On June 26, 2013, The Journal of Neuroscience published online a research article entitled "Tlx3 Controls Cholinergic Transmitter and Peptide Phenotypes in a Subset of Prenatal Sympathetic Neurons". This work was mainly carried out by graduate students Tianwen Huang and Jia Hu, under the supervision of Dr. Leping Cheng.

　　Mature sympathetic ganglia are composed of two types of neurons. The vast majority of sympathetic neurons are noradrenergic, while a small subset of sympathetic neurons use acetylcholine as transmitter. Substantial progress has been made in understanding how transcription factors control the acquisition of noradrenergic phenotype in sympathetic neurons. For cholinergic differentiation in sympathetic neurons, there are two modes by which sympathetic neurons could acquire their cholinergic phenotype. In the classic postnatal target-dependent cholinergic differentiation, the sudomotor neurons and the neurons that innervate the periosteum initially express noradrenergic phenotype, but acquire the cholinergic phenotype while losing functional adrenergic properties after contacting the target tissue. The switch from noradrenergic to cholinergic phenotype is initiated by signals released from the target (e.g. sweat glands). Prenatal expression of cholinergic properties has also been found in a very small population of developing sympathetic neurons. However, the molecular mechanism that controls the development of embryonic cholinergic neurons remains largely unknown.

　　Immunostaining for vesicular acetylcholine transporter (VAChT) showed that Tlx3 was highly expressed in cholinergic neurons at the late embryonic stage E18.5. Deletion of Tlx3 resulted in the loss of Vacht expression at E18.5 but not E12.5. By contrast, Tlx3 was required for expression of the cholinergic peptide vasoactive intestinal polypeptide (VIP), and somatostatin (SOM) at both E12.5 and E18.5. Furthermore, we found that, at E18.5 these putative cholinergic neurons expressed glial cell line-derived neurotrophic factor (GDNF) family co-receptor Ret but not tyrosine hydroxylase (Ret+/THa€“). Deletion of Tlx3 also resulted in disappearance of high-level Ret expression. Lastly, unlike Tlx3, Ret was required for the expression of VIP and SOM at E18.5 but not E12.5. Together, these results indicate that transcription factor Tlx3 is required for the acquisition of cholinergic phenotype at the late embryonic stage as well as the expression and maintenance of cholinergic peptides VIP and SOM throughout prenatal development of mouse sympathetic neurons. 

　　Transcription factor Tlx3 controls the expression of Ret, the GDNF family ligands co-receptor, in cholinergic neurons of the sympathetic ganglia