TASK-1 and TASK-3 are functional members of the tandem- pore K+ (K2P) channel family, and mRNAs for both channels are expressed together in many brain regions. Although TASK-1 and TASK-3 subunits are able to form heteromers when their complementary RNAs are injected into oocytes, whether functional heteromers are present in the native tissue is not known. Using cultured cerebellar granule (CG) neurons that express mRNAs of both TASK-1 and TASK-3, we studied the presence of heteromers by comparing the sensitivities of cloned and native K+ channels to extracellular pH (pHo) and ruthenium red. The single-channel conductance of TASK- 1, TASK-3 and a tandem construct (TASK-1/TASK-3) expressed in COS-7 cells were 14.2±0.4, 37.8 ±0.7, and 38.1±0.7 pS (-60 mV), respectively. TASK-3 and TASK-1/TASK-3 (and TASK-3/TASK- 1) displayed nearly identical single-channel kinetics. TASK-3 and TASK-1/TASK-3 expressed in COS-7 cells were inhibited by 26±4 and 38±6%, respectively, when pHo was changed from 8.3 to 7.3. In outside-out patches from CG neurons, the K+ channel with single channel properties similar to those of TASK-3 was inhibited by 38±6% by the same reduction in pHo. This suggested that native K+ channel is most similar to the TASK-1/TASK-3 heteromer. TASK-3 and TASK- 1/TASK-3 expressed in COS-7 cells were inhibited by 78 ±7 and 3±4%, respectively, when 5 µM ruthenium red was applied to outside-out patches. In outside-out patches from CG neurons containing a 38 pS channel, two types of responses to ruthenium red were observed. Ruthenium red inhibited the channel activity by 77±5% in 42% of patches, (range: 72-82%) and by 5±4% (range: 0-9%) in 58% of patches. When patches contained more than three 38 pS channels, the average response to ruthenium red was 47±6% inhibition (n=5). These electrophysiological studies show that native 38 pS K+ channels of the TASK family in cultured CG neurons consist of both homomeric TASK-3 and heteromeric TASK-1/TASK-3.