The human embryonic kidney cells (HEK-293) have been widely used as one mammalian expression system in the study of voltage-gated K⁺ (K_V) channels. Understanding the endogenous K_V channels in these cells is the prerequisite for the characterization of the heterogeneously expressed K_V channels in these cells. In the present study we screened the transcriptional expression of different K_V genes in HEK-293 cells using reverse transcribed DNA polymerase chain reaction (RT-PCR) method. Among 16 K_V genes examined in native HEK-293 cells 10 K_V genes were reproducibly amplified, including those Kv a subunits encoding for the delayed rectifier (I_K) [K_V1.1, K_V1.2, K_V1.3, K_V1.6, and K_V3.1], and for the transient outward K_V channels (I_A) [K_V1.4, K_V3.3, K_V3.4, and K_V4.1] as well as a K_Vb2. subunit. The whole-cell outward rectifier I_K currents in the native HEK-293 cells were recorded (203 ± 13 pA at +30 mV, n = 82) with the patch-clamp technique. In about 42% of the examined cells, I_A coexited with I_K currents. I_K currents were inhibited by tetraethylammonium chloride (TEA) at 1 and 10 mM by 39.5 and 48.4%, respectively. A 39.6% inhibition of I_K currents was also observed in the presence of 4-aminopyridine (4-AP, 5 mM). Interestingly, both TEA and 4-AP also inhibited I_A currents. 4-acelamido-4′-isothiocyanalostilbene-2, 2′-disulfonic acid (1 mM), a Cl^– channel blocker, had no effect on the endogenous outward currents. We concluded that multiple endogenous K_V genes were expressed in native HEK-293 cells, which possessed significant endogenous I_K and I_A currents with unique pharmacological properties.