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Bitter taste receptors on airway smooth muscle bronchodilate by localized calcium signaling and reverse obstruction.

Deshpande DA, Wang WC, McIlmoyle EL, Robinett KS, Schillinger RM, An SS, Sham JS, Liggett SB - Nat. Med. (2010)

Bottom Line: The relaxation induced by TAS2Rs is associated with a localized [Ca²(+)](i) response at the cell membrane, which opens large-conductance Ca²(+)-activated K(+) (BK(Ca)) channels, leading to ASM membrane hyperpolarization.Inhaled bitter tastants decreased airway obstruction in a mouse model of asthma.Given the need for efficacious bronchodilators for treating obstructive lung diseases, this pathway can be exploited for therapy with the thousands of known synthetic and naturally occurring bitter tastants.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA.

ABSTRACT
Bitter taste receptors (TAS2Rs) on the tongue probably evolved to evoke signals for avoiding ingestion of plant toxins. We found expression of TAS2Rs on human airway smooth muscle (ASM) and considered these to be avoidance receptors for inhalants that, when activated, lead to ASM contraction and bronchospasm. TAS2R agonists such as saccharin, chloroquine and denatonium evoked increased intracellular calcium ([Ca²(+)](i)) in ASM in a Gβγ-, phospholipase Cβ (PLCβ)- and inositol trisphosphate (IP₃) receptor-dependent manner, which would be expected to evoke contraction. Paradoxically, bitter tastants caused relaxation of isolated ASM and dilation of airways that was threefold greater than that elicited by β-adrenergic receptor agonists. The relaxation induced by TAS2Rs is associated with a localized [Ca²(+)](i) response at the cell membrane, which opens large-conductance Ca²(+)-activated K(+) (BK(Ca)) channels, leading to ASM membrane hyperpolarization. Inhaled bitter tastants decreased airway obstruction in a mouse model of asthma. Given the need for efficacious bronchodilators for treating obstructive lung diseases, this pathway can be exploited for therapy with the thousands of known synthetic and naturally occurring bitter tastants.

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Related in: MedlinePlus

Bitter tastants of diverse structures evoke increases in [Ca2+]i in human airway smooth muscle cells. Studies were performed with cultured primary ASM cells loaded with Fluo-4 AM. (a, b) [Ca2+]i transients and dose response curves to saccharin and chloroquine from 5−6 experiments. (c) Maximal [Ca2+]i responses to 1.0 mM of the bitter tastants aristocholic acid, chloroquine, colchicine, denatonium, quinine, saccharin, salicin, strychnine and yohimbine, and the bronchoconstrictive Gq-coupled agonists histamine (0.1 mM) and bradykinin (0.01 mM). Results are from 4−6 experiments. *, P < 0.01 vs. basal; #, P < 0.05 vs. denatonium. (d) The [Ca2+]i response to bitter tastant is ablated by the PLC inhibitor U73122 and the βγ antagonist gallein, and attenuated by the IP3 receptor antagonist 2APB. These studies were performed in the absence of extracellular calcium. Results shown are from a single representative experiment of at least three performed.
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Figure 1: Bitter tastants of diverse structures evoke increases in [Ca2+]i in human airway smooth muscle cells. Studies were performed with cultured primary ASM cells loaded with Fluo-4 AM. (a, b) [Ca2+]i transients and dose response curves to saccharin and chloroquine from 5−6 experiments. (c) Maximal [Ca2+]i responses to 1.0 mM of the bitter tastants aristocholic acid, chloroquine, colchicine, denatonium, quinine, saccharin, salicin, strychnine and yohimbine, and the bronchoconstrictive Gq-coupled agonists histamine (0.1 mM) and bradykinin (0.01 mM). Results are from 4−6 experiments. *, P < 0.01 vs. basal; #, P < 0.05 vs. denatonium. (d) The [Ca2+]i response to bitter tastant is ablated by the PLC inhibitor U73122 and the βγ antagonist gallein, and attenuated by the IP3 receptor antagonist 2APB. These studies were performed in the absence of extracellular calcium. Results shown are from a single representative experiment of at least three performed.

Mentions: Initial studies found that several known bitter taste receptor agonists (such as chloroquine, saccharin, and denatonium) evoked increased [Ca2+]i in cultured human ASM cells (Fig. 1a,b,c). The [Ca2+]i responses in ASM cells to these bitter tastants were found to be similar in magnitude to those for known bronchoconstrictive GPCR agonists such as histamine and bradykinin (Fig. 1c). These results prompted quantitative RT-PCR studies with primers for 25 TAS2R genes. Of note, the numerical designations of the TAS2Rs have very recently changed and here we utilize this new nomenclature (see http://www.genenames.org). Multiple TAS2R transcripts were found to be expressed in human ASM, with the TAS2R10, TAS2R14 and TAS2R31 subtypes being the most highly expressed (Table 1). Further screens with additional bitter tastants revealed [Ca2+]i responses to aristocholic acid, strychnine, quinine, colchicine, and yohimbine (Fig. 1c). We found a relatively low response in ASM to colchicine which activates TAS2R4 (a mid-level ASM expressor by RT-PCR) and no response to salicin which exclusively activates TAS2R1610 (which was not detected in ASM by RT-PCR). The robust response to strychnine (activates TAS2R10 and −46) is also consistent with TAS2R10 having high expression in ASM. Thus in ASM, the [Ca2+]i response to bitter tastants is concordant with a rank-order based on agonist specificity and the bitter taste receptor subtype expression in these cells. Immunofluorescence microscopy of human ASM cells using polyclonal antisera directed against four receptors found to have mRNA expressed by RT-PCR (TAS2R10, −14, −31, −19) and three receptors whose mRNAs were not detected (TAS2R7, −38, −43), revealed cell surface expression of the former four receptors but not for the latter three (Supplementary Fig. 1). These studies also revealed expression of the α subunit of gustducin in these cells (Supplementary Fig. 1). No [Ca2+]i response to the sweet receptor agonists sucralose and SC45647 was found, and the saccharin response was not blocked by the sweet receptor antagonist lactisole (Supplementary Fig. 2).


Bitter taste receptors on airway smooth muscle bronchodilate by localized calcium signaling and reverse obstruction.

Deshpande DA, Wang WC, McIlmoyle EL, Robinett KS, Schillinger RM, An SS, Sham JS, Liggett SB - Nat. Med. (2010)

Bitter tastants of diverse structures evoke increases in [Ca2+]i in human airway smooth muscle cells. Studies were performed with cultured primary ASM cells loaded with Fluo-4 AM. (a, b) [Ca2+]i transients and dose response curves to saccharin and chloroquine from 5−6 experiments. (c) Maximal [Ca2+]i responses to 1.0 mM of the bitter tastants aristocholic acid, chloroquine, colchicine, denatonium, quinine, saccharin, salicin, strychnine and yohimbine, and the bronchoconstrictive Gq-coupled agonists histamine (0.1 mM) and bradykinin (0.01 mM). Results are from 4−6 experiments. *, P < 0.01 vs. basal; #, P < 0.05 vs. denatonium. (d) The [Ca2+]i response to bitter tastant is ablated by the PLC inhibitor U73122 and the βγ antagonist gallein, and attenuated by the IP3 receptor antagonist 2APB. These studies were performed in the absence of extracellular calcium. Results shown are from a single representative experiment of at least three performed.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3066567&req=5

Figure 1: Bitter tastants of diverse structures evoke increases in [Ca2+]i in human airway smooth muscle cells. Studies were performed with cultured primary ASM cells loaded with Fluo-4 AM. (a, b) [Ca2+]i transients and dose response curves to saccharin and chloroquine from 5−6 experiments. (c) Maximal [Ca2+]i responses to 1.0 mM of the bitter tastants aristocholic acid, chloroquine, colchicine, denatonium, quinine, saccharin, salicin, strychnine and yohimbine, and the bronchoconstrictive Gq-coupled agonists histamine (0.1 mM) and bradykinin (0.01 mM). Results are from 4−6 experiments. *, P < 0.01 vs. basal; #, P < 0.05 vs. denatonium. (d) The [Ca2+]i response to bitter tastant is ablated by the PLC inhibitor U73122 and the βγ antagonist gallein, and attenuated by the IP3 receptor antagonist 2APB. These studies were performed in the absence of extracellular calcium. Results shown are from a single representative experiment of at least three performed.
Mentions: Initial studies found that several known bitter taste receptor agonists (such as chloroquine, saccharin, and denatonium) evoked increased [Ca2+]i in cultured human ASM cells (Fig. 1a,b,c). The [Ca2+]i responses in ASM cells to these bitter tastants were found to be similar in magnitude to those for known bronchoconstrictive GPCR agonists such as histamine and bradykinin (Fig. 1c). These results prompted quantitative RT-PCR studies with primers for 25 TAS2R genes. Of note, the numerical designations of the TAS2Rs have very recently changed and here we utilize this new nomenclature (see http://www.genenames.org). Multiple TAS2R transcripts were found to be expressed in human ASM, with the TAS2R10, TAS2R14 and TAS2R31 subtypes being the most highly expressed (Table 1). Further screens with additional bitter tastants revealed [Ca2+]i responses to aristocholic acid, strychnine, quinine, colchicine, and yohimbine (Fig. 1c). We found a relatively low response in ASM to colchicine which activates TAS2R4 (a mid-level ASM expressor by RT-PCR) and no response to salicin which exclusively activates TAS2R1610 (which was not detected in ASM by RT-PCR). The robust response to strychnine (activates TAS2R10 and −46) is also consistent with TAS2R10 having high expression in ASM. Thus in ASM, the [Ca2+]i response to bitter tastants is concordant with a rank-order based on agonist specificity and the bitter taste receptor subtype expression in these cells. Immunofluorescence microscopy of human ASM cells using polyclonal antisera directed against four receptors found to have mRNA expressed by RT-PCR (TAS2R10, −14, −31, −19) and three receptors whose mRNAs were not detected (TAS2R7, −38, −43), revealed cell surface expression of the former four receptors but not for the latter three (Supplementary Fig. 1). These studies also revealed expression of the α subunit of gustducin in these cells (Supplementary Fig. 1). No [Ca2+]i response to the sweet receptor agonists sucralose and SC45647 was found, and the saccharin response was not blocked by the sweet receptor antagonist lactisole (Supplementary Fig. 2).

Bottom Line: The relaxation induced by TAS2Rs is associated with a localized [Ca²(+)](i) response at the cell membrane, which opens large-conductance Ca²(+)-activated K(+) (BK(Ca)) channels, leading to ASM membrane hyperpolarization.Inhaled bitter tastants decreased airway obstruction in a mouse model of asthma.Given the need for efficacious bronchodilators for treating obstructive lung diseases, this pathway can be exploited for therapy with the thousands of known synthetic and naturally occurring bitter tastants.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, USA.

ABSTRACT
Bitter taste receptors (TAS2Rs) on the tongue probably evolved to evoke signals for avoiding ingestion of plant toxins. We found expression of TAS2Rs on human airway smooth muscle (ASM) and considered these to be avoidance receptors for inhalants that, when activated, lead to ASM contraction and bronchospasm. TAS2R agonists such as saccharin, chloroquine and denatonium evoked increased intracellular calcium ([Ca²(+)](i)) in ASM in a Gβγ-, phospholipase Cβ (PLCβ)- and inositol trisphosphate (IP₃) receptor-dependent manner, which would be expected to evoke contraction. Paradoxically, bitter tastants caused relaxation of isolated ASM and dilation of airways that was threefold greater than that elicited by β-adrenergic receptor agonists. The relaxation induced by TAS2Rs is associated with a localized [Ca²(+)](i) response at the cell membrane, which opens large-conductance Ca²(+)-activated K(+) (BK(Ca)) channels, leading to ASM membrane hyperpolarization. Inhaled bitter tastants decreased airway obstruction in a mouse model of asthma. Given the need for efficacious bronchodilators for treating obstructive lung diseases, this pathway can be exploited for therapy with the thousands of known synthetic and naturally occurring bitter tastants.

Show MeSH
Related in: MedlinePlus