TALK-1a, originally isolated from human pancreas, is a member of the tandem-pore K+ channel family. We identified and characterized three novel splice variants of TALK-1 from human pancreas. The cDNAs of TALK-1b, TALK-1c, and TALK-1d encode putative proteins of 294, 322, and 262 amino acids, respectively. TALK-1a and TALK-1b possessed all four transmembrane segments, whereas TALK-1c and TALK-1d lacked the fourth transmembrane domain because of deletion of exon 5. Northern blot analysis showed that among the 15 tissues examined, TALK-1 was expressed mainly in the pancreas. TALK-1a and TALK-1b, but not TALK-1c and TALK-1d, could be functionally expressed in COS-7 cells. Like TALK-1a, TALK-1b was a K+-selective channel that was active at rest. Single-channel openings of TALK-1a and TALK-1b were extremely brief such that the mean open time was <0.2 ms. In symmetrical 150 mM KCl, the apparent single-channel conductances of TALK-1a and TALK-1b were 23 +/- 3 and 21 +/- 2 pS at -60 mV and 11 +/- 2 and 10 +/- 2 pS at +60 mV, respectively. TALK-1b whole cell current was inhibited 31% by 1 mM Ba2+ and 71% by 1 mM quinidine but was not affected by 1 mM tetraethylammonium, 1 mM Cs+, and 100 microM 4-aminopyridine. Similar to TALK-1a, TALK-1b was sensitive to changes in external pH. Acid conditions inhibited and alkaline conditions activated TALK-1a and TALK-1b, with a K1/2 at pH 7.16 and 7.21, respectively. These results indicate that at least two functional TALK-1 variants are present and may serve as background K+ currents in certain cells of the human pancreas.