Simultaneous whole-cell patch-clamp and calcium imaging on myenteric neurons

Live calcium imaging is often used as a proxy for electrophysiological measurements and has been a valuable tool that allows simultaneous analysis of neuronal activity in multiple cells at the population level. In the enteric nervous system, there are two main electrophysiological classes of neurons, AH- and S-neurons, which have been shown to have different calcium handling mechanisms. However, they are rarely considered separately in calcium imaging experiments. A handful of studies have shown that in guinea pig, a calcium transient will accompany a single action potential in AH-neurons, but multiple action potentials are required to generate a calcium transient in S-neurons. How this translates to different modes of cellular depolarisation and whether this is consistent across species is unknown. In this study, we used simultaneous whole-cell patch-clamp electrophysiology together with calcium imaging to investigate how enteric neurons respond to different modes of depolarisation. Using both traditional (4Hz) and also high-speed (1000Hz) imaging techniques, we found that single action potentials elicit calcium transients in both AH-neurons and S-neurons. Sub-threshold membrane depolarisations were also able to elicit calcium transients, although calcium responses were generally amplified if an action potential was present. Further, we identified that responses to nicotinic acetylcholine receptor stimulation can be used to distinguish between AH- and S-neurons in calcium imaging.