Examining the function of serotonin receptor Htr4 on maturation of adult born dentate granule cells by single cell RNA interference technique
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New neurons are continuously generated in the dentate gyrus of the hippocampus throughout adulthood in a broad range of mammalian species. The birth of thousands of hippocampal neurons suggests that these new cells are important in the functions of the hippocampus. Newborn neurons are shown to become integrated into existing neural circuits and mature into functional neurons in the adult mammalian brain. During maturation these new neurons extend their axons, the mossy fibers, towards the CA3 region of the hippocampus, and their dendrites towards the molecular layer of the dentate gyrus. Newborn neurons are thought to make an important contribution to the function of learning and memory. Serotonin receptors are found to be expressed in the hippocampus and the dentate gyrus and have been shown to be involved in neurogenesis, regulating adult granule cell proliferation. In this thesis I aim to examine the function of the Htr4 on maturation and morphology of adult born dentate granule cells. The serotonin receptor Htr4 is found to be abundantly expressed in dentate granule cells and it is hypothesized that stimulation of the Htr4 may stimulate neurogenesis in the hippocampus. In neurogenesis in the enteric nervous system, the Htr4 is suggested to be required for growth, maintenance, development and survival, while other findings link the Htr4 to dematuration of dentate granule cells. The screening will be done in the dentate gyrus of mice with respect to two aspects of the complexity of the dendritic tree; the dendritic length and the number of branching points. In order to achieve this, I have used a fluorescence-based cell culture assay to identify two shRNA sequences that successfully down-regulate Htr4 by an average of 89%. These two sequences were then used to produce retroviral vectors which were injected in mice hippocampus, in order to down-regulate Htr4 in adult born granule cells, and examine the effect on their maturation. I found no significant difference, but a trend indicating that the Htr4 plays an inhibitory role in the maturation of adult born granule cells with respect to the dendritic length and the number of branching points. Power of the statistical analysis for these results were low, indicating that there might be a difference which were not detected due to low sample size.