Identification and spatiotemporal characterization of spontaneous Ca2+ sparks and global Ca2+ oscillations in retinal arteriolar smooth muscle cells.
OBJECTIVE
To identify spontaneous Ca(2+) sparks and global Ca(2+) oscillations in microvascular smooth muscle (MVSM) cells within intact retinal arterioles and to characterize their spatiotemporal properties and physiological functions.


METHODS
Retinal arterioles were mechanically dispersed from freshly isolated rat retinas and loaded with Fluo-4, a Ca(2+)-sensitive dye. Changes in [Ca(2+)](i) were imaged in MVSM cells in situ by confocal scanning laser microscopy in x-y mode or line-scan mode.


RESULTS
The x-y scans revealed discretely localized, spontaneous Ca(2+) events resembling Ca(2+) sparks and more global and prolonged Ca(2+) transients, which sometimes led to cell contraction. In line scans, Ca(2+) sparks were similar to those previously described in other types of smooth muscle, with an amplitude (DeltaF/F(0)) of 0.81 +/- 0.04 (mean +/- SE), full duration at half maximum (FDHM) of 23.62 +/- 1.15 ms, full width at half maximum (FWHM) of 1.25 +/- 0.05 mum, and frequency of 0.56 +/- 0.06 seconds(-1). Approximately 35% of sparks had a prolonged tail (>80 ms), similar to the Ca(2+)"embers" described in skeletal muscle. Sparks often summated to generate global and prolonged Ca(2+) elevations on which Ca(2+) sparks were superimposed. These sparks occurred more frequently (2.86 +/- 025 seconds(-1)) and spread farther across the cell (FWHM = 1.67 +/- 0.08 microm), but were smaller (DeltaF/F(0) = 0.69 +/- 0.04).


CONCLUSIONS
Retinal arterioles generate Ca(2+) sparks with characteristics that vary during different phases of the spontaneous Ca(2+)-signaling cycle. Sparks summate to produce sustained Ca(2+) transients associated with contraction and thus may play an important excitatory role in initiating vessel constriction. This deserves further study, not least because Ca(2+) sparks appear to inhibit contraction in many other smooth muscle cells.
Curtis
Tim M
TM
Ophthalmic Research Centre, The Queen's University of Belfast, Institute of Clinical Sciences, The Royal Victoria Hospital, Belfast, Northern Ireland.
Tumelty
James
J
Dawicki
Jennine
J
Scholfield
C Norman
CN
McGeown
J Graham
JG
D000818:Animals; D001160:Arterioles / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008297:Male; D018613:Microscopy, Confocal; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D009991:Oscillometry; D009994:Osmolar Concentration; D051381:Rats; D017207:Rats, Sprague-Dawley; D012171:Retinal Vessels / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D013997:Time Factors; D014018:Tissue Distribution; D014661:Vasoconstriction / Q000502:physiology
D002118:Calcium