Role of the Calcium Sensing Receptor in Meal Stimulated Gastrin Secretion

摄入的营养物质的分子识别导致胃肠激素的分泌，调节消化和肠道运动，这些营养物质的吸收是必要的。已知氨基酸、短链脂肪酸、碳水化合物和管腔内pH的变化是胃泌素、胆囊收缩素和胰泌素的有效刺激物，这些是调节消化、分泌和运动的一些最重要的胃肠激素。 推测负责这些营养物质的化学感受的细胞沿着衬在胃肠道的粘膜层分散，并且可能是肠神经系统内的神经元。这些化学感受细胞固有的分散性质和缺乏任何已知的标记物来识别它们，为解决营养识别的分子基础带来了困难。 作用于胃G细胞上的钙敏感受体的钙已显示刺激胃泌素的分泌。 CaSR已被证明是一个多模态传感器的氨基酸和pH值，除了二价阳离子。 因此，我们假设胃窦G细胞上表达的胃钙敏感受体（CaSR）是生理相关的传感器，介导蛋白胨、氨基酸（尤其是芳香族氨基酸）和pH刺激的体内胃泌素释放。 本工作利用Martin R. Pollak（JCI 111：1021-1028，2003）。 在CaSR基因或WT C57/B6小鼠的同窝纯合缺失（KO）（CaSR- /- /PTH- / -）、杂合（HET）（CaSR + /- /PTH- / -）和野生型（WT）（CaSR+ /+/PTH- / -）小鼠中评估血浆胃泌素对蛋白胨、苯丙氨酸（Phe）和pH的反应。 将小鼠禁食过夜，并随意喂食或以推注或肠胃外卡巴可（5 mg）或铃蟾肽（50 μ g/kg）的形式管饲蛋白胨（8%）、Phe（100 mM）或Hepes（150 mM，pH 7.0），并通过RIA测量每种试剂的峰值胃泌素反应。 测量响应于胃泌素-17（lmg/kg）、组胺（10 mg/kg）或卡巴胆碱（5 mg）的胃酸分泌。 使用兔抗胃泌素对来自胃的冷冻切片进行苏木精和伊红染色和免疫组织化学。 我们发现，CaSR缺陷的小鼠不分泌胃泌素在响应管腔内的刺激与钙，蛋白胨，L-苯丙氨酸和pH值升高。然而，CaSR缺陷的小鼠响应肠外蛙皮素和卡巴可WT小鼠相似。杂合子和WT同窝小鼠中自由采食和灌胃喂养的胃泌素反应与WT C57/B6对照小鼠中观察到的相似。高Ca++饮食校正的血浆Ca++在CaSR缺陷小鼠没有恢复他们的胃泌素反应。CaSR缺陷小鼠胃窦G细胞数量和胃泌素含量减少。他们也有一个静息胃pH值升高和基础酸分泌减少，虽然他们表现出正常的胃酸分泌反应外源性胃泌素和组胺。 从这些研究中，我们了解到胃CaSR对于管腔内营养物质如大鼠食物、蛋白胨、Phe和升高的pH的生理感测是必需的，这导致胃泌素从胃窦G细胞的刺激释放。胃窦G细胞上的CaSRs是否直接介导这些刺激物的感受，有待于使用分离的G细胞进行进一步的研究。 此外，我们发现了一个意想不到的作用，CaSR在确定胃窦G细胞的数量。

Molecular recognition of ingested nutrients results in the secretion of gastrointestinal hormones regulating digestion and gut motility necessary for absorption of these nutrients. Amino acids, short chain fatty acids, carbohydrates and changes in intraluminal pH are known potent stimulators of gastrin, cholecystokinin and secretin, some of the most important gastrointestinal hormones regulating digestion, secretion and motility. The cells responsible for chemosensation of these nutrients are presumed to be scattered along the mucosal layer lining the gastrointestinal tract and may possibly be neurons within the enteric nervous system. The inherent dispersed nature of these chemosensory cells and the lack of any known markers for their identification create a difficult problem for solving the molecular basis of nutrient recognition. Calcium acting at the calcium sensing receptor on gastric G cells has been shown to stimulate the secretion of gastrin. The CaSR has been shown to be a multimodal sensor for amino aicds and pH in addition to divalent cations. Therefore, we hypothesised that the gastric calcium sensing receptor (CaSR) expressed on antral G cells is the physiologically relevant sensor mediating peptone, amino acids (especially aromatic amino acids) and pH stimulated gastrin release in vivo. This work utilized CaSR and parathyroid hormone (PTH) gene deleted mice provided by Martin R. Pollak (JCI 111:1021-1028, 2003). Plasma gastrin response to peptone, phenylalanine (Phe) and pH was assessed in littermates homozygous deleted (KO) (CaSR- /- /PTH- / -), heterozygous (HET) (CaSR+ /- /PTH- / -) and wild type (WT) (CaSR+ /+/PTH- / -) for the CaSR gene or WT C57/B6 mice. Mice were fasted overnight and either fed ad libitum or gavaged with either peptone (8%), Phe (100 mM), or Hepes (150 mM, pH 7.0) as a bolus or parenteral carbacol (5 mg) or bombesin (50 ug/kg) and the peak gastrin response for each agent was measured by RIA. Gastric acid secretion were measured in response to gastrin -17 (1 mg/kg), histamine (10 mg/kg) or carbachol (5 mg). Hematoxylin and eosin staining and immunohistochemistry was performed on frozen sections from the stomach using rabbit anti-gastrin. We found that CaSR deficient mice do not secrete gastrin in response to intraluminal stimulation with calcium, peptone, L-phenylalanine and elevated pH. However, CaSR deficient mice responded to parenteral bombesin and carbacol similar to WT mice. The gastrin response for ad libitum and gavage feeding in the heterozygous and WT littermates was similar to that observed for WT C57/B6 control mice. High Ca++ dietary correction of plasma Ca++ in the CaSR deficient mice did not restore their gastrin response. CaSR deficient mice expressed a decreased number of antral G cells and gastrin content. They also had an elevated resting gastric pH and reduced basal acid secretion, although they displayed a normal gastric acid secretory response to exogenous gastrin and histamine. From these studies we learned that gastric CaSRs are necessary for the physiological sensing of intraluminal nutrients such as rat chow, peptone, Phe and elevated pH that results in the stimulated release of gastrin from antral G cells. Whether the sensing of these stimulants is directly mediated by CaSRs on gastric antral G cells awaits further studies using isolated G cells. In addition, we found an unexpected role of CaSR in determining antral G cell number.