Research in taste disorder and development of taste modulators

酸敏感离子通道-2a(ASIC2a)是一种可被阿米洛利阻断的质子门控阳离子通道，可能与大鼠味觉细胞的酸味检测有关。为了分离酸味受体的另一种亚型，我们筛选了大鼠环状乳头的cDNA文库，并鉴定了ASIC2b，它是ASIC2a的N端剪接变体。逆转录-聚合酶链式反应证实了ASIC2b在环状乳头中的表达，并证明了它在叶状乳头和菌状乳头中的表达。免疫组织化学分析表明，ASIC2b和ASIC2a在环状、叶状和菌状乳头中的味觉细胞亚群中都有表达，其中一些细胞同时显示ASIC2a和ASIC2b免疫反应。随后与环状乳头提取物的免疫共沉淀研究表明，ASIC2b与ASIC2a结合形成集合，结合我们的免疫组织化学结果，强烈表明这两个ASIC2亚单位在环状、叶状和菌状乳头中的味觉细胞中形成了异构体通道。卵母细胞电生理学研究表明，ASIC2a/ASIC2b通道在pH 2.0时产生最大的反向电流，这与体内大鼠味觉细胞对pH的敏感性一致；异构体对阿米洛利的敏感性随pH的降低而降低，在pH 2.0时几乎完全消失。这些发现为阿米洛利对酸诱导的大鼠味觉细胞反应不敏感提供了有说服力的解释。

Acid-sensing ion channel-2a (ASIC2a) is an amiloride-blockable proton-gated cation channel, probably contributing to sour-taste detection in rat taste cells. To isolate another subtype of the sour-taste receptor, we screened a rat circumvallate papilla cDNA library and identified ASIC2b, an N-terminal splice variant of ASIC2a, Reverse transcription-polymerase chain reaction analyses confirmed the expression of ASIC2b transcripts in the circumballate papilla and, moreover, demonstrated its expression in the foliate and fungiform papillae. Immunohistochemical analyses revealed that ASIC2b, as well as ASIC2a, was expressed in a subpopulation of taste cells in the circumvallate, foliate, and fungiform papillae, and some of the cells displayed both ASIC2a-and ASIC2b-immunoreactivities. Subsequent co-immunoprecipitation studies with circumvallate papillae extracts indicated that ASIC2b associated with ASIC2a to form assemblies, and, together with our immunohistochemical findings, strongly suggested that both ASIC2 subunits formed heteromeric channels in taste cells in the circumvallate, foliate, and fungiform papillae. Oocyte electrophysiology demonstrated that the ASIC2a/ASIC2b channel generated maximal in ward currents at pH 2.0, which is in good agreement with the in vivo pH-sensitivity of rat taste cells, and that the amiloride-sensitivity of the heteromer decreased with decreasing pH and was almost completely abolished at pH 2.0. These findings provide persuasive explanations for the amiloride-insensitivity of acid-induced responses of rat taste cells.