Background: The difference in spontaneous brain activity between acute subjective tinnitus patients (with or without hearing loss) and control participants was explored using the amplitude of low-frequency fluctuations and degree centrality methods through resting-state functional magnetic resonance imaging. The study aimed to provide an objective basis for clinical diagnosis and pathogenesis of patients with acute subjective tinnitus. Methods: Fourteen acute subjective tinnitus (AST) patients with hearing loss (AST-HL), 6 AST patients with no hearing loss (AST-NHL), and 14 healthy controls (HCs) with age, sex, and education status matched were recruited for this study. Resting-state functional magnetic resonance imaging (fMRI) examinations were performed in a resting state and the amplitude of low-frequency fluctuations (ALFF) and degree centrality (DC) values of each group were acquired. Statistical analysis was performed to assess the ALFF and DC values of different brain areas of the participants (AST-HL and AST-NHL were compared with HCs, but AST-HL and AST-NHL were not). Results: Patients with acute subjective tinnitus and hearing loss showed a significantly increased amplitude of low-frequency fluctuation values in the left middle temporal gyrus and bilateral frontal gyrus/marginal lobe/cingulate gyrus but a decreased amplitude of low-frequency fluctuations values in the bilateral superior temporal gyrus/anterior cerebellar lobe in comparison with healthy controls. The amplitude of low-frequency fluctuation values of patients with acute subjective tinnitus and hearing loss in the right posterior lobe of the cerebellum, bilateral temporal gyrus, bilateral lenticular nucleus, bilateral frontal gyrus, right inferior occipital gyrus, were higher, but were significantly lower in the bilateral anterior lobe of cerebellum/superior temporal gyrus and left posterior cerebellar lobe as compared with those of healthy controls. Degree centrality values in the healthy controls group were increased in the right superior marginal gyrus and decreased in the right thalamus in patients with acute subjective tinnitus and hearing loss, while patients with acute subjective tinnitus with no hearing loss presented significantly higher degree centrality values in the left frontal lobe and lower degree centrality values in the left center rear return. Conclusions: Aberrant amplitude of low-frequency fluctuations and values exist in various brain regions, indicating abnormal spontaneous brain activity in both acute subjective tinnitus and hearing loss and acute subjective tinnitus no hearing loss patients. The pathogenesis of acute subjective tinnitus may be related to abnormalities in both the auditory cortex and nonauditory cortex. These findings provide more evidence to help clarify the neuronal symptoms of acute subjective tinnitus patients.