Role for Gli3 in the formation of the major axonal tracts in the telencephalon

In the adult brain, the thalamocortical tract conveys sensory information from the external environment to the cortex. The cortex analyzes and integrates this information and sends neural responses back to the thalamus through the corticothalamic tract. To reach their final target both thalamocortical and corticothalamic axons have to cover long distances during embryogenesis, changing direction several times and passing through different brain territories. The ventral telencephalon plays a major role in the early development of these tracts. At least three main axon guidance mechanisms act in the ventral telencephalon. First, two different populations of pioneer neurons in the lateral ganglionic eminence (LGE) (LGE pioneer neurons) and medial ganglionic eminence (MGE) (MGE pioneer neurons) provide scaffolds which allow growing corticothalamic and thalamocortical axons to cross the pallium sub pallium boundary (PSPB) and the diencephalic telencephalic boundary (DTB), respectively. Second, the ventral telencephalon forms a permissive corridor for thalamic axons by tangential migration of Isl1 and Ebf1 expressing cells from the LGE into the MGE. Finally, thalamortical and corticothalamic axons guide each other once they have met in the ventral telencephalon (“handshake hypothesis”). The Gli3 transcription factor has been shown to be essential for normal early embryonic regionalization of the mammalian forebrain, although roles of Gli3 in later aspects of forebrain development, like the formation of axonal connections, have not been investigated previously. Here, I present the analysis of axonal tract development in the forebrain of the Gli3 hypomorphic mutant mouse Polydactyly Nagoja (Pdn). These animals lack the major axonal commissures of the forebrain: the corpus callosum, the hippocampal commissure, the anterior commissure and the fimbria. In addition, DiI injections and neurofilament (NF) staining showed defects in the formation of the corticothalamic and thalamocortical tracts. Although the Pdn/Pdn cortex forms early coticofugal neurons and their axons, these axons do not penetrate the LGE and instead run along the PSPB. Later in development, although a thick bundle of Pdn/Pdn cortical axons is still observed to project along the PSPB, some Pdn/Pdn cortical axons eventually enter the ventral telencephalon navigating along several abnormal routes until they reach thalamic regions. In contrast, Pdn/Pdn thalamic axons penetrate into the ventral telencephalon at early stages of thalamic tract development. However, rostrally they deviate from their normal trajectory, leaving the internal capsule prematurely and only few of them reach the developing cortex. Caudally, an ectopic Pdn/Pdn dorsal thalamic axon tract projects ventrally in the ventral telencephalon not entering the internal capsule at all. These defects are still observed in newborn Pdn/Pdn mutant mice. Next, I investigated the developmental mechanisms causing these pathfindings defects. No obvious defects are present in Pdn/Pdn cortical laminae formation and in the patterning of the Pdn/Pdn dorsal thalamus. In addition, Pdn/Pdn thalamocortical axons are able to respond to ventral telencephalic guidance cues when transplanted into wild type brain sections. However, these axonal pathfinding defects correlate with patterning defects of the Pdn/Pdn LGE. This region is partially ventralized and displays a reduction in the number of postmitotic neurons in the mantle zone due to an elongated cell cycle length of LGE progenitor cells. Finally, Pdn/Pdn mutant display an upregulation of Shh expression and Shh signalling in the ventral telencephalon. Interestingly, these patterning defects lead to the absence of DiI back-labelled LGE pioneer neurons, which correlates with the failure of corticothalamic axons to penetrate the ventral telencephalon. In addition, ventral telencephalic thalamocortical guidance mistakes happen at the same time of abnormal formation of the corridor cells. Taken together these data reveal a novel role for Gli3 in the formation of ventral telencephalic intermediate cues important for the development of the thalamocortical and corticothalamic connections. Indeed, Pdn animals are the first known mutants with defective development of the LGE pioneer neurons, and their study provides a link between early patterning defects and axon pathfinding in the developing telencephalon