Studies on cultured astrocytes: Their oxidative diversity and the effect of cytokines on their biochemical composition

Abstract

Astrocytes are the most common cell found in the central nervous system (CNS). Studies have demonstrated a number of subtypes of astrocytes based upon morphological, biochemical, and pharmacological differences in these cells. Since it is known that astrocytes from neonatal rat brain exhibit two morphologically-distinct subtypes in culture, polygonal and process-bearing, I decided to examine how these two types differ with respect to their oxidative metabolism. To assess the oxidative capacity of glial cells, mitochondrial malate dehydrogenase (mMDH) was visualized by indirect immunofluorescence in purified cultures of neonatal rat polygonal and process-bearing astrocytes as well as in oligodendrocytes. Both process-bearing astrocytes and oligodendrocytes showed uniformly intense anti-mMDH immunoreactivity whereas immunoreactivity of polygonal astrocytes varied from very weakly positive to intensely positive. The mMDH distribution pattern in cultures of polygonal astrocytes did not depend on the degree of morphological differentiation. Polygonal astrocytes cultivated from distinct regions of brain expressed variable but reproducible profiles of anti-mMDH immunoreactivity. Since certain cytokines such as interleukin (IL)-1, IL-2 and tumor necrosis factor (TNF) are present in the CNS and at the site of traumatic CNS lesions, I decided to assess whether these cytokines could influence the biochemical composition of astrocytes in culture. High affinity peripheral-type benzodiazepine binding sites (PTBBS) were present in polygonal astrocytes but barely detectable in process-bearing astrocytes. IL-1 and TNF increased PTBBS only within polygonal astrocytes and did so in both dose- and time-dependent manners. Scatchard analysis suggested the increase in PTBBS caused by TNF was due to an increased number of binding sites. These cytokines stimulated proliferation in cultures of polygonal astrocytes but not in cultures of process-bearing astrocytes. The cytokines did not significantly affect the level of glial fibrillary acidic protein (GFAP) but decreased the mRNA for GFAP by as much as 62%. IL-1 and TNF specifically increased the level of transferrin (Tf) and Tf mRNA within polygonal astrocytes in dose- and time-dependent manners. Process-bearing astrocytes were negative for Tf regardless of the treatment used. These studies may prove useful in understanding the cellular heterogeneity among astrocytes and their differential interactions with cytokines.