L‐type calcium channels (Cav1) represent one of the three major classes (Cav1–3) of voltage‐gated calcium channels. They were identified as the target of clinically used calcium channel blockers (CCBs; so‐called calcium antagonists) and were the first class accessible to biochemical characterization. Four of the 10 known α1 subunits (Cav1.1–Cav1.4) form the pore of L‐type calcium channels (LTCCs) and contain the high‐affinity drug‐binding sites for dihydropyridines and other chemical classes of organic CCBs. In essentially all electrically excitable cells one or more of these LTCC isoforms is expressed, and therefore it is not surprising that many body functions including muscle, brain, endocrine, and sensory function depend on proper LTCC activity. Gene knockouts and inherited human diseases have allowed detailed insight into the physiological and pathophysiological role of these channels. Genome‐wide association studies and analysis of human genomes are currently providing even more hints that even small changes of channel expression or activity may be associated with disease, such as psychiatric disease or cardiac arrhythmias. Therefore, it is important to understand the structure–function relationship of LTCC isoforms, their differential contribution to physiological function, as well as their fine‐tuning by modulatory cellular processes. WIREs Membr Transp Signal 2014,3:15–38. doi: 10.1002/wmts.102Conflict of interest: The authors have declared no conflicts of interest for this article.For further resources related to this article, please visit the WIREs website.

Cav1.3 voltage-gated L-type calcium channels (LTCCs) are part of postsynaptic neuronal signaling networks. They play a key role in brain function, including fear memory and emotional and drug-taking behaviors. A whole-exome sequencing study identified a de novo mutation, p.A749G, in Cav1.3 α1-subunits (CACNA1D), the second main LTCC in the brain, as 1 of 62 high risk-conferring mutations in a cohort of patients with autism and intellectual disability. We screened all published genetic information available from whole-exome sequencing studies and identified a second de novo CACNA1D mutation, p.G407R. Both mutations are present only in the probands and not in their unaffected parents or siblings. We functionally expressed both mutations in tsA-201 cells to study their functional consequences using whole-cell patch-clamp. The mutations p.A749G and p.G407R caused dramatic changes in channel gating by shifting (~15 mV) the voltage dependence for steady-state activation and inactivation to more negative voltages (p.A749G) or by pronounced slowing of current inactivation during depolarizing stimuli (p.G407R). In both cases, these changes are compatible with a gain-of-function phenotype. Our data, together with the discovery that Cav1.3 gain-of-function causes primary aldosteronism with seizures, neurologic abnormalities, and intellectual disability, suggest that Cav1.3 gain-of-function mutations confer a major part of the risk for autism in the two probands and may even cause the disease. Our findings have immediate clinical relevance because blockers of LTCCs are available for therapeutic attempts in affected individuals. Patients should also be explored for other symptoms likely resulting from Cav1.3 hyperactivity, in particular, primary aldosteronism.

Auxiliary channel subunits regulate membrane expression and modulate current properties of voltage-activated Ca²⁺ channels and thus are involved in numerous important cell functions, including muscle contraction. Whereas the importance of the α_1S, β_1a, and γ Ca²⁺ channel subunits in skeletal muscle has been determined by using null-mutant mice, the role of the α₂δ-1 subunit in skeletal muscle is still elusive. We addressed this question by small interfering RNA silencing of α₂δ-1 in reconstituted dysgenic (α_1S-null) myotubes and in BC3H1 skeletal muscle cells. Immunofluorescence labeling of the α_1S and α₂δ-1 subunits and whole cell patch clamp recordings demonstrated that triad targeting and functional expression of the skeletal muscle Ca²⁺ channel were not compromised by the depletion of the α₂δ-1 subunit. The amplitudes and voltage dependences of L-type Ca²⁺ currents and of the depolarization-induced Ca²⁺ transients were identical in control and in α₂δ-1-depleted muscle cells. However, α₂δ-1 depletion significantly accelerated the current kinetics, most likely by the conversion of slowly activating into fast activating Ca²⁺ channels. Reverse transcription-PCR analysis indicated that α₂δ-1 is the exclusive isoform expressed in differentiated BC3H1 cells and that depletion of α₂δ-1 was not compensated by the up-regulation of any other α₂δ isoform. Thus, in skeletal muscle the Ca²⁺ channel α₂δ-1 subunit functions as a major determinant of the characteristic slow L-type Ca²⁺ current kinetics. However, this subunit is not essential for targeting of Ca²⁺ channels or for their primary physiological role in activating skeletal muscle excitation-contraction coupling.

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 Parkinson9s 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 Parkinson9s 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.