Imaging acetylcholine-receptor-induced influx of inorganic ions at single-cell resolution with ion microscopy

Abstract

Ion microscopy was used to image neurotransmitter-induced tracer ion flux at single-cell resolution. A mammalian muscle cell line (BC 3H1) expressing the nicotinic acetylcholine receptor was exposed to 2 m m CsCl, with and without the acetylcholine analog carbamycholine. 133Cs +, 12C +, 40Ca +, 39K +, and 23Na + secondary ion images revealing intracellular distribution of these elements were recorded with a CAMECA IMS-3fion microscope from freeze-fractured freeze-dried BC 3H1 cells. The ion images were digitized directly from the microchannel plate/fluorescent screen detector assembly of the ion microscope using a charge-coupled device imager. Submillimolar concentrations of cesium were easily imaged. Cesium images were normalized to carbon images for a direct comparison of carbamylcholine-exposed and control cells. Carbamylcholine-exposed cells showed significantly higher cesium influx than controls. Within the carbamylcholine-exposed cells, cell-to-cell heterogenity for cesium influx was observed. Injured cells were identified by their potassium, sodium, and calcium signals and omitted from the quantitative analysis of the ion image data. This method should be useful for identifying cells from various regions of the nervous system containing receptors that control the translocation of monovalent cations, including Cs +. Among these neuronal receptors in the central nervous system are those activated by acetylcholine, glutamate, aspartate, or N-methyl- d-aspartate.