Baker Salah A, Drumm Bernard T, Saur Dieter, Hennig Grant W, Ward Sean M, Sanders Kenton M
Department of Physiology and Cell Biology, University of Nevada School of Medicine, Reno, NV, USA.
II. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar der TU München, München, Germany.
J Physiol. 2016 Jun 15;594(12):3317-38. doi: 10.1113/JP271699. Epub 2016 Mar 11.
Interstitial cells of Cajal at the level of the deep muscular plexus (ICC-DMP) in the small intestine generate spontaneous Ca(2+) transients that consist of localized Ca(2+) events and limited propagating Ca(2+) waves. Ca(2+) transients in ICC-DMP display variable characteristics: from discrete, highly localized Ca(2+) transients to regionalized Ca(2+) waves with variable rates of occurrence, amplitude, duration and spatial spread. Ca(2+) transients fired stochastically, with no cellular or multicellular rhythmic activity being observed. No correlation was found between the firing sites in adjacent cells. Ca(2+) transients in ICC-DMP are suppressed by the ongoing release of inhibitory neurotransmitter(s). Functional intracellular Ca(2+) stores are essential for spontaneous Ca(2+) transients, and the sarco/endoplasmic reticulum Ca(2+) -ATPase (SERCA) pump is necessary for maintenance of spontaneity. Ca(2+) release mechanisms involve both ryanodine receptors (RyRs) and inositol triphosphate receptors (InsP3 Rs). Release from these channels is interdependent. ICC express transcripts of multiple RyRs and InsP3 Rs, with Itpr1 and Ryr2 subtypes displaying the highest expression.
Interstitial cells of Cajal in the deep muscular plexus of the small intestine (ICC-DMP) are closely associated with varicosities of enteric motor neurons and generate responses contributing to neural regulation of intestinal motility. Responses of ICC-DMP are mediated by activation of Ca(2+) -activated Cl(-) channels; thus, Ca(2+) signalling is central to the behaviours of these cells. Confocal imaging was used to characterize the nature and mechanisms of Ca(2+) transients in ICC-DMP within intact jejunal muscles expressing a genetically encoded Ca(2+) indicator (GCaMP3) selectively in ICC. ICC-DMP displayed spontaneous Ca(2+) transients that ranged from discrete, localized events to waves that propagated over variable distances. The occurrence of Ca(2+) transients was highly variable, and it was determined that firing was stochastic in nature. Ca(2+) transients were tabulated in multiple cells within fields of view, and no correlation was found between the events in adjacent cells. TTX (1 μm) significantly increased the occurrence of Ca(2+) transients, suggesting that ICC-DMP contributes to the tonic inhibition conveyed by ongoing activity of inhibitory motor neurons. Ca(2+) transients were minimally affected after 12 min in Ca(2+) free solution, indicating these events do not depend immediately upon Ca(2+) influx. However, inhibitors of sarco/endoplasmic reticulum Ca(2+) -ATPase (SERCA) pump and blockers of inositol triphosphate receptor (InsP3 R) and ryanodine receptor (RyR) channels blocked ICC Ca(2+) transients. These data suggest an interdependence between RyR and InsP3 R in the generation of Ca(2+) transients. Itpr1 and Ryr2 were the dominant transcripts expressed by ICC. These findings provide the first high-resolution recording of the subcellular Ca(2+) dynamics that control the behaviour of ICC-DMP in situ.
小肠深层肌丛（ICC-DMP）中的Cajal间质细胞产生自发的Ca(2+)瞬变，其由局部Ca(2+)事件和有限传播的Ca(2+)波组成。ICC-DMP中的Ca(2+)瞬变表现出可变特征：从离散的、高度局部化的Ca(2+)瞬变到发生率、幅度、持续时间和空间传播可变的区域化Ca(2+)波。Ca(2+)瞬变随机发放，未观察到细胞或多细胞节律活动。相邻细胞的发放位点之间未发现相关性。ICC-DMP中的Ca(2+)瞬变受到抑制性神经递质持续释放的抑制。功能性细胞内Ca(2+)储存对于自发Ca(2+)瞬变至关重要，肌浆网/内质网Ca(2+) -ATP酶（SERCA）泵对于维持自发性是必需的。Ca(2+)释放机制涉及兰尼碱受体（RyRs）和三磷酸肌醇受体（InsP3 Rs）。从这些通道的释放是相互依赖的。ICC表达多种RyRs和InsP3 Rs的转录本，其中Itpr1和Ryr2亚型表达最高。
小肠深层肌丛中的Cajal间质细胞（ICC-DMP）与肠运动神经元的曲张体密切相关，并产生有助于肠道运动神经调节的反应。ICC-DMP的反应由Ca(2+)激活的Cl(-)通道激活介导；因此，Ca(2+)信号传导是这些细胞行为的核心。共聚焦成像用于表征在完整空肠肌肉中选择性在ICC中表达基因编码Ca(2+)指示剂（GCaMP3）的ICC-DMP中Ca(2+)瞬变的性质和机制。ICC-DMP表现出自发的Ca(2+)瞬变，范围从离散的局部事件到传播不同距离的波。Ca(2+)瞬变的发生率高度可变，并且确定发放本质上是随机的。在视野内的多个细胞中记录Ca(2+)瞬变，相邻细胞中的事件之间未发现相关性。TTX（1μm）显著增加Ca(2+)瞬变的发生率，表明ICC-DMP有助于抑制性运动神经元持续活动所传递的紧张性抑制。在无Ca(2+)溶液中12分钟后，Ca(2+)瞬变受到的影响最小，表明这些事件不立即依赖于Ca(2+)内流。然而，肌浆网/内质网Ca(2+) -ATP酶（SERCA）泵抑制剂以及三磷酸肌醇受体（InsP3 R）和兰尼碱受体（RyR）通道阻滞剂阻断了ICC的Ca(2+)瞬变。这些数据表明在Ca(2+)瞬变产生过程中RyR和InsP3 R之间存在相互依赖性。Itpr1和Ryr2是ICC表达的主要转录本。这些发现首次提供了控制原位ICC-DMP行为的亚细胞Ca(2+)动力学的高分辨率记录。
