0.80±0.02 to 0.22±0.01. Both, the changes in cell potential and the fractional resistance, persisted for at least 10 min and were dependent on the presence of Cl- in the medium. Subsequent addition of bumetanide (10-4m), an inhibitor of Na/K/2Cl cotransport, reduced the transepithelial potential, induced a repolarization of the cell potential and provoked a small increase of the transepithelial resistance and fractional apical resistance. Serosal Ba2+ (1 mm), a known inhibitor of basolateral K+ conductance, strongly reduced the electrical effects of forskolin. No evidence was found for a forskolin (cAMP)-induced modulation of basolateral K+ conductance. The results suggest that forskolin-induced Cl- secretion in the HT-29 cl.19A colonic cell line results mainly from a cAMP-provoked increase in the Cl- conductance of the apical membrane but does not affect K+ or Cl- conductance pathways at the basolateral pole of the cell. The sustained potential changes indicate that the capacity of the basolateral transport mechanism for Cl- and the basal Ba2+-sensitive K+ conductance are sufficiently large to maintain the Cl- efflux across the apical membrane. Furthermore, evidence is presented for an anomalous inhibitory action of the putative Cl- channel blockers NPPB and DPC on basolateral conductance rather than apical Cl- conductance.
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Bajnath, R.B, Augeron, C, Laboisse, C.L, Bijman, J, de Jonge, H.R, & Groot, J.A. (1991). Electrophysiological studies of forskolin-induced changes in ion transport in the human colon carcinoma cell line HT-29 cl.19A: Lack of evidence for a cAMP-activated basolateral K+ conductance. Journal of Membrane Biology, 122(3), 239–250. doi:10.1007/BF01871424