Abstract
Chronic monitoring of neuronal activity in the living brain with optical imaging techniques became feasible owing to the continued development of genetically encoded calcium indicators (GECIs). Here we report for the first time the successful generation of transgenic marmosets (Callithrix jacchus), an important nonhuman primate model in neurophysiological research, which were engineered to express the green fluorescent protein (GFP)-based family of GECIs, GCaMP, under control of either the CMV or the hSyn promoter. High titer lentiviral vectors were produced, and injected into embryos collected from donor females. The infected embryos were then transferred to recipient females. Eight transgenic animals were born and shown to have stable and functional GCaMP expression in several different tissues. Germline transmission of the transgene was confirmed in embryos generated from two of the founder transgenic marmosets that reached sexual maturity. These embryos were implanted into six recipient females, three of which became pregnant and are in advanced stages of gestation. We believe these transgenic marmosets will be invaluable non-human primate models in neuroscience, allowing chronic in vivo monitoring of neural activity with functional confocal and multi-photon optical microscopy imaging of intracellular calcium dynamics.
Highlights
Primates are ideal animal models for studying the complex human brain[17,18]
Marmoset embryos were collected from donor females either via nonsurgical uterine flushing of naturally fertilized (NAT) embryos, or alternatively via laparotomic follicular aspiration of unfertilized oocytes followed by in vitro maturation and fertilization (IVF)[20,23]
Number of procedures Procedures with successful collection (%) Total number of embryos collected (Mean number of embryos per procedure) Number of germinal vesicle-stage (GV) oocytes (Mean number of oocytes per procedure) Number of matured (MII) oocytes (% maturation rate) Number of fertilized oocytes (% fertilization rate) Number of embryos subjected to lentiviral injections Number of developed embryos(% cleavage rate) GCaMP expression confirmed after lentiviral injections (% expression rate) Number of embryos used for ET Number of surrogates Number of pregnancies (% pregnancy rate) Number of deliveries (% delivery rate) Number of TGs (% TG per ET)
Summary
Primates are ideal animal models for studying the complex human brain[17,18]. the recent feasibility of genome modification in nonhuman primates has significantly raised their role and importance in biomedical research[19]. The common marmoset (Callithrix jacchus) is a New World monkey that is a clear example of a nonhuman primate with ever growing interest as a model for neuroscience research, in great part due to the successful generation of transgenic marmosets with germline transmission of the transgene[20]. Germline transmission of the transgene was confirmed in oocytes collected from two of the founder transgenic marmosets that reached sexual maturity. These transgenic marmosets hold the promise to be a powerful tool for imaging neural activity in vivo that constitutes a closer and more accurate model in translational studies of aimed at understanding human brain function
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