Rhythmicity plays an important role in a number of biological systems. The habenular complex is reported to contain an intrinsic molecular clock and to show rhythmic expression of circadian clock genes and proteins including per2/PER2. In this study, we observed that there is a temporal rhythmicity in the presynaptic efficacy of the lateral habenula (LHb) neurons. We collected a substantial number of recordings at different time points of the day during the light phase. The frequency and amplitude of spontaneous excitatory transmission were increased in the afternoon compared to recordings performed in the morning. In addition, the paired-pulse ratio and the success rate of minimal stimulation were also significantly different depending on the time of the recording. We did not see any significant differences in recordings obtained from pyramidal neurons of the hippocampus in the same brain slices. Taken together, our data indicates that the LHb exhibits intrinsic temporal oscillation in basal neurotransmission and in presynaptic release probability. Given the rapidly growing interest on the function of the LHb, more careful examination of synaptic transmission in the LHb is thus required.