KATP channel formation by the sulphonylurea receptors SUR1 with Kir6.2 subunits in rat dorsal vagal neurons in situ

摘要

Functional and molecular properties of ATP-sensitive K+ (KATP) channels were studied in dorsal vagal neurons (DVNs) of rat brainstem slices using patch-clamp and single-cell antisense RNA amplification-polymerase chain reaction (PCR) techniques.In the cell-attached configuration, 1 mM cyanide resulted in block of spontaneous firing and concomitant opening of single channels with a mean single open time of 2-3 ms and a burst duration of up to several hundred milliseconds. Inhibition of such single-channel activity with 200 μM tolbutamide led to the reappearance of spontaneous discharge.Whole-cell recordings during anoxia revealed a hyperpolarization of the DVNs. Harvesting of cytoplasm, antisense RNA amplification and subsequent PCR showed coexpression for single DVNs of mRNA for the sulphonylurea receptor SUR1 isoform and for the inwardly rectifying K+ channel subunit Kir6.2, but not for the SUR2 or Kir6.1 isoforms of these channel/receptor subclasses.Upon anoxia, a stable depolarization by less than 10 mV was observed in non-excitable cells in the dorsal vagal nucleus. These cells, which expressed glial fibrillary acidic protein (GFAP), showed a high level of mRNA for Kir6.2, a weak signal for SUR1, whereas SUR2 or Kir6.1 were not detected.The results suggest that functional KATP channels in DVNs are constituted by the formation of Kir6.2 subunits with SUR1 receptors.