Purpose: To identify the changes of intrinsic brain activity and regional cerebral blood flow in patients with chronic neck and shoulder pain (CNSP) by using amplitude of low-frequency fluctuation (ALFF) analysis and arterial spin labelling study. Material and methods: In total, 28 CNSP patients and 25 age-matched and sex-matched healthy controls (HCs) participated in the study. Resting-state functional magnetic resonance imaging (rs-fMRI) and arterial spin labelling (ASL) MRI were acquired. Correlations between ALFF and cerebral blood flow (CBF) were analysed. Subsequently, the differences in ALFF and CBF were compared in the two groups. Finally, the visual analogue scale (VAS) was also assessed in the CNSP group. Results: Compared with HCs, CNSP patients showed significantly abnormal ALFF and CBF in several brain regions, including the cerebellum posterior lobe, middle orbitofrontal gyrus, medial superior frontal gyrus, middle temporal gyrus, precuneus, cingulate gyrus, middle occipital gyrus, middle frontal gyrus, postcentral gyrus, precentral gyrus, and superior parietal gyrus. Correlation analysis showed that the ALFF value of the medial superior frontal gyrus positively correlated with the VAS score. However, no correlation was found between the CBF values and the VAS score. Conclusions: The altered ALFF and CBF values in CNSP patients were observed in different pain-related brain regions that were involved in pain modulation and perception. The combination of rs-fMRI and ASL MRI might provide complementary information for increasing our understanding of the neuropathology in CNSP.