Subcellular Receptor Localization in the Substantia Nigra in a Superecliptic pHlourin-Tagged D2 Receptor Knockin Mouse

Abstract Dopamine neurons use autoregulation for appropriate modulation of goal-directed behaviors, and yet the mechanisms for D2 receptor (D2R)-mediated autoregulation are not fully understood. Electrophysiology suggests close proximity between dopamine release and receipt, but actual dendro-dendritic synapses are rare. This ultrastructural study used transgenic mice with a knockin of superecliptic green fluorescent protein (SEP) on the D2R (SEP-D2R) to determine how often autoreceptors are localized at directly apposed dendrites in the substantia nigra pars compacta (SNc). Silver-enhanced immunogold labeling for SEP-D2R was observed within dendrites, axon varicosities, astrocytes, and soma. Although most gold particles were intracellular, 28% of SEP-D2R gold was irregularly distributed along the plasma membrane. Structures immediately adjacent to dendritic membrane gold particles were axons (40%), astrocytes (19%), and other dendrites (7%), with the remaining structures unidentified in single sections. Known limitations in antibody penetration suggest the actual incidence of D2R localization at apposed dendrites is probably greater than 7%. Nevertheless, these results indicate that intercellular dopamine communication in the SNc is primarily extrasynaptic. The thin astrocytic processes often seen separating adjacent dendrites may provide channels along which transmitter diffuses to access dendritic D2Rs. Expression of D2Rs by the astrocytes themselves suggests they may participate in dopamine autoregulation. A novel finding of SEP-D2R on the axon initial segments (AISs) of SNc neurons was confirmed by immunofluorescence to involve dopamine cells. While some of this may represent axonal trafficking, membrane D2Rs might serve an autoregulatory function at the AIS yet to be physiologically characterized for dopamine neurons.