Block of kainate subtype glutamate receptor channels by internal polyamines was analysed using outside out patches from HEK 293 cells transiently transfected with GluR6(Q). Tetramines with different numbers and spacing of methylene groups between NH2 groups produced biphasic rectification well fit by the Woodhull model for a weakly permeable ion channel blocker. Such analysis revealed an increase in binding energy of 611 cal M(-1) for each methylene group added over the range 6-12 (CH2), suggesting that a major component of block by polyamines involves hydrophobic binding. Isomers with the same number of CH2 groups but different spacing between NH2 groups showed similar affinity. Due to differences in pKa values for protonation of NH2 groups, the average charge on the tetramines studied would be expected to vary from 3.98 to 2.22 at physiological pH; despite this, the voltage dependence of block was similar for all tetramines tested, with a mean value for ztheta of 1.82, similar to values for polyamines with five or six NH2 groups. In contrast, for 1,3-propane diamine (DA3 ztheta 0.83), and the N-propyl- (ztheta 1.42) and N,N'-diethyl- (ztheta 1.37) analogues of DA3, there was an increase in the voltage dependence of block on addition of hydrophobic groups.
