Feedback via Ca2+-Activated Ion Channels Modulates Endothelin 1 Signaling in Retinal Arteriolar Smooth Muscle

PURPOSE. To investigate the role of feedback by Ca2+-sensitive plasma-membrane ion channels in endothelin 1 (Et1) signaling in vitro and in vivo. METHODS. Et1 responses were imaged from Fluo-4-loaded smooth muscle in isolated segments of rat retinal arteriole using two-dimensional (2-D) confocal laser microscopy. Vasoconstrictor responses to intravitreal injections of Et1 were recorded in the absence and presence of appropriate ion channel blockers using fluorescein angiograms imaged using a confocal scanning laser ophthalmoscope. RESULTS. Et1 (10 nM) increased both basal [Ca2+](i) and the amplitude and frequency of Ca2+-waves in retinal arterioles. The Ca2+-activated Cl--channel blockers DIDS and 9-anthracene carboxylic acid (9AC) blocked Et1-induced increases in wave frequency, and 9AC also inhibited the increase in amplitude. Iberiotoxin, an inhibitor of large conductance (BK) Ca2+-activated K+-channels, increased wave amplitude in the presence of Et1 but had no effect on frequency. None of these drugs affected basal [Ca2+](i). The voltage-operated Ca2+-channel inhibitor nimodipine inhibited wave frequency and amplitude and also lowered basal [Ca2+] i in the presence of Et1. Intravitreal injection of Et1 caused retinal arteriolar vasoconstriction. This was inhibited by DIDS but not by iberiotoxin or penitrem A, another BK-channel inhibitor. CONCLUSIONS. Et1 evokes increases in the frequency of arteriolar Ca2+-waves in vitro, resulting in vasoconstriction in vivo. These responses, initiated by release of stored Ca2+, also require positive feedback via Ca2+-activated Cl--channels and L-type Ca2+-channels. (Invest Ophthalmol Vis Sci. 2012;53:30593066) DOI:10.1167/iovs.11-9192