Microscopic kinetics and energetics distinguish GABA(A) receptor agonists from antagonists

Although agonists and competitive antagonists presumably occupy overlapping binding sites on ligand-gated channels, these interactions cannot be ident ical because agonists cause channel opening whereas antagonists do not. One explanation is that only agonist binding performs enough work on the recep tor to cause the conformational changes that lead to gating. This idea is s upported by agonist binding rates at GABAA and nicotinic acetylcholine rece ptors that are slower than expected for a diffusion-limited process, sugges ting that agonist binding involves an energy-requiring event. This hypothes is predicts that competitive antagonist binding should require less activat ion energy than agonist binding. To test this idea, we developed a novel el econvolution-based method to compare binding and unbinding kinetics of GABA A receptor agonists and antagonists in outside-out patches from rat hippoca mpal neurons. Agonist and antagonist unbinding rates were steeply correlate d with affinity. Unlike the agonists, three of the four antagonists tested had binding rates that were fast, independent of affinity, and could be acc ounted for by diffusion- and dehydration-limited processes. In contrast, ag onist binding involved additional energy-requiring steps, consistent with t he idea that channel gating is initiated by agonist-triggered movements wit hin the ligand binding site. Antagonist binding does not appear to produce such movements, and may in fact prevent them.