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
Rb. Bajnath et al., 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, Pflugers Archiv, 425(1-2), 1993, pp. 100-108
Cell-attached patch-clamp studies with the human colon carcinoma HT-29
cl.19A cells revealed a small chloride channel with a unitary conducta
nce 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 patche
s the channels stayed active and displayed a linear I/V relation in sy
mmetrical (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 pro
bability was voltage-independent and was not different in the presence
of forskolin plus PDB or with forskolin alone. The conductance sequen
ce of the channel as deduced from outward currents carried by five dif
ferent anions including chloride was: Cl- > Br- > NO3- > gluconate ) I
-. The permeability sequence deduced from the reversal potentials was
NO(3)(-)greater than or equal to Br- > Cl- > I- > gluconate. With iodi
de in the pipette the conductance decreased strongly. Moreover, the in
ward 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 (I-sc) in confluent filter-grown monolaye
rs was strongly reduced by iodide in the apical perfusate. Iodide also
increased the fractional resistance of the apical membrane and repola
rized the membrane potential, indicating an inhibitory action on the f
orskolin-induced increase of the apical chloride conductance. The PDB-
induced I-sc was also reduced by iodide, suggesting that the same chlo
ride 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-co
nductance chloride channels in the HT-29cl.19A cells.