Influence of meningeal cells on the proliferation and maturation of rat neuroblasts in culture

Abstract

Neuronal cells were obtained by dissociating cells from the cerebral hemispheres of rat embryos (10 to 17-day-old), either cleaned entirely or only partially of their meningeal membranes. These cells were seeded on poly-lysine-coated Petri dishes in serum-containing medium. The cultures most enriched in neuronal cells were obtained from brains of 13- to 15-day-old embryos and after 2 h, the culture medium was switched to Dulbecco's modified Eagle's medium, without serum, supplemented with the N1 supplements as described by Bottenstein et al. (1980). The proliferation of neuroblasts from 13-day-old embryos in the presence or absence of meningeal cells was studied by using a combination of tritiated thymidine autoradiography and immuno-staining against neurofilament proteins. The neuroblasts seem to proliferate during the first 3 days. The proliferative activity was further enhanced in the presence of meningeal cells. The glioblasts multiply only after a period of one week in culture conditions as observed here. The subsequent development of the neuroblasts was followed over a period of 4 weeks and the ultrastructural appearance of these cells was investigated at 2 and 3 weeks. In the presence of meningeal cells, many neurons, intensely stained for neurofilament proteins, survived for 21 days, while in control cultures they underwent massive degeneration after 2 weeks. Synapses with numerous clear vesicles were abundant in cultures grown under the influence of meningeal cells; they were rare and possessed few vesicles in control cultures. The data indicate that meningeal cells affect the proliferation and maturation of rat neuroblasts in culture.

Influence of Basic Fibroblast Growth Factor and Astroglial Cells on the infrastructure of Developing Rat Brain Neuronal Precursors in vitro

“…We have previously shown at the light microscopic level that cultures derived from 14-day-old fetal rat cere bral hemispheres consisted essentially of dispersed neu roblasts (immature neurons) with short processes and of small round proliferating neuronal precursor cells arranged in clusters during the first 2 days [Gensburger et al, 1986], The present electron microscopic study confirms these findings and further indicates that between 3 and 5 days, the dispersed neuroblasts differ entiated into neurons containing the usual cytoplasmic organelles (fig. la).…”

Influence of Basic Fibroblast Growth Factor on the Development of Cholinoceptive Neurons from Fetal Rat Cerebrum in Culture

“…The neuronal cell culture system has been character ized previously [11]. Briefly, cultures derived from cere bral hemispheres of 14-day-old fetal rat contain essen tially neuronal cells during the 1st week in vitro and have been characterized by the presence of NF proteins.…”

“…Primary cultures of neuronal cells were prepared from cerebral hemispheres of 14-day-old fetal rats and grown in 35-mm diameter Falcon Petri dishes (8 x 105 cells/dish) for 6 days in a serum-free, chemically defined medium, as previously described [11] menial cultures were treated once o r several times with varying concentrations o f bFG F, as indicated below. bFG F was prepared in our laboratory from bovine brain by using heparin-Sepharose affinity chrom atography [12].…”

“…We found that bFGF affects GABAergic neurons in cul tures of fetal rat cerebral hemispheres [10], In the present work we investigated in this culture system the effect of bFGF on the in vitro development of acetylcholinester ase (AChE)-containing neurons. The bFGF was added to the culture medium either at a time when ncuroblasts still proliferate (between 4 h and 2 days after culture initiation) or at a time when neuronal multiplication has ceased (between 2 and 4 days after culture initia tion) [11].…”

Influence of Epidermal Growth Factor on the Maturation of Fetal Rat Brain Cells in Aggregate Culture

“…Monoclonal antibodies against GFAP (Bio Yeda; di luted 1:800) and against vimentin (Dakopatts; 1:10), as well as polyclonal antibodies against NSE (Polyscience; 1:250), against NF triplet as previously used by Gensburger et al [1986] (prepared and kindly provided by Dr. J.-F. Leterrier, CNRS Strasbourg, France; 1:100), and against GC (prepared and kindly provided by Dr. P. Gebicke-Harter, Pharmacology Institute, Freiburg i.Br.,, FRG; 1:25) [Gebicke-Harter et al, 1981] were applied to adjacent sections for 20 min, fol lowed either by rabbit antimouse immunoglobulins bound to fluorescein isothiocyanate (Dakopatts; 1:25), or by swine antirabbit immunoglobulins bound to fluorescein-isothiocyanate (Dakopatts; 1:25).…”