The kinetics of the CK reaction in the living rabbit brain was studied by three 31P magnetization transfer methods: inversion recovery (IR), inversion transfer (IT), and steady-state saturation transfer (SSST). Longitudinal relaxation rates (tau -1) were determined from IR and IT experiments. The values were 0.88 +/- 0.08 s-1 for PCr and 2.45 +/- 0.39 s-1 for gamma-ATP. Analysis of the results of SSST experiments in which gamma-ATP was saturated yielded a forward rate constant kF of 0.53 +/- 0.07 s-1. Upon saturation of PCr no change in the gamma-ATP signal could be detected in SSST. An average reverse rate constant (kR +/- S.D.) of 0.52 +/- 0.04 s-1 was estimated by analyzing IT data from three experiments with selective inversion of gamma-ATP. The standard error of kR was 50%. The average [PCr]/[ATP] of 1.21 +/- 0.16 together with the values of kF and kR yielded a forward-to-reverse flux ratio of 1.23. Within our limits of accuracy this ratio was not significantly different from 1.0, thus suggesting that in the brain the forward and reverse CK fluxes are equal.