¹H NMR study of cortex neurons and cerebellar granule cells on microcarriers and their PCA extracts: Lactate production under hypoxia

Abstract

Lactate production of 6-day-old cerebral cortex neurons and 7-day-old cerebellar granule cells from mouse brain attached to cytodex 3 microcarriers was studied as a function of time, under hypoxic conditions using 1H NMR. Perchloric acid extracts of both cell types were prepared and 1H NMR spectra showed compounds characteristic for these neurons. In particular the granule cell extracts showed a large amount of glutamate as expected from biochemical experiments, whereas the cortex neurons showed a large amount of 4-aminobutyric acid.

Proton Nuclear Magnetic Resonance Spectroscopy of Primary Cells Derived from Nervous Tissue

“…Cortical neurones were notable for a high GABA content, almost twice that of cerebellar granule neurones, with all other cell types containing still lower amounts of GABA. These data are in agreement with earlier work in which a qualitative comparison of 'H NMR spectra from cerebellar granule cells and cortical neurones was reported (Sonnewald et al, 1992). These data accord with the fact that cerebellar granule cells are predominately glutamatergic and would not be expected to express significant amounts of GABA.…”

“…One approach to solving this problem is to have prior knowledge of the spectra of individual cell types, and to this end we and others have previously used 'H NMR spectroscopy to study purified populations of different cell types derived from various cellular lineages of the CNS (Sonnewald et al, 1992;Leibfritz and Brand, 1993;Urenjak et al, 1993). Tn our (Urenjak et al, 1993) studies we found that it was possible to distinguish cerebellar granule neurones, meningeal cells, cortical astrocytes, oligodendrocyte type 2 astrocyte (O-2A) progenitor cells, and oligodendrocytes by analysis of their 'H NMR spectra.…”

Culturing primary brain astrocytes under a fully controlled environment in a novel bioreactor

“…Another critical issue in microcarrier technology is the inoculation step, because cellular proliferation occurs only after proper adhesion and a minimum amount of cells per microcarrier is necessary to avoid large lag phases (cell–cell interaction phenomena) (Hu et al, 1985). Different types of microcarriers have been used to grow primary rat brain astrocytes (Westergaard et al, 1991; Sonnewald et al, 1992; Alves et al, 1997). These studies were carried out in static conditions, however, where microcarriers were either placed on top of a confluent astrocytic monolayer growing in a culture dish or inoculated with cells and kept in T‐flasks.…”

Immobilization of Primary Astrocytes and Neurons for On-Line Monitoring of Biochemical Processes by NMR

“…Perfusion of primary brain cells has been reported using cells growing on top of microspheres [5,6]. Dis advantages of such a system are that only a limited num ber of cells can be accommodated for on the surface of the beads and shear stress due to perfusion removes the cells from their support and limits their lifetime.…”

“…As the brain consists of a complex mixture of different cell types it is impor tant to characterize each cell type separately. This can be done using NMR spectroscopy and cell culturing tech niques either recording brain cell extracts [1][2][3][4] or immobilized and perfused brain cells in the spectrometer [5,6], Extraction of cell cultures has the advantage that large quantities of cells can be pooled and the spectral resolu tion and sensitivity are superior with respect to spectra obtained from perfused cells. However, a disadvantage is that only one experiment is possible with an identical batch of cells.…”