Dlx transcription factors regulate differentiation of dopaminergic neurons of the ventral thalamus Gracie L. Andrews, a Kyuson Yun, b John L.R. Rubenstein, b and Grant S. Mastick* ,a a Biology and Biochemistry Departments, Cell and Molecular Biology Graduate Program, University of Nevada, Reno, NV 89557, USA b Nina Ireland Laboratory of Developmental Neurobiology, Center for Neurobiology and Psychiatry, Department of Psychiatry and Programs in Neuroscience and Developmental Biology, University of California at San Francisco, San Francisco, CA 94143-0984, USA Received 23 May 2002; revised 12 December 2002; accepted 27 December 2003 Abstract Recent studies have provided many lines of evidence that specific homeodomain factors act to regulate differentiation into specific neuron types. However, these studies have mainly focused on the caudal CNS, while in the forebrain, the regulation of neuron specification remains relatively unknown. To investigate the genetic regulatory networks that control neuron differentiation in the forebrain, we have analyzed the expression patterns and functions of DLX homeodomain factors in the ventral thalamus of early mouse embryos. During initial neurogenesis (E9.5–E10.5), DLX + cells are the first progenitors to make terminal divisions and differentiate as neurons. We have defined a set of regulatory genes coexpressed with DLX, in both progenitors (PAX6 and MASH1) and in the differentiating neurons (PAX6, along with a combination of LIM-type homeodomain factors, including ISL1, Lhx1/Lim1, and Lhx5/Lim2). These initial neurons express tyrosine hydroxylase (TH), and become the PAX6-expressing A13 dopaminergic neurons of the zona incerta. To test for DLX function, the initial differentiation of the ventral thalamic neurons was examined in embryos mutant for Dlx1 and Dlx2. Dlx1/2 double homozygous mutants formed ventral thalamic neurons, but these neurons lacked PAX6, ISL1, and TH expression. These data suggest that DLX genes act as forebrain-specific factors linking general neuron-inducing signals to region-specific neuron differentiation programs. © 2003 Elsevier Science (USA). All rights reserved. Introduction The central nervous system contains a great diversity of neuron types organized in precise spatial arrays. This spatial organization arises during development through establish- ment of local populations of neuronal progenitors that pro- duce restricted repertoires of region-specific neuron types. The current view is that these progenitor populations are set up through the influence of dorsal–ventral and rostral– cau- dal patterning signals that establish expression of specific combinations of transcription factors, largely of the ho- meodomain family (reviewed in Jessell, 2000; Lumsden and Krumlauf, 1996; Wilson and Rubenstein, 2000). Next, these progenitors produce differentiating neurons expressing a second set of transcription factors, predominately members of the LIM-homeodomain family, that regulate differentia- tion programs of neuron and axon migration and neurotrans- mitter phenotype. These mechanisms are best understood in the spinal cord, but several lines of evidence suggest that neuron specifica- tion programs are shared along the rostral– caudal axis of the CNS (Wilson and Rubenstein, 2000). In fact, the ex- pression of dorsal–ventral patterning signals (e.g., BMP4 and SHH) and homeodomain proteins that specify progen- itor domains (e.g., PAX6 and NKX2.2) are expressed in similar longitudinal domains from spinal cord up through the forebrain (Shimamura et al., 1995). Functional relation- ships appear to be conserved as well. For example, in the hindbrain the ventralizing signal Shh induces progenitors to produce motor neurons that express Islet-1, a LIM ho- meodomain transcription factor critical for motor neuron differentiation, while in ventral forebrain progenitors Shh induces the production of forebrain-specific interneurons that are also marked by Islet-1 expression (Ericson et al., * Corresponding author. University of Nevada, Reno, Biology/200, Reno, NV 89557. Fax: +1-775-784-1650. E-mail address: gmastick@unr.edu (G.S. Mastick). Molecular and Cellular Neuroscience 23 (2003) 107–120 www.elsevier.com/locate/ymcne 1044-7431/03/$ – see front matter © 2003 Elsevier Science (USA). All rights reserved. doi:10.1016/S1044-7431(03)00016-2