Progressive loss of dopaminergic neurons induced by unilateral rotenone infusion into the medial forebrain bundle

Rotenone, a mitochondrial complex 1 inhibitor, causes oxidative damage via production of reactive oxygen species. We examined the pathophysiology of neuronal and glial cells of the nigrostriatal pathway following unilateral infusion of varying doses of rotenone into the substantia nigra or medial forebrain bundle of adult male Sprague–Dawley rats, sacrificed 14 and 60 days after infusion. Immunofluorescence techniques were used to qualitatively and quantitatively assay dopaminergic neurons, their projections, glial cells, synapses, and oxidative stress. Rotenone infusion into the substantia nigra at all concentrations caused extensive damage and tissue necrosis, therefore of limited relevance for producing a Parkinson disease model. Infusion of 0.5 μg of rotenone targeting the medial forebrain bundle induced oxidative stress in dopaminergic neurons causing ongoing cell stress as defined by an elevation of stress granule and oxidative stress markers. This treatment resulted in the loss of tyrosine hydroxylase immunoreactive cells in the substantia nigra (p ≤ 0.01) and loss of tyrosine hydroxylase immunoreactive nerve fibres and synaptic specialisations in the striatum (p ≤ 0.01). The infusion of 0.5 μg of rotenone also caused an increase in astrocytes and microglial cells in the substantia nigra in comparison to control (p ≤ 0.01). We examined the time-dependent reduction of tyrosine hydroxylase-positive nerve fibres and cell bodies in the striatum and substantia nigra respectively, with a progressive reduction evident 60 days after infusion (p ≤ 0.01, p ≤ 0.05). Dopaminergic axons exposed to low-dose rotenone undergo oxidative stress, with a resultant ongoing loss of dopaminergic neurons, providing an animal model relevant to Parkinson disease.

Research Highlights
► Rotenone induces progressive nigrostriatal neurodegeneration.
► Low-dose rotenone induces markers for cellular stress G3BP and α-synuclein.
► Focal low-dose rotenone induced oxidative stress causes dopaminergic neuronal loss.
► Parkinson’s disease pathophysiology is mimicked by intracranial rotenone in rats.