Calcium entry via connexin hemichannels in lens epithelium. (March 2015)

• Record Type: Journal Article
• Authors: Mandal, Amritlal; Shahidullah, Mohammad; Delamere, Nicholas A.
• Is Part Of: Experimental eye research. Volume 132(2015:Mar.)
• Page Start: 52
• Page End: 58
• Publication Date: 2015-03
• Subjects: Calcium -- Connexin hemichannel -- Hyposmotic stress -- Lens epithelium -- Propidium iodide uptake; Ophthalmology -- Periodicals; Eye -- Periodicals; Œil -- Périodiques; Ophthalmology; Periodicals; Electronic journals; 612.8405
• DOI: 10.1016/j.exer.2015.01.012
• Languages: English
• ISSNs: 0014-4835

Abstract

Abstract: Exposure to hyposmotic solution causes release of ATP from lens cells via hemichannels. Because hemichannel opening feasibly could swamp the cells with calcium, we carried out studies to measure the magnitude of the increase in cytoplasmic calcium concentration caused by hemichannel opening. In studies on porcine lens epithelial cells in primary culture, propidium iodide (PI) uptake was measured as an index of hemichannel opening. PI uptake was increased significantly in cells exposed to hyposmotic solution. The PI increase under hyposmotic conditions was suppressed by GAP 27, a connexin inhibitor peptide. In studies on cells loaded with Fura-2, continuous exposure to hyposmotic solution caused a cytoplasmic calcium concentration increase that peaked within ∼30 s then remained elevated at or below the peak response for more than 60 min. The peak calcium concentration was 186 ± 2.3 nM compared to a baseline value of 98.0 ± 1.4 nM. The calcium concentration increased a lot further in cells exposed to A23187 (2.5 μM) or the sodium–calcium exchange inhibitor SN-6 (10 μM) added after the onset of the calcium rise in hyposmotic solution. The cytoplasmic calcium increase in hyposmotic solution was abolished by GAP 27. Calcium returned to baseline in cells exposed to hyposmotic solution then treated with GAP 27 starting 2 min after the onset of the calcium rise. The calcium increase in hyposmotic solution did not occur when calcium was eliminated from the bathing medium. The responses to hyposmotic and hyperosmotic stress were different. There was no detectable increase in calcium or PI entry in cells exposed to hyperosmotic solution (500 mOsm). In summary, GAP 27-sensitive accumulation of PI by cultured lens epithelium points to connexin hemichannel opening and associated calcium entry. Even though connexins form channels with a large carrying capacity, calcium entry does not increase the cytoplasmic calcium concentration beyond a tolerable physiological range. Graphical abstract: Exposure to hyposmotic solution was found to cause a sustained increase of calcium in lens epithelial cells. The ability of GAP 27, an inhibitory connexin mimetic peptide, to prevent the sustained calcium increase suggests it is the result of calcium entry via connexin hemichannels. Even though connexins form channels that can carry large solutes, the calcium response amounted to less than a doubling of the normal resting calcium concentration and calcium remains within a tolerable physiological range. Highlights: Sustained Ca 2+ rise in hyposmotic solution occurs via connexin hemichannel opening. Hemichannel opening increases [Ca 2+ ]i only to 186 nM from a baseline value of 97 nM. Although hemichannels open [Ca 2+ ]i keeps within a tolerable physiological range.