A SYNTHETIC PEPTIDE DERIVED FROM GLYCINE-GATED CL- CHANNEL INDUCES TRANSEPITHELIAL CL- AND FLUID SECRETION

M2GlyR is a synthetic 23-amino acid peptide that mimics the second mem brane-spanning region of the alpha-subunit of the postsynaptic glycine receptor. This peptide has been shown to form an anion-selective chan nel in phospholipid bilayers. We have investigated the possibility tha t the peptide may incorporate into the apical membrane of secretory ep ithelia and induce the secretion of Cl- and water. We improved the sol ubility of this peptide by adding four lysine residues to the carboxy terminus, C-K4-M2GlyR, and assayed its channel-forming activity using a subculture of Madin-Darby canine kidney (MDCK) cells. The addition o f 100 mu M C-K4-M2GlyR to the apical surface of MDCK monolayers signif icantly increased short-circuit current (I-sc), hyperpolarized transep ithelial potential difference, and induced fluid secretion. The increa se in I-sc was inhibited by 100 mu M bumetanide and by Cl- channel inh ibitors. The effectiveness of the channel blockers followed the sequen ce niflumic acid greater than or equal to 5-nitro-2-(3-phenylpropylami no )benzoate > diphenylamine-2-carboxylate (DPC) > glibenclamide. The effect of the peptide was not inhibited by 4,4'-diisothiocyanostilbene -2,2'-disulfonic acid. Removing Cl- from the bathing solutions also in hibited the effect of the peptide. The Cl- efflux pathway induced by C -K4-M2GlyR differs from the native pathway activated by the adenosine 3',5'-cyclic monophosphate (cAMP) agonist, forskolin. First, intracell ular cAMP levels were unaffected. Second, the concentration of DPC req uired to inhibit the effect of the peptide was much lower than that ne eded to block the forskolin response (100 mu M vs. 3 mM). These result s support the hypothesis that the synthetic peptide C-K4-M2GlyR can fo rm Cl--selective channels in the apical membrane of secretory epitheli al cells and can induce sustained transepithelial secretion of Cl- and fluid.