Feedback via Ca²⁺-activated ion channels modulates endothelin 1 signaling in retinal arteriolar smooth muscle

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.