L-type Calcium Channels Are Involved In The Degeneration Of Dopaminergic Neuron Of Parkinson's Disease

Parkinson's disease (PD) is pathologically characterized by the progressive loss of dopaminergic neurons in the substantia nigra (SN) and the depletion of dopamine (DA) in the striatum. The mechanisms underlying the pathological process of DA neuron degeneration are not fully understood, many factors have been implicated in the process, which includes but not limits to the increased iron level, oxidative stress, inflammation and excito-toxicity. Recent studies indicated that a dysregulation of L-type Cav1.3 calcium channels leads to calcium overload and neurodegeneration in the SN dopaminergic neurons. Moreover, iron selectivly accumulated in the SN area also plays a key role in the pathogenesis of PD, excessive free iron accumulation may generate hydroxyl radical (OH·), which could damage proteins, nuclear and unsaturated fatty acids, finally lead to the cell death. Previous studies showed that iron could enter cardiomyocytes via the L-type Cav1.2 calcium channels, indicating that L-type calcium channels participate in the process of iron transport. Recent studies also domenstrated that voltage-gated calcium channels provide an alternative rounte for iron uptake in neuronal cell cultures. As important subtypes of L-type calcium channels, L-type Cav1.2 and Cav1.3 calcium channels are widely expressed in the different types of neurons. However, effects of L-type calcium channels on the pathology of PD and the possibly mechanisms underlying the DA neuron degeneration is still uncertain. In the present study, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the release of lactate dehydrogenase (LDH) assay, reverse transcription polymerase chain reaction (RT-PCR), animal ethology, high-performance liquid chromatography-electrochemical detection (HPLC-ECD) and the iron contents of substantia nigra (SN) were applied to investigate effects of L-type calcium channels on the dopamine neuron degeneration in the pathogenesis of PD. The results were as follows:1. Different concentrations of 6-OHDA (5-100μmol/L) treatment for 24 h induced cell viability reduction in PC12 cells (P<0.01, compared with the control); cotreatment with the calcium channel blocker nifedipine (10μmol/L) may reduce the 6-OHDA-induced LDH release (P<0.01, compared with the 6-OHDA treatment group).2.10μmol/L 6-OHDA treatment for 24 h increased the mRNA expressions of L-type Cav1.2 calcium channelsα1 subunit (P<0.01, compared with the control). 3. The apomorphine-induced rotation numbers increased after 3 d,7 d,14 d and 28 d of 6-OHDA treatment (P<0.01, compared with the control); cotreatment with nifedipine could improve the apomorphine-induced rotation behavior in 6-OHDA-lesioned rats (P<0.01, compared with the 6-OHDA treatment group). No rotation behavior was observed in the control group.4. After 3 d of 6-OHDA lesions, the L-type Cav1.2 calcium channels al subunit mRNA expression showed a tendence of increase, however, no significance was observed (P>0.05, compared with the control); after 7 d,14 d and 28 d 6-OHDA lesions, the L-type Cav1.2 calcium channels al subunit mRNA expressions markedly increased (P<0.01, compared with the 6-OHDA treatment group).5. After 7 d of 6-OHDA lesions, the dopamine contents significantly decreased in the lesioned side of striatum in 6-OHDA treated group (P<0.01, compared wtih the control). Nifedipine conferred significant protection against 6-OHDA-induced dopamine depletion (P<0.05, compared with the 6-OHDA treatment group).6. The expressions of tyrosine hydroxylase (TH) mRNA decreased in the acute iron-overloaded SN of rats (P<0.01, compared with the control), pretreatment with nifedipine may partly reverse the iron-overloaded induced TH mRNA decrease (P<0.01, compared with the acute iron-overloaded group).7. The iron contents increased in the SN of acute iron-overloaded rats (P<0.01, compared with the control), this effect was partly restored by pretreatment with nifedipine (P<0.01, compared with the acute iron-overloaded group).The results suggested that: The expressions of L-type Cavl.2 calcium channels al subunit increased in the 6-OHDA lesioned rats and in the 6-OHDA treated PC 12 cells. The L-type calcium channel blocker nifedipine could partly protect against the 6-OHDA-induced injury. Nifedipine could also partly restore the iron-overloaded induced decrease of TH mRNA expression and the increase of iron contents in the SN of rats. All these results suggested that L-type calcium channels participate in the pathological process of DA neuron degeneration, which may play an important role in the process of iron accumulation and dopaminergic neuron death in SN. This work may provide a useful framework to advance our knowledge of L-type calcium channels mediated neurodegeneration in the pathogenesis of PD.