Effects of the synthetic glucocorticoid betamethasone on the survival of neurons in fetal rabbit brain cell culture

Abstract

Dissociated fetal rabbit brain cells were grown on petri dishes coated with collagen. Culture medium consisted of Dulbecco's Modified Eagles Medium plus 10% serum. The mitotic inhibitor 1-β- d-arabinofuranosylcytosine was added at 6 days for a 2 day period to inhibit over-growth by glial cells and fibroblasts. In some cases cultures were chronically exposed to 0.5, 1.0 or 2.0 μM betamethasone. Examination of cultures by phase microscopy and acetylcholinesterase (AChE) staining demonstrated that cultures incubated with 1.0 and 2.0 μM betamethasone contained 2–2.5 times as many neurons as compared to control cultures. Furthermore, there was an increase in the specific activities of both AChE and choline acetyltransferase (ChAT) which were proportional to the increase in neuronal cell numbers obtained from phase microscopy and ACHE staining. These results suggested that betamethasone enhanced survival of cholinergic neurons. Cultures were also examined for neuron specific γ-aminobutyric acid (GABA) uptake. Again GABA uptake was approximately 2–2.5 times as great in cultures incubated with 1–2 μM betamethasone when compared to controls. Thus, the increase in GABA uptake paralleled the increase in neurons observed by phase microscopy and AChE staining, suggesting that the survival effect of betamethasone was not specific to cholinergic neurons. While betamethasone treated cultures always contained greater numbers of neurons the percentage of neurons lost from all cultures after 2 weeks was the same. This suggested that the survival effect of betamethasone was transient and that the larger number of neurons detected at 3 weeks in betamethasone treated cultures may have been the result of larger numbers surviving during the first 1–2 weeks in culture.