Immunocytochemical analysis of the SNARE proteins syntaxin-1 and synaptobrevin-2 in rat circumvallate taste buds

Synaptic contacts between taste cells and nerve processes have been found to exist, but little is known about how synaptic vesicle docking and neurotransmitter release at taste cell synapses. The SNARE proteins SNAP-25, syntaxin, and synaptobrevin all play important roles during regulated synaptic vesicle docking and exocytosis of neurotransmitter at the CNS. We hypothesize that taste cell synapses utilize the same protein machinery as used by synapses in the CNS. Previously, we demonstrated that SNAP-25-LIR is present in taste cells with synapses (Yang et al., 2000a). At present, we are studying syntaxin-1 and synaptobrevin-2 in rat circumvallate taste buds. Our preliminary results indicate that:; Syntaxin-1 is present in a subset of taste cells and nerve processes in taste buds. Approximately 15% of the taste cells in a taste bud display cytoplasmic syntaxin-1-LIR. Double label studies show syntaxin-1-LIR cells also display immunoreactivity to PLCbeta2 (11%), synaptobrevin-2 (99%), PGP 9.5 (72%), and 5-HT (72%), respectively. No alpha-gustducin-LIR, however, is present in syntaxin-1-LIR cells. Only 4% of PLCbeta2-LIR cells display syntaxin-1-LIR, while syntaxin-1-LIR is present in 66% of synaptobrevin-2-, 75% of PGP 9.5-, and 94% of 5-HT-LIR cells. DAB immunoelectron microscopy reveals that type III taste cells with synapses displaying syntaxin-l-LIR onto nerve processes.; Synaptobrevin-2-LIR is present in approximately 35% of the taste cells in taste buds. Synaptobrevin-2-LIR colocalizes with syntaxin-1-, SNAP-25-, synaptotagmin-1-, serotonin-, and PGP 9.5-LIR. Synaptobrevin-2-LIR also colocalizes with immunoreactivity to taste signaling molecules alpha-gustducin, PLCbeta2, and IP3R3 in taste cells. All IP3R3-LIR taste cells express synaptobrevin-2-LIR, however, approximately 27% of the synaptobrevin-2-LIR taste cells do not display IP3R3-LIR. Based on ultrastructural and biochemical features, we believe that both type II and type III taste cells display synaptobrevin-2-LIR. All of the synapses that we observed from taste cells onto nerve processes express synaptobrevin-2-LIR. Using colloidal gold immunoelectron microscopy, we found that synaptobrevin-2-LIR is associated with synaptic vesicles at taste synapses.; In summary, the type III taste cells with synapses contain the SNARE proteins SNAP-25, syntaxin-1, and synaptobrevin-2; also contain the calcium sensor synaptotagmin-1. These results suggest that SNARE machinery may control synaptic vesicle fusion and exocytosis at taste cell synapses.