Calcium Homeostasis in Parkinson’s Disease: From Pathology to Treatment

Disturbances of Ca²⁺ homeostasis have been reported to be associated with the development of Parkinson’s disease (PD) (Fig. 1), which is characterized by the selective vulnerability of dopaminergic neurons in the substantia nigra pars compacta (SNpc). Different from the dopaminergic neurons in the ventral tegmental area (VTA), which mainly rely on hyperpolarization-activated cyclic nucleotide-gated/voltage-gated channels for pacing and have a strong Ca²⁺-buffering capacity, SNpc neurons mostly depend on L-type Ca²⁺ channels (LTCCs) for their basal activity. Also, the density of LTCCs is higher in the SNpc than in the VTA, which leads to a higher Ca²⁺ load and an increase the sensitivity of neurons to neurotoxins associated with PD [1]. Therefore, a difference in Ca²⁺ homeostasis has been proposed to be one of the most likely reasons for the selective degeneration of dopaminergic neurons in the SNpc rather than in the VTA region in PD [1]. Since α-synuclein aggregation, mitochondrial oxidative stress, and lysosome dysfunction are major risk factors of PD, potential interactions between Ca²⁺ and the above factors, and potential treatments for PD that target Ca²⁺ are discussed.

Fig. 1.

Calcium homeostasis in the physiological state and Parkinson’s disease Under physiological conditions, Ca²⁺ shuttles between the extracellular space, cytoplasm, endoplasmic reticulum (ER), mitochondria, lysosomes, and Golgi bodies, and maintains a stable amplitude and frequency of Ca²⁺ changes in different cellular compartments, which are strictly regulated by many types of Ca²⁺ channels or Ca²⁺ pumps. In the Parkinson’s disease state, aggregated α-synuclein, mutant PD-related genes (such as LRRK2, DJ-1, Parkin, and PINK), alter the levels of Ca²⁺ channels or damaged the cell membrane, causing imbalanced Ca²⁺ signaling between different cellular compartments, and thereby contribute to nigral dopaminergic neuron death. VGCC, voltage-gated Ca²⁺ channel; ROC, receptor-operated channel; TRP, transient receptor potential channel; ORAI, Ca²⁺ release-activated Ca²⁺ channel modulator; NCX, Na⁺/Ca²⁺ exchanger; PMCA, Ca²⁺ ATPase; ER, endoplasmic reticulum; SERCA, sarco-endoplasmic reticulum Ca²⁺-ATPase; RYR, ryanodine receptor; STIM, stromal interaction molecule; IP3, inositol-1, 4, 5-triphosphate; VDAC, voltage-dependent anion channel; MAM, mitochondria-associated endoplasmic reticulum membrane; MCU, mitochondrial Ca²⁺ uniporter; MICU1/2, mitochondrial Ca²⁺ uptake 1/2; OXPHOS, oxidative phosphorylation; TRC2/3, two-pore channels 2/3; P2X4, ATP-gated multi-ion channel; TRPML1, mucolipin-1; LRRK2, leucine-rich repetitive kinase 2; ROS, reactive oxygen species.

Calcium Homeostasis and α-Synuclein

Abnormal aggregation of α-synuclein is thought to be one of core pathogenic factors in PD. Ca²⁺ can bind to the C-terminus of α-synuclein, regulate the secretion of α-synuclein, and promote the formation of α-synuclein aggregates. Alpha-synuclein is mainly located at presynaptic terminals, where high Ca²⁺ fluctuations occur, and participates in synaptic vesicle endocytosis. Both the N-terminus and the C-terminus of α-synuclein interact with isolated synaptic vesicles; however, the binding with the C-terminus is regulated by Ca²⁺, which increases the lipid-binding capacity of α-synuclein [2]. Also, the C-terminus binding of Ca²⁺ to α-synuclein causes changes in the non-amyloid component domain and promotes the formation of β-folded structures, thus resulting in the formation of α-synuclein aggregates. Ca²⁺ and α-synuclein have been suggested to affect vesicle pool homeostasis through two mechanisms, one by promoting inter-vesicle interactions, and the other by the binding of synaptic vesicles to the plasma membrane, which may affect their access to voltage-gated Ca²⁺ channels. Also, α-synuclein has been reported to be secreted in the mouse striatum and in cultured neurons through a Ca²⁺-dependent mechanism, which is tightly regulated by gamma-aminobutyric acid transmission [3]. In PD patient-derived dopaminergic neurons with lost ATP13A2/PARK9 function, disrupted lysosomal Ca²⁺ homeostasis (decreased storage of lysosomal Ca²⁺ and increased cytosolic Ca²⁺) impairs lysosomal exocytosis, which impairs axonal and somatic α-synuclein secretion and promotes the accumulation of α-synuclein. When the lysosomal Ca²⁺ channel mucolipin-1 (TRPML1) is activated, lysosomal exocytosis is upregulated, which promotes α-synuclein secretion and prevents the cellular accumulation of α-synuclein [4]. Recently, inositol-1, 4, 5-triphosphate kinase B (ITPKB) has been shown to be associated with sporadic PD, and inhibition or knockdown of ITPKB reduces α-synuclein aggregation by affecting endoplasmic reticulum (ER)-to-mitochondria Ca²⁺ transport, which causes an accumulation of Ca²⁺ in mitochondria [5]. As a Ca²⁺-binding protein, calbindin also plays important roles in maintaining Ca²⁺ homeostasis. Both dementia with Lewy bodies and a unilateral rotenone mouse model have revealed that α-synuclein aggregates are not present in cells expressing calbinin-D28k, suggesting that calbinin-D28k is protective against α-synuclein aggregation and cell loss [6]. Similar evidence has been found in macaques expressing calbindin in the striatum by adenoviral vector injection: dopaminergic neurons express more α-synuclein on the non-calbindin-recruited control side after 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) administration; on the other hand, recruitment of calbindin into nigral dopaminergic neurons protects against MPTP-induced neurodegeneration [7].

Meanwhile, α-synuclein also affects Ca²⁺ signaling. Both the unfolded monomeric and oligomeric states of α-synuclein can induce an increase in intracellular Ca²⁺; however, only oligomeric α-synuclein induces Ca²⁺-dependent cell death. In neurons with α-synuclein triplication derived from human-induced pluripotent stem cells, excessive α-synuclein oligomers are incorporated into the membrane, which causes abnormal Ca²⁺ influx and derived ferroptosis [8].

Calcium Homeostasis and Mitochondria

In PD, mitochondrial Ca²⁺ signaling contributes to the death of nigral dopaminergic neurons, mainly by regulating the production of adenosine triphosphate (ATP) and mitochondrial oxidant stress. An α-synuclein-dependent increase of cytosolic Ca²⁺ in SNpc neurons has been found to be followed by an elevation of mitochondrial Ca²⁺ and mitochondrial oxidation [9]. In cultured hippocampal neurons, a cytosolic Ca²⁺ increase triggered by LTCC-mediated Ca²⁺ influx has reported to promote mitochondrial ATP synthesis [10]. As shown in primary cultured neurons and in mouse slices, dopaminergic neurons in the SNpc exhibit higher levels of mitochondrial oxidant stress under resting and pathology-free conditions. However, mitochondrial oxidative stress in VTA dopaminergic neurons is much lower [9, 11]. Mitochondrial oxidative stress has been reported to be enhanced by increased Ca²⁺ through Cav1, followed by the oxidation of dopamine, misfolding of α-synuclein, and impaired lysosomal function [12]. Systemic application of Cav1 channel inhibitors in mice, or inhibition of mitochondrial Ca²⁺ influx, could protect nigral dopaminergic neurons by reducing mitochondrial oxidative stress, slowing down mitophagy, and normalizing mitochondrial quality [11], suggesting that regulation of mitochondrial oxidative stress through Ca²⁺ signaling is beneficial for PD.

Mitophagy is a form of selective autophagy that degrades damaged mitochondria through autophagy. It is regulated by the PD-related gene PINK1 and the Parkin protein, and is important for mitochondrial quality control that is also affected by Ca²⁺. Loss of PINK1 is reported to be associated with reduced mitochondrial Ca²⁺ uptake or impaired mitochondrial Ca²⁺ outflow (promoting mitochondrial Ca²⁺ overload) due to depolarization [13]. Parkin can adjust the level and turnover of the mitochondrial Ca²⁺ uniporter (MCUC) regulator—MICU1/2 [14]. Also, PD-linked mutations in leucine-rich repetitive kinase 2 (LRRK2) increase the expression of MCU1 and MICU1 and reduce mitochondrial Ca²⁺ efflux [15].

Calcium Homeostasis and Lysosomes

Recently, abnormal lysosomal Ca²⁺ in PD has attracted increasing attention. Decreased content of the lysosomal Ca²⁺ store, followed by altered lysosomal morphology, has been reported in the fibroblasts of PD patient carrying the N370S mutation in β-glucocerebrosidase [16]. LRRK2 also functions as a regulator of lysosomal Ca²⁺; it is localized to the membranes of late endosomes and lysosomes, and regulates endocytic membrane trafficking in an Rab7-dependent manner. In fibroblasts from PD patients carrying the G2019S mutation in LRRK2, NAADP (nicotinic acid adenine dinucleotide phosphate) stimulates lysosomal Ca²⁺ release with the involvement of the two-pore channel and Lsm12 [17]. Meanwhile, an enlarged lysosomal system and increased perinuclear aggregation have also been reported, and these defects are corrected by the inhibition of NAADP. TRPML1 is another Ca²⁺-release channel on the lysosome that plays an essential role in lysosomal exocytosis and α-synuclein secretion, and the lysosomal Ca²⁺ release mediated by activated TRPML1 can rescue α-synuclein secretion and prevent the cellular α-synuclein accumulation in PD [4].

Calcium Homeostasis and Mitochondria-Associated ER Membranes (MAMs)

The transfer of Ca²⁺ between the ER and mitochondria depends on the Ca²⁺ transfer system located on the MAMs. Most of the Ca²⁺ in mitochondria originates from MAMs, which are closely connected to mitochondria and also associated with PD. Alpha-synuclein is also located on the MAMs, and the A30P mutation in α-synuclein shows reduced MAM localization. Both α-synuclein and its disease-causing mutants (A30P and A53T) have been shown to bind to the mitochondrial binding protein VAPB, thereby impeding its association with ER-specific PTPIP51 and disrupting the ER-mitochondrion contact site [18]. ITPKB (inositol-trisphosphate 3-kinase B), a new kinase that regulates MAMs, has recently been reported to negatively regulate α-synuclein aggregation by affecting ER-mitochondrion Ca²⁺ transport [5]. Recently, mutations in the Ca²⁺-binding EF and GTPase domains of Miro1, which is an outer mitochondrial membrane protein, have been identified in patients with sporadic PD, and these mutations cause reduced ER-mitochondrion contact sites, resulting in a reduced cytoplasmic Ca²⁺ buffering capacity of the mitochondria of fibroblasts [19]. Meanwhile, these mutations also increase mitophagy. Furthermore, the ER-mitochondrion connectivity is also regulated by familial PD-related proteins, such as DJ-1, which has been identified as an important component of the IP₃R-GRP 75-VDAC 1 complex that mediates ER-mitochondrion Ca²⁺ transfer [20]. The loss of DJ-1 function leads to destruction of the IP₃R-GRP 75-VDAC 1 complex and reduction of mitochondrial Ca²⁺ levels after IP₃R stimulation. In addition, the loss of DJ-1 also results in increased levels of the ER-mitochondrion contact site IP₃R3, suggesting that IP₃R3 forms aggregates when it loses its association with the IP₃R 3-GRP 75-VDAC 1 complex.

Treatment and Prospects

Accumulating evidence indicates the important role of imbalanced Ca²⁺ homeostasis in the development of PD, and Ca²⁺ channel blockers (such as nimodipine, isradipine, dihydropyridines, nifedipine, felodipine, and benidipine), and Ca²⁺-binding protein (such as S100B) have shown potential therapeutic effects in animal models of PD or clinical trials. However, treatment effects on PD that target lysosomal Ca²⁺ channels and MAMs are still limited. Also, the dynamic monitoring of Ca²⁺ is particularly important, especially for the storage and release of Ca²⁺ from organelles, such as the ER, mitochondria, and lysosomes. And specific drugs that target these processes may be beneficial for PD prevention or treatment. In addition, the interactions between Ca²⁺ and other metal ions, such as iron [21], which contribute to the development of PD, are also worthy of further investigation.