Ca 2+ signalling in mouse urethral smooth muscle in situ : role of Ca 2+ stores and Ca 2+ influx mechanisms

The Journal of Physiology

Hwang

Baker

et al. 2019

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Key points Colonic intramuscular interstitial cells of Cajal (ICC‐IM) exhibit spontaneous Ca2+ transients manifesting as stochastic events from multiple firing sites with propagating Ca2+ waves occasionally observed. Firing of Ca2+ transients in ICC‐IM is not coordinated with adjacent ICC‐IM in a field of view or even with events from other firing sites within a single cell. Ca2+ transients, through activation of Ano1 channels and generation of inward current, cause net depolarization of colonic muscles. Ca2+ transients in ICC‐IM rely on Ca2+ release from the endoplasmic reticulum via IP3 receptors, spatial amplification from RyRs and ongoing refilling of ER via the sarcoplasmic/endoplasmic‐reticulum‐Ca2+‐ATPase. ICC‐IM are sustained by voltage‐independent Ca2+ influx via store‐operated Ca2+ entry. Some of the properties of Ca2+ in ICC‐IM in the colon are similar to the behaviour of ICC located in the deep muscular plexus region of the small intestine, suggesting there are functional similarities between these classes of ICC. Abstract A component of the SIP syncytium that regulates smooth muscle excitability in the colon is the intramuscular class of interstitial cells of Cajal (ICC‐IM). All classes of ICC (including ICC‐IM) express Ca2+‐activated Cl− channels, encoded by Ano1, and rely upon this conductance for physiological functions. Thus, Ca2+ handling in ICC is fundamental to colonic motility. We examined Ca2+ handling mechanisms in ICC‐IM of murine proximal colon expressing GCaMP6f in ICC. Several Ca2+ firing sites were detected in each cell. While individual sites displayed rhythmic Ca2+ events, the overall pattern of Ca2+ transients was stochastic. No correlation was found between discrete Ca2+ firing sites in the same cell or in adjacent cells. Ca2+ transients in some cells initiated Ca2+ waves that spread along the cell at ∼100 µm s−1. Ca2+ transients were caused by release from intracellular stores, but depended strongly on store‐operated Ca2+ entry mechanisms. ICC Ca2+ transient firing regulated the resting membrane potential of colonic tissues as a specific Ano1 antagonist hyperpolarized colonic muscles by ∼10 mV. Ca2+ transient firing was independent of membrane potential and not affected by blockade of L‐ or T‐type Ca2+ channels. Mechanisms regulating Ca2+ transients in the proximal colon displayed both similarities to and differences from the intramuscular type of ICC in the small intestine. Similarities and differences in Ca2+ release patterns might determine how ICC respond to neurotransmission in these two regions of the gastrointestinal tract.

“…; Drumm et al . ). In some cells, Ca 2+ firing sites are believed to be due to the presence of a high density clusters of RyRs at these locations (Gordienko et al .…”

Section: Discussionmentioning

confidence: 97%

“…Photonics, Gräfelfing, Germany), as previously described (Drumm et al . , b ). The pixel size using this acquisition configuration was 0.225 µm.…”

Section: Methodsmentioning

confidence: 99%

Ca²⁺ signalling behaviours of intramuscular interstitial cells of Cajal in the murine colon

The Journal of Physiology

Hwang

Baker

et al. 2019

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“…While the bladder detrusor and internal urethral sphincter are both composed of smooth muscle cells and share common fundamental physiological traits [contractions are Ca 2ϩ dependent and can be affected by depolarization (10,11)], they receive differential neural input, which affects contractility. Detrusor smooth muscle (DSM) contraction occurs due to stimulation of the parasympathetic nervous system and subsequent binding of ACh to muscarinic receptors (3,4,14).…”

Section: Introductionmentioning

confidence: 99%

“…This action causes large contractions of the DSM, resulting in bladder emptying. In contrast, cholinergic innervation is sparse in USM (2,10), and USM is contracted by stimulation of the sympathetic nervous system via activation of ␣ 1 -adrenoceptors (1,8,10,12,15,20). In this laboratory practical, students will observe these differential effects on mammalian DSM and USM by performing recordings of contraction of bladder and urethra muscle tissues.…”

Section: Introductionmentioning

confidence: 99%

Laboratory practical to study the differential innervation pathways of urinary tract smooth muscle

Rembetski

Cobine

Advances in Physiology Education

2018

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In the mammalian lower urinary tract, there is a reciprocal relationship between the contractile state of the bladder and urethra. As the bladder fills with urine, it remains relaxed to accommodate increases in volume, while the urethra remains contracted to prevent leakage of urine from the bladder to the exterior. Disruptions to the normal contractile state of the bladder and urethra can lead to abnormal micturition patterns and urinary incontinence. While both the bladder and urethra are smooth-muscle organs, they are differentially contracted by input from cholinergic and sympathetic nerves, respectively. The laboratory practical described here provides an experiential approach to understanding the anatomy of the lower urinary tract. Several key factors in urinary tract physiology are outlined, e.g., the bladder is contracted by activation of the parasympathetic pathway via cholinergic stimulation on muscarinic receptors, whereas the urethra is contracted by activation of the sympathetic pathway via adrenergic stimulation on α-adrenoceptors. This is achieved by measuring the force generated by bladder and urethra smooth muscle to demonstrate that acetylcholine contracts the smooth muscle of the bladder, whereas adrenergic agonists contract the urethral smooth muscle. An inhibition of these effects is also demonstrated by application of the muscarinic receptor antagonist atropine and the α-adrenergic receptor blocker phentolamine. A list of suggested techniques and exam questions to evaluate student understanding on this topic is also provided.

“…As reported previously, imaging GCaMP for 20–30 s of consecutive recordings did not lead to a decrease in Ca 2+ transients (Drumm et al . ).…”

Section: Methodsmentioning

confidence: 97%

Pacemaker function and neural responsiveness of subserosal interstitial cells of Cajal in the mouse colon

The Journal of Physiology

Rembetski

Messersmith

et al. 2020

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Key pointsr Rhythmic action potentials and intercellular Ca 2+ waves are generated in smooth muscle cells of colonic longitudinal muscles (LSMC).r Longitudinal muscle excitability is tuned by input from subserosal ICC (ICC-SS), a population of ICC with previously unknown function.r ICC-SS express Ano1 channels and generate spontaneous Ca 2+ transients in a stochastic manner.r Release of Ca 2+ and activation of Ano1 channels causes depolarization of ICC-SS and LSMC, leading to activation of L-type Ca 2+ channels, action potentials, intercellular Ca 2+ waves and contractions in LSMC.r Nitrergic neural inputs regulate the Ca 2+ events in ICC-SS. r Pacemaker activity in longitudinal muscle is an emergent property as a result of integrated processes in ICC-SS and LSMC.Abstract Much is known about myogenic mechanisms in circular muscle (CM) in the gastrointestinal tract, although less is known about longitudinal muscle (LM). Two Ca 2+ signalling behaviours occur in LM: localized intracellular waves not causing contractions and intercellular waves leading to excitation-contraction coupling. An Ano1 channel antagonist inhibited intercellular Ca 2+ waves and LM contractions. Ano1 channels are expressed by interstitial cells of Cajal (ICC) but not by smooth muscle cells (SMCs). We investigated Ca 2+ signalling in a novel population of ICC that lies along the subserosal surface of LM (ICC-SS) in mice expressing GCaMP6f in ICC. ICC-SS fired stochastic localized Ca 2+ transients. Such events have been linked to activation of Ano1 channels in ICC. Ca 2+ transients in ICC-SS occurred by release from stores most probably via inositol trisphosphate receptors. This activity relied on influx via store-operated Bernard T. Drumm received his PhD in Physiology in 2013. His research investigates how Ca 2+ signalling impacts the functions of visceral smooth muscle organs in the gastrointestinal and urinary tracts. His doctoral work, studying Ca 2+ wave propagation in urethral cells, was followed by a postdoctoral position and later research track faculty position at the University of Nevada, Reno (UNR), where he studied the regulation of pacemaker and neuroeffector mechanisms in gastrointestinal interstitial cells. Dr Drumm took up a position as Lecturer in the