Pedunculopontine tegmentum (PPT) has GABA-ergic neurons and receives GABA-ergic projections from substantia nigra pars reticulata (SNrpr). Based on the recent studies from our and other laboratories, it was hypothesized that GABA in PPT promotes rapid eye movement (REM) sleep. In order to further study the role of GABA in PPT in REM sleep regulation, we microinjected GABA-A agonist, muscimol (200 nL, 3.5 mM), into the PPT. Muscimol in PPT significantly enhanced the amount of REM sleep by increasing the mean number of REM sleep bouts. Besides the local interneurons, GABA-ergic afferents from SNrpr are another source of GABA in PPT. In order to understand the contribution of GABA-ergic inputs from SNrpr into PPT for REM sleep regulation, SNrpr was electrically stimulated either alone or simultaneously along with the infusion of GABA-A antagonist, picrotoxin (200 nL, 0.86 mM), into the PPT. The experiment was designed with the premise that stimulation of SNrpr should increase GABA levels in PPT which should increase REM sleep comparable to that after muscimol microinjection in PPT. Further, the effect of stimulation of SNrpr on REM sleep should be antagonized by simultaneous infusion of picrotoxin into PPT. The electrical stimulation of SNrpr did not produce any significant change in sleep-wake states although it was sufficient to counter the effect of picrotoxin injection into the PPT. To overcome the limitations and confounds of electrical stimulation, SNrpr was pharmacologically stimulated by glutamate microinjection (200 nL, 5.34 mM). Infusion of glutamate into SNrpr enhanced REM sleep by increasing the mean number of REM sleep bouts, which was similar and comparable to the effect of muscimol injection into the PPT. The results confirm that GABA in PPT either from local neurons or from SNrpr promotes REM sleep.