Farncombe Family Digestive Health Research Institute, Department of Medicine, McMaster University, Hamilton, Ontario, Canada.
Am J Physiol Gastrointest Liver Physiol. 2011 Nov;301(5):G835-45. doi: 10.1152/ajpgi.00077.2011. Epub 2011 Aug 11.
Interstitial cells of Cajal associated with the myenteric plexus (ICC-MP) are pacemaker cells of the small intestine, producing the characteristic omnipresent electrical slow waves, which orchestrate peristaltic motor activity and are associated with rhythmic intracellular calcium oscillations. Our objective was to elucidate the origins of the calcium transients. We hypothesized that calcium oscillations in the ICC-MP are primarily regulated by the sarcoplasmic reticulum (SR) calcium release system. With the use of calcium imaging, study of the effect of T-type calcium channel blocker mibefradil revealed that T-type channels did not play a major role in generating the calcium transients. 2-Aminoethoxydiphenyl borate, an inositol 1,4,5 trisphosphate receptor (IP(3)R) inhibitor, and U73122, a phospholipase C inhibitor, both drastically decreased the frequency of calcium oscillations, suggesting a major role of IP(3) and IP(3)-induced calcium release from the SR. Immunohistochemistry proved the expression of IP(3)R type I (IP(3)R-I), but not type II (IP(3)R-II) and type III (IP(3)R-III) in ICC-MP, indicating the involvement of the IP(3)R-I subtype in calcium release from the SR. Cyclopiazonic acid, a SR/endoplasmic reticulum calcium ATPase pump inhibitor, strongly reduced or abolished calcium oscillations. The Na-Ca exchanger (NCX) in reverse mode is likely involved in refilling the SR because the NCX inhibitor KB-R7943 markedly reduced the frequency of calcium oscillations. Immunohistochemistry revealed 100% colocalization of NCX and c-Kit in ICC-MP. Testing a mitochondrial NCX inhibitor, we were unable to show an essential role for mitochondria in regulating calcium oscillations in the ICC-MP. In summary, ongoing IP(3) synthesis and IP(3)-induced calcium release from the SR, via the IP(3)R-I, are the major drivers of the calcium transients associated with ICC pacemaker activity. This suggests that a biochemical clock intrinsic to ICC determines the pacemaker frequency, which is likely directly linked to kinetics of the IP(3)-activated SR calcium channel and IP(3) metabolism.
与肌间神经丛相关的 Cajal 间质细胞（ICC-MP）是小肠的起搏细胞，产生特征性的普遍存在的电慢波，协调蠕动运动活动，并与节律性细胞内钙振荡相关。我们的目标是阐明钙瞬变的起源。我们假设 ICC-MP 中的钙振荡主要受肌浆网（SR）钙释放系统调节。通过钙成像研究 T 型钙通道阻滞剂米贝地尔的作用，发现 T 型通道在产生钙瞬变中不起主要作用。2-氨基乙氧基二苯硼酸盐，一种肌醇 1,4,5 三磷酸受体（IP3R）抑制剂，和 U73122，一种磷脂酶 C 抑制剂，都大大降低了钙振荡的频率，表明 IP3 和 IP3 诱导的钙从 SR 释放起主要作用。免疫组织化学证明了 IP3R 型 I（IP3R-I）的表达，但不是 IP3R 型 II（IP3R-II）和 III（IP3R-III）在 ICC-MP 中，表明 IP3R-I 亚基参与了钙从 SR 的释放。SR/内质网钙 ATP 酶泵抑制剂环匹阿尼酸强烈降低或消除钙振荡。SR 中的钙再填充可能涉及反向模式的 Na-Ca 交换体（NCX），因为 NCX 抑制剂 KB-R7943 显著降低了钙振荡的频率。免疫组织化学显示 NCX 和 c-Kit 在 ICC-MP 中 100%共定位。测试线粒体 NCX 抑制剂时，我们无法证明线粒体在调节 ICC-MP 中的钙振荡中起重要作用。总之，持续的 IP3 合成和 IP3 诱导的 SR 钙释放通过 IP3R-I 是与 ICC 起搏活动相关的钙瞬变的主要驱动因素。这表明 ICC 中的内在生化钟决定了起搏频率，这可能与 IP3 激活的 SR 钙通道和 IP3 代谢的动力学直接相关。
