Soil heterotrophic respiration (Rh) is the primary pathway for terrestrial ecosystem CO2 efflux. Herbivore grazing activity is known to affect grassland soil Rh, but the effects of grazing-related dung deposition on Rh and the mechanisms involved remain unknown. Here, we explored effects of dung on Rh via altered biotic and abiotic factors in a short-term experiment with yak dung application during a growing season in a Tibetan alpine meadow. Dung application significantly decreased soil pH, but increased soil Rh, concentrations of dissolved organic carbon (C), nitrogen (N), and phosphorus (P), microbial biomass, and activities of enzymes related to C-, N-, and P-degradation. Linear regression showed that Rh was positively correlated with labile substrates, microbial biomass, and enzyme activities, and negatively correlated with soil pH, suggesting that these factors are all potential drivers of Rh. Further, using piecewise structural equation modeling, we revealed that Rh was driven directly by altered microbial biomass and enzyme activities, and indirectly by altered pH and labile substrates. Particularly labile substrates and microbial biomass had more important roles in driving Rh than pH and enzyme activities. Taken together, our work revealed some mechanisms behind short-term dung deposition effects on Rh, which can be used to develop grassland management practices and forecast C feedback in grazed ecosystems.