L-type calcium channels regulate a diverse array of cellular functions within excitable cells. Of the four molecularly defined subclasses of L-type Ca channels, two are expressed ubiquitously in the mammalian nervous system (Ca_V1.2α₁ and Ca_V1.3α₁). Despite diversity at the molecular level, neuronal L-type channels are generally assumed to be functionally and pharmacologically similar, i.e., high-voltage activated and highly sensitive to dihydropyridines. We now show that Ca_V1.3α₁ L-type channels activate at membrane potentials ∼25 mV more hyperpolarized, compared with Ca_V1.2α₁. This unusually negative activation threshold for Ca_V1.3α₁ channels is independent of the specific auxiliary subunits coexpressed, of alternative splicing in domains I–II, IVS3–IVS4, and the C terminus, and of the expression system. The use of high concentrations of extracellular divalent cations has possibly obscured the unique voltage-dependent properties of Ca_V1.3α₁ in certain previous studies. We also demonstrate that Ca_V1.3α₁ channels are pharmacologically distinct from Ca_V1.2α₁. The IC₅₀for nimodipine block of Ca_V1.3α₁ L-type calcium channel currents is 2.7 ± 0.3 μm, a value 20-fold higher than the concentration required to block Ca_V1.2α₁. The relatively low sensitivity of the Ca_V1.3α₁ subunit to inhibition by dihydropyridine is unaffected by alternative splicing in the IVS3–IVS4 linker. Our results suggest that functional and pharmacological criteria used commonly to distinguish among different Ca currents greatly underestimate the biological importance of L-type channels in cells expressing Ca_v1.3α₁.