Permeation of Calcium through Purified Connexin 26 Hemichannels

Abstract

Indirect evidence suggests that connexin hemichannels are permeable to Ca(2+), but direct demonstration is lacking. Calcium moves into liposomes containing purified Cx26 in response to a concentration gradient. Cx26 hemichannels are permeable to Ca(2+). Cx26 hemichannels may play a role in Ca(2+) influx into cells under conditions that lead to hemichannel activation, such as ischemic damage. Gap junction channels communicate the cytoplasms of two cells and are formed by head to head association of two hemichannels, one from each of the cells. Gap junction channels and hemichannels are permeable to ions and hydrophilic molecules of up to M(r) 1,000, including second messengers and metabolites. Intercellular Ca(2+) signaling can occur by movement of a number of second messengers, including Ca(2+), through gap junction channels, or by a paracrine pathway that involves activation of purinergic receptors in neighboring cells following ATP release through hemichannels. Understanding Ca(2+) permeation through Cx26 hemichannels is important to assess the role of gap junction channels and hemichannels in health and disease. In this context, it is possible that increased Ca(2+) influx through hemichannels under ischemic conditions contributes to cell damage. Previous studies suggest Ca(2+) permeation through hemichannels, based on indirect arguments. Here, we demonstrate for the first time hemichannel permeability to Ca(2+) by measuring Ca(2+) transport through purified Cx26 hemichannels reconstituted in liposomes. We trapped the low affinity Ca(2+)-sensitive fluorescent probe Fluo-5N into the liposomes and followed the increases in intraliposomal [Ca(2+)] in response to an imposed [Ca(2+)] gradient. We show that Ca(2+) does move through Cx26 hemichannels and that the permeability of the hemichannels to Ca(2+) is high, similar to that for Na(+). We suggest that hemichannels can be a significant pathway for Ca(2+) influx into cells under conditions such as ischemia.