The subthalamic nucleus (STN) is one of the best targets for therapeutic deep brain stimulation (DBS) to control motor symptoms in Parkinson's disease. However, the precise circuitry underlying the effects of STN-DBS remains unclear. To understand how electrical stimulation affects STN projection neurons, we used a retrograde viral vector (AAV-retro-hSyn-eGFP) to label STN neurons projecting to the substantia nigra pars reticulata (SNr) (STN-SNr neurons) or the globus pallidus interna (GPi) (STN-GPi neurons) in mice, and performed whole-cell patch-clamp recordings from these projection neurons in ex vivo brain slices. We found that STN-SNr neurons exhibited stronger responses to depolarizing stimulation than STN-GPi neurons. In most STN-SNr and STN-GPi neurons, inhibitory synaptic inputs predominated over excitatory inputs and electrical stimulation at 20-130 Hz inhibited these neurons in the short term; its longer-term effects varied. 6-OHDA lesion of the nigrostriatal dopaminergic pathway significantly reduced inhibitory synaptic inputs in STN-GPi neurons, but did not change synaptic inputs in STN-SNr neurons; it enhanced short-term electrical-stimulation-induced inhibition in STN-SNr neurons but reversed the effect of short-term electrical stimulation on the firing rate in STN-GPi neurons from inhibitory to excitatory; in both STN-SNr and STN-GPi neurons, it increased the inhibition but attenuated the enhancement of firing rate induced by long-term electrical stimulation. Our results suggest that STN-SNr and STN-GPi neurons differ in their synaptic inputs, their responses to electrical stimulation, and their modification under parkinsonian conditions; STN-GPi neurons may play important roles in both the pathophysiology and therapeutic treatment of Parkinson's disease.