Effect Of Bone Morphogenetic Protein-2 On Cholinergic Neurons Development

It has become evident that cholinergic neurons serve a variety offunctions including sleep,memeory,cognitive and motivationalprocesses,and also are the target cells of many central systemdegeneration diseases. So the mechanisms of the generation andthe functional regulation of cholinergic neurons have always beenthe hot problems, which are also the basis to understand thefunction of CNS and to probe pathway of the therapy methods.Now the most important problem is to find the extracellularfactors and essential genes which play critical roles in theregulation of cholinergic neurons.To study the extracellular factors effecting the cholinergicneurons development, it's first to understand the developmentalprocess of them. As the marker protein of cholinergic neuronscholine acetyltransferase(ChAT) can show the generation ,transplantation and distribution of cholinergic neurons. Germinalzones for cholinergic cells were observed in all ventricular zonesof the forebrain with the ventral zones generating the earliest cellsby gestational day of the E13.5.On the E14, ChATimmunoreactive(IR) cells were visible in the germinal zones ofthe dorsolateral aspect of the lateral ventricle,lateral ganglioniceminence. ChAT IR neurons continurd to develop in these andadditional germinal zones on E15,including the medial,dorsal anddorsomedial walls of the lateral ventricle.Correlative informationmust regulate the distribution of the cholinergic neurons and theirdevelopmental fate, which is the main problem of our study.During the CNS development, it is the basic process to generatesome special neurons.Both the intracellular genes and theextracellular factors determine the essential process.It is the sameimportance for cholinergic neurons to follow this rule.Bone morphogenetic proteins (BMPs) are members of thetransforming growth factor-(TGF) superfamily of growth anddifferentiation factors. They elicit a broad range of celluarresponses both during development and in the adult animal,including cell proliferation,differentiaton,lineage determination,cell adhesion and positioning and apoptosis. BMPs is ahomologous proteins family, in which each member has specialtarget cells and expresses in different regions. On the E8.5,BMP-2 is exclusively expressed in the dorsal surface ectodermimmediately adjacent to the nruroectoderm, while BMP-5 andBMP-7 are weakly expressed throughout the dorsal surfaceectoderm; On the E11.5, BMP-2 expression is more strong in thedorsomedial telencephalon compared with the other BMPs; Onthe E13.5, BMP-2 expression is restricted to the thin layer of themedial walls of the lateral ventricles and the developing dentategyrus of the hippocampus, and others expressed in the choroidsplexus. So we can see that there are many domains where BMP-2and cholinergic neurons co-exist, which indicates that BMP-2 canregulate special genes expressions of cholinergic neurons topromote their proliferation and differentiation.To investigate BMP-2 functions during the cholinergic neuronsdevelopment , we treated primary cells derived from the dorsaltelencephlon of E14 rat brain with BMP-2. Experimental cellswere divided into three groups, which were given BMP-2 at20ng/ml,40ng/m,80ng/ml every 24 hours until the cells werecollected for analysis 4 days later and control group only withbFGF 20ng/ml. Immunofluorescence studies were performed with...