ABSTRACT: The lack of axonal growth after injury in the adult central nervous system (CNS) is partly due to the presence of growth-inhibitory molecules associated with myelin and the intrinsic growth-state of the injured neurons. To date, three inhibitors have been identified in myelin: Myelin- Associated Glycoprotein (MAG), Nogo A, and Oligodendrocyte-Myelin glycoprotein (OMgp). These three proteins all appear to be located in the periaxonal surface of the myelin membrane placing them in an optimal location to mediate axon-glial interaction. In addition, the three proteins have been shown to bind the same neuronal receptor, known as the Nogo-66 receptor (NgR). It has been hypothesized that inhibition of NgR may be a strategy to increase regeneration, plasticity and functional recovery of the lesioned central nervous system. Strong NgR mRNA expression is observed in the hippocampal pyramidal cell layers (CA1-3) and the granular layer of the dentate gyrus. It has been shown that animals exposed to entorhinal lesions show a biphasic regulation of NgR in the hippocampus, suggesting a tightly regulated mechanism mediated by this receptor. We have access to a transgenic model to over-express NgR in forebrain hippocampal neurons. Preliminary results have shown a phenotypic response in behavior and some molecular markers, as result of NgR overexpression. Knowledge of what genes are reacting in this novel transgenic model may provide insights into what pathways are affected by NgR to control synaptic plasticity in normal animals and during injury. We have access to a bitrasgenic conditional mouse model to everexpress the NgR receptor in forebrain neurons, through the CamKII promoter. This conditional model uses the tetracycline transactivator (tTA), which is expressed through the CamKII promoter. Upon removal of doxycycline, the TetOp target gene becomes active and over-expression of NgR in cells with an active CaMKII promoter is achieved. We will determine gene expression patterns in the dentate gyrus of transgenic animals engineered to over-express NgR through the CamKII promoter (present in several forebrain areas), and upon removal of doxyxycline. We hypothesize that over-expression of NgR in the hippocampus inhibits synaptic plasticity, thereby suppressing the formation of new synapses necessary for the formation of memory. EXPERIMENT 1 - Effects of NgR overexpression in the mature forebrain - 1) Experimental group: Bitrasgenic + water: Overexpresses NgR in forebrain neurons. In this group, doxyxycline was removed from the drinking water in adult animals. All the control groups were aged matched. 2) Control group: Bitransgenic + doxy: Should control for leakage by the TetOp target gene, or for any effects mediated by doxycycline, as compared to other control groups. 3) Control group: Monotransgenic + water: Should control for any leakage mediated by the TetOp target gene, as compared to other control groups. 4) Control group: Monotrangenic + water: Should control for leakage by the TetOp target gene, or for any effects mediated by doxycycline, as compared to other control groups. EXPERIMENT 2 - Effects of NgR overexpression in the developing forebrain - We will also analyze the effects of NgR overexpression in bitransgenic animals raised on water. In these animals, the NgR transgene is expressed in the forebrain upon activation of the CamKII promoter. Since neuronal differentiation and neuronal pathways are being formed during development, we anticipate that NgR overexpression (de)effects in these animals will be markedly different to those observed in adult animals. 1) Bitransgenic + water (through entire lifespan): The animals in this group were never treated with doxycycline, and therefore expressed the NgR transgene upon activity of the CamKII promoter. 2) Monotransgenic + water: This control group will control for leakage by the Tet Op gene, or for effects of the tTA expression. Keywords: NgR, forebrain, development
