The molecular determinants for G-protein regulation of neuronal calcium channels remain controversial. We have generated a series of α1B/α1E chimeric channels, since rat brain α1E (rbEII), unlike human α1E, showed no G-protein modulation. The study, carried out in parallel using D2 receptor modulation of calcium currents in Xenopus oocytes of Gβγ modulation of calcium currents in COS-7 cells, consistently showed an essential role for domain I (from the N terminus to the end of the I-II loop) of the α1B Ca2+ channel in G-protein regulation, with no additional effect of the C terminal of α1B. The I-II loop alone of α1B, or the I-II loop together with the C-terminal tail, was insufficient to confer G-protein modulation of α1E (rbEII). We have further observed that the α1E clone rbEII is truncated at the N-terminus compared to other a1 subunits, and we isolated a PCR product from rat brain equivalent to a longer N-terminal isoform. The long N-terminal α1E, unlike the short form, showed G-protein modulation. Furthermore, the equivalent truncation of α1B (ΔN1-55) abolished G-protein modulation of α1B. Thus, we propose that the N terminus of α1B and α1E calcium channels contains essential molecular determinants for membrane-delimited G-protein inhibition, and that other regions, including the I-II loop and the C terminus, do not play a conclusive role alone.