Fluoro-Jade C Stain Method Applied To Study Intervention Effect Of Neurokinin NK3 On Excitotoxic Injuries Of The Substantia Nigra Dopaminergic Neurons

Parkinson's disease (PD) is one of most common neurodegenerative diseases, which is characterized by serious motor sympotoms such as resting tremor, bradykinesia, rigidity, gait disturbance and postural instability. PD is typically resulted from the massive degenerative death of dopamine neurons in the substantia nigra. Current treatments can not effectively prevent nigral neurons from progressive death and pathogical progress of PD. Previous studies have shown that glutamate-driving excitotoxicity may be one of crucial factors implicating in degeneration of nigral neurons and pathogenesis of PD. It is necessary, thus, to investigate novel strategies on preventing nigral dapamine neurons from excitotoxic lesions and improving the treatment of PD.Mammalian neurokinins are a family of tachykinin peptides that include substance P (neurokinin-1, NK-1), neurokinin A (NK-2) and neurokinin B (NK-3). Their biological functions are mediated by three distinct G-protein coupled neurokinin receptors, namely SP receptor (NK-1 receptor, NK-1R), neurokinin A receptor (NK-2R) and neurokinin B receptor (NK-3R). Neurokinin peptides and neurokinin receptors are abundantly distributed in the basal ganglia regions and known to significantly interact with neurons of the basal ganglia. Previous evidences had suggested that they may be involved in the regulation of physiological and pathological processes in the basal ganglia circuitry. In previous studies we had found that neurokinin agonists had intervention effects on exitotoxic injuries of nigral dopamine neurons of mice, particularly, administration of NK3 receptor agonist had neurotoxic synergistic effect. Further study is devoted to confirm the synergistic effect of NK3 on excitotoxicity and degenerative death of nigral dopamine neurons.In the present study, MPTP or KA-induced animal models, Fluoro-Jade C stain, administration of neuronkinin NK3 receptor antagonist, double immunofluorescence, and behavior analysis were applied to confirm effectiveness of FJC staining in the MPTP or KA-lesioned animal models and intervention role of NK3 receptor antagonist in exitotoxic injuries of nigral neurons.Firstly, Fluoro-Jade C stain can be effectively applied to detect the neuronal degeneration in the substantia nigra of animals induced by MPTP or KA insults, which shows higher resolution and contrast to stain the fine dendrites, axons and cell bodies of the degenerative neurons in the midbrain substantia nigra. In the midbrain tissue sections, FJC-positive degenerative neurons were numerously detected in the substantia nigra pars compacta, whereas they were not seen in the nigral region of control animals.Secondly, NK3 receptor antagonist effectively decreased KA-induced exitotoxic injuries of nigral dopamine neurons. It revealed that comparison with that of single KA-treated group, pretreatment with NK3 receptor antagonist relieved symptom of animal turning behavior, decreased numbers of FJC-positive degenerative neurons and increased numbers of TH-positive neurons. The data have implied that NK3 receptor antagonist may have certain neuroprotection on dopamine neurons.Taken together with previous observations, the present results have indicated that neurokinin NK3 signals may play an important role in excitotoxic degeneration of nigral dopamine neurons and pathogenesis of PD. This study also further suggests that NK3 receptor may function as potential therapeutic intervention targets in protecting nigral dopamine neurons and improving treatment of PD in human beings.