Rapid warming and changing precipitation patterns in northern regions are alleviating temperature and nutrient limitations on microbial activity in soils and jeopardizing the stability of vast amounts of ancient organic carbon (C) stored in near-surface permafrost. Like fossil fuels, permafrost C is depleted in radiocarbon (14C) and its emission to the atmosphere contributes to climate change. While tundra systems continue to sequester C during the growing season, evidence is emerging that the non-growing season (fall through spring) is a critical period of microbial activity – turning the Arctic into a net C source.In my dissertation, I developed and deployed new technology to elucidate which C sources fuel microbial activity during the non-growing season and to quantify emissions of permafrost C. First, I present a new sampler for collecting depth-resolved, time-integrated soil-respired CO2 for 14C analysis from upland permafrost soils year-round. Second, I describe the first continuous, depth-resolved time series of microbial C emissions and annual flux-weighted signature of ecosystem-respired 14CO2 from graminoid tundra derived from year-round in situ measurements of soil 14CO2. Results show that soil microorganisms shift from using modern C during the growing season to increasingly older C pools in fall to winter. Third, I couple the sampler with bulk soil analyses, incubations, and soil thaw and land-atmosphere C flux observations to show that permafrost thaw, compaction and subsidence from 25 years of experimental snow augmentation has mobilized significant amounts of ancient C and profoundly impacted nitrogen cycling. These studies illustrate greater than expected seasonal dynamics in microbial C oxidation, further highlighting the role of the non-growing season for permafrost C emissions and paving the way for a regional-scale assessment of permafrost C emissions in the rapidly changing Arctic.