A NOVEL TYPE OF INTERNAL BARIUM BLOCK OF A MAXI-K-DEFERENS EPITHELIAL-CELLS( CHANNEL FROM HUMAN VAS)

We have recently shown that a maxi-K+ channel from vas deferens epithe lial cells contains two Ba2+-binding sites accessible from the externa l side: a ''flickering'' site located deep in the channel pore and a ' 'slow'' site located close to the extracellular mouth of the channel. Using the patch-clamp technique, we have now studied the effect of int ernal Ba2+ on this channel. Cytoplasmic Ba2+ produced a voltage-and co ncentration-dependent ''slow'' type of block with a dissociation const ant of similar to 100 mu M. However, based on its voltage dependence a nd sensitivity to K+ concentration, this block was clearly different f rom the external ''slow'' Ba2+ block previously described. Kinetic ana lysis also revealed a novel ''fast flickering'' block restricted to ch annel bursts, with an unblocking rate of similar to 310 s(-1), some 10 -fold faster than the external ''flickering'' block. Taken together, t hese results show that this channel contains multiple Ba2+-binding sit es within the conduction pore. We have incorporated this information i nto a new model of Ba2+ block, a novel feature of which is that intern al ''slow'' block results from the binding of at least two Ba2+ ions. Our results suggest that current models for Ba2+ block of maxi-K+ chan nels need to be revised.