AAV α-Synuclein Models for Parkinson’s Disease Drug Development

What is the AAV α-synuclein (AAV-Syn) model?

The progressive loss of dopaminergic neurons in the substantia nigra pars compacta, accumulation of α-synuclein-rich Lewy bodies, and associated motor dysfunction are hallmark features of Parkinson’s disease (PD). As a key contributor to PD pathophysiology, measurement of pathological α-synuclein is a key readout in preclinical PD models, and has been the rationale for the development of PD mice and rats overexpressing wild-type or mutant α-synuclein through transgenesis, pre-formed fibril (PFF) injection, or viral vector-based somatic transduction techniques.

The stereotaxic injection of adeno-associated virus (AAV) vectors overexpressing human α-synuclein (AAV-Syn) leads to the progressive loss of dopaminergic neurons in the substantia nigra pars compacta and nigrostriatal denervation. Using validated AAV vectors, AAV-Syn injections into the substantia nigra of the mouse or rat brains leads to stable α-synuclein transduction by 3-4 weeks post-injection. Typically, a progressive, greater than 50% loss of dopaminergic neurons in the substantia nigra pars compacta can be observed in these models. Additionally, AAV-Syn overexpression leads to α-synuclein aggregation, neuroinflammation, and a robust motor phenotype.

Microscopy Images

There are many α-synuclein-based animal models of Parkinson's disease available. However, there are several reasons why the AAV-Syn model is one of the preferred models for PD drug development. The disease phenotype appears in a relatively short time frame compared to α-synuclein-overexpressing transgenic animal models. AAV-Syn induced mouse and rat models have strong face and construct validity; they are superior in reproducing key molecular aspects that are in line with human PD, including the role of pathologic α-synuclein, progressive time course of dopamine (DA) dysfunction, and a predictable patten of neuroinflammation. Compared to traditional neurotoxin-based PD models (such as 6-hydroxydopamine [6-OHDA] and MPTP), AAV-Syn has improved predictive validity and is more effective at predicting therapeutic responses in human PD.

The AAV-Syn model is well-suited for drug development targeting nigrostriatal degeneration and motor dysfunction. It can also be used to assess experimental therapeutic agents targeting neuroinflammation. At Biospective, our stereotaxic delivery of AAV-Syn overexpressing human mutant A53T α-synuclein mice replicate most of the essential aspects of PD, including α-synuclein aggregation, neurodegeneration, neuroinflammation (microglia & astrocytes), and motor dysfunction.

Which motor & behavioral phenotypes can be observed in these models?

AAV-Syn models are typically generated by injecting human α-synuclein AAV vectors unilaterally into the substantia nigra of wild-type mice or rats, which induces quantifiable motor dysfunction in many motor and behavioral tests. A few commonly used, robust tests are highlighted below.

Elevated Body Swing Test (EBST) or Tail Suspension Swing Test (TSST)
The EBST has been typically used to evaluate experimental stroke in rodents, and has been adapted for use in unilateral lesion models of PD. Biospective has validated the EBST in the AAV-Syn mouse model, where we have shown significant swing asymmetry after injection of AAVs overexpressing A53T mutant human α-synuclein (AAV-hA53Tα-Syn).

Learn more about the Elevated Body Swing Test (EBST) or Tail Suspension Swing Test (TSST).

Amphetamine/Apomorphine-Induced Rotation
Unilaterally-injected AAV-Syn leads to increased rotational behavior following injection of amphetamine or apomorphine in rats starting at 8 weeks post-injection.

Learn more about Amphetamine/Apomorphine-Induced Rotation.

AAV-Syn models have also been shown to display non-motor disturbances. The forced swim test and sucrose preference test, which measure depressive-like behavior and anhedonia, respectively, are altered following bilateral AAV-Syn injections. It is also possible to use AAV-Syn injections to model gastrointestinal (GI) dysfunction.

What are the neuropathologic changes in these models?

AAV-Syn expression leads to the progressive loss of dopaminergic neurons in the substantia nigra pars compacta beginning at approximately 3-4 weeks post-injection. Preceding neurodegeneration is the rapid, early loss of striatal DA terminals and dopamine transporter dysfunction. This progressive time course of degeneration and DA dysfunction along the nigrostriatal axis closely replicates the patterns seen in human PD.

Apart from the abundant increase in transduced α-synuclein at the target site, Oliveras-Salva et al. and Bourdenx et al. showed that AAV-Syn can also lead to increased phosphorylated α-synuclein (pSer129). Daher and colleagues (Daher, 2014; Daher, 2015) identified nitrosylated α-synuclein, while Azaredo de Silviera et al. and Oliveras-Salva et al. showed increased proteinase K and urea-resistant α-synuclein aggregates in response to AAV-Syn injections.

AAV-Syn also induces a robust neuroinflammatory phenotype. Inflammatory changes include increased microglial activation, pro-inflammatory cytokine expression, innate immunity, T-cell infiltration, inflammasome expression, and astrocytosis (Theordore, 2008; Karikari, 2022; Grotemeyer, 2023).