Optical assessment of motoneuron function in a “twenty-four-hour” acute spinal cord slice model from fetal rats

Abstract

In acute slice preparations of most brain regions, neuronal functions are preserved for only few hours. Since the effects of growth factors or neurotoxic agents are often manifested beyond this time scale, corresponding studies are typically performed on cultured cells. However, cell cultures are generated and maintained under vastly different conditions that can grossly alter neuronal properties. For example, glutamate application to motoneuronal cultures has been reported to modulate neurite formation in some studies while in others it has been reported to kill cells. Here, we have examined whether acute spinal cord slices from rat fetuses can be used within a time window of 24 h for assessment of long-term effects of neuromodulators. In these slices, we have studied the action of glutamate on lumbar motoneurons loaded with fura-2 and rhodamine-123 to monitor intracellular Ca 2+ ([Ca 2+] i) and mitochondrial potential (Δ ψ), respectively. Further, loading with fura-2 or propidium iodide allowed for morphological assessment of cell viability and death, respectively. Pulses (15 s) or 1 h application of glutamate (300 μM) evoked a moderate (∼500 nM) [Ca 2+] i rise, but no change of Δ ψ. Even after 24 h, no glutamate-induced cell death was observed and glutamate pulse-evoked [Ca 2+] i transients were comparable to controls. The data demonstrate that glutamate does not deregulate [Ca 2+] i homeostasis in fetal motoneurons in situ. We propose that acute spinal cord slices from perinatal rodents are a robust model that allows for analysis of neuronal properties and cell viability within a time window of at least 24 h.