Abstract
In this article we demonstrate a general protocol for constructing systematically experimental steady-state bifurcation diagrams for electrophysiologically active cells. We perform our experiments on entorhinal cortex neurons, both excitatory (pyramidal neurons) and inhibitiory (interneurons). A slowly ramped voltage-clamp electrophysiology protocol serves as closed-loop feedback controlled experiment for the subsequent current-clamp open-loop protocol on the same cell. In this way, the voltage-clamped experiment determines dynamically stable and unstable (hidden) steady states of the current-clamp experiment. The transitions between observable steady states and observable spiking states in the current-clamp experiment provide partial evidence for stability and bifurcations of the steady states. This technique for completing steady-state bifurcation diagrams in a model-independent way expands support for model validation to otherwise inaccessible regions of the phase space. Overlaying the voltage-clamp and current-clamp protocols leads to an experimental validation of the classical slow-fast dissection method introduced by J. Rinzel in the 1980s and routinely applied ever since in order to analyse slow-fast neuronal models. Our approach opens doors to observing further complex hidden states with more advanced control strategies, allowing to control real cells beyond pharmacological manipulations.