Cholera Toxin Induces Sustained Hyperexcitability in Submucosal Secretomotor Neurons in Guinea Pig Jejunum

Background & Aims

Neural mechanisms underlying cholera toxin (CT)-induced intestinal hypersecretion remain unclear. We investigated long-term excitability changes in vasoactive intestinal peptide (VIP) and neuropeptide Y (NPY) secretomotor neurons after prolonged luminal exposure to CT.

Methods

Isolated segments of guinea pig jejunum were incubated with saline or CT ± neurotransmitter antagonist in the lumen; the submucosal plexus was then dissected clear, circumferentially adjacent to intact mucosa. Synaptic inputs and firing properties of S neurons in ganglia next to the mucosa in control saline were studied using intracellular recording. Neurons were processed for VIP and NPY immunoreactivity.

Results

Thirty S neurons (20 VIP⁺, 7 NPY⁺, 3 VIP⁻/NPY⁻) from CT-treated preparations and 27 control S neurons (19 VIP⁺, 4 NPY⁺, 4 VIP⁻/NPY⁻) in ganglia adjacent to intact mucosa were analyzed. VIP⁺ and NPY⁺ neurons in CT-treated preparations fired significantly more action potentials and for longer periods during injected depolarizing current pulses (50–350 pA) than control neurons. Addition of tetrodotoxin, hexamethonium, granisetron, or the neurokinin-1 (NK1) antagonist SR140333 during the CT incubation blocked CT-induced effects in both neuron types. The NK3 antagonist SR142801 blocked CT-induced effects in NPY⁺ neurons and reduced the number of action potentials in VIP⁺ neurons. Synaptic activity was unaffected by CT.

Conclusions

CT induces specific and sustained hyperexcitability of secretomotor neurons in enteric pathways. CT acts in the mucosa. Its effect is neurally mediated and depends on 5-hydroxytryptamine-3, nicotinic, and NK1 receptors. This system represents a unique model to understand the neural mechanisms of action of CT and to identify therapeutic targets.

Abbreviations used in this paper: ACh, acetylcholine, AH, after hyperpolarization, APs, action potentials, CFTR, cystic fibrosis transmembrane conductance regulator, CT, cholera toxin, CT-B, B-subunit of cholera toxin, ENS, enteric nervous system, EPSPs, excitatory postsynaptic potentials, IPSPs, inhibitory postsynaptic potentials, IR, input resistance, NK, neurokinin, NPY, neuropeptide Y, PPADS, pyridoxal phosphate-6-axophenyl-2′-4′-disulfonic acid, SMP, submucosal plexus, TTX, tetrodotoxin, VIP, vasoactive intestinal peptide