To measure the change in internal volume of channels, we have been subjecting perfused preparations to positive and negative osmotic stress. The squid axon K channel volume change inferred from hypertonic stress has an upper bound of about 1,300 cubic angstroms. The mitochondrial voltage-dependent anion channel (VDAC) reconstituted into planar lipid bilayers shows a volume change of 20 to 40 thousand cubic angstroms. These numbers are large if one expects a cork or turnstile mechanism, but are quite reasonable if one imagines a rearrangement involving the entire ionic path. The gap junction is the locus of direct transfer of ions and small molecules from cell to cell. We are attempting the incorporation of the gap junction channel into an artificial membrane system. Aliquots of the shifted vesicle fractions were added to a bilayer chamber under osmotic conditions known to promote fusion with a planar bilayer. At last three different types of channels were observed. We are presently undertaking steps to fractionate the channels further in order to isolate and identify the junctional channel, and to measure volume changes.
