alpha-Like toxins, a unique group designated among the scorpion alpha-toxin class that inhibit sodium channel inactivation, are highly toxic to mice but do not compete for alpha-toxin binding to receptor site 3 on rat brain sodium channels. We analysed the sequence of a new alpha-like toxin, which was also highly active on insects, and studied its action and binding on both mammalian and insect sodium channels. Action of the alpha-like toxin on isolated cockroach axon is similar to that of an alpha-toxin, and the radioactive toxin binds with a high affinity to insect sodium channels. Other sodium channel neurotoxins interact competitively or allosterically with the insect alpha-like toxin receptor site, similarly to alpha-toxins, suggesting that the alpha-like toxin receptor site is closely related to receptor site 3. Conversely, on rat brain sodium channels, specific binding of 125I-alpha-like toxin could not be detected, although at high concentration it inhibits sodium current inactivation on rat brain sodium channels. The difficulty in measuring binding to rat brain channels may be attributed to low-affinity binding due to the acidic properties of the alpha-like toxins that also impair the interaction with receptor site 3. The results suggest that alpha-like toxins bind to a distinct receptor site on sodium channels that is differentially related to receptor site 3 on mammalian and insect sodium channels.