The Cl -channel defect in cystic fibrosis

Several groups have shown that hormonal stimuli which lead to NaCl secretion in normal tissues fail to do so in CF tissues (reviewed in [17,18,60]). The production of the second messenger cAMP was unimpeded in these CF tissues. Hence it was 

In respiratory epithelial (RE) cells the Cl -conductance was attributed to the ICOR channel. In fact, it was reported by Frizzell et al. and Welsh s laboratories that catecholamines increased the incidence of ICOR channels in cell attached patches of normal RE cells but failed to do so in CF cells [110,111], Later both laboratories presented data on excised membrane patches of RE cells in which the protein kinase A which was added to the cytosolic side produced ICOR channel activity in the normal cells but not in the CF tissues [19,20]. This finding was reproduced by Guggino and coworkers [22] for RE cells and by others for lymphocytes [46]. Protein kinase C at physiological Ca -activities had a comparable effect in normal cells but also failed to function in CF cells [22,112]. 

Our own laboratory obtained different results. Not only were we unable to see a clear cut correlation between the incidence of ICOR channels in cell attached patches and the degree of hormonal stimulation [57], we were also unable to reproduce the activation studies in excised patches. In our hands, the activation of ICOR channels occurred simply by the excision, and this was equally true for the normal as for the CF cells [57]. We did note, however, that the other laboratories worked at room temperature whereas we always work at 37°C. Welsh s laboratory has shown meanwhile that excision activation of ICOR Cl -channels is a temperature-dependent process [113]. At low temperature, excision activation is largely delayed [113] but it is immediate in our experiments at 37°C [57]. We concluded that the activation of the ICOR channels has probably little to do with phosphorylation but is rather due to the fact that the excised patch faces a new environment on the cytosolic side [57,72], 

Meanwhile we have shown that the excision activation of ICOR channels is due to disinhibition [72]. The respective inhibitor, operationally named cytosolic inhibitor (Cl), is present in the cytosol of placenta trophoblast cells HT29- and Tg4-colonic carcinoma cells and RE cells of normal and CF patients. The molecule has an apparent molecular weight of 700-1 500 Da it is amphiphilic heat stable and not digested by trypsin, proteases, nucleotidases, lipases or amylase [72]. Burc-khardt, Fromter and their collaborators [114] have confirmed our results and extracted a similar or identical Cl from kidney cortex. 

In our patch clamp studies in excised membrane patches in which we attempted to characterize the Cl we have noted that Cl did not only inhibit the probability of the ICOR channel being open but we also found that the input conductance of the patch was reduced at the same time and with the same time course [72]. We have followed up on this observation and we were able to show that this reduction in input conductance is caused by an inhibition of small ( lOpS) Cl -channels. Hence, we postulate that the same patches containing ICOR channels also contain small (unresolved, cf. section 2.4) Cl -channels which are inhibited reversibly by CL It cannot be excluded at this stage that these small Cl -channels are responsible for the defect in CF. 

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