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Pflügers Archiv

, Volume 425, Issue 1–2, pp 100–108 | Cite as

Synergistic activation of non-rectifying small-conductance chloride channels by forskolin and phorbol esters in cell-attached patches of the human colon carcinoma cell line HT-29cl.19A

  • R. B. Bajnath
  • J. A. Groot
  • H. R. De Jonge
  • M. Kansen
  • J. Bijman
Transport Processes, Metabolism and Endocrinology; Kidney, Gastrointestinal Tract, and Exocrine Glands

Abstract

Cell-attached patch-clamp studies with the human colon carcinoma HT-29cl.19A cells revealed a small chloride channel with a unitary conductance of 6.5 pS at 70 mV and 4.6 pS at −70 mV clamp potential after cAMP was increased by activation of adenylyl cyclase by forskolin. Usually channels inactivated upon patch excision, but in a few excised patches the channels stayed active and displayed a linear I/V relation in symmetrical (150 mmol/l) chloride solutions with a conductance of 7.5 pS. A 16-fold increase in channel incidence was observed when forskolin and phorbol 12,13-dibutyrate (PDB) were present together. The open probability was voltage-independent and was not different in the presence of forskolin plus PDB or with forskolin alone. The conductance sequence of the channel as deduced from outward currents carried by five different anions including chloride was: Cl>Br>NO3>gluconate > I. The permeability sequence deduced from the reversal potentials was NO3≥Br>Cl>I>gluconate. With iodide in the pipette the conductance decreased strongly. Moreover, the inward current was reduced by 61%, indicating a strong inhibition of the chloride efflux by iodide. Similarly, the forskolin-induced increase of the short-circuit current (Isc) in confluent filter-grown monolayers was strongly reduced by iodide in the apical perfusate. Iodide also increased the fractional resistance of the apical membrane and repolarized the membrane potential, indicating an inhibitory action on the forskolin-induced increase of the apical chloride conductance. The PDB-induced Isc was also reduced by iodide, suggesting that the same chloride conductance is involved in the forskolin and in the PDB response. The results suggest that forskolin via cAMP-dependent protein kinase and PDB via protein kinase C regulate the same non-rectifying small-conductance chloride channels in the HT-29cl.19A cells.

Key words

Non-rectifying small-conductance Cl channels Cell-attached patch-clamp Protein kinase A Protein kinase C Forskolin Phorbol dibutyrate 

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Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • R. B. Bajnath
    • 1
  • J. A. Groot
    • 1
  • H. R. De Jonge
    • 2
  • M. Kansen
    • 3
  • J. Bijman
    • 3
  1. 1.Department of Experimental ZoologyUniversity of AmsterdamSM AmsterdamThe Netherlands
  2. 2.Department of BiochemistryErasmus UniversityDR RotterdamThe Netherlands
  3. 3.Department of Cell BiologyErasmus UniversityDR RotterdamThe Netherlands

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