Activation of ion transport by combined effects of ionomycin, forskolin and phorbol ester on cultured HT-29cl.19A human colonocytes.
The differentiated clone 19A of the HT-29 human colon carcinoma cell line was used as a model to study the intracellular electrophysiological effects of interaction of the cAMP, the protein kinase C (PKC) and the Ca2+ pathways. (a) A synergistic effect between ionomycin and forskolin was observed. From intracellular responses it was concluded that the synergistic effect is caused by activation of an apical Cl- conductance by protein kinase A and a basolateral K+ conductance by Ca2+. (b) A transient synergistic effect of ionomycin and the phorbol ester phorbol dibutyrate (PDB) was found. The decrease of the response appeared to be due to PKC-dependent inactivation of the basolateral K+ conductance. The synergism is caused by PKC-dependent increase of the apical Cl- conductance and Ca(2+)-dependent increase of the basolateral K+ conductance. (c) The effects of carbachol and PDB were not fully additive presumably because of their convergence on PKC activation. (d) Forskolin and PDB, when added in this order, had a less than additive effect. Results of cell-attached patch-clamp studies, presented in the accompanying paper, showed a synergistic effect of forskolin and PDB on non-rectifying small-conductance Cl- channels. Assuming that these channels are involved in the transepithelial responses it is suggested that forskolin and PDB induce a modulatory, synergistic increase of the apical Cl- conductance when both pathways are activated simultaneously. (e) The HT-29cl.19A cells differ from T84 cells in that the latter did n ot respond with an increase of the short-circuit current to addition of phorbol ester. this may be due to a very low expression of PKA alpha.
Bajnath
R B
RB
Department of Experimental Zoology, University of Amsterdam, The Netherlands.
van den Berghe
N
N
De Jonge
H R
HR
Groot
J A
JA
D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D018118:Chloride Channels / Q000502:physiology*; D005576:Colforsin / Q000494:pharmacology*; D003110:Colonic Neoplasms / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology; D004357:Drug Synergism; D004553:Electric Conductivity; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D015221:Potassium Channels / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism; D014407:Tumor Cells, Cultured
D018118:Chloride Channels; D015221:Potassium Channels; D005576:Colforsin; D015240:Phorbol 12,13-Dibutyrate; D015759:Ionomycin; D002217:Carbachol; D000242:Cyclic AMP; D011493:Protein Kinase C; D002118:Calcium