Previous saxitoxin binding studies indicated two forms of the sodium channel in the fetal rat brain; a low-affinity precursor located in an internal membrane compartment, present exclusively in growth cones and a high-affinity mature form present in the plasmalemma of growth cones and characteristic of synapses. This raises the questions (1) of the presence or absence of the beta2 subunit in these channel forms and (2) of the developmental regulation of the the beta2 subunit. Antibodies against the alpha and beta2 channel subunits were used to probe Western blots of subcellular fractions from rat brains at embryonic day 18 (E18), pups at postnatal (P) days 7-25, and adults, as well as purified sodium channels from adult brain. In both synaptosomes and the purified sodium channel the beta2 antibody recognized the expected band at 38 kDa under reducing conditions. However, in contrast to the alpha subunit, this band was absent at E18 and became apparent only from P7 onwards. At the earlier time intervals a very prominent immunoreactive band of unknown identity was evident at 260--300kDa, which declined in intensity concomitant with the appearance of the 38 kDa beta2 band. These data indicate that beta2 subunits are regulated independently from alpha subunits, are absent in differentiating neurons, and hence are not necessary for insertion of the sodium channel into the plasmalemma, at least during early development of the neuron.