Two firing modes and well-resolved Na+, K+, and Ca2+ currents at the cell-microelectrode junction of spontaneously active rat chromaffin cell on MEAs

Abstract

We recorded spontaneous extracellular action potentials (eAPs) from rat chromaffin cells (CCs) at 37 °C using microelectrode arrays (MEAs) and compared them with intracellularly recorded APs (iAPs) through conventional patch clamp recordings at 22 °C. We show the existence of two distinct firing modes on MEAs: a ~ 4 Hz irregular continuous firing and a frequent intermittent firing mode where periods of high-intraburst frequency (~ 8 Hz) of ~ 7 s duration are interrupted by silent periods of ~ 12 s. eAPs occurred either as negative- or positive-going signals depending on the contact between cell and microelectrode: either predominantly controlled by junction-membrane ion channels (negative-going) or capacitive/ohmic coupling (positive-going). Negative-going eAPs were found to represent the trajectory of the Na⁺, Ca²⁺, and K⁺ currents passing through the cell area in tight contact with the microelectrode during an AP (point-contact junction). The inward Nav component of eAPs was blocked by TTX in a dose-dependent manner (IC₅₀ ~ 10 nM) while the outward component was strongly attenuated by the BK channel blocker paxilline (200 nM) or TEA (5 mM). The SK channel blocker apamin (200 nM) had no effect on eAPs. Inward Nav and Cav currents were well-resolved after block of Kv and BK channels or in cells showing no evident outward K⁺ currents. Unexpectedly, on the same type of cells, we could also resolve inward L-type currents after adding nifedipine (3 μM). In conclusion, MEAs provide a direct way to record different firing modes of rat CCs and to estimate the Na⁺, Ca²⁺, and K⁺ currents that sustain cell firing and spontaneous catecholamines secretion.

Keywords: AP-clamp recording; Cell-electrode interface; Extracellular action potentials; Ion channel blockers; Na+, Ca2+, and K+ currents.