Evidence for multiple signaling pathways in single squid olfactory receptor neurons

Abstract

At least two different G-protein-mediated transduction cascades, the adenylate cyclase and phospholipase C (PLC) pathway, process chemosensory stimuli for various species. In squid olfactory receptor neurons (ORNs), physiological studies indicate that both pathways may be present; however, confirmation of the transduction molecules at the protein level is absent. Here we provide evidence that the G-proteins involved in both adenylate cyclase and PLC pathways are present in squid ORNs (Lolliguncula brevis). We used immunoblotting to show that Galpha(olf), Galpha(q), and a downstream effector, enzyme PLC140, are present in the squid olfactory epithelium (OE). To localize these proteins to one or more of the five morphological cell types described for squid OE, paraformaldehyde-fixed olfactory organs were cryosectioned (10 microm), double-labeled for Galpha(olf), Galpha(q), or PLC140, and imaged. Analysis of serial sections from entire olfactory organs for epithelial area and patterns of immunofluorescence revealed a region of highest immunoreactivity at the anterior half of the organ. At the cellular level, type 1 cells could not be distinguished morphologically and were not included in the analysis. The three labeling patterns observed in type 2 cells were Galpha(q) alone, PLC140 alone, and colocalization of Galpha(q) and PLC140. Subsets of cell types 3, 4, and 5 showed colocalization of Galpha(olf) with Galpha(q) but not with PLC140. These data suggest that the PLC pathway predominates in type 2 cells; however, coexpression of Galpha(olf) with Galpha(q) in cell types 3, 4, and 5 suggests that both pathways may participate in olfactory transduction in non-type 2 squid ORNs.