Ryanodine-Sensitive Stores Regulate the Excitability of AH Neurons in the Myenteric Plexus of Guinea-Pig Ileum

Abstract

Myenteric afterhyperpolarizing (AH) neurons are primary afferent neurons within the gastrointestinal tract. Stimulation of the intestinal mucosa evokes action potentials (AP) that are followed by a slow afterhyperpolarization (AHPslow) in the soma. The role of intracellular Ca2+ ([Ca2+]i) and ryanodine-sensitive Ca2+ stores in modulating the electrical activity of myenteric AH neurons was investigated by recording membrane potential and bis-fura-2 fluorescence from 34 AH neurons. Mean resting [Ca2+]i was ∼200 nM. Depolarizing current pulses that elicited APs evoked AHPslow and an increase in [Ca2+]i, with similar time courses. The amplitudes and durations of AHPslow and the Ca2+ transient were proportional to the number of evoked APs, with each AP increasing [Ca2+]i by ∼50 nM. Ryanodine (10 μM) significantly reduced both the amplitude and duration (by 60%) of the evoked Ca2+ transient and AHPslow over the range of APs tested (1–15). Calcium-induced calcium release (CICR) was graded and proportional to the number of APs, with each AP triggering a rise in [Ca2+]i of ∼30 nM Ca2+ via CICR. This indicates that CICR amplifies Ca2+ influx. Similar changes in [Ca2+]i and AHPslow were evoked by two APs in control and six APs in ryanodine. Thus, the magnitude of the change in bulk [Ca2+]i and not the source of the Ca2+ is the determinant of the magnitude of AHPslow. Furthermore, lowering of free [Ca2+]i, either by reducing extracellular Ca2+ or injecting high concentrations of Ca2+buffer, induced depolarization, increased excitability, and abolition of AHPslow. In addition, activation of synaptic input to AH neurons elicited a slow excitatory postsynaptic potential (sEPSP) that was completely blocked in ryanodine. These results demonstrate the importance of [Ca2+]i and CICR in sensory processing in AH neurons. Activity-dependent CICR may be a mechanism to grade the output of AH neurons according to the intensity of sensory input.