calpain and Parkinsonian-Disorders

Parkinson's disease (PD), a debilitating progressive degenerative movement disorder associated with loss of dopaminergic (DA) neurons in the substantia nigra (SN), afflicts approximately one million people in the U.S., including a significant number of Veterans. Disease characteristics include tremor, rigidity, postural instability, bradykinesia, and at a cellular level, glial cell activation and Lewy body inclusions in DA neurons. The most potent medical/surgical treatments do not ultimately prevent disease progression. Therefore, new therapies must be developed to halt progression of the disease. While the mechanisms of the degenerative process in PD remain elusive, chronic inflammation, a common factor in many neurodegenerative diseases, has been implicated with associated accumulation of toxic aggregated α-synuclein in neurons. Calpain, a calcium-activated cysteine neutral protease, plays a pivotal role in SN and spinal cord degeneration in PD via its role in α-synuclein aggregation, activation/migration of microglia and T cells, and upregulation of inflammatory processes. Here we report an increased expression of a subset of CD4+ T cells in rodent models of PD, including MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) mice and DSP-4 [N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride]/6-hydroxydopamine rats, which produced higher levels of perforin and granzyme B - typically found in cytotoxic T cells. Importantly, the CD4+ cytotoxic subtype was attenuated following calpain inhibition in MPTP mice, suggesting that calpain and this distinct CD4+ T cell subset may have critical roles in the inflammatory process, disease progression, and neurodegeneration in PD.

Topics: Animals; Calpain; CD4-Positive T-Lymphocytes; Disease Models, Animal; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Parkinsonian Disorders; Rats; Rats, Long-Evans; T-Lymphocyte Subsets; T-Lymphocytes, Cytotoxic

We have recently demonstrated neuroprotective abilities of nimodipine, an L-type voltage dependent calcium channel (VDCC) blocker in cellular and animal models of Parkinson's disease (PD). To understand the calcium regulatory mechanisms in the disease pathogenesis, the present study examined calcium regulatory proteins calbindin and calpain mRNA and protein levels employing quantitative PCR and western blot in 1-methyl-4-phenyl pyridinium ion (MPP

Topics: Animals; Calbindins; Calcium Channel Blockers; Calpain; Cell Line; Humans; Male; Mice; Mice, Inbred BALB C; MPTP Poisoning; Neuroprotective Agents; Nimodipine; Parkinsonian Disorders

Parkinson's disease (PD) is the most common neurodegenerative movement disorder, resulting in dopaminergic (DA) neuronal loss in the substantia nigra pars compacta (SNpc) and damage to the extranigral spinal cord neurons. Current therapies do not prevent the disease progression. Hence, developing efficacious therapeutic strategies for treatment of PD is of utmost importance. The goal of this study is to delineate the involvement of calpain-mediated inflammation and neurodegeneration in SN and spinal cord in MPTP-induced parkinsonian mice (C57BL/6 N), thereby elucidating potential therapeutic target(s). Increased calpain expression was found localized to tyrosine hydroxylase (TH(+)) neurons in SN with significantly increased TUNEL-positive neurons in SN and spinal cord neurons in MPTP mice. Inflammatory markers Cox-2, caspase-1, and NOS-2 were significantly upregulated in MPTP mouse spinal cord as compared to control. These parameters correlated with the activation of astrocytes, microglia, infiltration of CD4(+)/CD8(+) T cells, and macrophages. We found that subpopulations of CD4(+) cells (Th1 and Tregs) were differentially expanded in MPTP mice, which could be regulated by inhibition of calpain with the potent inhibitor calpeptin. Pretreatment with calpeptin (25 μg/kg, i.p.) attenuated glial activation, T cell infiltration, nigral dopaminergic degeneration in SN, and neuronal death in spinal cord. Importantly, calpeptin ameliorated MPTP-induced altered gait parameters (e.g., reduced stride length and increased stride frequency) as demonstrated by analyses of spatiotemporal gait indices using ventral plane videography. These findings suggest that calpain plays a pivotal role in MPTP-induced nigral and extranigral neurodegenerative processes and may be a valid therapeutic target in PD.

Topics: Animals; Astrocytes; Calpain; Dipeptides; Dopaminergic Neurons; Gait Disorders, Neurologic; Inflammation; Lymphocyte Subsets; Macrophages; Male; Mice; Mice, Inbred C57BL; Microglia; Nerve Degeneration; Nerve Tissue Proteins; Parkinsonian Disorders; Spinal Cord; Substantia Nigra

It is widely known that the pathophysiology of idiopathic Parkinson's disease (PD) is associated with neurodegeneration and inflammatory responses in the midbrain substantia nigra. However, the possibility of neurodegeneration and inflammatory responses in other areas of the central nervous system (CNS) in course of the pathogenesis of PD remains to be explored. In this investigation, we provide evidence in support of the hypothesis that spinal cord, the final coordinator of movement, is also involved during parkinsonian degeneration using two distinct experimental parkinsonism models induced by the neurotoxin 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine (MPTP) and the environmental toxin rotenone. A key focus of our study is the role that calpain, a Ca(2+)-activated neutral protease, plays in disrupting the structural-functional integrity of the spinal cord in the context of spinal cord degeneration in experimental parkinsonism. We examined the mechanisms of calpain-mediated neuronal death in differentiated spinal cord motoneuron cultures following exposure to the active parkinsonian toxins 1-methyl-4-phenyl-pyridinium ion (MPP(+)) and rotenone and also tested the neuroprotective efficacy of calpeptin, a calpain inhibitor, in these cell culture models of experimental parkinsonism. Our results implied that spinal cord motoneurons could be a potential extranigral target of neurodegeneration during pathogenesis of PD in the CNS and that calpain inhibition could provide neuroprotection.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Apoptosis; Calpain; Cell Line; Dipeptides; Humans; Mice; Mice, Inbred C57BL; Nerve Degeneration; Neuroprotective Agents; Neurotoxins; Parkinson Disease; Parkinsonian Disorders; Rotenone; Spinal Cord; Substantia Nigra; Uncoupling Agents

The mechanisms underlying dopamine neuron loss in Parkinson's disease (PD) are not clearly defined. Here, we delineate a pathway by which dopaminergic loss induced by 1-methyl-4-phenyl 1,2,3,6 tetrahydropyridine (MPTP) is controlled in vivo. We reported previously that calpains play a central required role in dopamine loss after MPTP treatment. Here, we provide evidence that the downstream effector pathway of calpains is through cyclin-dependent kinase 5 (cdk5)-mediated modulation of the transcription factor myocyte enhancer factor 2 (MEF2). We show that MPTP-induced conversion of the cdk5 activator p35 to a pathogenic p25 form is dependent on calpain activity in vivo. In addition, p35 deficiency attenuates MPTP-induced dopamine neuron loss and behavioral outcome. Moreover, MEF2 is phosphorylated on Ser444, an inactivating site, after MPTP treatment. This phosphorylation is dependent on both calpain and p35 activity, consistent with the model that calpain-mediated activation of cdk5 results in phosphorylation of MEF2 in vivo. Finally, we provide evidence that MEF2 is critical for dopaminergic loss because "cdk5 phosphorylation site mutant" of MEF2D provides neuroprotection in an MPTP mouse model of PD. Together, these data indicate that calpain-p35-p25/cdk5-mediated inactivation of MEF2 plays a critical role in dopaminergic loss in vivo.

Topics: Animals; Apoptosis; Calcium-Binding Proteins; Calpain; Corpus Striatum; Cyclin-Dependent Kinase 5; Dopamine; Enzyme Activation; Male; MEF2 Transcription Factors; Mice; Mice, Inbred C57BL; Myogenic Regulatory Factors; Nerve Tissue Proteins; Neurons; Parkinsonian Disorders; Phosphorylation; Point Mutation; Protein Processing, Post-Translational; Recombinant Fusion Proteins; Signal Transduction; Substantia Nigra

Parkinson's disease (PD) is a movement disorder characterized by progressive degeneration of primarily the dopaminergic neurons in the substantia nigra (SN). The present study briefly describes our findings to support the hypothesis that there is a possibility of degeneration of spinal cord (SC) motoneurons in course of parkinsonism. In cell culture models of experimental parkinsonism, we examined the degeneration of ventral SC motoneuron cell line (VSC4.1) following exposure to two different toxins, such as 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and rotenone. Our studies suggested calpain activation in the apoptosis of VSC4.1 motoneurons due to exposure to these parkinsonian toxins. Furthermore, our study showed the toxic effects of the dopaminergic toxin methamphetamine (METH) on VSC4.1 cells. The results strongly implicated a possible role for calpain in the mechanism of motoneuron apoptosis during parkinsonian degeneration, at large. Hence, we examined the neuroprotective efficacy of calpeptin, a specific inhibitor of calpain, in cell culture model of experimental parkinsonism.

Topics: Animals; Apoptosis; Calpain; Cell Culture Techniques; Dipeptides; Disease Models, Animal; DNA Fragmentation; Humans; In Situ Nick-End Labeling; Motor Neurons; Parkinsonian Disorders; Rats

Parkinson's disease (PD) is a movement disorder characterized by rigidity, tremor, and bradykinesia, originating from degeneration of dopaminergic neurons in the substantia nigra (SN), retrorubral area, and locus ceoruleus (LC). Calpain has been implicated in the pathophysiology of neurodegenerative diseases. Since the spinal cord (SC) and brain are integrally connected and calpain is involved in cell death and mitochondrial dysfunction, we hypothesized that SC neurons are also affected in PD. In order to examine this hypothesis, we examined both brain and SC from mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). To identify cells expressing calpain, double immunofluorescent labeling was performed with antibodies specific for calpain and a cell type (OX-42, GFAP, or NeuN). Combined terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) and double immunofluorescent labeling were used to identify death of specific cells in the central nervous system (CNS). There was an increase in calpain expression in microglia, astrocytes, and neurons in the SC of MPTP-treated mice at 1 and 7 days, as compared to controls. TUNEL-positive neurons in the SC and SN showed apoptotic characteristics. These results demonstrated that neuronal death occurred not only in SN but also in the SC of MPTP-treated mice and has provided evidence for a possible calpain-mediated SC neuronal death in MPTP-induced parkinsonism in mice.

Topics: Analysis of Variance; Animals; Antigens, CD; Antigens, Neoplasm; Antigens, Surface; Avian Proteins; Basigin; Blood Proteins; Calpain; Cell Count; Cell Death; Disease Models, Animal; Glial Fibrillary Acidic Protein; Immunohistochemistry; In Situ Nick-End Labeling; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Neuroglia; Neurons; Parkinson Disease; Parkinsonian Disorders; Spinal Cord; Time Factors

Parkinson's disease (PD) is a neurodegenerative disorder resulting in slowness, tremors, and imbalance. Treatment of mice with 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) is one of several models used to mimic PD in humans. Administration of MPTP leads to the production of 1-methyl-4-phenyl-2,3 dihydropyridinium (MPP(+)). MPP(+) is taken up by dopaminergic neurons, causing mitochondrial dysfunction and cell death. Because calpain is involved in neuronal cell death and mitochondrial dysfunction, we examined the level of calpain in neurons in the substantia nigra (SN) and hippocampus of MPTP-treated C57BL/6 mice. Because of the interconnections between spinal cord and upper central nervous system neurons, we examined morphology, calpain activity, and calpain expression in neurons by double immunofluorescence using calpain and neuron marker (NeuN) antibodies. In controls, calpain expression was low in SN, hippocampus, and spinal cord NeuN(+) cells, and the NeuN stain was concentrated around the nucleus. In mice sacrificed 24 h after administration of three 20 mg/kg doses of MPTP, calpain expression was slightly increased in SN and hippocampal neurons and moderately increased in spinal cord neurons. In these animals, the NeuN stain was less concentrated around the nuclear membrane. One week after MPTP treatment, calpain content in NeuN(+) cells was greatly increased in SN, hippocampus, and spinal cord. Morphologically, SN and spinal cord neurons, treated for one week, were necrotic with a granular cytoplasmic NeuN content. Also, MPTP treatment upregulated calpain activity and mRNA level in spinal cord. These data suggest that following MPTP treatment, calpain causes neuronal death in SN as well as in spinal cord.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Calpain; Cell Death; Male; Mice; Mice, Inbred C57BL; Neurons; Parkinsonian Disorders; Spinal Cord

Frontal temporal dementia and Parkinsonism linked to chromosome 17 (FTDP-17) is caused by splice site and missense mutations in the tau gene, and characterized by the accumulation of filamentous tau in cerebral neurons and glia. The missense mutations reduce the ability of tau to promote microtubule assembly and increase the ability of tau to form filaments. In this report we demonstrate that mutants V337M and R406W are less susceptible than mutant P301L or corresponding wild type tau to degradation by calpain I. The differences were at least in part due to changes in accessibility of a cleavage site located about 100 amino acids off the carboxy-terminus. The results suggest that the pathogenesis of some forms of FTDP-17 may involve tau accumulation due to decreased proteolytic degradation.

Topics: Calpain; Chromosomes, Human, Pair 17; Dementia; Frontal Lobe; Humans; Immunoblotting; Kinetics; Mutation; Parkinsonian Disorders; Recombinant Proteins; Silver Staining; tau Proteins; Time Factors