Examination of the number of spinal motor neurons by using selective elimination of motor neurons

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To clear how the number of spinal motor neurons is controlled during development (neurogenesis and elimination by cell death), selective elimination of motor neurons has been experimentally developed in this study. Bax, a pro-apoptotic member of the Bcl-2 family, was examined as executioner for motor neurons. To introduce Bax specifically into motor neurons during the period of early PCD, we have used in ovo electroporation coupled with the tet regulatory system. The reverse tetracycline-controlled transactivator was expressed under the control of the motor neuron specific HB9 promoter to target gene expression to spinal motor neurons. When gain-of-function Bax mutants were introduced into chick cervical motor neurons by this method, the number of dying motor neurons was significantly increased compared with those of control chick embryos. Naive Bax did not affect motor neuron survival by this method but significantly induced a large amount of cell death as soon as expressed in the spinal cord using the constitutive promoter such as the CMV promoter for their expression. Taken together, these strategies examined in this study will allow spatially and temporally controlled elimination of spinal motor neurons and will give specific insight into how the number of spinal motor neurons is regulated during development.