DEVELOPMENT OF MIDBRAIN AND ANTERIOR HINDBRAIN OCULAR MOTONEURONS IN NORMAL AND WNT-1 KNOCKOUT MICE

The effect of homozygotic Wnt-1(-/-) mutations on the development of o cular motoneurons was examined with the lipophilic dye DiI and compare d to control and phenotypic wild-type mouse embryos. A piece of DiI-so aked filter paper was inserted into the orbit, the midbrain, or rhombo mere 5 of the hindbrain in six paraformaldehyde-fixed litters (10.5, 1 2.5, and 14.5 days postcoitum) containing Wnt-1, Wnt(+/-), and Wnt-1(/+) individuals and three control litters. We labeled all ocular moton eurons retrogradely and all relevant nerves anterogradely in all contr ol and phenotypic wild-type animals. In all phenotypically identified Wnt-1(-/-) mutants we could always label the abducens nerve and motone urons and the optic fibers to the thalamus, but we were unable to labe l oculomotor or trochlear nerves or motoneurons. In addition to Wnt-1 knockout mutants, we also labeled mice from the WZT9B transgenic line carrying a lacZ reporter gene driven by the Wnt-1 gene enhancer. In th ese embryos we tested for co-localization of Wnt-1 expression in bioti nylated dextran amine-labeled ocular motoneurons using a newly develop ed technique. In younger embryos we obtained evidence for co-localizat ion of the beta-galactosidase reaction product derived from lacZ gene activity in some retrogradely filled oculomotor motoneurons and adjace nt to other oculomotor and the trochlear motoneurons. Acetylcholine es terase, a marker of early differentiating cholinergic neurons, showed a similar topology with respect to the lacZ reaction product. Thus, at least some future oculomotor motoneurons express Wnt-1, whereas other s and the trochlear motoneurons caudal to the ventral midbrain express ion of Wnt-1 may be exposed to the short range diffusion of the Wnt-1 gene product. Thus, the Wnt-1(-/-) mutation precludes formation or sur vival of midbrain and anterior hindbrain neurons, including oculomotor and trochlear motoneurons. (C) 1995 John Wiley and Sons, Inc.