We used optically detected magnetic resonance (ODMR) technique to directly probe electron-spin resonance transitions in the excited state of negatively-charged boron vacancy (VB-) defects in hexagonal boron nitride (hBN) at room temperature. The data showed that the excited state has a zero-field splitting of ~ 2.1 GHz, a g factor similar to the ground state and two types of hyperfine splitting ~ 90 MHz and ~ 18.8 MHz respectively. Pulsed ODMR experiments were conducted to further verify observed resonant peaks corresponding to spin transitions in the excited state. In addition, negative peaks in photoluminescence and ODMR contrast as a function of magnetic field magnitude and angle at level anti-crossing were observed and explained by coherent spin precession and anisotropic relaxation. This work provided significant insights for studying the structure of VB- excited states, which might be used for quantum information processing and nanoscale quantum sensing.