TREK-1, is a member of the two-pore-domain potassium channel family that is regulated by polyunsaturated fatty acids, mechanical stretch of the membrane, intracellular acidification and other physical and chemical stimuli. We describe here three novel splice variants of rat TREK-1 differing in exon 1 at their extreme N-terminus or in the number of exons. TREK-1c and TREK-1d proteins possess four transmembrane domains whereas in TREK-1e the last two transmembrane domains are lacking because exon 5 is skipped. Expression analysis in seven different tissues revealed that rat TREK-1e is mainly transcribed in the brain and in the kidney. The splice variants TREK-1a-d, but not TREK-1e, could be functionally expressed in Xenopus oocytes. Co-expression of TREK-1e with TREK-1a-d in Xenopus oocytes resulted in reduced current amplitudes for TREK-1a-d. Using a yeast-two hybrid assay we found that the C-terminus of TREK-1e interacted with the N-termini of TREK-1a-d. Surface expression of HA-tagged TREK-1a channels was reduced by co-expression of TREK-1e in Xenopus oocytes. Furthermore, fusion of the cytoplasmic C-terminus of TREK-1e to the reporter channel protein Kir2.1 caused intracellular retention of the reporter protein. We conclude that TREK-1e is a negative regulator of the splice variants TREK-1a-d.