Regulation of glycogen in the dentate gyrus of the in vitro guinea pig hippocampus; effect of combined deprivation of glucose and oxygen.
- P Lipton
- Journal of neuroscience methods
INTRODUCTION With its preservation of cytoarchitecture and synaptic circuitry, the hippocampal slice preparation has been a critical tool for studying the electrophysiological effects of pharmacological and genetic manipulations. To analyze the maximum number of slices or readouts per dissection, long incubation times postslice preparation are commonly used. We were interested in how slice integrity is affected by incubation postslice preparation. METHODS Hippocampal slices were prepared by three different methods: a chopper, a vibratome, and a rotary slicer. To test slice integrity, we compared glycogen levels and immunohistochemistry of selected proteins in rat hippocampal slices immediately after dissection and following 2 and 4 hr of incubation. RESULTS We found that immunoreactivity of the dendritic marker microtubule-associated protein 2 (Map2) drastically decreased during this incubation period, whereas immunoreactivity of the glutamate transporter VGlut1 did not significantly change with incubation time. Astrocytic and microglial cell numbers also did not significantly change with incubation time whereas glycogen levels markedly increased during incubation. CONCLUSION Immunoreactivity of the dendritic marker Map2 quickly decreased after dissection with all the slicing methods. This work highlights a need for caution when using long incubation periods following slice preparation.