Cav1.3

Cav1.3, encoded by the gene cacna1d, is a calcium channel, voltage-dependent, L type, alpha 1D subunit. It is expressed throughout the organism and is responsible for brain function, pacemaking, secretion, and hair cell architecture. Mutations of the channel are the cause of certain psychiatric disorders, deafness, and bradycardia.

In humans, cacna1d, the gene which encodes Cav1.3, is composed of 55 exons located on chromosome 3 (3p21.1).

The α1 subunit of Cav1.3 channels is subject to alternative splicing, which cannot only affect channel gating itself, but also interaction with protein partners or tissue expression.

Some important exons, often involved in alternative splicing are [2393]:

• Exons 8a and 8b
  • Encodes the transmembrane segment S6 of Domain I
  • Expression of 8a or 8b is mutually exclusive
  • 8b is abundantly present in cochlear inner hair cells, sinoatrial node and in the brain
• Exon 11
  • Encodes the cytoplasmic loop connecting domain I and domain II
• Exon 32
  • Extracellular S3-S4 connecting loop of domain IV
• Exons 39 to 49
  • Encode various sections of the C terminus

The human Cav1.3 protein is composed of 2161 amino acid (aa) and has a molecular weight of 245 Kda.
There exists a number of protein isoforms that arise from the translation of the aforementioned transcript variants.

Protein isoforms resulting from the alternative splicing of the C-terminus have been predominantly studied. The C-terminus is a strong target for alternative splicing due to the C-terminus gating modulator’s ability to prevent Ca2+ inactivation of the channels [2394].

• The full-length (250 kD) protein isoform (Cav1.342L) possesses all the regulatory domains and is predominant in the prenatal stages.
• Cav1.3 42A and Cav1.3 43S are short isoforms lacking distal C-terminal domains. Cav1.3 42A activates at more negative voltages and inactivates faster due to enhanced Ca2+-dependent inactivation. These isoforms' C-terminal domains compete with calmodulin (CaM) for IQ domain binding.
• Splicing of exon 41 removes the IQ motif resulting in a truncated Cav1.3 protein with diminished inactivation
• Splicing of exons 44 and 48 in-frame causes disruption of the distal modulator binding to the IQ domain

Isoforms containing exon 8b resulted in a six-amino acid difference in the pore region of repeat. They also displayed lower sensitivity to DHP antagonists, faster activation kinetics, slower inactivation compared to α1C-a currents [506]

Like most mammalian proteins, Cav1.3 is subject to a series of post translational modifications (PTM).

Multiple phosphorylation sites were identified in Cav1.3, with phosphorylation mediated by proteins such as AKAPS, CaMKII, PKA, and PKC, depending on channel expression location [2395]. Phosphorylation of by PKA in the proximal C-terminus cardiac neurons was shown to increase Cav1.3 activity and thereby current by 25%. [2394] Phosphorylation by PKC in the N-terminal domain reduces up to 50% Cav1.3 mediated current by reducing the open probability of the open channel [2394].

Cav1.3 calcium channels undergo A-to-I RNA editing. This PTM occurs within exon 41, and is mediated by ADAR2. The result is slower channel inactivation kinetics, ultimately influencing the firing frequency of action potentials in neurons. The edited Cav1.3 channels have been implicated in various physiological and pathological processes, including their role in the superchiasmatic nucleus and potential relevance in Alzheimer's disease. [2396]

Lika most Cav channels, Cav1.3 is made up of a single protein composed of 4 homologous domains (DI-DIV). Each domain is made up of 6 transmembrane subunits (S1-S6) connected by extracellular loops. S1-4 form the voltage sensing domain (VSD) whereas the S5-6 form the pore module (PM). The S4 subunit of each domain contains a series of positively charged residues.
When membrane depolarization occurs, these charged residues cause the movement of the S4 subunit. This translocation of the S4 helices causes further conformational rearrangements within CaV1.3 that activate the receptor and open the channel pore. [2361]

The structure of human Cav1.3 complex, comprising the pore-forming α1 and auxiliary α2δ-1 and β3 subunits, bound to cinnarizine were determined at 3.0 Å and 3.1 Å, giving us a detailed insight into the specific structural features the protein. However, it is worth noting that these experiments may not represent the canonical native conformation [2397]. However, some Cav1.3 structural specificities were highlighted:

• Cav1.3 has minor structural differences compared to Cav1.1 in the α1-interacting domain (AID). AID is responsible for the association of β subunit to α1 and serves as a lever for β subunit to regulate the gating properties of Cav channels.[2397]
• The C-terminus of Cav1.3 channels contains a modulatory domain (CTM) that interferes with CaM modulation. The CTM has two α-helices: DCRD (C-terminal end) and PCRD (after the IQ motif). These helices bind to each other, reducing open probability and CDI, and shift voltage dependence to more positive voltages. These effects are strong in Cav1.3 and Cav1.4 but weaker in Cav1.2 channels [2395]

Cav1.3, along with other L-type channels, has 3 broadly-defined gating modes [2362]:

• Mode 0: the closed state of the channel
• Mode 1: brief ~1 ms opening
• Mode 2: longer opening duration due to strong depolarisation or interaction with other actors

Each of CaV1.3's four voltage sensors play a different role in the kinetics of the channel: one (VSD-I) is crucial for CaV1.3 activation, while the others (VSD-II, VSD-III, VSD-IV) trigger calcium release more rapidly. [2365]

In comparison to other L-type calcium channels, Cav1.3 opens and closes with faster kinetics. Cav1.3 was shown to activate a more negative potential compared (~70 mV) to Cav1.2 and have much faster activation kinetics. [2397] [2398] The channel also displays slower current inactivation during depolarizations, allowing for the mediation of long lasting Ca2+ influx during weak depolarization, and showed a very low open probability (about 0.15 at 20 mV: near the peak of an action potential) [506] [2398]

Single channel unitary conductance

The single channel unitary conductance of Cav1.3 is approximately 14.4 pS [2398]

Models

There are currently no genetic ion channel models available

Tissue & cellular

Cav1.3 is found expressed in both the PNS and CNS.

In the PNS, Cav1.3 has been identified in the sinoatrial node (SAN), atrioventricular node (AVN), the atria (A) of cardiac tissue [2399], endocrine cells [2395], sensory cells of the inner ear (inner and outer cochlear hair cells and vestibular hair cells [2395]

In the CNS, Cav1.3 is present in the brain, accounting for 10% of Cav current and in somatodendritic neurons [2395] [2395]

In the CNS, Cav1.3 channels are primarily located postsynaptically in the spines and shafts of dendrites [2393] [2395]

Cerebral function

Given their postsynaptic location throughout the brain, Cav1.3 shape neuronal excitability and dendritic spine morphology [[2393]. They also activate Ca2+ signaling pathways involved in ETC. ETC transforms synaptic activity patterns into neuronal remodeling associated with learning, memory, drug addiction, and neuronal development [2395]

Indeed, knock out mouse models of cacna1d have been used to study Cav1.3 LTCCs' roles in anxiety, depressive-like behavior, and fear processing. The key findings were [2400]: Anxiety: Knockdown of Cav1.3 in the prefrontal cortex (PFC) does not affect anxiety, indicating these channels do not mediate anxiety-like behavior. Depressive-like Behavior: Cav1.3 knockout mice show an antidepressant-like phenotype in the forced swim test (FST). Mutant mice insensitive to dihydropyridine (DHP) blockers exhibit depressive-like behavior when treated with the DHP activator BayK8644, suggesting Cav1.3's role in depression modulation. Fear Processing: Homozygous cacna1d knockout mice show impaired fear memory consolidation but normal extinction in contextual fear conditioning. Another study identified three significant CACNA1D SNPs associated with cocaine dependence, indicating Cav1.3 potential role in addiction. [2400]

Heart Pacemaking

Cav1.3 is also present in the heart and plays a vital role in pacemaking. Knockout mouse models showed bradycardia and arrhythmia due to sinoatrial node dysfunction [[2394]. Further experiments, causing the Inactivation of the Ca(v)1.3 gene in mice, also slowed pacemaker activity, resulting in arrhythmia in SAN cells due to the abolishing of the L-type current during diastolic depolarization [2401].

Adrenal hormone secretion

Cav1.3 is thought to play an important role in the secretion of aldosterone, a steroid hormone secreted by adrenal glands. Its main role is to regulate salt and water in the body, thus having an effect on blood pressure. The mechanisms by which this is done are poorly understood. However, as will be discussed later, disruption of Cav1.3 in the adrenal glands, leads to a number of channelopathies.

Sensory hair cells

Inner hair cells (IHCs) contain about 1700 Ca2+ channels, mainly CaV1.3, localized at ribbon-type active zones. Each active zone holds around 80 Ca2+ channels. This tight control of Ca2+ entry enables precise exocytosis signaling, particularly at low sound intensities [2402].

Cav1.3 were also shown to regulate ribbon synapse architecture in developing zebrafish sensory hair cells. Disrupting or blocking Ca(V)1.3a enlarges synaptic ribbons, while activation reduces ribbon size and intact synapses. Ca(2+) influx via Ca(V)1.3 fine-tunes ribbon size and is crucial for synaptic maintenance (23197719).

Outer hair cells (OHCs), like IHCs, fire spontaneous Ca2+-induced action potentials during immature development, driven by CaV1.3 Ca2+ channels (31661723).

Channelopathies

Psychiatric disorders

As Cav1.3 plays an important role in proper brain function, deregulation of the protein is the cause of a number of psychiatric disorders. These include:

• Bipolar disorder [2400]
• Autism spectrum disorder [2400]
• Long term depression [2403]

Cav1.3 may also have a role in parkinson's disease development as Dopamine-containing neurons in the substantia nigra pars compacta rely on L-type Cav1.3 Ca2+ channels for rhythmic pacemaking, making them vulnerable to Parkinson's disease stressors. This reliance increases with age. Juvenile neurons use different pacemaking mechanisms common to unaffected neurons. Blocking Cav1.3 channels in adult neurons reverts them to juvenile pacemaking, protecting against Parkinson's disease in vitro and in vivo, suggesting a new therapeutic strategy [2404].

Deafness & bradycardia
Pathologies associated with a homozygous loss of Cav1.3 function are well-studied and generally result in a number of heart conditions, including [2394]:

• Autoimmune-associated congenital heart block
• Sinoatrial node dysfunction: Autoimmune-associated sinus bradycardia
• Atrioventricular node dysfunction: Autoimmune-associated atrioventricular block
• Atrioventricular node dysfunction
• Atrial fibrillation
• Heart failure

Interestingly, these seem to go hand in hand with congenital deafness In mouse models, genetic deletion of Cav1.3 also caused congenital deafness, sinus bradycardia, and various degrees of atrioventricular (AV) block consistent with region-specific expression. Furthermore, Cav1.3−/− mice display impaired Ca2+ homeostasis associated with atrial fibrillation (AF) [2394]. These pathologies are unsurprising given the expression location of Cav1.3 in both the heart and auditory sensory cells.

Adrenal disease
Mutations to CACNA1D are linked to a common subtype of aldosteronism, Idiopathic hyperaldosteronism (IHA) [2405]. The pathology manifests itself with seizures, neurological abnormalities, and intellectual disability [2406]

Aberrant Cav1.3 may also lead to the formation of Adrenal aldosterone-producing adenomas (APAs) and often result in severe hypertension due to aldosterone overproduction. Five somatic mutations in the CACNA1D gene were found, altering Gly403 or Ile770 in the channel pore's S6 segments. This lead to lower depolarized potential activation and impaired inactivation, increasing Ca(2+) influx, and stimulating aldosterone production and cell proliferation [2407].

Auxiliary subunit

Cav1.3 channels typically exist as multi-subunit complexes comprised of the main pore-forming α1 subunit, as previously described, and auxiliary subunits α2δ-2, β3 subunits [2408] [2395]

Calcium & CaM

Cav1.3 allows the passage of Ca2+ ions in and out of the cell. However, it is itself sensitive to the fluctuation concentrations of the ion and can undergo Ca2+ dependent inactivation thanks to its interaction with certain proteins, namely CAM.

• Upon Ca2+ binding CaM undergoes a conformational change that promotes inactivation: Ca2+ unbound CAM, apoCAM, binds not just to the IQ domain but also to a central midsection of the channel.
• When Ca2+ binds to the N-lobe of CaM, CaM shifts to bind a different part of the channel (NSCaTE module), causing inactivation (N-lobe CDI).
• If Ca2+ binds to the C-lobe, CaM repositions again, leading to another configuration that also results in inactivation (C-lobe CDI).

These multiple CAM C- & N-terminal binding sites are a particularity of Cav1.3, as other L-type channels generally interact only via the C-terminal by interaction with the C-terminal and, in case of Cav1.3, also N-terminal. CDI is an important autoinhibitory mechanism preventing excessive Ca2+ influx. The strength of CDI itself can be adjusted by regulating the strength of CaM binding. [2409] [2395]

Interestingly, Cav1.3 CDI is location dependent. Cav1.3 channels auditory inner hair cells (IHCs) exhibit very little CDI compared to heterologously expressed Ca(V)1.3 channels, that exhibit intense CDI. This was shown to be caused by CaM-like calcium-binding protein (CaBP) molecules, present within the organ de Corti, which eliminated the baseline CDI of the proteins. These results further highlight the importance of Ca2+ control in Cav1.3 function [2410]

Dihydropyridine

3 large classes of drugs are known to bind to Cav1.2: phenylalkylamines (PAAs), verapamil, benzothiazepines (BTZ), diltiazem, and dihydropyridines (DHP), isradipine and nifedipine. [2395]

Other proteins

A host of other proteins are known to interact with Cav1.3 and aid in the proper function of the channel. These include [2395]:
AKAP-MAP2B: Targets PKA, which is required for efficient phosphorylation and physiological regulation
AKAP-15: Enhances channel activity by recruiting PKA
Shank: Mediates synaptic clustering of Cav1.3, and interaction plays an important role in pCREB signaling
Densin: Recruits CaMKII, which enhances activity by inducing CDF
Erbin: Increases activity by enhancing VDF
Actinin2: Crosslinks SK2 K+-channels to both LTCCs

Contributors: Rajnish Ranjan, Michael Schartner