Mechanisms inhibiting sympathetic neurotransmitter release during gastrointestinal inflammation
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Inflammatory bowel disease (IBD) alters neuronal regulation of the gastrointestinal (GI) tract. The superior mesenteric ganglia (SMG) contain sympathetic neurons that modulate GI functions such, as motility and blood flow. IBD reduces the release of noradrenaline, a sympathetic neurotransmitter. We hypothesized that the reduction in NA release is due to inhibition of voltage-gated calcium current (ICa), as calcium influx is a regulator of neurotransmitter release. We also hypothesized that tumor necrosis factor α (TNFα), a proinflammatory cytokine elevated during IBD, can also inhibit the ICa of SMG neurons. Therefore, we compared ICa amplitude in neurons from normal mice and mice with dextran sulphate sodium (DSS; 5% w/v)-induced colitis. Neurons dissociated from the SMG were cultured overnight and changes to ICa were investigated using electrophysiological, Ca2+ imaging, PCR and neurotransmitter release techniques. Colitis significantly reduced ICa of SMG neurons by selectively inhibiting N-type Ca2+ channels. This was accompanied by a reduction in mRNA encoding the N-type channel alpha subunit (CaV 2.2) and a rightward shift in the voltage dependence of activation of ICa. Colitis reduced the NA release from the colon and jejunum. Depolarization-induced release of tritiated-NA was inhibited by ω-Conotoxin GVIA (300 nM). These results suggest that the changes in VGCC observed at the cell bodies of SMG neurons were also occurring at the nerve terminals during colitis. Similar experimental techniques were performed using SMG neurons incubated overnight in TNFα (1nM). TNFα decreased ICa and depolarization-induced Ca2+ influx in SMG neurons. Similar to DSS-induced colitis, the reduction in ICa was limited to N-type Ca2+ channels. Preincubation of neurons with SC 514 (20μM) and Bay 11 7082 (1µM), inhibitors of nuclear factor kappa B signaling, prevented the reduction in ICa. Preincubation with the p38 MAPK inhibitor, PD 169316 (30µM), recovered a smaller portion of the reduction in Ca2+ influx. These data suggest that DSS colitis and TNFα inhibit N-type VGCC ICa in sympathetic neurons and identify a novel role for NF-κB and p38 MAPK in the regulation of neurotransmitter release. These findings also suggest that DSS colitis inhibits NA release by altering sympathetic N-type VGCC in the colon and jejunum.