Ca²⁺-influx through L-type Ca²⁺-channels (LTCCs) is associated with activity-related stressful oscillations of Ca²⁺ levels within dopaminergic (DA) neurons in the substantia nigra (SN), which may contribute to their selective degeneration in Parkinson's disease (PD). LTCC blockers were neuroprotective in mouse neurotoxin models of PD, and isradipine is currently undergoing testing in a phase III clinical trial in early PD. We report no evidence for neuroprotection by in vivo pretreatment with therapeutically relevant isradipine plasma levels, or Ca_v1.3 LTCC deficiency in 6-OHDA-treated male mice. To explain this finding, we investigated the pharmacological properties of human LTCCs during SN DA-like and arterial smooth muscle (aSM)-like activity patterns using whole-cell patch-clamp recordings in HEK293 cells (Ca_v1.2 α1-subunit, long and short Ca_v1.3 α1-subunit splice variants; β3/α2δ1). During SN DA-like pacemaking, only Ca_v1.3 variants conducted Ca²⁺ current (I_Ca) at subthreshold potentials between action potentials. SN DA-like burst activity increased integrated I_Ca during (Ca_v1.2 plus Ca_v1.3) and after (Ca_v1.3) the burst. Isradipine inhibition was splice variant and isoform dependent, with a 5- to 11-fold lower sensitivity to Ca_v1.3 variants during SN DA-like pacemaking compared with Ca_v1.2 during aSM-like activity. Supratherapeutic isradipine concentrations reduced the pacemaker precision of adult mouse SN DA neurons but did not affect their somatic Ca²⁺ oscillations. Our data predict that Ca_v1.2 and Ca_v1.3 splice variants contribute differentially to Ca²⁺ load in SN DA neurons, with prominent Ca_v1.3-mediated I_Ca between action potentials and after bursts. The failure of therapeutically relevant isradipine levels to protect SN DA neurons can be explained by weaker state-dependent inhibition of SN DA LTCCs compared with aSM Ca_v1.2.SIGNIFICANCE STATEMENT The high vulnerability of dopamine (DA) neurons in the substantia nigra (SN) to neurodegenerative stressors causes Parkinson's disease (PD). Ca²⁺ influx through voltage-gated L-type Ca²⁺ channels (LTCCs), in particular Ca_v1.3, appears to contribute to this vulnerability, and the LTCC inhibitor isradipine is currently being tested as a neuroprotective agent for PD in a phase III clinical trial. However, in our study isradipine plasma concentrations approved for therapy were not neuroprotective in a PD mouse model. We provide an explanation for this observation by demonstrating that during SN DA-like neuronal activity LTCCs are less sensitive to isradipine than Ca_v1.2 LTCCs in resistance blood vessels (mediating dose-limiting vasodilating effects) and even at supratherapeutic concentrations isradipine fails to reduce somatic Ca²⁺ oscillations of SN DA neurons.

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