Abstract：
:Using the low-affinity fluorescent Ca(2+) indicators, Mag-Fura-2 and Mag-Fura Red, we studied light- and InsP(3)-induced Ca(2+) release in permeabilized microvillar photoreceptors of the medicinal leech, Hirudo medicinalis. Two major components of the phosphoinositide signaling pathway, phospholipase-C and the InsP(3) receptor, were characterized immunologically and appropriately localized in photoreceptors. Whereas phospholipase-C was abudantly expressed in photoreceptive microvilli, InsP(3) receptors were found mostly in submicrovillar endoplasmic reticulum (SER). Permeabilization of the peripheral plasma membrane with saponin allowed direct measurements of luminal free Ca(2+) concentration (Ca(L)) changes. Confocal Ca(2+) imaging using Mag-Fura Red demonstrated that Ins(1,4,5)P(3) mobilizes Ca(2+) from SER. As detected with Mag-Fura-2, a brief 50ms light flash activated rapid Ca(2+) depletion of SER, followed by an effective refilling within 1min of dark adaptation after the light flash. Sensitivity to Ins(1,4,5)P(3) of the Ca(2+) release from SER in leech photoreceptors was accompanied by irreversible uncoupling of phototransduction from Ca(2+) release. Depletion of Ca(2+) stores was induced by Ins(1,4,5)P(3)(EC(50)= 4.75 microM) and the hyper-potent agonist adenophostin A (EC(50)/40nM) while the stereoisomer L-myo Ins(1,4,5)P(3) was totally inactive. Ins(1,4,5)P(3)- or adenophostin A-induced Ca(2+) release was inhibited by 0.1-1mg/ml heparin. The Ca(2+) pump inhibitors, cyclopiazonic acid and thapsigargin, in the presence of Ins(1,4,5)P(3), completely depleted Ca(2+) stores in leech photoreceptors.
journal_name
Cell Calciumjournal_title
Cell calciumauthors
Ukhanov K,Mills SJ,Potter BV,Walz Bdoi
10.1054/ceca.2001.0195keywords:
subject
Has Abstractpub_date
2001-05-01 00:00:00pages
335-45issue
5eissn
0143-4160issn
1532-1991pii
S0143-4160(01)90195-2journal_volume
29pub_type
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