Background: Blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI) and diffusion tensor imaging (DTI) are useful methods for investigating the morphology and function of the kidneys, including revealing unilateral renal damage. Nevertheless, these techniques have not yet been applied for bilateral renal function. The aim of this study was to investigate whether the combination of DTI and BOLD could be used to examine different degrees of contrast-induced acute kidney injury (CI-AKI) in bilateral kidneys compared to standard methods such as serum creatinine (SCr) detection. Methods: Forty-Two New Zealand white rabbits were divided into two groups: the experimental group and the control group. Physiological saline and iodine contrast agent (iohexol, 1.0 g iodine/kg, 1.0 ml/sec) were injected via the right renal artery. DTI and BOLD-MR data were acquired longitudinally at the baseline and 1, 24, 48, and 72 h after high-pressure syringe injection to measure the apparent diffusion coefficient (ADC), fractional anisotropy (FA) and relative transverse relaxation rate (R2*). After the MR scan at each time point, three rabbits in each group were sacrificed, and changes in SCr and hypoxia-inducible factor-1α (HIF-1α) were analyzed using histopathology and immunochemistry. Results: Twenty-four hours after iohexol administration, the values of ADC and FA decreased significantly (P < 0.05), while R2* values increased (P < 0.05) in the renal cortex (CO), outer medulla (OM) and inner medulla (IM). Besides, significant negative correlations were observed among ADC, FA, and R2* in CO, OM, and IM (all P < 0.001, r = − 0.654-0.828). Conclusions: DTI and BOLD can simultaneously and non-invasively assess different degrees of CI-AKI in bilateral kidneys.