To clarify seasonal and inter-annual variations in the contribution of heterotrophic respiration to soil respiration and influences of such changes on the carbon balance in a forest ecosystem, relationships between respiration rates, soil temperature and soil water content, and the contribution ratio of heterotrophic respiration, were estimated over almost 3 years. Heterotrophic respiration was separated from soil respiration by the trenching method; CO2 emissions from the soil surface were measured along with soil temperature and volumetric soil water content in a cool-temperate deciduous forest from November 2009 to September 2012. Also, the root bag method and two decay models were used to accurately measure CO2 emissions from decomposing dead roots. The soil and heterotrophic respiration responded differently to changes in soil temperature and soil water content. An increase in soil temperature caused increases in respiration, and the response of heterotrophic respiration was lower than that of soil respiration. Also, an increase in the soil water content caused a slight increase in soil respiration and a decrease in heterotrophic respiration. These responses of the respirations to environmental factors caused a decrease in the contribution ratio of heterotrophic respiration with an increase in soil temperature and soil water content. These results suggest that the contribution ratio could experience complex changes when both factors change simultaneously in field conditions. Annual contribution ratios of heterotrophic respiration were estimated at 62% in 2010 and 2011, and the contribution ratio showed seasonal variation ranging from 60% to 100%. The estimated annual heterotrophic respiration rate in 2010 without such seasonal variation ranged from 1.50 to 2.51 kg CO2 m -2 yr -1, and these rates were 96% to 161% of that with seasonal variation (1.56 kg CO2 m -2 yr -1 ) . Also, the annual contribution ratio in 2010 estimated without the effect of soil water content (using respiration and soil temperature curve) was 80%. The resulting annual heterotrophic respiration rate was 2.01 kg CO2 m -2 yr -1, and this rate was 128% of that with the effects of soil water content (using respiration, soil temperature and soil water content curve) . In contrast, the effect of inter-annual variation in the contribution ratio on annual heterotrophic respiration rate was small. Therefore, it is important to take into account the seasonal variation in the contribution ratio and the effects of the soil water content on the contribution ratio for more accurate estimation of heterotrophic respiration and net ecosystem production.