Measuring Connexin Hemichannel Opening in Response to an InsP3-Mediated Cytosolic Ca < sup > 2+ < /sup > Increase

Methods Mol Biol. 2024;2801:189-197. doi: 10.1007/978-1-0716-3842-2_14.ABSTRACTThe opening of connexin hemichannels (HCs) expressed at the plasma membrane of mammalian cells is regulated by a number of physiological parameters, including extracellular and intracellular Ca2+ ions. Submicromolar variations of the cytosolic Ca2+ concentration ([Ca2+]c) are per se sufficient to trigger extracellular bursts of messenger molecules through connexin HCs, thus mediating paracrine signaling. In this chapter, we present a quantitative method to measure the opening dynamics of connexin HCs expressed in a single HeLa cell upon stimulation by a canonical InsP3-mediated [Ca2+]c transient. The protocol relies on a combination of Ca2+ imaging and patch-clamp techniques. The insights gained from our method are expected to make a significant contribution to understanding the structure-function relationship of connexin HCs. The protocol is also suitable to screen candidate therapeutic compounds to treat connexin-related diseases linked to HC dysfunction.PMID:38578422 | DOI:10.1007/978-1-0716-3842-2_14
Source: Mol Biol Cell - Category: Molecular Biology Authors: Source Type: research