The properties of the rat brain alpha1E Ca2+ channel subunit and its modulation by accessory rat brain alpha2-delta and beta1b subunits were studied by transient transfection in a mammalian cell line in order to attempt to reconcile the debate as to whether alpha1E forms a low-voltage-activated (LVA) or high-voltage-activated (HVA) Ca2+ channel and to examine its pharmacology in detail. alpha1E alone was capable of forming an ion-conducting pore in COS-7 cells. The properties of heteromultimeric alpha1E/alpha2-delta/beta1b channels were largely dictated by the presence of the beta1b subunit, which increased current density and tended to produce a hyperpolarizing shift in the voltage dependence of activation and inactivation. alpha1E/alpha2-delta/beta1b channels did not appear to be regulated by Ca2+-induced inactivation. alpha1E was shown to exhibit a unique pharmacological profile. omega-Agatoxin IVA blocked the current in a dose-dependent manner with an IC50 of approximately 50 nM and a maximum inhibition of about 80%, whilst omega-conotoxin MVIIC was without effect. The 1,4-dihydropyridine (DHP) antagonist nicardipine (1 micro;M) produced an inhibition of 51 +/- 7%, whereas the DHP agonist S-(-)BAY K 8644 was without effect. Our findings suggest a re-evaluation of the classification of the alpha1E Ca2+ channel subunit; we propose that rat brain alpha1E forms a novel Ca2+ channel with properties more similar to a subtype of LVA than HVA Ca2+ current.