Our research aims to understand how channel proteins work by exploiting naturally occuring neurotoxins as molecular probes of channel structure and function. Two ion channels under current investigation are the voltage-gated Na⁺-channel and the Ca²⁺-activated K⁺ channel which are both expressed in many different types of electrically excitable cells. A key question in the Na⁺-channel field is: How does this channel selectively discriminate among inorganic cations such as Na⁺, K⁺ and Ca²⁺? We are beginning to obtain molecular insights to this mechanism by analyzing channel mutations that are defective in ionic selectivity. In the case of the K(Ca) channel, we are studying how intracellular domains of the channel protein control ion permeation and gating. We have found that a class of small proteins (Kunitz inhibitors), which include mamba snake dendrotoxins and bovine pancreatic trypsin inhibitor, bind to an internal site on the channel protein and induce discrete subconductance events at the single-channel level. In our work, we use a diverse combination of techniques that include single-channel analysis, planar bilayer, whole-cell and patch recording, as well as molecular biological approaches. Our work has also led to the discovery of an interesting saxitoxin-binding protein called saxiphilin. Saxiphilin is a homolog of transferrin that does not bind Fe³⁺. It is also a potent inhibitor of cysteine proteinases such as papain and cathepsins B and L. The long term goal of this project is to determine the physiological function of saxiphilin.