Analysis of whole-cell currents by patch clamp of guinea-pig myenteric neurones in intact ganglia

Whole-cell patch-clamp recordings taken from guinea-pig duodenal myenteric neurones within intact ganglia were used to determine the properties of S and AH neurones. Major currents that determine the states of AH neurones were identified and quantified. S neurones had resting potentials of −47 ± 6 mV and input resistances ( R in ) of 713 ± 49 MΩ at voltages ranging from −90 to −40 mV. At more negative levels, activation of a time-independent, caesium-sensitive, inward-rectifier current ( I Kir ) decreased R in to 103 ± 10 MΩ. AH neurones had resting potentials of −57 ± 4 mV and R in was 502 ± 27 MΩ. R in fell to 194 ± 16 MΩ upon hyperpolarization. This decrease was attributable mainly to the activation of a cationic h current, I h , and to I Kir . Resting potential and R in exhibited a low sensitivity to changes in [K + ] o in both AH and S neurones. This indicates that both cells have a low background K + permeability. The cationic current, I h , contributed about 20 % to the resting conductance of AH neurones. It had a half-activation voltage of −72 ± 2 mV, and a voltage sensitivity of 8.2 ± 0.7 mV per e-fold change. I h has relatively fast, voltage-dependent kinetics, with on and off time constants in the range of 50–350 ms. AH neurones had a previously undescribed, low threshold, slowly inactivating, sodium-dependent current that was poorly sensitive to TTX. In AH neurones, the post-action-potential slow hyperpolarizing current, I AHP , displayed large variation from cell to cell. I AHP appeared to be highly Ca 2+ sensitive, since its activation with either membrane depolarization or caffeine (1 m m ) was not prevented by perfusing the cell with 10 m m BAPTA. We determined the identity of the Ca 2+ channels linked to I AHP . Action potentials of AH neurones that were elongated by TEA (10 m m ) were similarly shortened and I AHP was suppressed with each of the three Ω-conotoxins GVIA, MVIIA and MVIIC (0.3–0.5 μ m ), but not with Ω-agatoxin IVA (0.2 μ m ). There was no additivity between the effects of the three conotoxins, which indicates the presence of N- but not of P/Q-type Ca 2+ channels. A residual Ca 2+ current, resistant to all toxins, but blocked by 0.5 m m Cd 2+ , could not generate I AHP . This patch-clamp study, performed on intact ganglia, demonstrates that the AH neurones of the guinea-pig duodenum are under the control of four major currents, I AHP , I h , an N-type Ca 2+ current and a slowly inactivating Na +