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1. The% reduction in size of the externally recorded action potential produced by concentrations of tetrodotoxin (TTX) in the range 6-300 nM was determined for the small non-myelinated fibres of the rabbit cervical vagus nerve and of the walking leg nerves of crab and lobster. The concentration of TTX for 50% reduction was around 80 nM for rabbit vagus and 14 nM for crab nerve.
2. Bio-assay procedures were devised to measure the amount of TTX taken up by a nerve when it was exposed to a very small volume of a solution whose TTX content was just great enough to produce 100% block of conduction. The extracellular space of each nerve was determined with [14C]sugar so that an allowance could be made for extracellular dilution.
3. The TTX binding by rabbit, crab and lobster nerve was respectively 0·064, 0·053 and 0·036 p-mole/mg wet weight of nerve.
4. The binding of saxitoxin was measured in rabbit vagus nerve, and found to be much the same as that of TTX.
5. Control experiments on rabbit sciatic nerve, where the area of excitable membrane was much smaller, showed that there was relatively little unspecific binding of TTX.
6. In view of the evidence presented here and elsewhere that the blocking of sodium conductance by TTX involves the attachment of only one TTX molecule at each sodium site, and that unspecific binding of TTX does not cause serious errors, these results suggest that in 1 µm2 of nerve membrane the number of sodium sites is 75 for rabbit, 49 for crab, and 36 for lobster nerve.
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