Limits...
Expression of GABAergic receptors in mouse taste receptor cells.

Starostik MR, Rebello MR, Cotter KA, Kulik A, Medler KF - PLoS ONE (2010)

Bottom Line: Taste receptor cells (TRCs) in the circumvallate papillae express multiple subunits of the GABA(A) and GABA(B) receptors as well as multiple GATs.Both GABA(A)-and GABA(B)- immunoreactivity were detected in the peripheral taste receptor cells.Thus, we were able to identify that GABAergic receptors are expressed in some Type II and Type III taste cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University at Buffalo, The State University of New York, Buffalo, New York, United States of America.

ABSTRACT

Background: Multiple excitatory neurotransmitters have been identified in the mammalian taste transduction, with few studies focused on inhibitory neurotransmitters. Since the synthetic enzyme glutamate decarboxylase (GAD) for gamma-aminobutyric acid (GABA) is expressed in a subset of mouse taste cells, we hypothesized that other components of the GABA signaling pathway are likely expressed in this system. GABA signaling is initiated by the activation of either ionotropic receptors (GABA(A) and GABA(C)) or metabotropic receptors (GABA(B)) while it is terminated by the re-uptake of GABA through transporters (GATs).

Methodology/principal findings: Using reverse transcriptase-PCR (RT-PCR) analysis, we investigated the expression of different GABA signaling molecules in the mouse taste system. Taste receptor cells (TRCs) in the circumvallate papillae express multiple subunits of the GABA(A) and GABA(B) receptors as well as multiple GATs. Immunocytochemical analyses examined the distribution of the GABA machinery in the circumvallate papillae. Both GABA(A)-and GABA(B)- immunoreactivity were detected in the peripheral taste receptor cells. We also used transgenic mice that express green fluorescent protein (GFP) in either the Type II taste cells, which can respond to bitter, sweet or umami taste stimuli, or in the Type III GAD67 expressing taste cells. Thus, we were able to identify that GABAergic receptors are expressed in some Type II and Type III taste cells. Mouse GAT4 labeling was concentrated in the cells surrounding the taste buds with a few positively labeled TRCs at the margins of the taste buds.

Conclusions/significance: The presence of GABAergic receptors localized on Type II and Type III taste cells suggests that GABA is likely modulating evoked taste responses in the mouse taste bud.

Show MeSH

Related in: MedlinePlus

Localization of GABAAα1 receptors in the IP3R3-GFP expressing circumvallate taste buds.A Z-stack of 4 laser scanning confocal micrographs (LSCM, 0.5 µm each, collected 1 µm apart) of circumvallate taste buds from an IP3R3-GFP mouse labeled with an antibody directed against the GABAAalpha1 subunit is shown. Panel A shows the GFP fluorescence with the corresponding anti-GABAAα1 immunoreactivity of the same section shown in panel B (red labeling). A DIC bright field image of the taste buds is shown in C. An overlay of the images from A, B, and C is shown in D and demonstrates that some IP3R3-GFP expressing taste cells were immunoreactive for GABAAα1 (see arrowheads for example cells). The lack of labeling when the section is incubated with primary antibody that has been pre-incubated with blocking peptide is shown in E. All staining is eliminated when the blocking peptide is present. F shows an overlay of the panel from E with the corresponding DIC image and GFP expression in the taste buds. Scale bars  = 20 µm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2964312&req=5

pone-0013639-g003: Localization of GABAAα1 receptors in the IP3R3-GFP expressing circumvallate taste buds.A Z-stack of 4 laser scanning confocal micrographs (LSCM, 0.5 µm each, collected 1 µm apart) of circumvallate taste buds from an IP3R3-GFP mouse labeled with an antibody directed against the GABAAalpha1 subunit is shown. Panel A shows the GFP fluorescence with the corresponding anti-GABAAα1 immunoreactivity of the same section shown in panel B (red labeling). A DIC bright field image of the taste buds is shown in C. An overlay of the images from A, B, and C is shown in D and demonstrates that some IP3R3-GFP expressing taste cells were immunoreactive for GABAAα1 (see arrowheads for example cells). The lack of labeling when the section is incubated with primary antibody that has been pre-incubated with blocking peptide is shown in E. All staining is eliminated when the blocking peptide is present. F shows an overlay of the panel from E with the corresponding DIC image and GFP expression in the taste buds. Scale bars  = 20 µm.

Mentions: Immunocytochemical studies provide further evidence for the presence of the ionotropic GABA receptors in mouse circumvallate papillae. Staining of TRCs with the GABA Aα1 antibody showed a spotty distribution of GABA immunoreactivity throughout the taste cell in a subpopulation of TRCs (Figure 3A–D) with no corresponding labeling in the negative control sample (Figure 3E–F). GABAA immunoreactivity was detected in all taste buds that were labeled with anti-GABAA antibodies (n = 173 taste buds from IP3R3-GFP mice and n = 64 from GAD-GFP mice). Some GABA Aα1-immunoreactivity was detected on the IP3R3-GFP taste cells indicating that these receptors are expressed on some Type II taste cells. There was also GABA Aα1 labeling in the cells surrounding the taste bud. This labeling was abolished when the blocking peptide was incubated with the primary antibody (see Figure 3E) and immunoreactivity was not readily apparent below the taste bud (see Figure 3B), so we concluded the antibody labeling is specific and that GABA Aα1 expression is not restricted to the taste bud. This agrees with our RT-PCR results that found GABA Aα1 expression in both the taste buds and non-gustatory lingual epithelium. We also evaluated GABA Aα1 expression in Type III cells using the GAD67-GFP to identify this sub-population of taste cells (Figure 4). Some labeling in the taste bud was detected but the strongest labeling was in the surrounding epithelial cells. We saw a few GAD67-GFP labeled TRCs that had some immunoreactivity for GABA Aα1, but most fluorescent cells were not labeled.


Expression of GABAergic receptors in mouse taste receptor cells.

Starostik MR, Rebello MR, Cotter KA, Kulik A, Medler KF - PLoS ONE (2010)

Localization of GABAAα1 receptors in the IP3R3-GFP expressing circumvallate taste buds.A Z-stack of 4 laser scanning confocal micrographs (LSCM, 0.5 µm each, collected 1 µm apart) of circumvallate taste buds from an IP3R3-GFP mouse labeled with an antibody directed against the GABAAalpha1 subunit is shown. Panel A shows the GFP fluorescence with the corresponding anti-GABAAα1 immunoreactivity of the same section shown in panel B (red labeling). A DIC bright field image of the taste buds is shown in C. An overlay of the images from A, B, and C is shown in D and demonstrates that some IP3R3-GFP expressing taste cells were immunoreactive for GABAAα1 (see arrowheads for example cells). The lack of labeling when the section is incubated with primary antibody that has been pre-incubated with blocking peptide is shown in E. All staining is eliminated when the blocking peptide is present. F shows an overlay of the panel from E with the corresponding DIC image and GFP expression in the taste buds. Scale bars  = 20 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2964312&req=5

pone-0013639-g003: Localization of GABAAα1 receptors in the IP3R3-GFP expressing circumvallate taste buds.A Z-stack of 4 laser scanning confocal micrographs (LSCM, 0.5 µm each, collected 1 µm apart) of circumvallate taste buds from an IP3R3-GFP mouse labeled with an antibody directed against the GABAAalpha1 subunit is shown. Panel A shows the GFP fluorescence with the corresponding anti-GABAAα1 immunoreactivity of the same section shown in panel B (red labeling). A DIC bright field image of the taste buds is shown in C. An overlay of the images from A, B, and C is shown in D and demonstrates that some IP3R3-GFP expressing taste cells were immunoreactive for GABAAα1 (see arrowheads for example cells). The lack of labeling when the section is incubated with primary antibody that has been pre-incubated with blocking peptide is shown in E. All staining is eliminated when the blocking peptide is present. F shows an overlay of the panel from E with the corresponding DIC image and GFP expression in the taste buds. Scale bars  = 20 µm.
Mentions: Immunocytochemical studies provide further evidence for the presence of the ionotropic GABA receptors in mouse circumvallate papillae. Staining of TRCs with the GABA Aα1 antibody showed a spotty distribution of GABA immunoreactivity throughout the taste cell in a subpopulation of TRCs (Figure 3A–D) with no corresponding labeling in the negative control sample (Figure 3E–F). GABAA immunoreactivity was detected in all taste buds that were labeled with anti-GABAA antibodies (n = 173 taste buds from IP3R3-GFP mice and n = 64 from GAD-GFP mice). Some GABA Aα1-immunoreactivity was detected on the IP3R3-GFP taste cells indicating that these receptors are expressed on some Type II taste cells. There was also GABA Aα1 labeling in the cells surrounding the taste bud. This labeling was abolished when the blocking peptide was incubated with the primary antibody (see Figure 3E) and immunoreactivity was not readily apparent below the taste bud (see Figure 3B), so we concluded the antibody labeling is specific and that GABA Aα1 expression is not restricted to the taste bud. This agrees with our RT-PCR results that found GABA Aα1 expression in both the taste buds and non-gustatory lingual epithelium. We also evaluated GABA Aα1 expression in Type III cells using the GAD67-GFP to identify this sub-population of taste cells (Figure 4). Some labeling in the taste bud was detected but the strongest labeling was in the surrounding epithelial cells. We saw a few GAD67-GFP labeled TRCs that had some immunoreactivity for GABA Aα1, but most fluorescent cells were not labeled.

Bottom Line: Taste receptor cells (TRCs) in the circumvallate papillae express multiple subunits of the GABA(A) and GABA(B) receptors as well as multiple GATs.Both GABA(A)-and GABA(B)- immunoreactivity were detected in the peripheral taste receptor cells.Thus, we were able to identify that GABAergic receptors are expressed in some Type II and Type III taste cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University at Buffalo, The State University of New York, Buffalo, New York, United States of America.

ABSTRACT

Background: Multiple excitatory neurotransmitters have been identified in the mammalian taste transduction, with few studies focused on inhibitory neurotransmitters. Since the synthetic enzyme glutamate decarboxylase (GAD) for gamma-aminobutyric acid (GABA) is expressed in a subset of mouse taste cells, we hypothesized that other components of the GABA signaling pathway are likely expressed in this system. GABA signaling is initiated by the activation of either ionotropic receptors (GABA(A) and GABA(C)) or metabotropic receptors (GABA(B)) while it is terminated by the re-uptake of GABA through transporters (GATs).

Methodology/principal findings: Using reverse transcriptase-PCR (RT-PCR) analysis, we investigated the expression of different GABA signaling molecules in the mouse taste system. Taste receptor cells (TRCs) in the circumvallate papillae express multiple subunits of the GABA(A) and GABA(B) receptors as well as multiple GATs. Immunocytochemical analyses examined the distribution of the GABA machinery in the circumvallate papillae. Both GABA(A)-and GABA(B)- immunoreactivity were detected in the peripheral taste receptor cells. We also used transgenic mice that express green fluorescent protein (GFP) in either the Type II taste cells, which can respond to bitter, sweet or umami taste stimuli, or in the Type III GAD67 expressing taste cells. Thus, we were able to identify that GABAergic receptors are expressed in some Type II and Type III taste cells. Mouse GAT4 labeling was concentrated in the cells surrounding the taste buds with a few positively labeled TRCs at the margins of the taste buds.

Conclusions/significance: The presence of GABAergic receptors localized on Type II and Type III taste cells suggests that GABA is likely modulating evoked taste responses in the mouse taste bud.

Show MeSH
Related in: MedlinePlus