Sympathoadrenal Progenitor Research Articles

Membrane depolarization induces p140trk and NGF responsiveness, but not p75LNGFR, in MAH cells.

Nerve growth factor (NGF) is required for the maturation and survival of sympathetic neurons, but the mechanisms controlling expression of the NGF receptor in developing neuroblasts have not been defined. MAH cells, an immortalized sympathoadrenal progenitor cell line, did not respond to NGF and expressed neither low-affinity NGF receptor (p75) nor p140trk messenger RNAs. Depolarizing concentrations of potassium chloride, but none of a variety of growth factors, induced expression of p140trk but not p75 messenger RNA. A functional response to NGF was acquired by MAH cells under these conditions, suggesting that expression of p75 is not essential for this response. Depolarization also permitted a relatively high proportion of MAH cells to develop and survive as neurons in fibroblast growth factor and NGF. These data establish a relation between electrical activity and neurotrophic factor responsiveness in developing neurons, which may operate in the functioning of the mature nervous system as well.

Changes in competence determine the timing of two sequential glucocorticoid effects on sympathoadrenal progenitors

We have studied the control of adrenal chromaffin cell development by glucocorticoids (GCs), in a reconstituted in vitro system. The development of the chromaffin phenotype involves two sequential, GC-dependent events: the decision not to become a sympathetic neuron, and the decision to express the epinephrine-synthesizing enzyme, phenylethanolamine-N-methyltransferase (PNMT). Both decisions appear to be mediated by the type II GC receptor. Competence to express PNMT develops on a schedule in vitro which parallels that seen in vivo, but only in progenitors that have first failed to undergo neuronal differentiation. The schedule of PNMT induction is thus controlled by the time of appearance of neither the inducing signal nor its receptor, as previously suggested, but rather by a cell-intrinsic timed process in chromaffin precursors. The two effects of GCs are pharmacologically distinct, suggesting that the GC receptor may interact differently with different genes in the same cell, in a manner that changes with development.

Evidence that enteric neurons may derive from the sympathoadrenal lineage

The first neurons that differentiate in the embryonic foregut of mammals transiently express catecholamine biosynthetic enzymes and accumulate catecholamine. Since this transmitter is found predominantly in cells of the sympathoadrenal (SA) lineage, it has been suggested that enteric and sympathetic neurons may derive from the same progenitor. Enteric neurons would then lose the catecholamine phenotype during further development, as the two lineages diverge. We have further investigated this possibility using the SA1 monoclonal antibody that binds selectively to SA progenitor cells in the embryonic rat. We find that SA1 binds to the tyrosine hydroxylase +, neurofilament +, and SCG10 + cells of the Embryonic Day 14.5 (E14.5) rat foregut. We also find that a marker for later neuronal differentiation in the SA lineage, B2, also appears in the myenteric plexus concomitant with the loss of SA1 staining. Thus, at least some enteric neuronal precursors may exhibit the SA1 → B2 antigenic switch previously observed in developing sympathetic neurons at E14.5. SA1 staining in the foregut partially overlaps with staining for neuropeptide Y, vasoactive intestinal polypeptide, and serotonin. These results support the hypothesis that enteric and sympathetic neurons derive from a common progenitor and that as the markers for the SA lineage are down-regulated, the many types of enteric neurons begin to differentiate.

Co-expression of multiple neurotransmitter enzyme genes in normal and immortalized sympathoadrenal progenitor cells

We have examined the expression of mRNAs encoding five major neurotransmitter-synthesizing enzymes in MAH cells, a clonal cell line derived by retroviral immortalization of a rat embryonic sympathoadrenal progenitor cell. These mRNAs include tyrosine hydroxylase (TH), choline acetyltransferase (ChAT), tryptophan hydroxylase (TpH), and glutamic acid decarboxylases (GADs) 1 and 2. We find that MAH cells express high levels of TH mRNA and low levels of ChAT and TpH mRNAs. Neither GAD1 nor GAD2 mRNAs are detectable using an RNase protection assay with a detection limit of less than one transcript per cell. A similar pattern of mRNA expression is observed in postnatal superior cervical ganglia, adrenal medulla, and in PC12 cells. Transmitter synthesis and accumulation assays indicate that MAH cells can synthesize both catecholamines and acetylcholine. Thus the TH and ChAT mRNAs detected in these cells are likely to be translated into active enzyme. To corroborate these data obtained using MAH cells, we performed similar transmitter synthesis and accumulation assays on sympathoadrenal progenitors directly isolated from E14.5 fetal adrenal glands by fluorescence-activated cell sorting. These progenitor cells also synthesize and accumulate both catecholamines and acetylcholine, albeit to different extents than MAH cells. Both MAH cells and their nonimmortal counterparts are able to increase slightly their cholinergic function upon short-term exposure to CDF/LIF, a factor known to induce acetylcholine synthesis in postmitotic sympathetic neurons. Taken together, these data suggest that progenitor cells in the sympathoadrenal lineage acquire the ability to simultaneously transcribe several different neurotransmitter enzyme genes early in development, prior to their choice of final cell fate. At the same time, the progenitors possess receptors which regulate expression of these genes in response to environmental factors. This ability may permit the cells to choose from several different transmitter phenotypes in response to different environments, as they migrate through the embryo. The persistent transcription of these genes in adult cells, moreover, may in part account for the phenotypic plasticity of cells in this lineage.

A v-myc-immortalized sympathoadrenal progenitor cell line in which neuronal differentiation is initiated by FGF but not NGF

Sympathetic neurons differentiate from a developmentally restricted progenitor cell in the neural crest-derived sympathoadrenal lineage. We have isolated these progenitors by fluorescence-activated cell sorting and immortalized them using a v-myc-containing retrovirus. The complement of antigenic markers expressed by these lines suggests that they have retained many of the properties of their normal counterparts.These lines initiate neuronal differentiation in response to basic FGF, but not to NGF, and do not contain NGF receptor mRNA. In NGF plus FGF, however, a small percentage of the cells differentiate to NGF-dependent postmitotic neurons. Furthermore, an induction of NGF receptor mRNA can be observed in response to FGF Thus, the development of sympathetic neurons may involve a relay, in which FGF both initiates differentiation and induces the NGF receptor, which in turn controls further maturation and survival.

Role of glucocorticoids in the chromaffin-neuron developmental decision

Chromaffin cells and sympathetic neurons develop from a common neural crest-derived progenitor cell. The developmental fate of this cell differs depending upon whether it migrates to the sympathetic ganglion or to the adrenal gland primordium, suggesting that local environmental signals control its differentiation. Glucocorticoid (GC) is a good candidate for an important adrenal environmental signal. These steroids are known to regulate PNMT, an adrenal-specific enzyme. However, in vivo observations suggest that the adrenal microenvironment influences the phenotype of sympathoadrenal progenitor cells as early as E14.5, 2 days before PNMT is first expressed by developing chromaffin cells. Using cDNA probes, we find that GC receptor mRNA can be detected in the embryonic adrenal at least one full day before the initial appearance of PNMT mRNA. This observation is compatible with the idea that the apparent early influence of the adrenal microenvironment reflects the action of GC on progenitors which have migrated into this environment. In support of this, we show that similar influences can be exerted by GC on PC12 cells, which contain GC receptor mRNA but do not express or induce PNMT mRNA. Taken together, these data suggest that other factors in addition to the presence of the GC receptor may be necessary for the developmental appearance of PNMT expression.