Pronounced cellular diversity and extrasynaptic location of nicotinic acetylcholine receptor subunit immunoreactivities in the chicken pretectum.

The diversity of nicotinic ACh receptor (AChR) expression in the chick lateral spiriform nucleus (SpL) was assessed using subunit-specific monoclonal antibodies (mAbs) and laser scanning confocal microscopy. The late embryonic SpL was immunoreactive for mAbs against the alpha 2, alpha 5, alpha 7, alpha 8, and beta 2 AChR subunits. Distinct neuronal cell classes were determined using pair-wise staining of mAbs. Approximately 90% of the neurons in the SpL contained both alpha 5-like immunoreactivity (LI) and beta 2-LI, with no neurons having only one of these subunit-LIs. Approximately 70% of the neurons contained alpha 2-LI. All alpha 2-LI neurons contained alpha 5/beta 2-LI; thus, neurons having alpha 2-LI are a subset of those having alpha 5- and beta 2-LI. Fewer neurons, approximately 20%, contained alpha 7-LI. A subset of alpha 7-positive neurons were immunoreactive for other subunits; for example, some alpha 7-positive neurons also contained alpha 2-LI. Fewer than 15% of the neurons contained alpha 8-LI. Some of the alpha 8-LI-containing neurons contained alpha 7-LI. The 14 week post-hatch SpL resembles the late embryonic nucleus in the percentage of neurons immunoreactive for alpha 2, alpha 5, alpha 7, alpha 8, and beta 2 AChR subunits, and in the presence of multiple classes based on AChR subunit immunoreactivity. In addition, alpha 4-LI was found in about 20% of the 14 week SpL neurons. Double-label immunofluorescence experiments with mAbs to AChRs and to synaptic vesicle antigens showed that most clusters of alpha 5-LI and beta 2-LI are extrasynaptic. The pronounced diversity of AChR subunit expression and the extrasynaptic location of AChR-LI suggest that AChR-like molecules in the SpL do not function solely to respond to transmitter focally released from presynaptic terminals.