ATP Release through Connexin Hemichannels in Peripheral Nerve

Abstract

Schwann cells wrapping peripheral nerves express purinergic receptors and may also be a source of extracellular ATP through hemichannels. Peripheral nerve fibers contain areas of non-compact myelin, Schmidt-Lantermann incisures and paranodes, where Connexin32 (Cx32) is located and could form hemichannels. ATP release from cultured RT4 Schwann cells and mice sciatic nerve trunks through Cx32 hemichannels was assessed by triggering the liberation of ATP under several conditions: electrical stimuli, perfusion of low divalent solutions and hypotonic shock. The extracellular ATP was monitored as the light signal emitted by the luciferase assay. Low divalent solutions triggered the release of ATP from isolated nerve trunks, with continuous ATP release observed for several minutes that could be blocked by the well characterized connexin blockers, 1.5 mM octanol and 50 μM flufenamic acid. To further investigate whether this release occurs in vivo we carried out electrical stimulation of the sciatic nerve in anesthetized animals. Trains of 10 Hz stimuli were delivered and subsequent ATP release recorded. ATP release could also be recorded from both Schwann cells and sciatic nerves after hypotonic shock. ATP release was blocked pharmacologically by CBX 100 μM and in the Cx32 KO, suggesting that Cx32 hemichannels are the structures responsible for the exchange of ATP between the intracellular space of Schwann cells and extracellular media. As in the demyelinating X-linked Charcot-Marie-Tooth disease (CMTX) Cx32 is mutated, these findings suggest a possible relationship between changes in Cx32 hemichannel permeability for ATP and peripheral nerve injury.This work is supported by SAF SAF2011-28485 of the Spanish Government,by FEDER Founds of EU and by Generalitat de Catalunya, SGR 2009-152.