Background: We endeavored to examine the impact of reactive oxygen species (ROS) on the degeneration of dopaminergic neurons by employing the medial forebrain bundle (MFB) axotomy model, a recognized animal model of Parkinson&rsquo;s disease. 
 
Methods: The MFB of Wistar rats was lesioned and animals were sacrificed at 7, 14 and 28 days post-lesion for immunofluorescent staining. Additionally, in some experimental animals, hydroethidine was injected into the abdominal cavity 30 minutes before sacrifice to examine the presence of ROS within the brain tissue and characterize the cell types responsible for their generation throughout the pathological progression. 
 
Result: At 7 days post-surgery, an excessive accumulation of activated microglia was observed in the ipsilateral substantia nigra (SN) using antibodies OX42 and OX6, indicating their strong immunopositive reaction. Although the reactivity decreased somewhat in the 14-day and 28-day groups compared to the 7-day group, a significant number of activated microglia still congregated. Additionally, these cells exhibited strong immunopositive reactions to ED1, indicating their active phagocytic activity. Furthermore, many activated microglia showing positive reactions in OX6 immunofluorescent staining simultaneously exhibited ethidium fluorescence activity, indicating their vigorous generation and secretion of ROS.

The Activation of Microglia and the Generation of Reactive Oxygen Species Induced by Medial Forebrain Bundle Axotomy

Background: We aimed to investigate the role of microglia in the degeneration of dopaminergic neurons by using the medial forebrain bundle (MFB) axotomy model, one of the animal models of Parkinson&rsquo;s disease.

Methods: We performed immunohistochemical staining, immunofluorescence staining, and Western blot assays on rats sacrificed 7, 14, and 28 days after the axotomy of their MFB to study tissue immune responses. Animal sacrifice was performed 30 minutes prior to injection of hydroethidine into the abdominal cavity to investigate the presence of oxidative stress in the brain tissue and the characteristics of cells that produce reactive oxygen species (ROS).

Result: Using OX42 and OX6 antibodies, excessive activation of microglia occurred in the surgical site of the substantia nigra on 7 days after the procedure. Activated microglia were still present and clustered even in the 14-day and 28-day groups, although their reactivity had slightly decreased compared to the 7-day group. These cells showed strong immunopositive reactions to ED1, indicating that they were actively engaged in phagocytic activity. Many activated microglia showing positive immunofluorescence staining for OX6 also expressed ethidium fluorescence, indicating that these cells were actively generating and secreting ROS.

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The Activation of Microglia and the Generation of Reactive Oxygen Species Induced by Medial Forebrain Bundle Axotomy

Medial Forebrain Bundle Axotomy Induced Dopaminergic Neuronal Degeneration is Accompanied with Microglial Activation