Quantification of Dopaminergic Neuron Density and Phenotype in Human Midbrain Tissue v1

Parkinson’s disease (PD) is characterised by the abnormal intraneuronal accumulation of α-synuclein (αSyn) and the selective degeneration of neuromelanin (NM)-containing dopaminergic neurons in the substantia nigra pars compacta (SNpc). However, the relationship between NM pigmentation, dopaminergic phenotype and neuronal vulnerability in PD remains unclear. This protocol describes a method for quantifying neuronal populations in human midbrain sections based on NM pigmentation and tyrosine hydroxylase (TH) immunoreactivity. Neurons are classified into three phenotypes: NM⁺ only, TH⁺NM⁺, and TH⁺ only. The density and relative proportion of each phenotype are quantified in the ventral (SNv) and dorsal (SNd) tiers of the SNpc and the ventral tegmental area (VTA). Comparisons between PD and control tissue allow assessment of regional differences in neuronal density and changes in the relative abundance of these neuronal phenotypes.