Taste bud cells of adult mice are responsive to Wnt/β-catenin signaling: implications for the renewal of mature taste cells

Abstract

Wnt/β-catenin signaling initiates taste papilla development in mouse embryos, however, its involvement in taste cell turnover in adult mice has not been explored. Here we used the BATGAL reporter mouse model, which carries an engineered allele in which the LacZ gene is expressed in the presence of activated β-catenin, to determine the responsiveness of adult taste bud cells to canonical Wnt signaling. Double immunostaining with markers of differentiated taste cells revealed that a subset of Type I, II, and III taste cells express β-galactosidase. Using in situ hybridization, we showed that β-catenin activates the transcription of the LacZ gene mainly in intragemmal basal cells that are immature taste cells, identified by their expression of Sonic Hedgehog (Shh). Finally, we showed that β-catenin activity is significantly reduced in taste buds of 25-week-old mice compared with 10-week-old animals. Our data suggest that Wnt/β-catenin signaling may influence taste cell turnover by regulating cell differentiation. Reduced canonical Wnt signaling in older mice could explain in part the loss of taste sensitivity with aging, implicating a possible deficiency in the rate of taste cell renewal. More investigations are now necessary to understand if and how Wnt signaling regulates adult taste cell turnover.

Fig. 1. A subset of each fusiform taste cell type is responsive to β-catenin in both the circumvallate and fungiform papillae of adult mice

Double labeled cells are identified with a white asterisk on 0.75 µm optical section micrographs of co-labeling of β-galactosidase (green nuclei) and cell type markers (Type I NTPdase2; Type II PLCβ2 and α-gustducin; and Type III NCAM, red) in (a) the circumvallate papilla and (b) fungiform papillae from 10 week-old BATGAL reporter mice. Data express the percentage of β-catenin responsive cells within each taste cell type per taste bud section. N = 3 mice. Scale bars = 10 µm in all panels.

Fig. 2. In situ hybridization reveals that β-catenin responsive cells are primarily basal cells and often express Sonic Hedgehog (Shh)

Representative 0.75 µm optical sections of taste buds (encircled by white dash) in the circumvallate papilla reveal both Shh (green), and LacZ expressing (red) cells within taste buds; co-labeled cells are yellow. a) At 10 weeks of age, numerous double labeled cells are evident in taste buds of young mice (white arrowheads), as are cells singly labeled for Shh (open arrowhead: Shh-only expressing type IV cell), and cells labeled only for LacZ (white arrow: LacZ-expressing fusiform cell). b) In addition to Shh/LacZ double labeled cells within taste buds (white arrowhead), occasional basal cells outside of taste buds express LacZ (yellow arrow); Blue arrow: LacZ expressing intragemmal basal cell. c) At 25 weeks, LacZ expression is greatly diminished in circumvallate taste buds; we detected only sparse and very week expression of the reporter transcript (dim red; blue arrow), and sparsely encountered cells also double labeled for Shh. d) Shh expression (green; white arrowhead) in taste buds of 25 week-old mice, by contrast, was comparable to that detected in taste buds from mice at 10 weeks of age. Scale bars = 20 µm in all panels.

Fig. 3. Aging significantly reduces β-galactosidase expression in differentiated taste cells in circumvallate but not fungiform taste buds

a) The proportion of β-galactosidase positive type II (PLCβ2 and α-gustducin) and type III (NCAM) taste cells is significantly reduced in circumvallate taste buds of older mice (black bars: 10 week-old; white bars: 25 week-old mice). b) In fungiform taste buds, a similar trend of fewer β-galactosidase immunoreactive type II and III taste cells is evident, but this shift is not significant. Examples of taste buds from 25 week-old with limited β-galactosidase expressing type II (c,d,f,g) and type III (e,h) cells in 0.75 µm optical sections from circumvallate (c,d,e) and fungiform (f,g,h) papillae. Student’s t-test, n = 3 mice. *, p<0.05; **, p<0.01. Scale bars = 10 µm in all panels.

Fig. 4. Aging reduces the number of β-catenin responsive cells per taste bud, but not the number of each taste cell type, nor the proportion of each cell type expressing β-galactosidase

a) Left: The number of β-galactosidase positive cells per taste bud section of circumvallate and fungiform papilla from 10 week-old is significantly greater than that of 25 week-old mice. Right: representative stacks of confocal optical sections reveal the much greater extent of BATGAL reporting (green) in young versus old mice. Student’s t-test, n=5–6 mice. *, p<0.05; ***, p<0.001. Scale bars = 20 µm in all panels. b) By contrast, the number of differentiated type II and III cells per taste bud section does not differ between 10 and 25 week-old mice. Student’s t-test, n=3 mice. c) The proportion of β-galactosidase positive cells expressing each of the cell type markers per taste bud section also does not differ between 10 and 25 week-old mice.