Development of phenylethanolamine N-methyltransferase (PNMT) in cultures of dissociated embryonic rat medulla oblongata

Abstract

The adrenergic phenotypic marker, phenylethanolamine N-methyltransferase (PNMT) is expressed in a subgroup of catecholaminergic neurons in the brain, as well as in the chromaffin cells of the adrenal medulla. Although PNMT in the rat adrenal is regulated by glucocorticoids, PNMT in the rat brainstem appears not to be regulated by glucocorticoids. Furthermore, little is known about factors required for the differentiation of this specific class of central neuron. The identification of such factors has been hampered not only by the heterogeneity of cell types in the brainstem, of which only a smaller number express PNMT, but also by the lack of a well characterized in vitro system in which the development of these neurons can be studied under defined conditions. The present study addresses this issue by establishing and characterizing a culture system for the study of adrenergic neurons. Dissociated cultures were prepared from embryonic rat medulla oblongata and the expression and development of PNMT was studied using immunocytochemistry and radioisotopic assay of PNMT activity. The survival of PNMT-immunoreactive (IR) neurons in vitro was found to be critically dependent on embryonic age. Numerous PNMT-IR neurons were observed in cultures prepared only from embryos of 46-51 somites (embryonic day E13-13.5). In contrast, cultures containing numerous neurons immunoreactive for tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis, could be successfully established from medulla oblongata of any age between E13 and E16.(ABSTRACT TRUNCATED AT 250 WORDS)