Distinct expression of cold receptors (TRPM8 and TRPA1) in the rat nodose-petrosal ganglion complex.

TRPM8 and TRPA1 are cold-activated transient receptor potential (TRP) cation channels. TRPM8 is activated by moderate cooling, while TRPA1 is activated by extreme, noxious cold temperatures. These cold receptors are expressed in different subpopulations of primary afferent neurons. TRPA1 is co-expressed in a subpopulation of somatosensory neurons expressing TRPV1, which is activated by heat. However, the distribution and co-expression of these channels in the nodose-petrosal ganglion complex, which contains the jugular (JG), petrosal (PG), and nodose ganglia (NG) (mainly involved in putative somatic, chemo- and somato-sensation, and somato and visceral sensation, respectively), remain unknown. Here, we conducted in situ hybridization analysis of the rat nodose-petrosal ganglion complex using specific riboprobes for TRPM8, TRPA1, and TRPV1 to compare the features of the cranial sensory ganglia. Hybridization signals for TRPA1 were diffusely observed throughout these ganglia, whereas TRPM8 transcripts were seen in the JG and PG but not in the NG. We retrogradely labeled cranial nerve X with Fast Blue (fluorescent dye) and found TRPM8 transcripts in the jugular-vagal ganglion but not the NG neurons. TRPA1 transcripts were not detected in TRPM8-expressing neurons but were present in the subpopulation of TRPV1-expressing visceral sensory neurons. Taken together, these findings support that in the vagal system the expression of cold-activated TRP channels differs between nodose- and jugular-ganglion neurons suggesting different mechanisms of cold-transduction and that the TRPA1 distribution is consistent with its proposed function as a cold-sensing receptor in the visceral system.

Development of generic calcium imaging assay for monitoring Gi-coupled receptors and G-protein interaction.

G-protein-coupled receptors (GPCRs) are important therapeutic targets for many areas of drug research and development. Although chimeric Galpha16 proteins are valuable tools for detecting the activation of Galpha(i/o)-coupled receptors, the details of the activation process remain unclear. The authors introduce a series of chimeras that combine both Galpha16 and Galpha(i/o) (Galpha(16/o), Galpha(16/i2), and Galpha(16/i3)) into a well-established transient expression system to examine the ability of these chimeras to interact with D2 long-form (D2L) dopamine and 5-HT1A serotonin receptors. The pEC50 data obtained for known agonists were similar to results from previous studies that used other cell-based assays, thus indicating sufficient sensitivity for the assay. Moreover, quinpirole exhibited similar intrinsic activity to dopamine at the D2L receptor, whereas S-(-)-3-PPP displayed partial activity of dopamine and quinpirole in the presence of the Galpha(16/o) chimera. The potency of dopamine for D2L receptors was similar among Galpha(16/o), Galpha(16/i2), and Galpha(16/i3). In contrast, the 5-HT1A receptor exhibited a significantly preferential coupling for Galpha(16/i3) compared with Galpha(16/i2) when serotonin was used as a ligand. This finding was in close agreement with the results of previous reports. The present system could therefore be used as a rapid functional assay for high-throughput screening and deorphanization.

P2X(2)- and P2X(3)-positive fibers in fungiform papillae originate from the chorda tympani but not the trigeminal nerve in rats and mice.

The subtype 2 and subtype 3 ionotropic purinergic receptors (P2X receptors) are crucial for gustation, but the distribution of these receptors in the geniculate ganglion (GG) and their colocalization in tongue papillae remain unknown. Here we investigated the expression and colocalization of P2X(2) and P2X(3) receptors in the GG and fungiform papillae in rats and mice by using in situ hybridization and immunohistochemistry. In both species, P2X(2) transcripts and immunoreactivity were detected in approximately 50-60% of GG neuronal somata, whereas those of P2X(3) were observed in almost all neurons. In each fungiform papilla, immunoreactivity for both receptors was mostly colocalized and was seen in nerve fibers and their bundles concentrated in the taste buds. Because it is well known that the P2X receptors are involved in not only taste but also nociception, we determined whether the expression originated from the chorda tympani nerve (CT, gustatory) or trigeminal nerve (somatosensory) by cutting the CT in both animals. Most P2X(2) and P2X(3) immunoreactivity in the fungiform papillae was abolished after transection, although the nerve fiber immunoreactivity of transient receptor potential V1 (a marker of somatosensory nerve fibers) remained unchanged, indicating that most fungiform papillae nerve fibers with P2X(2) and P2X(3) receptors were derived from CT. Taken together, these findings suggest that most P2X(2) and P2X(3) receptors in fungiform papillae are used for gustation rather than somatosensation.