Parkinson's disease (PD) is a common neurodegenerative disorder characterized by gradual and irreversible loss of dopaminergic neurons in the substantia nigra (SN) and the formation of Lewy bodies, which are cytoplasmic proteinaceous aggregates containing α-synuclein (α-syn) [1,2]. In particular, insoluble α-syn aggregates are a significant contributors to PD progression; this abnormal deposition results from a complex interplay of various pathways, including mitochondrial dysfunction, energy depletion-induced oxidative stress, and glial activation-derived neuroinflammation [3,4]. α-syn aggregates, for example, exacerbate mitochondrial complex I deficiency and oxidative stress in cells overexpressing α-syn, as well as in PD-model transgenic mice bearing mutant A53T or A30T genes [5,6]. Therefore, reducing oxidative stress while increasing bioenergetics may be an effective strategy for treating PD pathology.

Most neurodegenerative disorders share a common molecular mechanism that eventually culminates in neuronal death in specific areas. Heterogeneous programmed cell death, including apoptosis and necroptosis, occurs with disease progression. Necroptosis refers to a form of inflammatory cell breakdown that generates unrestrained damage to a limited location by releasing cellular components into the extracellular environment and generating a robust immune response [7,8]. The necroptotic pathway is strictly regulated by the expression and activity of receptor-interacting protein kinase (RIPK)1 and RIPK3, as well as of mixed lineage kinase domain-like protein (MLKL). The final executor MLKL, activated by phosphorylated RIPK3, causes plasma membrane rupture and leakage of damage-related molecular patterns, such as high mobility group box 1 protein and mitochondrial DNA [9,10]. Increasing evidence suggests that necroptosis is involved in the etiology of several neurodegenerative disorders, including Alzheimer's disease (AD) and PD [11], [12], [13], [14]. In the context of necroptosis and PD pathogenesis, necrostatin-1 (Nec-1), a selective RIPK1 inhibitor, has shown neuroprotective effects in neural cells produced from induced pluripotent stem cells of patients with PD carrying OPA1 gene mutations [13]. Furthermore, Parkin activation, mediated by AMP-activated protein kinase (AMPK), reduces the formation of the RIPK1/RIPK3 complex, whereas parkin deficiency increases the interaction between RIPK1 and RIPK3, promoting necroptotic cell lysis [15]. A recent study found that genetic deletion of ripk3 or mlkl reduced dopaminergic degeneration and pro-inflammatory responses in mice treated with 6-hydroxydopamine (6-OHDA) or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) [14,16]. These findings support the critical involvement of necroptosis in the loss of dopaminergic cells in PD. Consequently, inhibition of necroptotic activity may be an effective treatment for nigrostriatal dopaminergic degeneration.

Physical activity is well-known for promoting neuroplasticity and eliciting neuroprotection [17]. In PD, it reduces mitochondrial abnormalities and increases trophic factor expression and striatal dopamine transmission [17], [18], [19]. We recently revealed that motor learning-based exercise reduces nigrostriatal dopaminergic degeneration in a PD mouse model by reducing α-syn aggregation [20].

Creatine is a nitrogenous organic molecule that acts as a powerful activator of cellular bioenergetics by boosting the availability of high-energy phosphate in energy-demanding tissues, such as the muscle and brain [21,22]. Chronic energy deficit leads to mitochondrial/cell integrity disruption, cell death, and oxidative stress in neurodegenerative disorders, such as AD, PD, and Huntington's disease (HD) [23,24]. Increasing creatine levels in the brain may thus be a viable therapeutic technique against neurodegeneration. In particular, combining creatine with rofecoxib (a cyclooxygenase 2 inhibitor) or coenzyme Q10 enhances the neuroprotective effects of creatine in neurodegenerative-linked transgenic mice with PD, HD, and amyotrophic lateral sclerosis (ALS) [25,26]. Although the underlying process is poorly understood, these findings imply that combined treatment may be a superior option. The purpose of this study was to determine the possible role of combined creatine administration and exercise in the subacute MPTP mouse model as well as the underlying mechanism.