Visualization of elementary mechanosensitive Ca<sup>2+</sup>‐influx events, Ca<sup>2+</sup> spots, in bovine lens epithelial cells

Ohata, Hisayuki; Tanaka, Kenichi; Maeyama, Naoto; Yamamoto, Masayuki; Momose, Kazutaka

doi:10.1111/j.1469-7793.2001.0031g.x

Cited by 13 publications

(10 citation statements)

References 37 publications

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“…Fluorescence images of fluo 4 were collected by use of a real-time confocal imaging system as previously described. 26 With use of the imaging system, fluo 4 images with 60ϫ60 pixels with x-y resolution of 1.3 to 2 m per pixel were acquired with 55-to 110-ms exposure at 32°C. The intensity of fluo 4 fluorescence for each region of interest was divided by the averaged resting fluorescence intensity of Ͼ10 frames before stimulation of the same region, after which the background fluorescence was subtracted.…”

Section: Confocal Imaging Of [Ca 2؉ ] I With Fluomentioning

confidence: 99%

“…25 In addition, using real-time confocal microscopy, we recently demonstrated that LPA enhanced the mechanical stress-induced local increase in [Ca 2ϩ ] i , termed Ca 2ϩ spots, in cultured lens epithelial cells. 26 Considering those facts, we have proposed that the Ca 2ϩ spot is an elementary Ca 2ϩ -influx event through MS ion channels that is directly coupled with the first step in mechanoreception, and we have suggested that LPA functions as an endogenous factor affecting mechanotransduction systems.…”

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confidence: 99%

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Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells

Ohata

Ikeuchi

Kamada

et al. 2001

Circulation Research

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Section: Confocal Imaging Of [Ca 2؉ ] I With Fluomentioning

confidence: 99%

mentioning

confidence: 99%

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells

Ohata

Ikeuchi

Kamada

et al. 2001

Circulation Research

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“…As presented in Fig. 3B, the spatiotemporal properties were similar to those of Ca 2+ spots observed in lens epithelial cells (16). In addition, the pharmacological properties of the local increase in [Ca 2+ ]i showed that the response was due to Ca 2+ influx through MS channels (13) …”

Section: +mentioning

confidence: 51%

“…This mechanical stress did not affect acquisition of fluorescent images and induced a reproducible response; however, a combination of both flow and pressure stress may have been involved. Estimated intensities of the shear and the pressure stresses occurred within physiological range (16 ]i in the presence of LPA in ileum smooth muscle cells (7), lung epithelial cells (10) and lens epithelial cells (11,16) (Fig. 1A).…”

Section: +mentioning

confidence: 99%

Physiological and Pharmacological Role of Lysophosphatidic Acid as Modulator in Mechanotransduction

Ohata

Tanaka

Maeyama

et al. 2001

Japanese Journal of Pharmacology

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ABSTRACT-The mechanotransduction mechanism is believed to play an important role in maintenance of cellular homeostasis in a wide variety of cell types. In particular, the mechanotransduction system in vascular endothelial cells may be an essential mechanism for local hemodynamic control. Elevations in intracellular free Ca]i) are an important signal in the initial step of mechanotransduction and mechanosensitive (MS) cation channels are thought to be a putative pathway; however, the molecular mechanisms remain unclear. We found that lysophosphatidic acid (LPA), a bioactive phospholipid, sensitizes the response of [Ca 2+ ]i to mechanical stress in several cell types. Employing real-time confocal microscopy, local increases in [Ca 2+ ]i in several regions within the cell during application of mechanical stress were clearly visualized in bovine lens epithelial and endothelial cells in the presence of LPA. The phenomenon was termed "Ca 2+ spots". Pharmacological studies revealed that Ca 2+ spots arise due to influx through MS channels. In this report, our data indicating the possible significance of LPA as an endogenous factor involved in regulation of mechanotransduction is reviewed. Furthermore, our findings suggest that the Ca 2+ spot is a novel phenomenon occurring as an elementary Ca 2+-influx event through MS channels directly coupled with the initial step in mechanotransduction.

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“…-influx event through mechanosensitive channels directly coupled with the initial step in mechanotransduction in cultured endothelial (4 -6) and cultured lens epithelial cells (6,7). The Ca 2+ spots, which develop sporadically, exhibit a spatiotemporal pattern distinct from Ca 2+ sparks, the elementary [Ca…”

Section: +mentioning

confidence: 99%

Optical Bioimaging: From Living Tissue to a Single Molecule: Calcium Imaging in Blood Vessel In Situ Employing Two-Photon Excitation Fluorescence Microscopy

et al. 2003

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Abstract. Recent developments in optoelectronics permit real-time Ca2+ imaging of thin planes within cells utilizing laser scanning confocal microscopy (LSCM). However, a major complication associated with this imaging system involves increased phototoxicity with improved spatiotemporal resolution. Two-photon excitation microscopy (TPEM) helps to minimize phototoxicity due to the restriction of this technique to the volume proximal to the geometric focus of the light. In this study, the capability of Ca 2+ imaging was investigated employing recently developed real-time TPEM, RTS2000MP (Bio-Rad, Tokyo) with a mode-locked Ti-sapphire laser. Z-axis resolution of RTS2000MP with high NA objectives defined as full-width at half maximum (FWHM) with a 0.5-mm fluorescent bead provided values nearly identical to those obtained with LSCM at a small pinhole (0.2 mm) (approximately 0.6 mm). When serial sectioning of 21 sequential images at 0.3-mm intervals in cultured endothelial cells loaded with calcein and tetramethyl-rhodamine methylester were performed with TPEM, the z-axis resolution was higher than that observed with LSCM; moreover, the photobleaching rate was significantly lower than that obtained with LSCM. Maximum fluorescence intensities were detected at 780 nm in excitation spectra of fluo-3 and fluo-4 Ca ] i of endothelial cells; subsequently, relaxation along the major axis of smooth muscle cells was evident. Furthermore, consecutive long-lasting experiments could be repeated with identical response in the same microscopic field. In conclusion, fluorescence imaging employing TPEM is useful for Ca 2+ imaging in blood vessels in situ.

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Visualization of elementary mechanosensitive Ca²⁺‐influx events, Ca²⁺ spots, in bovine lens epithelial cells

Cited by 13 publications

References 37 publications

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells

Physiological and Pharmacological Role of Lysophosphatidic Acid as Modulator in Mechanotransduction

Optical Bioimaging: From Living Tissue to a Single Molecule: Calcium Imaging in Blood Vessel In Situ Employing Two-Photon Excitation Fluorescence Microscopy

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Visualization of elementary mechanosensitive Ca2+‐influx events, Ca2+ spots, in bovine lens epithelial cells

Cited by 13 publications

References 37 publications

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca 2+ Influx in Bovine Aortic Endothelial Cells

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca 2+ Influx in Bovine Aortic Endothelial Cells

Physiological and Pharmacological Role of Lysophosphatidic Acid as Modulator in Mechanotransduction

Optical Bioimaging: From Living Tissue to a Single Molecule: Calcium Imaging in Blood Vessel In Situ Employing Two-Photon Excitation Fluorescence Microscopy

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Visualization of elementary mechanosensitive Ca²⁺‐influx events, Ca²⁺ spots, in bovine lens epithelial cells

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells

Lysophosphatidic Acid Positively Regulates the Fluid Flow–Induced Local Ca ²⁺ Influx in Bovine Aortic Endothelial Cells