Multicellular recordings of cultured brainstem neurons in microelectrode arrays

Several vital systemic functions are controlled by the brainstem, which has been studied in a variety of experimental preparations and by various techniques, including in-vitro electrophysiological preparations. Although these in-vitro approaches have greatly advanced the understanding of brainstem neurons, most recording methods with microelectrodes and patch pipettes are invasive. To take advantage of in-vitro approaches but avoid their potential problems, we have studied brainstem neurons in microelectrode arrays (MEA). Neurons were isolated from the medulla oblongata and cultured in DMEM. Extracellular recordings were performed with no evident perturbations to the cellular environment. Neurons started firing after 24-48 h in culture, reached stable activity in 3-4 weeks, and retained this activity for at least 3 months. From their firing patterns, these neurons could be divided into tonic and bursting units. The latter could be further divided into regular and irregular bursters based on their burst intervals. Cells were stimulated or inhibited by exposure to 10% CO2. The stimulatory effect of CO2, though smaller, was still seen after selective ablation of serotonergic neurons or with low Ca++ and high Mg++ in the extracellular medium. Similar treatments had no significant effect on CO2-inhibited units. The abundance of units with respect to their firing patterns and CO2 responses, together with the long-term stable non-invasive recordings with no evident perturbation to cellular environments, suggests that MEA represent another promising in-vitro approach for studying brainstem neurons.