Abstract
In addition to the T2R bitter taste receptors, neuronal nicotinic acetylcholine receptors (nAChRs) have recently been shown to be involved in the bitter taste transduction of nicotine, acetylcholine and ethanol. However, at present it is not clear if nAChRs are expressed in enteroendocrine cells other than beta cells of the pancreas and enterochromaffin cells, and if they play a role in the synthesis and release of neurohumoral peptides. Accordingly, we investigated the expression and functional role of nAChRs in enteroendocrine STC-1 cells. Our studies using RT-PCR, qRT-PCR, immunohistochemical and Western blotting techniques demonstrate that STC-1 cells express several α and β nAChR subunits. Exposing STC-1 cells to nicotine acutely (24h) or chronically (4 days) induced a differential increase in the expression of nAChR subunit mRNA and protein in a dose- and time-dependent fashion. Mecamylamine, a non-selective antagonist of nAChRs, inhibited the nicotine-induced increase in mRNA expression of nAChRs. Exposing STC-1 cells to nicotine increased intracellular Ca2+ in a dose-dependent manner that was inhibited in the presence of mecamylamine or dihydro-β-erythroidine, a α4β2 nAChR antagonist. Brain-derived neurotrophic factor (BDNF) mRNA and protein were detected in STC-1 cells using RT-PCR, specific BDNF antibody, and enzyme-linked immunosorbent assay. Acute nicotine exposure (30 min) decreased the cellular content of BDNF in STC-1 cells. The nicotine-induced decrease in BDNF was inhibited in the presence of mecamylamine. We also detected α3 and β4 mRNA in intestinal mucosal cells and α3 protein expression in intestinal enteroendocrine cells. We conclude that STC-1 cells and intestinal enteroendocrine cells express nAChRs. In STC-1 cells nAChR expression is modulated by exposure to nicotine in a dose- and time-dependent manner. Nicotine interacts with nAChRs and inhibits BDNF expression in STC-1 cells.
Highlights
Our sense of taste helps us to determine whether the food is nutritious and should be ingested or is potentially toxic and should be rejected [1]
The mouse primers used to detect the presence of mRNAs for the nicotinic acetylcholine receptors (nAChRs) subunits, TRPM5, αENaC, TRPV1, GAPDH, Brain-derived neurotrophic factor (BDNF) and β-actin are shown in Table 1 and were synthesized by Thermo Fisher Scientific
The mRNAs for chrna3, chrna4, chrna5, chrna6, chrna7, chrnb2, and chrnb4 nAChR subunits were detected from RNA isolated from STC-1 cells by RT-PCR (Fig 1A)
Summary
Our sense of taste helps us to determine whether the food is nutritious and should be ingested or is potentially toxic and should be rejected [1]. Taste contributes to palatability [2], satiation, thermogenic effects [3] and the reward value of food [4]. A distinct subset of taste receptor cells (TRCs) in the taste buds on the tongue detect taste stimuli representing the five primary taste qualities salty, sour, sweet, bitter, and umami [5]. Sweet and umami taste is detected by TRCs that express G-protein coupled taste receptors (GPCRs; T1Rs and T2Rs), PLCβ2 and TRPM5. Salty taste is perceived by TRCs that express the amiloride- and Bz-sensitive epithelial Na+ channel (ENaC) [6,7,8]. Sour taste is perceived by TRCs that express PKD2L1 channels, carbonic anhydrase-4 [9, 10] and Zn2+-sensitive proton channels [8, 11, 12]
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