Several recent studies have reported a correlation between regional glutamate concentration and BOLD activation. In this study, we combined resting-state fMRI and MRS to investigate whether this association maintain in spontaneous brain activity in the thalamus. Significant positive correlation was found between glutamate concentration and the ALFF and ReHo in left thalamus; negative correlation between glutamate and DC in left thalamus. Furthermore, the ALFF of left primary motor cortex and bilateral auditory cortex were affected by left thalamus glutamate. This provides insight into better understanding the neuronal and biochemical mechanisms of thalamus function.