The proton transverse-relaxation spectrum of peripheral nerve has been reported to consist of three components arising from microanatomical water compartments within the nerve. The component with the shortest decay time has been consistently seen to be a result of the myelin. In this study, experiments were performed to reveal the microanatomical sources of the two longest-lived components. Transverse-relaxation spectra were repeatedly obtained from samples of sciatic nerve of Xenopus laevis, the African clawed frog. These samples were maintained in vitro in a perfusion chamber, allowing for "on-the-fly" changes in perfusate composition. The changes in composition involved the addition of either 10-mM Mn(2+) or 30-mM Gd-DTPA to the perfusate, or the replacement of H(2)O in the perfusate by D(2)O. The D(2)O experiments revealed that >98% of the signal attributed to the nerve resulted from exchangeable water. Following the introduction of a paramagnetic agent into the perfusate, it was found that the intermediate-lived component of the nerve T(2) spectra disappeared within 10-50 min. However, the shortest- and longest-lived components remained, experiencing much more gradual long-term changes. It was thus concluded that the intermediate-lived component corresponds to the interaxonal compartment of nerve tissue based upon its closer proximity to the outside environment of the nerve.