Netrin 1 regulates ventral tangential migration of guidepost neurons in the lateral olfactory tract

Abstract

MiceTime-pregnant ICR wild-type mice were purchased from SLC (Shizuoka, Japan). Mice expressing green fluorescent protein (GFP; green mice) were provided by Okabe et al. (Okabe et al., 1997). Xt J /Xt J mutant embryos, which have deletions in the Gli3 gene, were generated as described previously (Tomioka et al., 2000). When the Xt J mutation was introduced into green mice, green mice were crossed with Xt J /+ mice and the resulting Xt J /+ mice expressing GFP were intercrossed to obtain Xt J /Xt J green mouse embryos. Heterozygous netrin 1 mutant mice (Serafini et al., 1996) and Dcc mutant mice (Fazeli et al., 1997) were crossed to obtain homozygous embryos. The day on which a vaginal plug was found was designated embryonic day 0.5 (E0.5). All experimental protocols were approved by the Animal Committee of National Institute of Genetics. Organotypic culture of telencephalon stripsEach telencephalic hemisphere of the E10.5 mouse embryo was dissected along the dorsoventral axis into a strip ~1.5 mm wide, including the neocortex, the presumptive LOT area and the lateral and medial ganglionic eminences (GE) (see Fig. 1A). In the strip, the presumptive LOT area was located around the middle along the dorsoventral axis. In a standard culture, the strip was labeled by injection of dextran tetramethylrhodamine In the developing nervous system, functional neural networks are constructed with intricate coordination of neuronal migrations and axonal projections. We have previously reported a ventral tangential migration of a special type of cortical neurons, lot cells, in the mouse embryo. These neurons originate from the ventricular zone of the entire neocortex, tangentially migrate in the surface layer of the neocortex into the ventral direction, align in the future pathway of the lateral olfactory tract (LOT) and eventually guide the projection of LOT axons. In this study, we developed an organotypic culture system to investigate the regulation of this cell migration in the developing telencephalon. Our data show that the neocortex contains the signals that direct lot cells ventrally, that the ganglionic eminence excludes lot cells by repelling the migration and that lot cells are attracted to netrin 1, an axon guidance factor. Furthermore, we demonstrate that mutations in the genes encoding netrin 1 and its functional receptor Dcc lead to inappropriate distribution of lot cells and subsequent partial disruption of LOT projection. These results suggest that netrin 1 regulates the migration of lot cells and LOT projections, possibly by ensuring the correct distribution of these guidepost neurons. Molecular Probes, Eugene, OR) or 1,1-dioctadecyl-3,3,3Ј3Ј-tetrametylindocarbacyanine perchlorate (DiI; Molecular Probes) into a small area of the dorsal neocortex, unfolded on a collagen-coated membrane filter (Transwell-COL inserts #3492, Corning, Acton, MA) and cultured in Dulbecco's modified Eagle's medium (DMEM)/F-12 (Sigma-Aldrich, St Louis, MO) containing 10% fetal bovine serum (Cansera, Rexdale, ONT, Canada) and 5% hor...

Robo1 and Robo2 Control the Development of the Lateral Olfactory Tract

“…First, in Slit1;Slit2 double-deficient mice, the number of lot-1 expressing cells is slightly reduced near the exit point of medial OB axons, suggesting that this subtle disruption of the distribution of lot cells might play a part in the ectopic projection of OB axons. Second, it has been shown previously that in deleted in colorectal cancer (DCC) mutant embryos, the perturbation of lot cell migration leads to defects of LOT axons pathfinding (Kawasaki et al, 2006). It is unlikely in this system that a direct effect of the DCC on the guidance of olfactory axons contributes to the misguidance, as netrin-1 does not exert any effect on OB axons in vitro (de Castro et al, 1999;Li et al, 1999).…”

Systemic human Netrin-1 gene delivery by adeno-associated virus type 8 alters leukocyte accumulation and atherogenesis in vivo

“…Its major function may be in brain embryogenesis by functioning as an axon attractant molecule involved in the generation of the normal neuronal lattice development. [23][24][25] As with Mo/MF, it is a chemorepellent for oligodendrocyte precursor cells in the embryonic spine, as well as trochlear motor axons. However, it is a chemoattractant for certain other axons.…”

“…Yet, another function of Netrin-1, as expressed in endothelial capillary tip cells, is in the formation of the developing capillary tree, somewhat similar to axon guidance, during embryogenesis. 24,26,27 It may also mildly promote the development of microvessels in the brain by direct AAV2/Netrin-1 gene injection into the brain. 28 Moreover, Netrin-1 may also protect the brain by limiting leukocyte migration and invasion.…”

Role of a transcription factor Pax6 in the developing vertebrate olfactory system

“…It has been shown that a specific neuronal population known as 'lot cells' is involved in LOT development (Sato et al 1998). During early embryonic stages, lot cells are born at the dorsal part of the telencephalon, and migrate ventrally, then populate at the presumptive LOT region, similar to the olfactory cortex neurons (Tomioka et al 2000;Kawasaki et al (Hirata et al 2002), suggesting that LOT defects in the mutant are independent of the lot cells. Interestingly, collateral formation of the mammlio-tegmental tract is also compromised in the Pax6 mutant (Valverde et al 2000), suggesting that Pax6 regulates common mechanism(s) underlying collateral axonal branching in the developing brain.…”