Department of Physiology and Biophysics, Virginia Commonwealth University, Richmond, VA, United States of America.
NYU College of Dentistry, New York, NY, United States of America.
PLoS One. 2018 Mar 7;13(3):e0194089. doi: 10.1371/journal.pone.0194089. eCollection 2018.
In rodents, CHRNs are involved in bitter taste transduction of nicotine and ethanol. Currently, it is not clear if CHRNs are expressed in human taste cells and if they play a role in transducing the bitter taste of nicotine and ethanol or in the synthesis and release of neurohumoral peptides. Accordingly, we investigated the expression and functional role of CHRNs in HBO cells. Using molecular techniques, we demonstrate that a subset of HBO cells express CHRNs that also co-express TRPM5, T1R3 or T2R38. Exposing HBO cells to nicotine or ethanol acutely or to nicotine chronically induced a differential increase in the expression of CHRN mRNA and protein in a dose- and time-dependent manner. Acutely exposing HBO cells to a mixture containing nicotine plus ethanol induced a smaller increase in CHRN mRNAs relative to nicotine or ethanol treatment alone. A subset of HBO cells responded to nicotine, acetylcholine and ATP with a transient increase in [Ca2+]i. Nicotine effects on [Ca2+]i were mecamylamine sensitive. Brain-derived neurotrophic factor (BDNF) protein was detected in HBO cells using ELISA. Acute nicotine exposure decreased BDNF in HBO cells and increased BDNF release in the medium. CHRNs were also detected in HEK293 cells by RT-PCR. Unlike HBO cells, CHRNs were localized in most of HEK293 cells and majority of HEK293 cells responded to nicotine and ethanol stimulation with a transient increase in [Ca2+]i. BDNF levels in HEK293 cells were significantly higher than in HBO cells but the nicotine induced release of BDNF in the media was a fraction of the BDNF cellular content. We conclude that CHRNs are expressed in TRPM5 positive HBO cells. CHRN mRNA expression is modulated by exposure to nicotine and ethanol in a dose- and time-dependent manner. Nicotine induces the synthesis and release of BDNF in HBO cells.
在啮齿动物中,CHRN 参与了尼古丁和乙醇的苦味味觉转导。目前尚不清楚 CHRN 是否在人味觉细胞中表达,以及它们是否在转导尼古丁和乙醇的苦味味觉或神经激素肽的合成和释放中发挥作用。因此,我们研究了 CHRN 在 HBO 细胞中的表达和功能作用。使用分子技术,我们证明了一组 HBO 细胞表达 CHRN,这些细胞也共同表达 TRPM5、T1R3 或 T2R38。急性暴露于尼古丁或乙醇或慢性暴露于尼古丁会导致 CHRN mRNA 和蛋白的表达以剂量和时间依赖的方式产生差异增加。急性将包含尼古丁加乙醇的混合物暴露于 HBO 细胞会导致相对于单独的尼古丁或乙醇处理,CHRN mRNA 的增加较小。HBO 细胞的一部分对尼古丁、乙酰胆碱和 ATP 做出反应,导致 [Ca2+]i 短暂增加。尼古丁对 [Ca2+]i 的作用是美卡拉明敏感的。使用 ELISA 在 HBO 细胞中检测到脑源性神经营养因子 (BDNF) 蛋白。急性尼古丁暴露降低了 HBO 细胞中的 BDNF,并增加了培养基中的 BDNF 释放。通过 RT-PCR 也在 HEK293 细胞中检测到 CHRN。与 HBO 细胞不同,CHRN 定位于大多数 HEK293 细胞中,大多数 HEK293 细胞对尼古丁和乙醇刺激的反应是 [Ca2+]i 的短暂增加。HEK293 细胞中的 BDNF 水平明显高于 HBO 细胞,但尼古丁诱导的培养基中 BDNF 的释放仅为 BDNF 细胞内含量的一小部分。我们得出结论,CHRN 在表达 TRPM5 的 HBO 细胞中表达。CHRN mRNA 表达受尼古丁和乙醇暴露的剂量和时间依赖性调节。尼古丁诱导 HBO 细胞中 BDNF 的合成和释放。